The conjugate of the drug substance / ligand, methods for creating this conjugate, and pharmaceuticals containing this conjugate.

Abstract

The invention relates to conjugates of biologically active compounds and methods for their preparation. The invention relates to drug-ligand conjugates, as represented by formula XV, and methods for their preparation. These drug-ligand conjugates are useful in the prevention and / or treatment of diseases involving abnormal cellular activity, including but not limited to the prevention and / or treatment of tumor diseases. The invention also relates to an antibody or its antigen-binding fragment that binds to B7H3, the isolated nucleic acid molecule encoding the antibody or its antigen-binding fragment, the vector and host cell containing the isolated nucleic acid molecule, and the method for generating the antibody or its antigen-binding fragment. The invention also relates to a pharmaceutical containing them.

Description

Bioactive substance conjugate and its preparation method and use

[0001] This application claims priority to Chinese patent application No. 2021101781361, filed on February 9, 2021; claims priority to Chinese patent application No. 2021103408065, filed on March 30, 2021; claims priority to Chinese patent application No. 202110825932X, filed on July 21, 2021; claims priority to Chinese patent application No. 2021108259226, filed on July 21, 2021, and a sequence listing (txt) was filed concurrently with Chinese patent application No. 2021108259226 on July 21, 2021; and claims priority to Chinese patent application No. 2021108259067, filed on July 21, 2021, and a sequence listing (txt) was filed concurrently with Chinese patent application No. 2021108259067 on July 21, 2021. This application incorporates the entire text of the aforementioned Chinese patent applications. The sequence listing contained in this application is part of the specification and is herein incorporated by reference in its entirety. Technical Field

[0002] The present disclosure belongs to the field of medical technology and relates to bioactive substance conjugates (ligand-drug conjugates), compounds, drug-linker conjugates and preparation methods thereof, as well as their use in preventing and / or treating diseases associated with abnormal cell activity, including but not limited to preventing and / or treating tumor diseases. Background Art

[0003] Cytotoxic chemotherapy was once the standard treatment for cancer, but highly potent cytotoxic molecules can kill normal cells and cause severe side effects. Targeted anti-tumor drugs, due to their combined targeting and anti-tumor activity, have become a hot topic in oncology research. However, these drugs often produce significant side effects due to their target selectivity, limiting their therapeutic efficacy. Biological macromolecule drugs, such as antibodies or antibody fragments, while highly targeted, have limited or no therapeutic effect against solid tumors. ADCs, conjugates of antibodies and small molecule drugs, combine the targeting properties of antibodies with the activity of bioactive molecules, creating a biological missile with highly promising efficacy and safety advantages. Antibodies guide ADCs to target cells, where they are then internalized. The small molecule drug is then released through enzymatic degradation by specific enzymes to treat the disease.

[0004] ADC drugs have developed very rapidly in recent years, and there are 14 ADCs on the market: (1) Mylotarg (Gemtuzumab Ozogamicin, Gemtuzumab (CD33)-Cazitamycin), CD33-positive acute myeloid leukemia (AML), approved by the FDA in 2000, withdrawn from the market in 2010, and re-launched in 2017; (2) Adcetris (Brentuximab Vedotin, CD30 monoclonal antibody-MMAE), classical Hodgkin lymphoma and anaplastic large cell lymphoma, approved by the FDA in 2011; (3) Kadcyla (Trastuzumab Emtansine, Trastuzumab (Her2)-Maytansine alkaloid TM1), HER2-positive breast cancer, approved by the FDA in 2013; (4) Besponsa (Inotuzumab Vedotin, CD30 monoclonal antibody-MMAE), HER2-positive breast cancer, approved by the FDA in 2013; (5) (5) Lumoxiti (Moxetumomab pasudotox-tdfk, CD22-pseudomonas exotoxin), relapsed or refractory hairy cell leukemia (HCL), launched in 2018; (6) Polivy (Polatuzumab vedotin-piiq, CD79b-MMAE), relapsed or refractory diffuse large B-cell lymphoma (R / RDLBCL), launched in 2019; (7) Padcev (Enfortumab vedotin-ejfv, Nectin4-MMAE), locally advanced or metastatic urothelial carcinoma (UC), launched in 2019; (8) Enhertu (Famtrastuzumab vedotin-piiq, CD79b-MMAE), locally advanced or metastatic urothelial carcinoma (UC), launched in 2019; (1) deruxtecan-nxki (Her2-topoisomerase I inhibitor), breast cancer, launched in 2019; (2) Trodelvy (Trop2-topoisomerase I inhibitor ADC), triple-negative breast cancer, approved by the FDA in April 2020; (3) BCMA ADC Belantamab mafodotin (GSK2857916); (4) CD19 ADC Zynlonta (loncastuximab tesirine); (5) EGFR ADC Akalux; (6) Her2 ADC vedicizumab RC48; (7) Tissue Factor ADC Tivadak (tisotumab vedotin).

[0005] ADC drugs are composed of three parts: antibodies, bioactive molecules (drug molecules) and linkers. The bioactive molecules are covalently coupled to the antibodies through the linker. In addition to targeting and bioactivity, the coupling method of the bioactive molecules and antibodies is also one of the core features of ADC drugs. It determines the uniformity and stability of the drug and greatly affects the pharmacokinetic properties and toxicity of the ADC. Improving the stability of the linker between the bioactive molecules and the antibody will increase the stability of the ADC systemic circulation and reduce the shedding of the drug in non-target tissues, thereby relatively increasing the amount of drugs brought into the target cells and around the target cells, increasing the release of the bioactive molecules in and around the target cells, and further enhancing efficacy and reducing toxicity. Therefore, increasing the stability of the connection between the bioactive molecules and the antibody is one of the extremely important technical fields in ADC drug research.

[0006] There are two main ways to couple the 10 ADC antibodies and linkers currently on the market: (1) Lysine is the most common linking site in antibodies, and its ε-amino group can react with the activated carboxyl group of the linker to form an amide bond. Currently, there is a technology that can achieve site-specific coupling, that is, using an activating group to activate the carboxyl group of the linker, and then forming an amide bond with the specific lysine ε-amino group in the antibody to complete the coupling. On the one hand, the site-specific quantitative coupling achieved through this type of amide bond does not have broad practicality and can only be used in certain specific situations; on the other hand, it is easy to hydrolyze under the action of enzymes in the body, causing the bioactive molecules and antibodies to fall off before reaching the target cells, increasing toxicity. (2) The sulfhydryl group (SH) of the cysteine ​​of the antibody exists in the form of a disulfide bond. Opening the disulfide bond in the antibody can provide multiple free sulfhydryl groups as coupling sites. One method of coupling with the antibody sulfhydryl group is to react the free sulfhydryl group on the antibody with maleimide through Michael addition reaction, or a specific substrate can react with the free sulfhydryl group on the antibody through two Michael addition reactions to form a unique structural sulfur bridge bond. However, many literature reports show that 30% or more of the ADCs obtained by the thiol Michael addition method will undergo reverse Michael addition in the systemic circulation, resulting in premature toxin shedding and toxic reactions.

[0007] Antibody-drug conjugates can be divided into four generations based on their development history:

[0008] The first generation of antibody-drug conjugates are based on mouse antibodies, which can produce complex immunogenicity and thus have strong toxic side effects;

[0009] Second-generation antibody-drug conjugates (ADCs) utilize humanized or fully human antibodies based on the first generation, resulting in significantly improved drug properties. The first and second generation ADCs are relatively random in the connection between the antibody and the drug, resulting in a multi-component mixture. These drugs are not only highly toxic but also difficult to control in terms of quality.

[0010] The third generation of antibody-drug conjugates uses humanized or fully human antibodies to simultaneously achieve site-specific and quantitative conjugation of toxins, linkers, and antibodies, thereby generating uniform single-molecule antibody-drug conjugates. The toxicity and quality control of this type of antibody-drug conjugate are significantly improved compared to the second generation.

[0011] The fourth generation of antibody-drug conjugates is based on the third generation and has innovated the toxin-linker. The fourth generation of antibody-drug conjugates changed the previous antibody-drug conjugate strategy based on highly toxic toxins, and selected relatively low-toxic camptothecin molecules as bioactive ingredients. At the same time, it adopted a high toxin-antibody ratio (DAR), such as a DAR value of 8. This type of ADC can also produce a good therapeutic effect on tumors with low expression of tumor surface antigens. Although the fourth generation of antibody-drug conjugates started late, due to its significant efficacy, it emerged as a new force and quickly gained clinical recognition. Its representative drugs Enhertu and Trodelvy have been approved for marketing by the FDA. However, the fourth generation of ADCs still have problems with poor stability or low solubility, and existing technologies lack universality. Further development of new toxin-linkers is still needed to solve these problems.

[0012] Existing antibody-drug conjugates (ADCs) typically bind to tumor cell surface antigens through the binding of the antibody in the ADC. The ADC then undergoes endocytosis and enters the endosome, where it is converted to the lysosome. Lysosomal hydrolases then dissociate the bioactive molecule (toxin or payload) from the ADC. The dissociated bioactive molecule then enters the cytoplasm and kills the tumor cell. The bioactive molecule that escapes from the killed tumor cell can further kill surrounding tumor cells that do not express or express low levels of the antigen (the so-called bystander effect). Alterations in any of these steps can lead to drug resistance and loss of therapeutic efficacy. These include changes in antigen expression, decreased or even loss of endocytosis, or altered endosome or lysosomal function. Therefore, it is crucial to develop an ADC that is stable in the bloodstream and capable of cleaving the toxin in the tumor microenvironment without endocytosis, thereby eliminating the toxin and achieving tumor killing. Such an ADC would overcome the various resistance mechanisms associated with traditional ADCs and would be of great clinical significance.

[0013] Although there is evidence that Immunomedics and Gilead's Trodelvy can be cleaved outside tumor cells without the need for endocytosis, Trodelvy is very unstable in plasma and aqueous solutions, with a plasma half-life of less than 24 hours. Therefore, this type of ADC can only be used for ADCs composed of toxins with low biological activity to avoid non-targeted toxic side effects caused by premature shedding of ADC toxins.

[0014] In addition, the function of the linker of traditional ADC is to complete enzymatic cleavage in cells or use it to regulate water solubility. How to make the linker meet the requirements of enzymatic cleavage while using the linker to achieve ADC enrichment in tumor tissue is also of great significance for reducing the toxic side effects of ADC in normal tissues.

[0015] B7H3 is a type I transmembrane protein of the B7 family, with two isoforms in humans: 2Ig-B7H3 and 4Ig-B7H3. B7H3 is widely expressed in normal tissues, constitutively expressed only in non-immune resting fibroblasts, endothelial cells, osteoblasts, and amniotic fluid stem cells, and inducibly expressed on activated T cells, NK cells, DCs, and macrophages, with no positive expression 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 about three times higher than those of healthy controls (60.79±19.45 vs. 20.52±8.46 ng / mL), but its correlation with the disease progression remains 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).

[0016] Currently, there are therapeutic strategies targeting B7H3 in preclinical studies. For example, antibodies targeting mouse B7H3 enhance CD8-positive T cells infiltrating tumors and inhibit tumor growth (Mod Pathol. 2010 Aug; 23(8): 1104-12.). In addition, WO2008 / 066691 shows that antibodies that recognize the B7H3 variant B7H3a have anti-tumor effects on adenocarcinoma in vivo. In clinical studies, a combination drug of mouse B7H3 antibodies and radioactive iodine-131 can significantly inhibit the growth of neuroblastoma in patients [J Neufoocol 97(3): 409-18 (2010)].

[0017] Summary of the Invention

[0018] In order to improve the therapeutic effect of antibody-drug conjugates (ADCs) or other ligand-drug conjugates (PDCs), reduce drug toxicity and side effects, and increase the therapeutic window, the present disclosure provides a ligand-drug conjugate represented by formula (XV) or a pharmaceutically acceptable salt or solvate thereof, which contains a bioactive molecule (drug molecule), a linker, and a targeting moiety, wherein the targeting moiety is connected to the linker via an active group (e.g., a sulfhydryl group) to form a ligand-drug conjugate. The present disclosure also develops antibodies comprising a variable light chain domain and / or a variable heavy chain (VH) domain that binds to a B7H3 molecule, wherein the variable light chain domain and / or variable heavy chain (VH) domain antibodies are already derived from human antibodies.

[0019] To this end, in the first aspect of the present disclosure, the present disclosure provides a ligand-drug conjugate as shown in Formula XV,

[0020]

[0021] or a stereoisomer of the ligand-drug conjugate, a prodrug thereof, a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable solvate thereof,

[0022] in:

[0023] Tb is a ligand or targeting moiety that binds to the target;

[0024] q is the drug-ligand coupling ratio;

[0025] D is a bioactive molecular fragment;

[0026] L1 is the extension unit;

[0027] L2 does not exist or is a linker unit; L3 is selected from an amino acid residue or a short peptide consisting of 2-10 amino acid residues;

[0028] L4 is absent or present. When L4 is present, L4 is selected from

[0029] Bit 1 is connected to L3 and bit 2 is connected to D.

[0030] In addition, it should be noted that regarding "position 1 of L1 is connected to Tb via an S atom", those skilled in the art will understand that position 1 of L1 is connected to the sulfhydryl group contained in Tb (such as an antibody) after the disulfide bond is opened (for example, the disulfide bond can be opened by reducing the reducing agent TCEP to generate a sulfhydryl group -SH). In other words, the -S- between L1 and Tb is not an additional external sulfur atom. For example, In the example, -S- is not an additional external sulfur atom, but the thiol group contained in Tb itself after the disulfide bond is opened and the thiol group of L1 is connected. -S- is formed by connecting the 1 bits of

[0031] The Stretcher unit is a component of a ligand-drug conjugate, drug-linker conjugate, or linker that connects the target-binding ligand or targeting moiety to the remainder of the ligand-drug conjugate or linker. The Stretcher unit can connect the Tb unit to L2 (if present) or L3. Specific examples include, but are not limited to, (where position 1 is connected to the target-binding ligand or targeting moiety, and position 2 is connected to L2 or L3):

[0032] In some embodiments, L1 is selected from:

[0033]

[0034]

[0035]

[0036] Each Z is independently selected from a direct bond, a carbon-carbon triple bond, a carbon-carbon double bond, a C6-10 aryl group, a 5-10 membered heteroaryl group, an amide group, a sulfonamide group, an imine group, and CF2;

[0037] Rx and Ry are independently selected from H and C1-4 alkyl;

[0038] Each m is independently selected from 0, 1, 2, 3, 4, 5 and 6;

[0039] y1, y2, y3 and y4 are independently selected from any integer between 0 and 20;

[0040] Position 1 is connected to Tb via an S atom, and position 2 is connected to L2 or L3.

[0041] The Linker is a component of a ligand-drug conjugate, drug-linker conjugate, or linker that serves to bind the Stretcher to an amino acid residue or a short peptide consisting of 2-10 amino acid residues. When present, the Linker can link L1 to L3. Specific examples include, but are not limited to, (wherein position 1 is linked to the Stretcher and position 2 is linked to L3):

[0042] In some embodiments, L2 is absent or present, and when present, L2 is selected from:

[0043]

[0044]

[0045] y1, y2, y3 and y4 are independently selected from any integer between 0 and 20, the 1st position is connected to L1, and the 2nd position is connected to L3.

[0046] In some embodiments, L1 is

[0047] In some embodiments, L1 is selected from Each Z is independently selected from a direct bond, a carbon-carbon triple bond, a carbon-carbon double bond, a C6-10 aromatic group, a 5-10 membered heteroaromatic group and an amide group (preferably selected from a direct bond, a carbon-carbon triple bond and a carbon-carbon double bond); Rx and Ry are independently selected from H and a C1-4 alkyl group; 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 (such as 4, 5, 6); each y2 is independently selected from any integer between 0 and 15 (such as 6-15); each y3 is independently selected from 1, 2 and 3; each y4 is independently selected from 0 and 1; position 1 is connected to Tb through an S atom, and position 2 is connected to L2 or L3.

[0048] In some embodiments, L1 is selected from m is selected from 2, 3, and 4, y1 is selected from any integer between 1 and 6 (such as 4, 5, and 6), each y2 is independently selected from any integer between 0 and 10 (such as 6 and 10), each y3 is independently selected from 1 or 2, position 1 is connected to Tb through an S atom, and position 2 is connected to L2 or L3.

[0049] In some embodiments, L1 is selected from Position 1 is connected to Tb via an S atom, and position 2 is connected to L2 or L3.

[0050] In some embodiments, L1 is selected from

[0051] In some embodiments, L1 is selected from Position 1 is connected to Tb via an S atom, and position 2 is connected to L2 or L3.

[0052] In some embodiments, L1 is selected from Position 1 is connected to Tb via an S atom, and position 2 is connected to L2 or L3.

[0053] In some embodiments, L1 is selected from Position 1 is connected to Tb via an S atom, and position 2 is connected to L2 or L3.

[0054] In some embodiments, L2 is absent or present, and when L2 is present, L2 is selected from y1 is selected from any integer between 1-6 (such as 4, 5, 6), each y2 is independently selected from any integer between 0-10 (such as 6-10), each y3 is independently selected from 1 or 2, each y4 is independently selected from 0 or 1, position 1 is connected to L1, and position 2 is connected to L3.

[0055] In some embodiments, L2 is absent or present, and when L2 is present, L2 is selected from Bit 1 is connected to L1 and bit 2 is connected to L3.

[0056] In some embodiments, L2 is absent or present, and when L2 is present, L2 is selected from Bit 1 is connected to L1 and bit 2 is connected to L3.

[0057] In some embodiments, L2 is absent or present, and when L2 is present, L2 is selected from Bit 1 is connected to L1 and bit 2 is connected to L3.

[0058] In some embodiments, L2 is absent.

[0059] In some embodiments, L2 is selected from

[0060] In some embodiments, L3 is selected from an amino acid residue or a short peptide consisting of 2-10 amino acid residues; the amino acid residue is selected from a natural amino acid residue, a non-natural amino acid residue, or an amino acid residue represented by AA1 or a stereoisomer thereof.

[0061] In some embodiments, L3 is selected from the group consisting of amino acid residues Val, D-Val, Cit, Phe, Lys, Lys(Ac), Leu, Gly, Ala, Asn, Asp, Arg, AA 1 or 2-10 selected from Val, Cit, Phe, Lys, D-Val, Leu, Gly, Ala, Asn, Asp, AA 1 A short peptide composed of amino acid residues.

[0062] In some embodiments, L3 is selected from 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 AA 1 -Gly, Ala-Ala-Ala, Ala-Ala-Asn, Ala-Ala-Asp, Val-AA 1 -Gly, Ala-AA 1 -Gly, Gly-AA 1 -Gly, Lys-Ala-Ala-Asn, Lys-Ala-Ala-Asp, Gly-Phe-Gly, Gly-Gly-Phe-Gly, D-Val-Leu-Lys, Gl y-Gly-Arg, Ala-Ala-Asn, Gly-Gly-Phe, Val-Lys-Gly, Val-Lys-Gly-Gly, Val-Lys and Lys-Ala-Asn.

[0063] In some embodiments, L3 is selected from AA 1 AA 1 -Gly, Val-Cit, Val-AA 1 -Gly, AA 1 -Ala-Asn and Gly-Gly-Phe-Gly.

[0064] In some embodiments, L3 is selected from AA 1and Val-AA 1 -Gly.

[0065] In some embodiments, L3 is selected from Val-AA 1 -Gly.

[0066] In some embodiments, L3 is selected from X - Selected from halogen ions, carboxylate ions, sulfate ions, hydrogen sulfate ions and OH - , bit 1 is connected to L1 or L2, bit 2 is connected to L4 or D.

[0067] In some embodiments, L3 is selected from

[0068] X - Selected from halogen ions, carboxylate ions, sulfate ions, hydrogen sulfate ions and OH - , bit 1 is connected to L1 or L2, bit 2 is connected to L4 or D.

[0069] In some embodiments, L3 is selected from X - Selected from halogen ions, carboxylate ions, sulfate ions, hydrogen sulfate ions and OH - , bit 1 is connected to L1 or L2, bit 2 is connected to L4 or D.

[0070] In some embodiments, L3 is selected from Bit 1 is connected to L1 or L2, and bit 2 is connected to L4 or D.

[0071] In some embodiments, L3 is selected from Bit 1 is connected to L1 or L2, and bit 2 is connected to L4 or D.

[0072] In some embodiments, AA 1 The structures of the indicated amino acid residues are shown below,

[0073]

[0074] in:

[0075] R a 、R b Each independently selected from H, And R a 、R b Not at the same time H;

[0076] Or, R a With R b Together with the carbon atoms to which they are attached, they form a 4-10 membered heterocyclic ring, which is optionally substituted by one or more R 0 replaced by;

[0077] r、r 1 are each independently selected from any integer from 0 to 20;

[0078] R m1 、R n1 Each independently selected from H, C1-6 alkyl, C3-6 cycloalkyl and -COOR x1 ;

[0079] R x1 Selected from C1-6 alkyl;

[0080] Or, R m1 With R n1 Together with the nitrogen atom to which they are attached, they form a 4-10 membered heterocyclic ring, which is optionally substituted by one or more R 0’ replaced by;

[0081] R z Selected from C1-6 alkyl;

[0082] R 0 、R 0’ Each independently selected from C1-6 alkyl, C3-6 cycloalkyl, -NR m2 R n2 and a 4-10 membered heterocyclic group optionally substituted by a C1-6 alkyl group;

[0083] R m2 、R n2 Each is independently selected from H and C1-6 alkyl.

[0084] In some embodiments, R a 、R b Among them, any one is H, and the other is selected from

[0085] In some embodiments, R a 、R b Among them, any one is H, and the other is selected from

[0086] In some embodiments, R a With R bTogether with the carbon atoms they are connected to, they form the 0 Substituted 5-6 membered heterocycle.

[0087] In some embodiments, R a With R b Together with the carbon atoms they are connected to, they form the 0 a substituted piperidine ring or a piperazine ring.

[0088] In some embodiments, R a With R b Together with the carbon atoms they are connected to, they form the 0 Substituted piperidine ring.

[0089] In some embodiments, R a With R b Together with the carbon atoms they are connected to, they form Carbon atom No. 1 is R a and R b carbon atoms that are linked together.

[0090] In some embodiments, R a With R b Together with the carbon atoms they are connected to, they form Carbon atom No. 1 is R a and R b carbon atoms that are linked together.

[0091] In some embodiments, r, r 1 Each is independently selected from 0, 1, 2, 3, 4 and 5.

[0092] In some embodiments, r, r 1 are each independently selected from 0 and 4.

[0093] In some embodiments, r, r 1 If one of them is 0, the other one is 4.

[0094] In some embodiments, R m1 、R n1 Each is independently selected from H, methyl, ethyl, n-propyl, n-butyl, -COOCH3, -COOCH2CH3, -COOCH2CH2CH3, -COOCH(CH3)2, -COOC(CH3)3 and -COOCH2CH2CH2CH3.

[0095] In some embodiments, R m1 、R n1 Each is independently selected from H, C1-6 alkyl, C3-6 cycloalkyl and tert-butyloxycarbonyl.

[0096] In some embodiments, R m1 、R n1 Each is independently selected from H and C1-6 alkyl.

[0097] In some embodiments, R m1 、R n1 Each is independently selected from H, methyl, ethyl and n-propyl.

[0098] In some embodiments, r, r 1 In the example, r is 4, r 1 When R is 0, m1 、R n1 Each is independently selected from H, C1-6 alkyl (such as H, methyl); r is 0, r 1 When R is 4, m1 、R n1 Each is independently selected from C1-6 alkyl (such as methyl, ethyl, n-propyl), preferably selected from C2-6 alkyl (such as ethyl, n-propyl).

[0099] In some embodiments, R m1 With R n1 Together with the nitrogen atom to which they are commonly attached, they form an optionally R 0’ Substituted 5-6 membered heterocycle.

[0100] In some embodiments, R m1 With R n1 Together with the nitrogen atom to which they are commonly attached, they form an optionally R 0’ a substituted piperidine ring or a piperazine ring.

[0101] In some embodiments, R m1 With R n1 Together with the nitrogen atoms they are connected to, they form Nitrogen atom No. 1 is R m1 and R n1 Commonly linked nitrogen atoms.

[0102] In some embodiments, R z It is a methyl group.

[0103] In some embodiments, R 0 、R 0’ Each independently selected from C1-6 alkyl, -NR m2 R n2 and a 5-6 membered heterocyclic group optionally substituted by a C1-6 alkyl group.

[0104] In some embodiments, R 0is selected from C1-6 alkyl and a 5-6 membered heterocyclic group substituted by a C1-6 alkyl group, wherein the 5-6 membered heterocyclic group is selected from piperidinyl and piperazinyl.

[0105] In some embodiments, R 0 is selected from methyl, ethyl and a 5-6 membered heterocyclic group substituted by a methyl group, wherein the 5-6 membered heterocyclic group is a piperidinyl group.

[0106] In some embodiments, R 0 is selected from methyl and a 5-6 membered heterocyclic group substituted by methyl, wherein the 5-6 membered heterocyclic group is piperidinyl.

[0107] In some embodiments, R 0 Selected from methyl, ethyl and

[0108] In some embodiments, R 0 Selected from methyl and

[0109] In some embodiments, R 0’ Selected from C1-6 alkyl and -NR m2 R n2 .

[0110] In some embodiments, R 0’ Selected from methyl and -NR m2 R n2 .

[0111] In some embodiments, R m2 、R n2 It is a methyl group.

[0112] In some embodiments, AA 1 The amino acid residues shown are selected from

[0113] In some embodiments, AA 1 The amino acid residues shown are selected from

[0114] In some embodiments, AA 1 The amino acid residues shown are selected from

[0115] In some embodiments, AA 1 The amino acid residues shown are selected from

[0116] In some embodiments, L4 is absent or present, and when L4 is present, L4 is selected from Bit 1 is connected to L3 and bit 2 is connected to D.

[0117] In some embodiments, L4 is absent or present, and when L4 is present, L4 is Bit 1 is connected to L3 and bit 2 is connected to D.

[0118] In some embodiments, L4 is absent.

[0119] In some embodiments, L4 is selected from Bit 1 is connected to L3 and bit 2 is connected to D.

[0120] In some embodiments, L4 is selected from Bit 1 is connected to L3 and bit 2 is connected to D.

[0121] In some embodiments, The structure is selected from the following structural fragments:

[0122]

[0123] In some embodiments, Tb is an antibody or an antigen-binding fragment thereof.

[0124] In some embodiments, the 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.

[0125] In some embodiments, Tb is an antibody or an antigen-binding fragment thereof having endocytic or non-endocytic activity.

[0126] In some embodiments, Tb is an antibody or an antigen-binding fragment thereof having endocytic activity.

[0127] In some embodiments, Tb is an antibody or an antigen-binding fragment thereof that has the activity of binding to free antigens in tumor tissues and / or tumor cell surface antigens.

[0128] In some embodiments, Tb is an antibody or an antigen-binding fragment thereof that has the activity of being internalized by tumor cells and has the activity of binding to free antigens in tumor tissues or antigens on the surface of tumor cells.

[0129] In some embodiments, Tb is an antibody or an antigen-binding fragment thereof that has the activity of being endocytosed by tumor cells and has the activity of binding to free antigens in tumor tissues and antigens on the surface of tumor cells.

[0130] In some embodiments, Tb is an antibody or an antigen-binding fragment thereof that has the activity of being endocytosed by tumor cells and has no free antigen-binding activity in tumor tissues.

[0131] In some embodiments, Tb is an antibody or an antigen-binding fragment thereof that has the activity of being endocytosed by tumor cells and has no activity of binding to tumor cell surface antigens.

[0132] In some embodiments, Tb is an antibody or an antigen-binding fragment thereof that has no endocytosis activity or has weak endocytosis activity.

[0133] In some embodiments, Tb is an antibody or an antigen-binding fragment thereof that does not have the activity of binding to free antigens in tumor tissues and / or tumor cell surface antigens.

[0134] In some embodiments, Tb is an antibody or an antigen-binding fragment thereof that has no or weak endocytosis activity by tumor cells and has binding activity to free antigens in tumor tissues or antigens on the surface of tumor cells.

[0135] In some embodiments, Tb is an antibody or an antigen-binding fragment thereof that has no or weak endocytosis activity by tumor cells and has binding activity to free antigens in tumor tissues and antigens on the surface of tumor cells.

[0136] In some embodiments, Tb is an antibody or antigen-binding fragment thereof that has no or weak endocytosis activity by tumor cells and no binding activity to antigens on the surface of tumor cells. In some embodiments, Tb is an antibody or antigen-binding fragment thereof that has no or weak endocytosis activity by tumor cells and no binding activity to free antigens in tumor tissue.

[0137] In some embodiments, Tb is an antibody or an antigen-binding fragment thereof that has no endocytosis activity by tumor cells and has no corresponding antigen in the human body.

[0138] In some embodiments, Tb is an antibody that does not bind to a tumor cell-associated antigen.

[0139] In some embodiments, the antibody or antigen-binding fragment thereof is selected from an IgG isotype antibody.

[0140] In some embodiments, the antibody or antigen-binding fragment thereof is selected from the group consisting of isotype IgG1, isotype IgG2, isotype IgG3, and isotype IgG4.

[0141] In some embodiments, the antibody or antigen-binding fragment thereof is an anti-chicken lysozyme human IgG1 isotype antibody.

[0142] In some embodiments, Tb is an antibody or an antigen-binding fragment thereof that has the activity of binding to non-endocytic antigens (eg, ALCAM / CD166) on the surface of tumor cells.

[0143] In some embodiments, Tb is an antibody that binds to a tumor cell-associated antigen.

[0144] In some embodiments, Tb is an antibody or an antigen-binding fragment thereof that has binding activity to free antigens in tumor tissues or antigens on the surface of tumor cells.

[0145] In some embodiments, Tb is an antibody or an antigen-binding fragment thereof that has binding activity to free antigens in tumor tissues and antigens on the surface of tumor cells.

[0146] In some embodiments, Tb is an antibody or antigen-binding fragment thereof having B7H3-2Ig and / or B7H-4Ig.

[0147] In some embodiments, Tb is an antibody or antigen-binding fragment thereof having higher binding activity for B7H3-4Ig than for B7H3-2Ig.

[0148] In some embodiments of the present disclosure, Tb is a ligand or targeting moiety that binds to a target.

[0149] In some embodiments, the target of Tb is selected from targets that are expressed higher in tumor cells than in normal cells.

[0150] In some embodiments, the target of Tb is selected from targets that are highly expressed in tumor cells and lowly expressed in normal cells.

[0151] In some embodiments, the target of Tb is selected from the group consisting of: B7H3, CD20, CD19, CD30, GPNMB, Her2, Trop-2, EGFR, Her3, GD-2, CD79b and BCMA, etc.

[0152] In some embodiments, the Tb is an antibody or an antigen-binding fragment thereof.

[0153] In some embodiments, Tb is a non-human antibody, a humanized antibody, a chimeric antibody, or a fully human antibody.

[0154] In some embodiments, Tb is a monoclonal antibody, a bispecific antibody, or a multispecific antibody.

[0155] In some embodiments, Tb is a monoclonal antibody or an antigen-binding fragment thereof.

[0156] 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.

[0157] In some embodiments, Tb is an anti-B7H3 antibody or an antigen-binding fragment thereof, such as 1D1, 1D1-01, 2E3, 2E3-02 antibodies, enoblituzumab, mirzotamab, omburtamab, or an antigen-binding fragment thereof.

[0158] The sequence of 1D1 is shown in SEQ ID NO: 1. The VH sequence of 1D1-01 is shown in SEQ ID NO: 3, and the VL sequence is shown in SEQ ID NO: 13. The heavy chain sequence of 1D1-01 is shown in SEQ ID NO: 45, and the light chain sequence is shown in SEQ ID NO: 46. The sequence of 2E3 is shown in SEQ ID NO: 2. The VH sequence of 2E3-02 is shown in SEQ ID NO: 23, and the VL sequence is shown in SEQ ID NO: 33. The heavy chain sequence of 2E3-02 is shown in SEQ ID NO: 47, and the light chain sequence is shown in SEQ ID NO: 48.

[0159] In some embodiments, Tb is an anti-B7H3 monoclonal antibody or an antigen-binding fragment thereof.

[0160] 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.

[0161] In some embodiments, Tb is an anti-Trop-2 monoclonal antibody or an antigen-binding fragment thereof.

[0162] In some embodiments, Tb is an anti-Her 2 antibody or an antigen-binding fragment thereof, such as anbenitamab, coprelotamab, disitamab, gancotamab, margetuximab, pertuzumab, timigutuzumab, zanidatamab, trastuzumab, pertuzumab or an antigen-binding fragment thereof.

[0163] In some embodiments, Tb is an anti-Her 2 monoclonal antibody or an antigen-binding fragment thereof, such as trastuzumab, pertuzumab, or an antigen-binding fragment thereof.

[0164] In some embodiments, Tb is an anti-Her3 antibody or an antigen-binding fragment thereof, such as barecetamab, duligotuzumab, elgemtumab, istiratumab, lumretuzumab, patritumab, seribantumab, zenocutuzumab, 202-2-1 antibody or an antigen-binding fragment thereof.

[0165] In some embodiments, Tb is an anti-Her3 monoclonal antibody or an antigen-binding fragment thereof.

[0166] In some embodiments, Tb is an anti-EGFR antibody or an antigen-binding fragment thereof, such as demupitamab, depatuxizumab, futuximab, imgatuzumab, laprituximab, losatuxizumab, matuzumab, modotuximab, necitumumab, nimotuzumab, panitumumab, pimurutamab, serclutamab, tomozutuximab, zalutumumab, Cetuximab or an antigen-binding fragment thereof.

[0167] In some embodiments, Tb is an anti-EGFR monoclonal antibody or an antigen-binding fragment thereof.

[0168] In some embodiments, the antibodies include antibodies that bind to antigens but do not have endocytosis activity. In some embodiments, the antibodies include antibodies that bind to antigens such as B7H3, CD20, CD19, CD30, GPNMB, Her2, Trop-2, EGFR, Her3, GD-2, CD79b, and BCMA but do not have endocytosis activity.

[0169] In some embodiments, the antibody has an antibody that binds to the B7H3 antigen but does not have endocytosis activity, such as INV721 and I7-01 of 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.

[0170] In some embodiments, the antibody has an antibody that binds to the GD2 antigen but does not have endocytosis activity, such as INV721 and GD2-5 of 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.

[0171] In some embodiments, the antibody has an antibody that binds to the HER3 antigen but does not have endocytosis activity, such as the anti-HER3 21F06 antibody shown in SEQ ID NO: 22 of US10808032B2.

[0172] In some embodiments, the antibody has the ability to bind to the CD20 antigen but lacks endocytosis activity. Although so-called "type II" CD20-specific antibodies have been shown to be poorly internalized 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 expression levels of activating and inhibitory FcγRs on the target cells with which they interact. In some embodiments, the antibody has the ability to bind to the CD20 antigen but lacks endocytosis activity is a "type II" CD20-specific antibody, such as obinutuzumab.

[0173] In some embodiments, the antibody has the activity of binding to non-endocytic antigens (e.g., ALCAM / CD166). In some embodiments, the antibody is an antibody comprising the VH of SEQ ID No: 73 and the VL of SEQ ID No: 74, the VH of SEQ ID No: 75 and the VL of SEQ ID No: 76, the VH of SEQ ID No: 77 and the VL of SEQ ID No: 78, or the VH of SEQ ID No: 79 and the VL of SEQ ID No: 88 in EP3911682A1.

[0174] In some embodiments, the Tb is a targeting moiety, such as a ligand, a protein, a polypeptide, a non-protein agent (such as a sugar, RNA or DNA), an antibody analog, etc.

[0175] In some embodiments, q is selected from any integer between 0.1 and 16.0; in a preferred embodiment, q is selected from any integer between 0.1 and 16.0.

[0176] In some embodiments, q is selected from any integer between 0.1 and 8.0. In a preferred embodiment, q is selected from any integer between 0.1 and 8.0.

[0177] In some embodiments, q is selected from any value between 2-8.

[0178] In some embodiments, q is selected from any value between 3-8.

[0179] In some embodiments, q is selected from any value between 4-8.

[0180] In some embodiments, q is selected from any value between 6-8.

[0181] In some embodiments, q is selected from any integer between 2-8.

[0182] In some embodiments, q is selected from any integer between 3-8.

[0183] In some embodiments, q is selected from any integer between 4-8.

[0184] In some embodiments, q is selected from any integer between 6-8.

[0185] In some embodiments, q is selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, and 12.

[0186] In some embodiments, q is selected from 2, 4, 6, and 8.

[0187] In this disclosure, the bioactive molecular fragments referred to are parts (fragments or groups) of antibody-drug conjugates (or antibody-drug conjugates, ADCs) known in the art, which can form bioactive drugs (such as small molecule cytotoxic drugs, including groups after losing an atom or group of atoms) or their derivatives (such as precursors) after the linker cleavage / degradation / enzymatic cleavage between tumor tissues or within tumor cells. For the avoidance of doubt, "drugs" do not only refer to "drugs" that have been approved by medical regulatory authorities, but also include any molecules with potential therapeutic biological activity in clinical practice, or in R&D and academic research.

[0188] In some embodiments, D is a molecular fragment having anti-tumor biological activity.

[0189] In some embodiments, D is a fragment of a molecule with anti-tumor biological activity, wherein the biologically active molecule is selected from a cytotoxic agent or a derivative thereof, such as a DNA topoisomerase inhibitor (e.g., a camptothecin-type biologically active molecule, such as camptothecin, DXD, a substituent-modified camptothecin, or a substituent-modified DXD) or a microtubule inhibitor (e.g., MMAF-type microtubule inhibitor, MMAE-type microtubule inhibitor).

[0190] In some embodiments, the Ligand Drug Conjugate has the structure of Formula I:

[0191]

[0192] in,

[0193] R1 and R2 are each independently selected from H, halogen, -OH, optionally substituted C1-6 alkyl and optionally substituted C1-6 alkoxy, or,

[0194] R1 and R2 together with the carbon atoms to which they are attached form a 5-7 membered carbocyclic ring or a 5-7 membered heterocyclic ring, wherein the heterocyclic ring contains one or more O, S, N, carbonyl, sulfoxide or sulfone groups or any combination thereof;

[0195] R3 is selected from H, halogen, -OH, -NH2, optionally substituted C1-6 alkyl and optionally substituted C1-6 alkoxy, or,

[0196] R3 and X together with the carbon atom to which they are attached form a 5-7 membered carbocyclic ring or a 5-7 membered heterocyclic ring containing one or more O, S, N, carbonyl, sulfoxide or sulfone groups or any combination thereof, or

[0197] R3 and R2 together with the carbon atom to which they are attached form a 5-7 membered carbocyclic ring or a 5-7 membered heterocyclic ring, wherein the heterocyclic ring contains one or more O, S, N, carbonyl, sulfoxide or sulfone groups or any combination thereof;

[0198] W is absent or present. When W is present, W is selected from -O-, -S-, -NR4-, Bit 1 is connected to X, bit 2 is connected to L4 or L3;

[0199] X is selected from a direct bond, an optionally substituted -O-(CH2) n3 -、-N(R4)-(CH2) n3 -、-S-(CH2) n3 -, carbonyl-(CH2) n3 、-SO2-(CH2) n3 -、 -(CH2) n1 -, C3-6 cycloalkyl, C6-10 aryl, 5-10 membered heteroaryl and 4-10 membered heterocyclic group, the 1 position is connected to the parent ring, and the 2 position is connected to W or L4; the substituent is selected from one or more C1-4 alkyl, C3-6 cycloalkyl, or multiple C1-4 alkyl and the carbon atoms to which they are simultaneously connected together form a C3-6 cycloalkyl;

[0200] Each M is independently selected from a direct bond and -CR 5a R 5b -;

[0201] 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;

[0202] n, n', n1, n2, n3 are each independently selected from any integer between 0 and 6;

[0203] L4 is absent or present. When L4 is present, L4 is selected from Bit 1 is connected to L3, and bit 2 is connected to W or X.

[0204] Tb, L1, L2, L3 and q have the meanings provided above and in any of the embodiments specifically described herein.

[0205] In some embodiments, R1 and R2 are each independently selected from H, halogen, and C1-4 alkyl.

[0206] In some embodiments, R1 and R2, together with the carbon atom 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.

[0207] In some embodiments, R1 is selected from H and halogen, and R2 is selected from H and C1-4 alkyl.

[0208] In some embodiments, R1 and R2 and the carbon atom to which they are attached form The dotted line indicates the position where the heterocyclic ring is fused to the benzene ring.

[0209] In some embodiments, R1 is H or F, and R2 is H or methyl.

[0210] In some embodiments, R1 is F, R2 is methyl, or R1 and R2 together with the carbon atom to which they are attached form

[0211] In some embodiments, R1 is F and R2 is methyl.

[0212] In some embodiments, R1 and R2 together with the carbon atom to which they are attached form

[0213] In some embodiments, R3 is selected from H and C1-4 alkyl.

[0214] In some embodiments, R3 and X, together with the carbon atom to which they are attached, form a 5-6 membered carbocyclic ring.

[0215] In some embodiments, R3 is H or R3 and X together with the carbon atom to which they are attached form The dotted lines indicate where the carbocyclic ring is fused to the benzene and pyridine rings.

[0216] In some embodiments, R3 is H.

[0217] In some embodiments, W is absent or present, and when W is present, W is selected from -O-, -S-, -NR4-, Bit 1 is connected to X, and bit 2 is connected to L4 or L3.

[0218] In some embodiments, W is absent or present, and when W is present, W is selected from -O-, -S-, -NR4-, Bit 1 is connected to X, and bit 2 is connected to L4 or L3.

[0219] In some embodiments, W is selected from -O-, -NR4- and Bit 1 is connected to X, and bit 2 is connected to L4 or L3.

[0220] In some embodiments, W is selected from -O- and -NR4-, position 1 is connected to X, and position 2 is connected to L4 or L3.

[0221] In some embodiments, X is selected from optionally substituted -(CH2) n1 -、 10-membered aryl, 5-10-membered heteroaryl and 4-10-membered heterocyclic group, the 1 position is connected to the parent ring, and the 2 position is connected to W or L4; the substituent is selected from 1 or 2 C1-4 alkyl groups, or 2 C1-4 alkyl groups and the carbon atoms to which they are simultaneously connected form a C3-6 cycloalkyl group.

[0222] In some embodiments, X is selected from optionally substituted Position 1 is connected to the parent ring, and position 2 is connected to W or L4; the substituent is selected from 1 or 2 C1-4 alkyl groups (such as methyl), or 2 C1-4 alkyl groups (such as methyl) and the carbon atoms to which they are simultaneously connected form a C3-6 cycloalkyl group (such as cyclopropyl).

[0223] In some embodiments, X is selected from Position 1 is connected to the mother ring, and position 2 is connected to W or L4.

[0224] In some embodiments, X is selected from Position 1 is connected to the mother ring, and position 2 is connected to W.

[0225] In some embodiments, when W is absent, X is selected from 1 is connected to the parent ring, 2 is connected to L4; when W exists, X is selected from Position 1 is connected to the mother ring, and position 2 is connected to W.

[0226] In some embodiments, W is selected from -O-, -NR4- and Position 1 is connected to X, position 2 is connected to L4 or L3; X is selected from Position 1 is connected to the mother ring, and position 2 is connected to W.

[0227] In some embodiments, R4 and R5 are each independently selected from H, C1-4 alkyl and C3-6 cycloalkyl.

[0228] In some embodiments, each R4 is independently selected from H, C1-4 alkyl, and C3-6 cycloalkyl, and R5 is H.

[0229] In some embodiments, each R4 is independently selected from H, methyl, ethyl, n-propyl, isopropyl, tert-butyl, and cyclopropyl, and R5 is H.

[0230] In some embodiments, R 5a 、R 5b Each is independently selected from H and C1-4 alkyl.

[0231] In some embodiments, R 5a 、R 5b are each independently selected from H and methyl.

[0232] In some embodiments, each R7 is independently selected from H and C1-4 alkyl.

[0233] In some embodiments, R7 is H.

[0234] In some embodiments, n is selected from 1, 2, and 3.

[0235] In some embodiments, n is 1.

[0236] In some embodiments, n1 is selected from 1, 2, 3, and 4.

[0237] In some embodiments, n2 is 1.

[0238] In some embodiments, n3 is 0.

[0239] In some embodiments, L3 is selected from X - Selected from halogen ions, carboxylate ions, sulfate ions, hydrogen sulfate ions and OH - , bit 1 is connected to L1 or L2, bit 2 is connected to L4 or W.

[0240] In some embodiments, L3 is selected from X -Selected from halogen ions, carboxylate ions, sulfate ions, hydrogen sulfate ions and OH - , bit 1 is connected to L1 or L2, bit 2 is connected to L4 or W.

[0241] In some embodiments, L3 is selected from X - Selected from halogen ions, carboxylate ions, sulfate ions and hydrogen sulfate ions, OH - , bit 1 is connected to L1 or L2, bit 2 is connected to L4 or W.

[0242] In some embodiments, L3 is selected from Bit 1 is connected to L1 or L2, and bit 2 is connected to L4 or W.

[0243] In some embodiments, L3 is selected from Bit 1 is connected to L1 or L2, and bit 2 is connected to L4 or W.

[0244] In some embodiments, L4 is absent or present, and when L4 is present, L4 is selected from Bit 1 is connected to L3, and bit 2 is connected to W or X.

[0245] In some embodiments, L4 is absent or present, and when L4 is present, L4 is Bit 1 is connected to L3, and bit 2 is connected to W or X.

[0246] In some embodiments, L4 is absent.

[0247] In some embodiments, L4 is selected from Bit 1 is connected to L3, and bit 2 is connected to W or X.

[0248] In some embodiments, L4 is selected from Bit 1 is connected to L3, and bit 2 is connected to W or X.

[0249] It should be noted that, as described above, W is absent or present. Therefore, when W is absent, position 1 of L4 is connected to L3 and position 2 is connected to X. When W is present, position 1 of L4 is connected to L3 and position 2 is connected to W. The following connection relationship of L4 can be understood with reference to the above content.

[0250] In some embodiments, The structure is selected from the following structural fragments:

[0251]

[0252]

[0253]

[0254]

[0255] Among them, bit 1 is connected to Tb and bit 2 is connected to W.

[0256] In some embodiments, D is The structural fragment shown; position 1 is connected to L3 or L4; for example

[0257] In some embodiments, The structure is selected from the following structural fragments:

[0258]

[0259]

[0260]

[0261] Among them, bit 1 is connected to L4; when L4 does not exist, bit 1 is connected to L3.

[0262] In some embodiments,

[0263] W is absent or present. When W is present, W is selected from -O-, -S-, -NR4-, For example, there is no, -O-, -NR4- or R4 and R5 are each independently selected from H and C1-4 alkyl; n is independently selected from 0, 1, 2, 3 and 4;

[0264] X is selected from

[0265] R1 is selected from H, halogen, R2 is selected from H, C1-4 alkyl, or R1 and R2 and the carbon atom to which they are connected form The dotted line indicates the position where the heterocyclic ring is fused to the benzene ring;

[0266] R3 is selected from H and C1-4 alkyl, or R3 and X together with the carbon atom to which they are attached form a 5-6 membered carbocyclic ring;

[0267] Preferably, W is absent or present, and when W is present, W is selected from -O-, -NR4- (eg, -NH-, -N(CH3)-, -N(C2H5)-), R4 is independently selected from H, methyl, ethyl, isopropyl, n-propyl, tert-butyl and cyclopropyl;

[0268] When W does not exist, X is selected from 1 is connected to the parent ring; when W exists, X is selected from

[0269] AA1 wherein r is selected from 0, 1, 2, 3, 4 and 5;

[0270] R a 、R b Among them, any one is H, and the other is selected from Or, R a With R b Together with the carbon atoms they are connected to, they form the 0 substituted 5-6 membered heterocycle;

[0271] R1 is selected from H, halogen, R2 is selected from H, C1-4 alkyl, or R1 and R2 and the carbon atom to which they are connected form

[0272] R3 is selected from H and C1-4 alkyl, or R3 and X together with the carbon atom to which they are attached form R3 and X together with the carbon atom to which they are attached form

[0273] More preferably, W is selected from -O- and -NR4-, position 1 is connected to X, and position 2 is connected to L4 or L3;

[0274] X is selected from Position 1 is connected to the mother ring, and position 2 is connected to W; For example

[0275] R1 is F,

[0276] R2 is methyl or R1 and R2 together with the carbon atom to which they are attached form

[0277] R3 is H.

[0278] In some embodiments, the Ligand Drug Conjugate has the structure of Formula I-1:

[0279]

[0280] wherein Tb, L1, L2, L3, L4, X, R1, R2, R3, R4 and q have the meanings provided above and in any of the embodiments specifically described herein.

[0281] In some embodiments, the Ligand Drug Conjugate has the structure of Formula I-1A or I-1B:

[0282]

[0283] wherein Tb, L2, L3, L4, X, R1, R2, R3, R4 and q have the meanings provided above and in any of the embodiments specifically described herein.

[0284] In some embodiments, the Ligand Drug Conjugate has the structure of Formula I-2:

[0285]

[0286] wherein Tb, L1, L2, L3, L4, X, R1, R2, R3 and q have the meanings provided above and in any of the embodiments specifically described herein.

[0287] In some embodiments, the Ligand Drug Conjugate has the structure of Formula I-2A or I-2B:

[0288]

[0289] wherein Tb, L2, L3, L4, X, R1, R2, R3 and q have the meanings provided above and in any of the embodiments specifically described herein.

[0290] In some embodiments, the Ligand Drug Conjugate has the structure of Formula I-3:

[0291]

[0292] wherein Tb, L1, L2, L3, L4, X, R1, R2, R3, R4, R5, n, and q have the meanings provided above and in any of the embodiments specifically described herein.

[0293] In some embodiments, the Ligand Drug Conjugate has the structure of Formula I-3A or I-3B:

[0294]

[0295] wherein Tb, L2, L3, L4, X, R1, R2, R3, R4 and q have the meanings provided above and in any of the embodiments specifically described herein.

[0296] In some embodiments, the Ligand Drug Conjugate has the structure of Formula IA:

[0297]

[0298] Among them, Tb, X, R1, R2, R3, R a 、R b and q have the meanings provided above and in any embodiments specifically described herein.

[0299] In some embodiments, the Ligand Drug Conjugate has the structure of Formula IB:

[0300]

[0301] Among them, Tb, X, R1, R2, R3, R a 、R b and q have the meanings provided above and in any embodiments specifically described herein.

[0302] In some embodiments, the ligand drug conjugate is selected from the group consisting of:

[0303]

[0304]

[0305]

[0306]

[0307]

[0308]

[0309]

[0310]

[0311]

[0312]

[0313]

[0314]

[0315]

[0316]

[0317]

[0318]

[0319]

[0320]

[0321]

[0322]

[0323]

[0324] The Tb antibodies or antigen-binding fragments thereof can be prepared by various methods known in the art, such as by genetic engineering and recombinant techniques. For example, DNA molecules encoding the heavy and light chain genes of the disclosed antibodies can be obtained by chemical synthesis or PCR amplification. The resulting DNA molecules are inserted into expression vectors, which are then transfected into host cells. The transfected host cells are then cultured under specific conditions to express the disclosed antibodies.

[0325] 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, omburtamab, and see CN112521512, WO2021027674, WO2021021543, WO2021006619, CN111662384, CN111454357, WO2020151384, WO2020140094, WO2020103100, WO2020102779, WO2020063673, WO2020047257, WO2020041626, CN1 10684790, CN110642948, WO2019225787, WO2019226017, US20190338030, CN110305213, WO2018209346, WO2018177393, US9150656, WO2016106004, WO2016044383, WO2016033225, WO2015181267, US20120294796, WO2011109400, CN101104639, WO2004093894, WO2002010187, WO2001018021. In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof is selected from the group consisting of: antibody numbered M30-H1-L4 in CN 103687945B and antibody numbered mAb-C-DUBA in CN 109069633 A.

[0326] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises the following complementarity determining regions (CDRs), wherein the CDRs are defined according to the IMGT numbering system:

[0327] (a) HCDR1 of SEQ ID NO: 10, HCDR2 of SEQ ID NO: 11, and HCDR3 of SEQ ID NO: 12; and / or,

[0328] LCDR1 with the sequence of SEQ ID NO: 20, LCDR2 with the sequence of GTF, and LCDR3 with the sequence of SEQ ID NO: 22;

[0329] (b) HCDR1 of SEQ ID NO: 10, HCDR2 of SEQ ID NO: 11, and HCDR3 of SEQ ID NO: 12; and / or,

[0330] LCDR1 with the sequence of SEQ ID NO: 40, LCDR2 with the sequence of GAS, and LCDR3 with the sequence of SEQ ID NO: 42;

[0331] (c) HCDR1 of SEQ ID NO: 30, HCDR2 of SEQ ID NO: 31, and HCDR3 of SEQ ID NO: 32; and / or,

[0332] LCDR1 with the sequence of SEQ ID NO: 40, LCDR2 with the sequence of GAS, and LCDR3 with the sequence of SEQ ID NO: 42;

[0333] or

[0334] (d) HCDR1 of SEQ ID NO: 30, HCDR2 of SEQ ID NO: 31, and HCDR3 of SEQ ID NO: 32; and / or,

[0335] The sequence of LCDR1 is SEQ ID NO: 20, the sequence of LCDR2 is GTF, and the sequence of LCDR3 is SEQ ID NO: 22.

[0336] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof, as defined by the IMGT numbering system, comprises:

[0337] a VH comprising (a) a heavy chain HCDR1, HCDR2, HCDR3 and a VL comprising (a) a light chain LCDR1, LCDR2, LCDR3;

[0338] a VH comprising (b) the heavy chain HCDR1, HCDR2, and HCDR3 and a VL comprising (b) the light chain LCDR1, LCDR2, and LCDR3;

[0339] A VH comprising (c) heavy chain HCDR1, HCDR2, HCDR3 and a VL comprising (c) light chain LCDR1, LCDR2, LCDR3; or

[0340] (d) VH containing heavy chain HCDR1, HCDR2, HCDR3 and (d) VL containing light chain LCDR1, LCDR2, LCDR3.

[0341] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises the following complementarity determining regions (CDRs), wherein the CDRs are defined according to the Chothia numbering system:

[0342] (a) HCDR1 of SEQ ID NO: 4, HCDR2 of SEQ ID NO: 5, and HCDR3 of SEQ ID NO: 6; and / or,

[0343] The sequence of LCDR1 is SEQ ID NO: 14, the sequence of LCDR2 is SEQ ID NO: 15, and the sequence of LCDR3 is SEQ ID NO: 16:

[0344] (b) HCDR1 of SEQ ID NO: 24, HCDR2 of SEQ ID NO: 25, and HCDR3 of SEQ ID NO: 26; and / or,

[0345] The sequence of LCDR1 is SEQ ID NO: 34, the sequence of LCDR2 is SEQ ID NO: 35, and the sequence of LCDR3 is SEQ ID NO: 36:

[0346] (c) HCDR1 of SEQ ID NO: 4, HCDR2 of SEQ ID NO: 5, and HCDR3 of SEQ ID NO: 6; and / or,

[0347] LCDR1 with the sequence of SEQ ID NO: 34, LCDR2 with the sequence of SEQ ID NO: 35, and LCDR3 with the sequence of SEQ ID NO: 36;

[0348] or

[0349] (d) HCDR1 of SEQ ID NO: 24, HCDR2 of SEQ ID NO: 25, and HCDR3 of SEQ ID NO: 26; and / or,

[0350] The sequence of LCDR1 is SEQ ID NO: 14, the sequence of LCDR2 is SEQ ID NO: 15, and the sequence of LCDR3 is SEQ ID NO: 16.

[0351] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof, as defined by the Chothia numbering system, comprises:

[0352] a VH comprising (a) a heavy chain HCDR1, HCDR2, HCDR3 and a VL comprising (a) a light chain LCDR1, LCDR2, LCDR3;

[0353] a VH comprising (b) the heavy chain HCDR1, HCDR2, and HCDR3 and a VL comprising (b) the light chain LCDR1, LCDR2, and LCDR3;

[0354] A VH comprising (c) heavy chain HCDR1, HCDR2, HCDR3 and a VL comprising (c) light chain LCDR1, LCDR2, LCDR3; or

[0355] (d) VH containing heavy chain HCDR1, HCDR2, HCDR3 and (d) VL containing light chain LCDR1, LCDR2, LCDR3.

[0356] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises the following complementarity determining regions (CDRs), wherein the CDRs are defined according to the Kabat numbering system:

[0357] (a) HCDR1 of SEQ ID NO: 7, HCDR2 of SEQ ID NO: 8, and HCDR3 of SEQ ID NO: 9; and / or,

[0358] LCDR1 with the sequence of SEQ ID NO: 17, LCDR2 with the sequence of SEQ ID NO: 18, and LCDR3 with the sequence of SEQ ID NO: 19;

[0359] (b) HCDR1 of SEQ ID NO: 27, HCDR2 of SEQ ID NO: 28, and HCDR3 of SEQ ID NO: 29; and / or,

[0360] The LCDR1 sequence is SEQ ID NO: 37, the LCDR2 sequence is SEQ ID NO: 38, and the LCDR3 sequence is SEQ ID NO: 39:

[0361] (c) HCDR1 of SEQ ID NO: 7, HCDR2 of SEQ ID NO: 8, and HCDR3 of SEQ ID NO: 9; and / or,

[0362] LCDR1 with the sequence of SEQ ID NO: 37, LCDR2 with the sequence of SEQ ID NO: 38, and LCDR3 with the sequence of SEQ ID NO: 39;

[0363] or

[0364] (d) HCDR1 of SEQ ID NO: 27, HCDR2 of SEQ ID NO: 28, and HCDR3 of SEQ ID NO: 29; and / or,

[0365] The sequence of LCDR1 is SEQ ID NO: 17, the sequence of LCDR2 is SEQ ID NO: 18, and the sequence of LCDR3 is SEQ ID NO: 19.

[0366] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof, as defined by the Kabat numbering system, comprises:

[0367] a VH comprising (a) a heavy chain HCDR1, HCDR2, HCDR3 and a VL comprising (a) a light chain LCDR1, LCDR2, LCDR3;

[0368] a VH comprising (b) the heavy chain HCDR1, HCDR2, and HCDR3 and a VL comprising (b) the light chain LCDR1, LCDR2, and LCDR3;

[0369] A VH comprising (c) heavy chain HCDR1, HCDR2, HCDR3 and a VL comprising (c) light chain LCDR1, LCDR2, LCDR3; or

[0370] (d) VH containing heavy chain HCDR1, HCDR2, HCDR3 and (d) VL containing light chain LCDR1, LCDR2, LCDR3.

[0371] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises:

[0372] (a) a heavy chain variable region (VH) comprising an amino acid sequence selected from the group consisting of:

[0373] (i) the sequence shown in SEQ ID NO: 3 or 23;

[0374] (ii) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence shown 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

[0375] (iii) 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;

[0376] and / or

[0377] (b) a light chain variable region (VL) comprising an amino acid sequence selected from the group consisting of:

[0378] (iv) the sequence shown in SEQ ID NO: 13 or 33;

[0379] (v) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence shown 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

[0380] (vi) 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: 13 or 33.

[0381] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises the VH set forth in SEQ ID NO: 3, and / or the VL set forth in SEQ ID NO: 13.

[0382] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises the VH set forth in SEQ ID NO: 23, and / or the VL set forth in SEQ ID NO: 33.

[0383] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises the VH set forth in SEQ ID NO: 3, and / or the VL set forth in SEQ ID NO: 33.

[0384] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises the VH set forth in SEQ ID NO: 23, and / or the VL set forth in SEQ ID NO: 13.

[0385] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises:

[0386] (a) VH of SEQ ID NO: 3 and VL of SEQ ID NO: 13;

[0387] (b) VH of SEQ ID NO: 23 and VL of SEQ ID NO: 33;

[0388] (c) VH of the sequence shown in SEQ ID NO: 3 and VL of the sequence shown in SEQ ID NO: 13;

[0389] (d) VH of SEQ ID NO: 23 and VL of SEQ ID NO: 13;

[0390] (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 described in any one of groups (a) to (f), respectively; or

[0391] (f) 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 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 any one of groups (a) to (d), respectively. Preferably, the substitutions are conservative substitutions.

[0392] In some embodiments, the heavy chain of the anti-B7H3 antibody comprises a heavy chain constant region (CH) of a human immunoglobulin or a variant thereof having up to 50 conservative substitutions of amino acids compared to the wild-type sequence from which it is derived (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 substitutions of amino acids; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 conservative substitutions of amino acids). In certain embodiments, the anti-B7H3 antibody light chain comprises a light chain constant region (CL) of a human immunoglobulin or a variant thereof having 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.

[0393] In some embodiments, the constant region is altered, for example, mutated, to modify the properties of the anti-B7H3 antibody molecule (e.g., to alter one or more of the following: Fc receptor binding, antibody glycosylation, the number of cysteine ​​residues, effector cell function, or complement function). Functional changes can be produced by replacing at least one amino acid residue in the antibody constant region with a different residue, for example, by altering the affinity of the antibody for an effector ligand (such as FcR or complement C1q), thereby altering (e.g., reducing) the effector function. The Fc region of an antibody mediates several important effector functions, such as ADCC, phagocytosis (ADCP), CDC, etc.

[0394] In some embodiments, the 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 selected from, for example, the heavy chain constant regions of IgG1, IgG2, IgG3, and IgG4, and more particularly selected from the heavy chain constant region of IgG1 (e.g., human IgG1). In some embodiments, the human IgG1 heavy chain constant region is as shown in SEQ ID NO: 43. In some embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure have 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 shown in SEQ ID NO: 44.

[0395] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises the CH set forth in SEQ ID NO:43 or a variant thereof having 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; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 conservative amino acid substitutions) compared to SEQ ID NO:43, or 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 SEQ ID NO:43.

[0396] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises a light chain constant region or a 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) as set forth in SEQ ID NO: 44 or a variant thereof, wherein the variant has up to 20 conservative amino acid substitutions (e.g., up to 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: 44, or 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 SEQ ID NO: 44.

[0397] 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.

[0398] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises:

[0399] (a) a heavy chain comprising an amino acid sequence selected from the group consisting of:

[0400] (i) a sequence comprising the VH sequence shown in SEQ ID NO: 3 and the CH sequence shown in SEQ ID NO: 43;

[0401] (ii) a sequence having one or several amino acid substitutions, deletions or additions or any combination thereof compared to the sequence shown 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; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof); or

[0402] (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 shown in (i); and

[0403] (b) a light chain comprising an amino acid sequence selected from the group consisting of:

[0404] (iv) a sequence comprising the VL sequence shown in SEQ ID NO: 13 and the CL sequence shown in SEQ ID NO: 44;

[0405] (v) a sequence having one or several amino acid substitutions, deletions or additions or any combination thereof compared to the sequence shown in (iv) (e.g., at most 50, at most 45, at most 40, at most 35, at most 30, at most 25, at most 20, at most 15, at most 10 or at most 5 amino acid substitutions, deletions or additions or any combination thereof; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof); or

[0406] (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 shown in (iv).

[0407] In some embodiments, the substitution described in (ii) or (v) is a conservative substitution.

[0408] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises:

[0409] (a) a heavy chain comprising an amino acid sequence selected from the group consisting of:

[0410] (i) a sequence comprising the VH sequence shown in SEQ ID NO: 23 and the CH sequence shown in SEQ ID NO: 43;

[0411] (ii) a sequence having one or several amino acid substitutions, deletions or additions or any combination thereof compared to the sequence shown 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; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof); or

[0412] (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 shown in (i); and

[0413] (b) a light chain comprising an amino acid sequence selected from the group consisting of:

[0414] (iv) a sequence comprising the VL sequence shown in SEQ ID NO: 33 and the CL sequence shown in SEQ ID NO: 44;

[0415] (v) a sequence having one or several amino acid substitutions, deletions or additions or any combination thereof compared to the sequence shown in (iv) (e.g., at most 50, at most 45, at most 40, at most 35, at most 30, at most 25, at most 20, at most 15, at most 10 or at most 5 amino acid substitutions, deletions or additions or any combination thereof; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof); or

[0416] (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 shown in (iv).

[0417] In some embodiments, the substitution described in (ii) or (v) is a conservative substitution.

[0418] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises a heavy chain and a light chain,

[0419] The heavy chain comprises:

[0420] (i) the sequence shown in SEQ ID NO: 45;

[0421] (ii) a sequence having one or several amino acid substitutions, deletions or additions or any combination thereof compared to the sequence shown 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; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof); or

[0422] (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 shown in (i); and

[0423] The light chain comprises:

[0424] (iv) the sequence shown in SEQ ID NO: 46;

[0425] (v) a sequence having one or several amino acid substitutions, deletions or additions or any combination thereof compared to the sequence shown 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; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof); or

[0426] (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 to the sequence set forth in (iv);

[0427] Preferably, the substitutions described in (ii) or (v) are conservative substitutions.

[0428] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises a heavy chain and a light chain,

[0429] The heavy chain comprises:

[0430] (i) the sequence shown in SEQ ID NO: 47;

[0431] (ii) a sequence having one or several amino acid substitutions, deletions or additions or any combination thereof compared to the sequence shown 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; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof); or

[0432] (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); and

[0433] The light chain comprises:

[0434] (iv) the sequence shown in SEQ ID NO: 48;

[0435] (v) a sequence having one or several amino acid substitutions, deletions or additions or any combination thereof compared to the sequence shown in (iv) (e.g., at most 50, at most 45, at most 40, at most 35, at most 30, at most 25, at most 20, at most 15, at most 10 or at most 5 amino acid substitutions, deletions or additions or any combination thereof; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof); or

[0436] (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 to the sequence set forth in (iv);

[0437] Preferably, the substitutions described in (ii) or (v) are conservative substitutions.

[0438] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof is a chimeric antibody, a humanized antibody, or a fully human antibody.

[0439] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof is selected from scFv, Fab, Fab', (Fab')2, Fv fragment, disulfide-linked Fv (dsFv), and diabody.

[0440] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof is a scFv. In certain embodiments, the scFv of the present disclosure comprises:

[0441] (a) VH of SEQ ID NO: 3 and VL of SEQ ID NO: 13;

[0442] (b) VH of SEQ ID NO: 23 and VL of SEQ ID NO: 33;

[0443] (c) VH of SEQ ID NO: 3 and VL of SEQ ID NO: 33;

[0444] (d) VH of SEQ ID NO: 23 and VL of SEQ ID NO: 13;

[0445] (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 described in any one of groups (a) to (d), respectively; or

[0446] (f) 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 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 any one of groups (a) to (d), respectively. Preferably, the substitutions are conservative substitutions.

[0447] In some embodiments, the antibodies of the present disclosure are scFvs. In certain embodiments, the scFvs of the present disclosure comprise:

[0448] (i) the sequence shown in SEQ ID NO: 1 or 2;

[0449] (ii) a sequence having one or several amino acid substitutions, deletions or additions or any combination thereof compared to the sequence shown 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; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof); or

[0450] (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 shown in (i);

[0451] Preferably, the substitutions described in (ii) are conservative substitutions.

[0452] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof is a scFv. In certain embodiments, the scFv of the present disclosure comprises the sequence shown in SEQ ID NO: 1 or 2.

[0453] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof is selected from the group consisting of: 1D1 antibody, 1D1-01 antibody, 2E3 antibody, and 2E3-02 antibody.

[0454] In some embodiments of the present disclosure, the targeting moiety is trastuzumab or pertuzumab. Trastuzumab is an anti-Her 2 monoclonal antibody, and its amino acid sequence is known to those skilled in the art. Its schematic sequence can be found, for example, in CN103319599. The terminal Lys is easily deleted without affecting biological activity, see Dick, LW et al., Biotechnol. Bioeng., 100: 1132-1143.

[0455] Exemplary heavy and light chain sequences of trastuzumab can be found, for example, in IMGT / mAb-DB ID 97. Exemplary heavy and light chain sequences of pertuzumab can be found in SEQ ID No. 16 and SEQ ID No. 15 of US7560111, and also in IMGT / mAb-DB ID 80.

[0456] 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 prior art anti-Her3 antibodies, for example, barecetamab, duligotuzumab, elgemtumab, istiratumab, lumretuzumab, patritumab, seribantumab, zenocutuzumab, 202-2-1 antibody, antibody CN 103189392 B with a heavy chain represented by SEQ ID NO: 10 and a light chain represented by SEQ ID NO: 14, and the antibody represented by IMGT / mAb-DB SEQ ID NO: 546.

[0457] In some embodiments, the targeting moiety is an anti-Her3 antibody, wherein the anti-Her3 antibody or antigen-binding fragment thereof comprises the following complementarity determining regions (CDRs), wherein the CDRs are defined according to the IMGT numbering system:

[0458] HCDR1 of SEQ ID NO: 56, HCDR2 of SEQ ID NO: 57, HCDR3 of SEQ ID NO: 58; and / or,

[0459] The sequence of LCDR1 is SEQ ID NO: 41, the sequence of LCDR2 is AAS, and the sequence of LCDR3 is SEQ ID NO: 21.

[0460] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof, as defined by the IMGT numbering system, comprises: the aforementioned VH comprising the aforementioned heavy chain HCDR1, HCDR2, and HCDR3 and the aforementioned VL comprising the aforementioned light chain LCDR1, LCDR2, and LCDR3.

[0461] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof comprises the following complementarity determining regions (CDRs), wherein the CDRs are defined according to the Kabat numbering system:

[0462] HCDR1 of SEQ ID NO: 53, HCDR2 of SEQ ID NO: 54, HCDR3 of SEQ ID NO: 55; and / or,

[0463] The sequence of LCDR1 is SEQ ID NO: 59, the sequence of LCDR2 is SEQ ID NO: 60, and the sequence of LCDR3 is SEQ ID NO: 21.

[0464] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof, as defined by the Kabat numbering system, comprises: the aforementioned VH comprising the aforementioned heavy chain HCDR1, HCDR2, and HCDR3 and the aforementioned VL comprising the aforementioned light chain LCDR1, LCDR2, and LCDR3.

[0465] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof comprises:

[0466] (a) a heavy chain variable region (VH) comprising an amino acid sequence selected from the group consisting of:

[0467] (i) the sequence shown in SEQ ID NO: 49;

[0468] (ii) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence shown 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

[0469] (iii) 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: 49;

[0470] and / or

[0471] (b) a light chain variable region (VL) comprising an amino acid sequence selected from the group consisting of:

[0472] (iv) the sequence shown in SEQ ID NO: 50;

[0473] (v) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence shown 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

[0474] (vi) 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:50.

[0475] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof comprises:

[0476] (a) VH of SEQ ID NO: 49 and VL of SEQ ID NO: 50;

[0477] (b) 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 described in group (a), respectively; or

[0478] (c) 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 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), respectively. Preferably, the substitutions are conservative substitutions.

[0479] In some embodiments, the heavy chain of the anti-Her3 antibody comprises a heavy chain constant region (CH) of a human immunoglobulin or a variant thereof, which has up to 50 conservative substitutions of amino acids compared to the wild-type sequence from which it is derived (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 substitutions of amino acids; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 conservative substitutions of amino acids). In certain embodiments, the anti-B7H3 antibody light chain comprises a light chain constant region (CL) of a human immunoglobulin or a variant thereof having 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.

[0480] In some embodiments, the constant region is altered, for example, mutated, to modify the properties of the anti-Her3 antibody molecule (e.g., to alter one or more of the following: Fc receptor binding, antibody glycosylation, the number of cysteine ​​residues, effector cell function, or complement function). Functional changes can be produced by replacing at least one amino acid residue in the antibody constant region with a different residue, for example, by changing the affinity of the antibody for an effector ligand (such as FcR or complement C1q), thereby altering the effector function (e.g., reducing it). The Fc region of an antibody mediates several important effector functions, such as ADCC, phagocytosis (ADCP), CDC, etc.

[0481] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof has a heavy chain constant region (CH), which is selected from, for example, the heavy chain constant region of IgG1, IgG2, IgG3, IgG4, IgM, IgA1, IgA2, IgD and IgE; in particular, it is selected from, for example, the heavy chain constant region of IgG1, IgG2, IgG3 and IgG4, more particularly, it is selected from the heavy chain constant region of IgG1 (for example, human IgG1). In some embodiments, the antibody or antigen-binding fragment thereof of the present disclosure has a light chain constant region, which is selected from, for example, a kappa or lambda light chain constant region, preferably a kappa light chain constant region (for example, a human kappa light chain constant region).

[0482] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof comprises a heavy chain constant region (CH) as shown in SEQ ID NO: 43 and a light chain constant region (CL) as shown in SEQ ID NO: 44.

[0483] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof comprises a heavy chain and a light chain,

[0484] The heavy chain comprises:

[0485] (i) the sequence shown in SEQ ID NO: 51;

[0486] (ii) a sequence having one or several amino acid substitutions, deletions or additions or any combination thereof compared to the sequence shown 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; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof); or

[0487] (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 shown in (i); and

[0488] The light chain comprises:

[0489] (iv) the sequence shown in SEQ ID NO: 52;

[0490] (v) a sequence having one or several amino acid substitutions, deletions or additions or any combination thereof compared to the sequence shown in (iv) (e.g., at most 50, at most 45, at most 40, at most 35, at most 30, at most 25, at most 20, at most 15, at most 10 or at most 5 amino acid substitutions, deletions or additions or any combination thereof; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof); or

[0491] (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 to the sequence set forth in (iv);

[0492] Preferably, the substitutions described in (ii) or (v) are conservative substitutions.

[0493] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof is a chimeric antibody, a humanized antibody, or a fully human antibody.

[0494] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof is selected from scFv, Fab, Fab', (Fab')2, Fv fragment, disulfide-linked Fv (dsFv), and diabody.

[0495] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof is a scFv. In certain embodiments, the scFv of the present disclosure comprises:

[0496] (a) VH of SEQ ID NO: 49 and VL of SEQ ID NO: 50;

[0497] (b) 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 described in group (a), respectively; or

[0498] (c) 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 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), respectively. Preferably, the substitutions are conservative substitutions.

[0499] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof is selected from: 202-2-1 antibody.

[0500] In some embodiments of the present disclosure, the targeting moiety is cetuximab. Cetuximab is an anti-EGFR monoclonal antibody, whose amino acid sequence is known to those skilled in the art. For its schematic sequence, see the antibody shown in IMGT / mAb-DB ID 151.

[0501] In some embodiments, the ligand drug conjugate is selected from:

[0502]

[0503]

[0504]

[0505] Wherein, Tb1 is an anti-B7H3 antibody or an antigen-binding fragment thereof, such as enoblituzumab, mirzotamab, omburtamab, 1D1-01, or 2E3-02 antibody, preferably 1D1-01 or 2E3-02 antibody; and q is selected from any value between 0.1 and 16.0, preferably any value 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.

[0506] In some embodiments, the ligand drug conjugate is selected from:

[0507]

[0508]

[0509] Wherein, Tb2 is an anti-Trop-2 antibody or an antigen-binding fragment thereof, such as datopotamab or sacituzumab, preferably sacituzumab; and q is selected from any value between 0.1 and 16.0, preferably any value 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.

[0510] In some embodiments, the ligand drug conjugate is selected from:

[0511]

[0512]

[0513] Wherein, Tb3 is an anti-Her2 antibody or an antigen-binding fragment thereof, such as anbenitamab, coprelotamab, disitamab, gancotamab, margetuximab, pertuzumab, timigutuzumab, zanidatamab, trastuzumab, pertuzumab, preferably trastuzumab or pertuzumab; q is selected from any value between 0.1 and 16.0, preferably any value 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.

[0514] In some embodiments, the ligand drug conjugate is selected from:

[0515]

[0516] Wherein, Tb4 is an anti-Her3 antibody or an antigen-binding fragment thereof, for example, barecetamab, duligotuzumab, elgemtumab, istiratumab, lumretuzumab, patritumab, seribantumab, zenocutuzumab, the 202-2-1 antibody, the antibody with the heavy chain represented by SEQ: 10 and the light chain represented by SEQ: 14 in CN 103189392 B, the antibody represented by IMGT / mAb-DB ID: 564, or the 202-2-1 antibody; preferably, the 202-2-1 antibody; q is selected from any value between 0.1 and 16.0, preferably any value 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.

[0517] In some embodiments, the ligand drug conjugate is selected from:

[0518]

[0519] Wherein, Tb5 is an anti-EGFR antibody or an antigen-binding fragment thereof, such as demupitamab, depatuxizumab, futuximab, imgatuzumab, laprituximab, losatuxizumab, matuzumab, modotuximab, necitumumab, nimotuzumab, panitumumab, pimurutamab, serclutamab, tomozutuximab, zalutumumab, or cetuximab; preferably cetuximab; and q is selected from any value between 0.1 and 16.0, preferably any value 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.

[0520] In some embodiments, the ligand drug conjugate is selected from:

[0521]

[0522] Wherein, Tb6 is an antibody without tumor cell endocytosis activity or an antibody with binding activity to non-endocytic antigens (e.g., ALCAM / CD166), for example, an IgG isotype antibody for which the human body lacks the corresponding cell surface antigen, an anti-CD166 antibody, preferably an anti-chicken lysozyme human IgG1 isotype antibody; q is selected from any value between 0.1 and 16.0, preferably any value 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.

[0523] In some embodiments, the ligand drug conjugate is selected from:

[0524]

[0525] Wherein, Tb7 is an antibody with weak or no tumor cell endocytosis activity but with tumor cell surface antigen binding activity; q is selected from any value between 0.1 and 16.0, preferably any value 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.

[0526] In some embodiments, the antibody has an antibody that binds to the B7H3 antigen but does not have endocytosis activity, such as INV721 and I7-01 of 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.

[0527] In some embodiments, the antibody has an antibody that binds to the GD-2 antigen but does not have endocytosis activity, such as INV721 and GD2-5 of WO2021168379A1. More specifically, the anti-GD-2 antibody has a VH of SEQ ID NO: 4 and a VL of SEQ ID NO: 3 as described in WO2021168379A1.

[0528] In some embodiments, the antibody has an antibody that binds to the HER3 antigen but does not have endocytosis activity, such as the anti-HER3 21F06 antibody shown in SEQ ID NO: 22 of US10808032B2.

[0529] In some embodiments, the antibody has the ability to bind to the CD20 antigen but lacks endocytosis activity. Although so-called "type II" CD20-specific antibodies have been shown to be poorly internalized 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 expression levels of activating and inhibitory FcγRs on the target cells with which they interact. In some embodiments, the antibody has the ability to bind to the CD20 antigen but lacks endocytosis activity is a "type II" CD20-specific antibody, such as obinutuzumab.

[0530] In some embodiments, the antibody has the activity of binding to non-endocytic antigens (e.g., ALCAM / CD166). In some embodiments, the antibody is an antibody comprising the VH of SEQ ID No: 73 and the VL of SEQ ID No: 74, the VH of SEQ ID No: 75 and the VL of SEQ ID No: 76, the VH of SEQ ID No: 77 and the VL of SEQ ID No: 78, or the VH of SEQ ID No: 79 and the VL of SEQ ID No: 88 in EP3911682A1.

[0531] In some embodiments, the ligand drug conjugate is selected from:

[0532]

[0533] Wherein, Tb8 is an antibody that has tumor cell endocytosis activity and tumor cell surface antigen binding activity, such as 1D1-01, 2E3-02, sacituzumab, pertuzumab, trastuzumab, or cetuximab; q is selected from any value between 0.1 and 16.0, preferably any value 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.

[0534] It should also be noted that those skilled in the art will understand that L is linked to the sulfhydryl group contained in Tb (e.g., an antibody) after disulfide bond opening (for example, disulfide bond reduction by the reducing agent TCEP can open the disulfide bond to form a sulfhydryl group -SH). In other words, the -S- between L and Tb is not a separately attached sulfur atom. For example, the -S- is not a separately attached sulfur atom, but rather the -S- formed by the sulfhydryl group contained in Tb after disulfide bond opening connecting with L.

[0535] In the second aspect of the present disclosure, the present disclosure provides a compound represented by formula II:

[0536]

[0537] or a stereoisomer of the compound, a prodrug thereof, a pharmaceutically acceptable salt thereof, a pharmaceutically acceptable solvate thereof, or a ligand-drug conjugate thereof,

[0538] in:

[0539] R1 and R2 are each independently selected from H, halogen, -OH, optionally substituted C1-6 alkyl, optionally substituted C1-6 alkoxy, or,

[0540] R1 and R2 together with the carbon atoms to which they are attached form a 5-7 membered carbocyclic ring or a 5-7 membered heterocyclic ring, wherein the heterocyclic ring contains one or more O, S, N, carbonyl, sulfoxide or sulfone groups or any combination thereof;

[0541] R3 is selected from H, halogen, -OH, -NH2, optionally substituted C1-6 alkyl, optionally substituted C1-6 alkoxy, or,

[0542] R3 and X together with the carbon atom to which they are attached form a 5-7 membered carbocyclic ring or a 5-7 membered heterocyclic ring, wherein the heterocyclic ring contains one or more O, S, N, carbonyl, sulfoxide or sulfone groups or any combination thereof;

[0543] R3 and R2 together with the carbon atom to which they are attached form a 5-7 membered carbocyclic ring or a 5-7 membered heterocyclic ring, wherein the heterocyclic ring contains one or more O, S, N, carbonyl, sulfoxide or sulfone groups or any combination thereof;

[0544] W is absent or present. When W is present, W is selected from -OH, -SH, -NHR4, 1 bit is connected to X;

[0545] X is selected from a direct bond, an optionally substituted -O-(CH2) n3 -、-N(R4)-(CH2) n3 -、-S-(CH2) n3 -, carbonyl-(CH2) n3 、-SO2-(CH2) n3 -, -(CH2) n1 -、 C3-6 cycloalkyl, C6-10 aryl, 5-10 membered heteroaryl and 4-10 membered heterocyclyl, the 1 position is connected to the parent ring, and the 2 position is connected to W; the substituent is selected from one or more C1-4 alkyl, C3-6 cycloalkyl, or multiple C1-4 alkyl groups and the carbon atoms to which they are simultaneously connected together form a C3-6 cycloalkyl;

[0546] Each M is independently selected from a direct bond, -CR 5a R5b -;

[0547] R4, R5, R 5a 、R 5b , R6, R 7 Each is independently selected from H, optionally substituted C1-4 alkyl, optionally substituted C1-4 alkoxy, optionally substituted C3-6 cycloalkyl;

[0548] n, n', n1, n2, n3 are each independently selected from any integer between 0 and 6;

[0549] Among them, when R1 and R2 are both H, and X is -(CH2) n1 -, n1 is 1, 2, 3, 4, W is not -OH or -NHR4; and the compound represented by formula II does not contain

[0550]

[0551] In some embodiments, R1 and R2 are each independently selected from H, halogen, or C1-4 alkyl.

[0552] In some embodiments, R1 and R2, together with the carbon atom 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.

[0553] In some embodiments, R1 is selected from H and halogen, and R2 is selected from H and C1-4 alkyl.

[0554] In some embodiments, R1 and R2 and the carbon atom to which they are attached form The dotted line indicates the position where the heterocyclic ring is fused to the benzene ring.

[0555] In some embodiments, R1 is H or F, and R2 is H or methyl.

[0556] In some embodiments, R1 and R2 together with the carbon atom to which they are attached form

[0557] In some embodiments, R1 is F, R2 is methyl; or R1 and R2 together with the carbon atom to which they are attached form

[0558] In some embodiments, R1 is F and R2 is methyl.

[0559] In some embodiments, R1 and R2 together with the carbon atom to which they are attached form

[0560] In some embodiments, R3 is selected from H and C1-4 alkyl.

[0561] In some embodiments, R3 is H.

[0562] In some embodiments, W is absent or present, and when W is present, W is selected from -OH, -SH, -NHR4, 1 bit is connected to X.

[0563] In some embodiments, W is absent or present, and when W is present, W is selected from -OH, -SH, -NHR4, 1 bit is connected to X.

[0564] In some embodiments, W is selected from -OH, -NHR4 and 1 bit is connected to X.

[0565] In some embodiments, W is selected from -OH and -NHR4, and position 1 is connected to X.

[0566] In some embodiments, X is selected from optionally substituted -(CH2) n1 -、 C6-10 aryl, 5-10 membered heteroaryl and 4-10 membered heterocyclic group, the 1 position is connected to the parent ring, and the 2 position is connected to W; the substituent is selected from 1 or 2 C1-4 alkyl groups, or 2 C1-4 alkyl groups and the carbon atoms to which they are simultaneously connected form a C3-6 cycloalkyl group.

[0567] In some embodiments, X is selected from optionally substituted Position 1 is connected to the parent ring, and position 2 is connected to W; the substituent is selected from 1 or 2 C1-4 alkyl groups (such as methyl), or 2 C1-4 alkyl groups (such as methyl) and the carbon atoms to which they are simultaneously connected form a C3-6 cycloalkyl group (such as cyclopropyl).

[0568] In some embodiments, X is selected from Position 1 is connected to the mother ring, and position 2 is connected to W.

[0569] In some embodiments, X is selected from Position 1 is connected to the mother ring, and position 2 is connected to W.

[0570] In some embodiments, when W is absent, X is selected from 1 is connected to the parent ring; when W exists, X is selected from Position 1 is connected to the mother ring, and position 2 is connected to W.

[0571] In some embodiments, W is selected from -OH, -NHR4 and Position 1 is connected to X, position 2 is connected to L4 or L3; X is selected from Position 1 is connected to the parent ring, and position 2 is connected to W. In some embodiments, R4 and R5 are each independently selected from H, C1-4 alkyl, and C3-6 cycloalkyl.

[0572] In some embodiments, each R4 is independently selected from H, C1-4 alkyl, and C3-6 cycloalkyl, and R5 is H.

[0573] In some embodiments, each R4 is independently selected from H, methyl, ethyl, isopropyl, n-propyl, tert-butyl, and cyclopropyl, and R5 is H.

[0574] In some embodiments, R 5a and R 5b Each independently selected from H and C1-4 alkyl;

[0575] In some embodiments, R 5a and R 5b are each independently selected from H and methyl;

[0576] In some embodiments, each R 7 Independently selected from H and C1-4 alkyl;

[0577] In some embodiments, R 7 For H.

[0578] In some embodiments, n is 1, 2, or 3.

[0579] In some embodiments, n is 1.

[0580] In some embodiments, n1 is 1, 2, 3, or 4.

[0581] In some embodiments, n2 is 1;

[0582] In some embodiments, n3 is 0.

[0583] In some embodiments, the compound represented by Formula II is selected from any one of the following compounds:

[0584]

[0585]

[0586]

[0587] In the third aspect of the present disclosure, the present disclosure provides a drug-linker conjugate shown in formula III,

[0588]

[0589] or a stereoisomer of the drug-linker conjugate, a prodrug thereof, a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable solvate thereof,

[0590] in:

[0591] R1 and R2 are each independently selected from H, halogen, -OH, optionally substituted C1-6 alkyl or optionally substituted C1-6 alkoxy, or,

[0592] R1 and R2 together with the carbon atoms to which they are attached form a 5-7 membered carbocyclic ring or a 5-7 membered heterocyclic ring, wherein the heterocyclic ring contains one or more O, S, N, carbonyl, sulfoxide or sulfone groups or any combination thereof;

[0593] R3 is selected from H, halogen, -OH, -NH2, optionally substituted C1-6 alkyl and optionally substituted C1-6 alkoxy, or,

[0594] R3 and X together with the carbon atom to which they are attached form a 5-7 membered carbocyclic ring or a 5-7 membered heterocyclic ring containing one or more O, S, N, carbonyl, sulfoxide or sulfone groups or any combination thereof, or

[0595] R3 and R2 together with the carbon atom to which they are attached form a 5-7 membered carbocyclic ring or a 5-7 membered heterocyclic ring, wherein the heterocyclic ring contains one or more O, S, N, carbonyl, sulfoxide or sulfone groups or any combination thereof;

[0596] W is absent or present. When W is present, W is selected from -O-, -S-, -NR4-, Bit 1 is connected to X, bit 2 is connected to L4 or L3;

[0597] X is selected from a direct bond, an optionally substituted -O-(CH2) n3 -、-N(R4)-(CH2) n3 -、-S-(CH2) n3 -, carbonyl-(CH2) n3 、-SO2-(CH2) n3 -、 -(CH2) n1 -, C3-6 cycloalkyl, C6-10 aryl, 5-10 membered heteroaryl and 4-10 membered heterocyclic group, the 1 position is connected to the parent ring, and the 2 position is connected to W or L4; the substituent is selected from one or more C1-4 alkyl, C3-6 cycloalkyl, or multiple C1-4 alkyl and the carbon atoms to which they are simultaneously connected together form a C3-6 cycloalkyl;

[0598] Each M is independently selected from a direct bond, -CR 5a R 5b -;

[0599] R4, R5, R 5a 、R 5b , R6 and R7 are each independently selected from H, optionally substituted C1-4 alkyl, optionally substituted C1-4 alkoxy and optionally substituted C3-6 cycloalkyl;

[0600] n, n', n1, n2, n3 are each independently selected from any integer between 0 and 6;

[0601] L1 is selected from:

[0602]

[0603]

[0604]

[0605] Each Z is independently selected from a direct bond, a carbon-carbon triple bond, a carbon-carbon double bond, a C6-10 aryl group, a 5-10 membered heteroaryl group, an amide group, a sulfonamide group, an imine group, or CF2; Rx and Ry are each independently selected from H or a C1-4 alkyl group; each m is independently selected from 0, 1, 2, 3, 4, 5, or 6; y1, y2, y3, and y4 are each independently selected from any integer between 0 and 20; position 1 is connected to Lg, and position 2 is connected to L2 or L3;

[0606] L2 is absent or present. When L2 is present, L2 is selected from:

[0607]

[0608]

[0609] y1, y2, y3, and y4 are each independently selected from any integer between 0 and 20, with position 1 connected to L1 and position 2 connected to L3;

[0610] L3 is selected from an amino acid residue or a short peptide consisting of 2-10 amino acid residues;

[0611] L4 is absent or present. When L4 is present, L4 is selected from Bit 1 is connected to L3, bit 2 is connected to W or X;

[0612] Lg is a leaving group, which is selected from halogen, sulfone, tertiary amine salt (Me3N + 、Et3N +), diazonium base, -OMs, MeSO2-, CF3SO3-.

[0613] In some embodiments, R1 and R2 are each independently selected from H, halogen, and C1-4 alkyl.

[0614] In some embodiments, R1 and R2, together with the carbon atom 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.

[0615] In some embodiments, R1 is selected from H, halogen, and R2 is selected from H, C1-4 alkyl.

[0616] In some embodiments, R1 and R2 and the carbon atom to which they are attached form The dotted line indicates the position where the heterocyclic ring is fused to the benzene ring.

[0617] In some embodiments, R1 is H or F, and R2 is H or methyl.

[0618] In some embodiments, R1 is F, R2 is methyl, or R1 and R2 together with the carbon atom to which they are attached form

[0619] In some embodiments, R1 is F and R2 is methyl.

[0620] In some embodiments, R1 and R2 together with the carbon atom to which they are attached form

[0621] In some embodiments, R3 is selected from H, C1-4 alkyl.

[0622] In some embodiments, R3 and X, together with the carbon atom to which they are attached, form a 5-6 membered carbocyclic ring.

[0623] In some embodiments, R3 is H or R3 and X together with the carbon atom to which they are attached form The dotted lines indicate where the carbocyclic ring is fused to the benzene and pyridine rings.

[0624] In some embodiments, R3 is H.

[0625] In some embodiments, R3 and X together with the carbon atom to which they are attached form The dotted lines indicate where the carbocyclic ring is fused to the benzene and pyridine rings.

[0626] In some embodiments, W is absent or present, and when W is present, W is selected from -O-, -S-, -NR4-, Bit 1 is connected to X, and bit 2 is connected to L4 or L3.

[0627] In some embodiments, W is absent or present, and when W is present, W is selected from -O-, -S-, -NR4-, Bit 1 is connected to X, and bit 2 is connected to L4 or L3.

[0628] In some embodiments, W is selected from -O-, -NR4-, Bit 1 is connected to X, and bit 2 is connected to L4 or L3.

[0629] In some embodiments, W is selected from -O-, -NR4-, position 1 is connected to X, and position 2 is connected to L4 or L3.

[0630] In some embodiments, X is selected from optionally substituted -(CH2) n1 -、 C6-10 aryl, 5-10 membered heteroaryl and 4-10 membered heterocyclic group, the 1 position is connected to the parent ring, and the 2 position is connected to W or L4; the substituent is selected from 1 or 2 C1-4 alkyl groups, or 2 C1-4 alkyl groups and the carbon atoms to which they are simultaneously connected form a C3-6 cycloalkyl group.

[0631] In some embodiments, X is selected from optionally substituted Position 1 is connected to the parent ring, and position 2 is connected to W or L4; the substituent is selected from 1 or 2 C1-4 alkyl groups (such as methyl), or 2 C1-4 alkyl groups (such as methyl) and the carbon atoms to which they are simultaneously connected form a C3-6 cycloalkyl group (such as cyclopropyl).

[0632] In some embodiments, X is selected from Position 1 is connected to the mother ring, and position 2 is connected to W or L4.

[0633] In some embodiments, X is selected from Position 1 is connected to the mother ring, and position 2 is connected to W.

[0634] In some embodiments, when W is absent, X is selected from 1 is connected to the parent ring, 2 is connected to L4; when W exists, X is selected from Position 1 is connected to the mother ring, and position 2 is connected to W.

[0635] In some embodiments, W is selected from -O-, -NR4-, Position 1 is connected to X, position 2 is connected to L4 or L3; X is selected from Position 1 is connected to the mother ring, and position 2 is connected to W.

[0636] In some embodiments, R4 and R5 are each independently selected from H, C1-4 alkyl, and C3-6 cycloalkyl.

[0637] In some embodiments, each R4 is independently selected from H, C1-4 alkyl, C3-6 cycloalkyl, and R5 is H.

[0638] In some embodiments, each R4 is independently selected from H, methyl, ethyl, n-propyl, isopropyl, tert-butyl, cyclopropyl, and R5 is H.

[0639] In some embodiments, R 5a 、R 5b Each is independently selected from H, C1-4 alkyl.

[0640] In some embodiments, R 5a 、R 5b Each is independently selected from H, methyl.

[0641] In some embodiments, each R 7 Independently selected from H, C1-4 alkyl.

[0642] In some embodiments, R 7 For H.

[0643] In some embodiments, n is selected from 1, 2, or 3.

[0644] In some embodiments, n is 1.

[0645] In some embodiments, n1 is selected from 1, 2, 3 or 4.

[0646] In some embodiments, n2 is 1.

[0647] In some embodiments, n3 is 0.

[0648] In some embodiments, L1 is selected from 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 group (preferably selected from a direct bond, a carbon-carbon triple bond, a carbon-carbon double bond), Rx, Ry are each independently selected from H, a C1-4 alkyl group, each m is independently selected from 0, 1, 2, 3, 4, 5, 6, y1 is selected from any integer between 1-6 (such as 4, 5, 6), each y2 is independently selected from any integer between 0-15 (such as 6-15), each y3 is independently selected from 1, 2 or 3, each y4 is independently selected from 0 or 1, 1 is independently selected from 1, 2 or 3, Position 1 is connected to Lg, and position 2 is connected 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, a C1-4 alkyl group, each m is independently selected from 0, 1, 2, 3, 4, 5, or 6, y1 is selected from any integer between 1 and 6 (e.g., 4, 5, or 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, each y4 is independently selected from 0 or 1, position 1 is connected to Lg through an S atom, and position 2 is connected to L2 or L3.

[0649] In some embodiments, L1 is selected from m is selected from 2, 3, and 4, y1 is selected from any integer between 1 and 6 (such as 4, 5, and 6), each y2 is independently selected from any integer between 0 and 10 (such as 6 and 10), and each y3 is independently selected from 1 or 2. Position 1 is connected to Lg, and position 2 is connected to L2 or L3.

[0650] In some embodiments, L1 is selected from Position 1 is connected to Lg, and position 2 is connected to L2 or L3.

[0651] In some embodiments, L1 is selected from Position 1 is connected to Lg, and position 2 is connected to L2 or L3.

[0652] In some embodiments, L1 is selected from Position 1 is connected to Lg, and position 2 is connected to L2 or L3.

[0653] In some embodiments, L1 is selected from Position 1 is connected to Lg, and position 2 is connected to L2 or L3.

[0654] In some embodiments, L2 is absent or present, and when L2 is present, L2 is selected from y1 is selected from any integer between 1-6 (such as 4, 5, 6), each y2 is independently selected from any integer between 0-10 (such as 6-10), each y3 is independently selected from 1 or 2, each y4 is independently selected from 0 or 1, position 1 is connected to L1, and position 2 is connected to L3.

[0655] In some embodiments, L2 is absent or present, and when L2 is present, L2 is selected from Bit 1 is connected to L1 and bit 2 is connected to L3.

[0656] In some embodiments, L2 is absent or present, and when L2 is present, L2 is selected from Bit 1 is connected to L1 and bit 2 is connected to L3.

[0657] In some embodiments, L2 is absent or present, and when L2 is present, L2 is selected from Bit 1 is connected to L1 and bit 2 is connected to L3.

[0658] In some embodiments, L2 is absent.

[0659] In some embodiments, L2 is selected from

[0660] In some embodiments, L3 is selected from an amino acid residue or a short peptide consisting of 2-10 amino acid residues; the amino acid residue is selected from a natural amino acid residue, a non-natural amino acid residue, an AA 1 The indicated amino acid residues or stereoisomers thereof.

[0661] In some embodiments, L3 is selected from the group consisting of amino acid residues Val, D-Val, Cit, Phe, Lys, Lys(Ac), Leu, Gly, Ala, Asn, Asp, Arg, AA 1 or 2-10 selected from Val, Cit, Phe, Lys, D-Val, Leu, Gly, Ala, Asn, Asp, AA 1 A short peptide composed of amino acid residues.

[0662] In some embodiments, L3 is selected from 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-AA1 、Ala-AA 1 、Gly-AA 1 AA 1 -Gly, Ala-Ala-Ala, Ala-Ala-Asn, Ala-Ala-Asp, Val-AA 1 -Gly, Ala-AA 1 -Gly, Gly-AA 1 -Gly, Lys-Ala-Ala-Asn, Lys-Ala-Ala-Asp, Gly-Phe-Gly, Gly-Gly-Phe-Gly, D-Val-Leu-Lys, Gl y-Gly-Arg, Ala-Ala-Asn, Gly-Gly-Phe, Val-Lys-Gly, Val-Lys-Gly-Gly, Val-Lys, Lys-Ala-Asn.

[0663] In some embodiments, L3 is selected from AA 1 AA 1 -Gly, Val-Cit, Val-AA 1 -Gly, AA 1 -Ala-Asn, Gly-Gly-Phe-Gly.

[0664] In some embodiments, L3 is selected from AA 1 、Val-AA 1 -Gly.

[0665] In some embodiments, L3 is selected from Val-AA 1 -Gly.

[0666] In some embodiments, L3 is selected from X - Selected from halogen ions, carboxylate ions, sulfate ions, hydrogen sulfate ions, OH - , bit 1 is connected to L1 or L2, bit 2 is connected to L4 or W.

[0667] In some embodiments, L3 is selected from X - Selected from halogen ions, carboxylate ions, sulfate ions, hydrogen sulfate ions, OH - , bit 1 is connected to L1 or L2, bit 2 is connected to L4 or W.

[0668] In some embodiments, L3 is selected from X - Selected from halogen ions, carboxylate ions, sulfate ions, hydrogen sulfate ions, OH - , bit 1 is connected to L1 or L2, bit 2 is connected to L4 or W.

[0669] In some embodiments, L3 is selected from Bit 1 is connected to L1 or L2, and bit 2 is connected to L4 or W.

[0670] In some embodiments, L3 is selected from Bit 1 is connected to L1 or L2, and bit 2 is connected to L4 or W.

[0671] In some embodiments, AA 1 The structures of the indicated amino acid residues are shown below,

[0672]

[0673] in:

[0674] R a 、R b Each independently selected from H, And R a 、R b Not at the same time H;

[0675] Or, R a With R b Together with the carbon atoms to which they are attached, they form a 4-10 membered heterocyclic ring, which is optionally substituted by one or more R 0 replaced by;

[0676] r、r 1 are each independently selected from any integer from 0 to 20;

[0677] R m1 、R n1 Each independently selected from H, C1-6 alkyl, C3-6 cycloalkyl, -COOR x1 ;

[0678] R x1 Selected from C1-6 alkyl;

[0679] Or, R m1 With R n1 Together with the nitrogen atom to which they are attached, they form a 4-10 membered heterocyclic ring, which is optionally substituted by one or more R 0’ replaced by;

[0680] R z Selected from C1-6 alkyl;

[0681] R 0 、R 0’ Each independently selected from C1-6 alkyl, C3-6 cycloalkyl, -NR m2 R n2 , a 4-10 membered heterocyclic group optionally substituted by a C1-6 alkyl group;

[0682] R m2 、R n2 Each is independently selected from H, C1-6 alkyl.

[0683] In some embodiments, R a 、R b Among them, any one is H, and the other is selected from

[0684] In some embodiments, R a 、R b Among them, any one is H, and the other is selected from

[0685] In some embodiments, R a With R b Together with the carbon atoms they are connected to, they form the 0 Substituted 5-6 membered heterocycle.

[0686] In some embodiments, R a With R b Together with the carbon atoms they are connected to, they form the 0 a substituted piperidine ring or a piperazine ring.

[0687] In some embodiments, R a With R b Together with the carbon atoms they are connected to, they form the 0 Substituted piperidine ring.

[0688] In some embodiments, R a With R b Together with the carbon atoms they are connected to, they form Carbon atom No. 1 is R a and R b carbon atoms that are linked together.

[0689] In some embodiments, R a With R b Together with the carbon atoms they are connected to, they form Carbon atom No. 1 is R aand R b carbon atoms that are linked together.

[0690] In some embodiments, r, r 1 Each is independently selected from 0, 1, 2, 3, 4, and 5.

[0691] In some embodiments, r, r 1 Each is independently selected from 0, 4.

[0692] In some embodiments, r, r 1 If one of them is 0, the other one is 4.

[0693] In some embodiments, R m1 、R n1 Each is independently selected from H, methyl, ethyl, n-propyl, n-butyl, -COOCH3, -COOCH2CH3, -COOCH2CH2CH3, -COOCH(CH3)2, -COOC(CH3)3, -COOCH2CH2CH2CH3.

[0694] In some embodiments, R m1 、R n1 Each is independently selected from H, C1-6 alkyl, C3-6 cycloalkyl, and tert-butyloxycarbonyl.

[0695] In some embodiments, R m1 、R n1 Each is independently selected from H, C1-6 alkyl.

[0696] In some embodiments, R m1 、R n1 Each is independently selected from H, methyl, ethyl, and n-propyl.

[0697] In some embodiments, r, r 1 In the example, r is 4, r 1 When R is 0, m1 、R n1 Each is independently selected from H, C1-6 alkyl (such as H, methyl); r is 0, r 1 When R is 4, m1 、R n1 Each is independently selected from C1-6 alkyl (such as methyl, ethyl, n-propyl), preferably selected from C2-6 alkyl (such as ethyl, n-propyl).

[0698] In some embodiments, R m1 With R n1 Together with the nitrogen atom to which they are commonly attached, they form an optionally R 0’ Substituted 5-6 membered heterocycle.

[0699] In some embodiments, R m1 With R n1 Together with the nitrogen atom to which they are commonly attached, they form an optionally R 0’ a substituted piperidine ring or a piperazine ring.

[0700] In some embodiments, R m1 With R n1 Together with the nitrogen atoms they are connected to, they form Nitrogen atom No. 1 is R m1 and R n1 Commonly linked nitrogen atoms.

[0701] In some embodiments, R z It is a methyl group.

[0702] In some embodiments, R 0 、R 0’ Each independently selected from C1-6 alkyl, -NR m2 R n2 , a 5-6 membered heterocyclic group optionally substituted by a C1-6 alkyl group.

[0703] In some embodiments, R 0 The 5-6 membered heterocyclic group is selected from C1-6 alkyl and substituted by C1-6 alkyl, wherein the 5-6 membered heterocyclic group is selected from piperidinyl and piperazinyl.

[0704] In some embodiments, R 0 It is selected from methyl, ethyl, and a 5-6 membered heterocyclic group substituted by a methyl group, wherein the 5-6 membered heterocyclic group is a piperidinyl group.

[0705] In some embodiments, R 0 The 5- to 6-membered heterocyclic group is selected from methyl and methyl-substituted 5- to 6-membered heterocyclic group, wherein the 5- to 6-membered heterocyclic group is piperidinyl.

[0706] In some embodiments, R 0 Selected from methyl, ethyl,

[0707] In some embodiments, R 0 Selected from methyl,

[0708] In some embodiments, R 0’ Selected from C1-6 alkyl, -NR m2 R n2 .

[0709] In some embodiments, R 0’ Selected from methyl, -NR m2 R n2 .

[0710] In some embodiments, R m2 、R n2 It is a methyl group.

[0711] In some embodiments, AA 1 The amino acid residues shown are selected from

[0712] In some embodiments, AA 1 The amino acid residues shown are selected from

[0713] In some embodiments, AA 1 The amino acid residues shown are selected from

[0714] In some embodiments, AA 1 The amino acid residues shown are selected from

[0715] In some embodiments, L4 is absent or present, and when L4 is present, L4 is selected from Bit 1 is connected to L3, and bit 2 is connected to W or X.

[0716] In some embodiments, L4 is absent or present, and when L4 is present, L4 is Bit 1 is connected to L3, and bit 2 is connected to W or X.

[0717] In some embodiments, L4 is absent.

[0718] In some embodiments, L4 is selected from Bit 1 is connected to L3, and bit 2 is connected to W or X.

[0719] In some embodiments, L4 is selected from Bit 1 is connected to L3, and bit 2 is connected to W or X.

[0720] In some embodiments, Lg is selected from F, Cl, MeSO2-.

[0721] In some embodiments, Lg is selected from F, MeSO2-.

[0722] In some embodiments, The structure is selected from the following:

[0723]

[0724]

[0725]

[0726]

[0727] Among them, bit 1 is connected to Lg and bit 2 is connected to W.

[0728] In some embodiments, The structure is selected from the following:

[0729]

[0730]

[0731]

[0732]

[0733] Among them, bit 1 is connected to L4; when L4 does not exist, bit 1 is connected to L3.

[0734] In some embodiments, the drug-linker conjugate has the structure of Formula III-(1):

[0735]

[0736] wherein L1, L2, L3, L4, X, R1, R2, R3, R4 and Lg have the meanings provided above and in any embodiment specifically described herein.

[0737] In some embodiments, the drug-linker conjugate has a structure represented by Formula III-(1A) or III-(1B):

[0738]

[0739] wherein L2, L3, L4, X, R1, R2, R3, R4 and Lg have the meanings provided above and in any of the embodiments specifically described herein.

[0740] In some embodiments, the drug-linker conjugate has the structure of Formula III-(2):

[0741]

[0742] wherein L1, L2, L3, L4, X, R1, R2, R3 and Lg have the meanings provided above and in any embodiment specifically described herein.

[0743] In some embodiments, the drug-linker conjugate has the structure of Formula III-(2A) or III-(2B):

[0744]

[0745] wherein L2, L3, L4, X, R1, R2, R3 and Lg have the meanings provided above and in any of the embodiments specifically described herein.

[0746] In some embodiments, the drug-linker conjugate has the structure of Formula III-(3):

[0747]

[0748] wherein L1, L2, L3, L4, X, R1, R2, R3, R4, R5, n, and Lg have the meanings provided above and in any of the embodiments specifically described herein.

[0749] In some embodiments, the drug-linker conjugate has a structure represented by Formula III-(3A) or III-(3B):

[0750]

[0751] wherein Lg, L2, L3, L4, X, R1, R2, R3, R4 and Lg have the meanings provided above and in any of the embodiments specifically described herein.

[0752] In some embodiments, the drug-linker conjugate has the structure of Formula III-A:

[0753]

[0754] Among them, Lg, X, R1, R2, R3, R a 、R b and q have the meanings provided above and in any embodiments specifically described herein.

[0755] In some embodiments, the drug-linker conjugate has the structure shown in Formula III-B:

[0756]

[0757] Among them, Lg, X, R1, R2, R3, R a 、R b and q have the meanings provided above and in any embodiments specifically described herein.

[0758] In some embodiments, the drug-linker conjugate represented by Formula III is selected from the following structures:

[0759]

[0760]

[0761]

[0762]

[0763]

[0764]

[0765]

[0766]

[0767]

[0768]

[0769]

[0770]

[0771]

[0772]

[0773]

[0774]

[0775]

[0776]

[0777]

[0778]

[0779]

[0780]

[0781] In some embodiments, compounds are provided wherein the L1-L2-L3 unit of the drug-linker conjugate of Formula III has been partially cleaved, thereby leaving the drug moiety bonded to the amino acid residue.

[0782] In some embodiments, the partially released free drug is a compound of Formula III-(A):

[0783]

[0784] or a stereoisomer or mixture of stereoisomers thereof, or a pharmaceutically acceptable salt thereof, wherein L4, X, W, R1, R2 and R3 have the meanings provided above and in any of the embodiments specifically described herein.

[0785] In some embodiments, The structure is selected from the following:

[0786]

[0787]

[0788]

[0789]

[0790]

[0791] The present disclosure also provides a linker in a ligand-drug conjugate, the structure of which comprises the following fragments:

[0792]

[0793] Among them, position 2 is connected to the bioactive molecule fragment; (position 1 is connected to the ligand end, for example, when it is connected to the linker unit (such as L2), and then connected to the ligand or targeting portion (Tb) through the extension unit (such as L1); or, when the linker unit does not exist, it is directly connected to the extension unit and then further connected to the ligand);

[0794] The definitions of L3 and L4 are as described in any embodiment of the present disclosure.

[0795] In some embodiments, the structure of the linker in the ligand drug conjugate is as follows:

[0796]

[0797] Among them, position 1 is connected to the ligand or targeting portion that binds to the target, and position 2 is connected to the bioactive molecular fragment;

[0798] The definitions of L1, L2, L3, and L4 are as described in any embodiment of the present disclosure;

[0799] Preferably, the target-binding ligand or targeting portion and the bioactive molecule fragment are defined as Tb and D in any embodiment of the present disclosure, respectively.

[0800] In some embodiments, the linker is connected to the antibody or antigen-binding fragment thereof at position 1 and to the biologically active molecule fragment at position 2.

[0801] In some embodiments, the linker is connected to a cysteine ​​or lysine on the antibody or antigen-binding fragment thereof at position 1, and is connected to a biologically active molecule fragment at position 2;

[0802] Preferably, the antibody or antigen-binding fragment thereof is as described in any embodiment of the present disclosure.

[0803] In the fourth aspect of the present disclosure, the present disclosure further provides a linker represented by formula III-1:

[0804] Lg-L1-L2-L3-L4-Lg1

[0805] III-1

[0806] Lg, L1, L2, L3 and L4 have the meanings provided above and in any of the embodiments specifically described herein; Lg1 is a leaving group when reacting with a drug molecule.

[0807] In some embodiments, Lg1 is preferably -OH, or halogen.

[0808] In the fifth aspect of the present disclosure, the present disclosure further provides a linker represented by formula III-2:

[0809] Lg-L1-L2-L3-Lg2

[0810] III-2

[0811] Lg, L1, L2 and L3 have the meanings provided above and in any of the embodiments specifically described herein; Lg2 is a leaving group when reacting with L4 or a fragment containing a drug molecule.

[0812] In some embodiments, Lg2 is preferably -OH, or halogen.

[0813] In a sixth aspect of the present disclosure, the present disclosure provides a method for preparing the compound (drug molecule, bioactive molecule) represented by Formula II and the drug-linker conjugate represented by Formula III.

[0814] In one aspect, the present disclosure provides a method for preparing a compound (drug molecule, biologically active molecule) represented by Formula II. Specifically:

[0815] Compound (VI) can be obtained by alkylating compound (IV) with compound (V) in the presence of an iron compound or other related Minisci reactions.

[0816]

[0817] Alternatively, compound (IV) is oxidized to compound (VII), which is then treated with a phosphorus oxyhalide to produce compound (VIII), a halogenated product of compound (IV). Compound (VIII) can be converted to compound (VI) through various reactions, such as Heck reaction, Suzuki reaction, Buchwald reaction, Nigishi reaction, Stille reaction, and the like.

[0818]

[0819] In compound (VI), R 11 Different or more complex molecules can be synthesized through a variety of chemical transformations, such as oxidation, reduction, substitution, and other methods commonly seen in textbooks.

[0820] Alternatively, compound (VI) and compound (IV) can also be obtained by a ring-closure reaction of compound (IX) and compound (X) as shown below.

[0821]

[0822] In another aspect, the present disclosure provides a method for preparing the fragments represented by Formula III-1 and Formula III-2. Specifically:

[0823] Compound III-a can react with the functional group Fg1 of compound III-b to obtain compound III-c, and compound III-c can react with the functional group Fg2 of compound III-b to obtain compound III-c. a Compound III-d can be obtained, and similar transformations can be used to obtain compounds of formula III-2 and III-1. a , Lg b , Lg1, Lg2 are reactive leaving groups, such as -OH, Or halogen, etc.

[0824]

[0825] In yet another aspect, the present disclosure provides a method for preparing a drug-linker conjugate represented by Formula III. Specifically:

[0826] The target product can be obtained by coupling fragments B and C as shown in the following reaction formula. The coupling product is subjected to some commonly used chemical modifications, such as oxidation and deprotection, to obtain the drug-linker conjugate shown in general formula III.

[0827]

[0828] In a seventh aspect of the present disclosure, the present disclosure provides a method for preparing the aforementioned ligand-drug conjugate, comprising:

[0829] Conjugate Tb with the drug linker shown in formula III Carry out the coupling reaction in a suitable solvent and conditions;

[0830] in:

[0831] Tb has the meaning provided above and in any embodiment specifically described herein;

[0832] R1, R2, R3, X, W, L1, L2, L3, L4, and Lg have the meanings provided above and in any of the embodiments specifically described herein.

[0833] In some embodiments, the method comprises conjugating Tb with a drug linker of formula III The step of forming a C-S bond by coupling reaction is carried out in a suitable solvent and conditions.

[0834] In some embodiments, the ratio of the amount of Tb to the amount of the drug-linker conjugate is 1:(1-20), such as 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).

[0835] In some embodiments, the coupling reaction is carried out in water and / or an organic solvent.

[0836] In some embodiments, the organic solvent is selected from N,N-dimethylformamide, dimethyl sulfoxide, N-methylpyrrolidone, nitriles (such as acetonitrile), alcohols (such as methanol, ethanol) or any combination thereof.

[0837] In some embodiments, the method further comprises the step of purifying the coupling product.

[0838] In some embodiments, the coupled product is purified by chromatography.

[0839] In some embodiments, the chromatography method comprises one or more of ion exchange chromatography, hydrophobic chromatography, reverse phase chromatography, or affinity chromatography.

[0840] 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 thereof comprises the following complementarity determining regions (CDRs):

[0841] HCDR1 or a variant thereof, HCDR2 or a variant thereof, and HCDR3 or a variant thereof contained in the heavy chain variable region (VH) shown in SEQ ID NO: 3 or 23; and / or

[0842] LCDR1 or a sequence variant thereof, LCDR2 or a sequence variant thereof, and LCDR3 or a sequence variant thereof contained in the light chain variable region (VL) shown in SEQ ID NO: 13 or 33.

[0843] In certain embodiments, the antibody or antigen-binding fragment thereof comprises HCDR1 or a variant of its sequence, HCDR2 or a variant of its sequence, and HCDR3 or a variant of its sequence contained in the VH shown in SEQ ID NO: 3; and / or LCDR1 or a variant of its sequence, LCDR2 or a variant of its sequence, and LCDR3 or a variant of its sequence contained in the VL shown in SEQ ID NO: 13.

[0844] In certain embodiments, the antibody or antigen-binding fragment thereof comprises HCDR1 or a variant of its sequence, HCDR2 or a variant of its sequence, and HCDR3 or a variant of its sequence contained in the VH shown in SEQ ID NO: 3; and / or LCDR1 or a variant of its sequence, LCDR2 or a variant of its sequence, and LCDR3 or a variant of its sequence contained in the VL shown in SEQ ID NO: 33.

[0845] In certain embodiments, the antibody or antigen-binding fragment thereof comprises HCDR1 or a variant of its sequence, HCDR2 or a variant of its sequence, and HCDR3 or a variant of its sequence contained in the VH shown in SEQ ID NO: 23; and / or LCDR1 or a variant of its sequence, LCDR2 or a variant of its sequence, and LCDR3 or a variant of its sequence contained in the VL shown in SEQ ID NO: 33.

[0846] In certain embodiments, the antibody or antigen-binding fragment thereof comprises HCDR1 or a variant of its sequence, HCDR2 or a variant of its sequence, and HCDR3 or a variant of its sequence contained in the VH shown in SEQ ID NO: 23; and / or LCDR1 or a variant of its sequence, LCDR2 or a variant of its sequence, and LCDR3 or a variant of its sequence contained in the VL shown in SEQ ID NO: 13.

[0847] In certain preferred embodiments, the sequence variant is a CDR having one or several amino acid substitutions, deletions or additions (eg, 1, 2 or 3 amino acid substitutions, deletions or additions) compared to its source CDR.

[0848] In certain preferred embodiments, the substitutions are conservative substitutions.

[0849] Preferably, the CDRs are defined according to the AbM, Chothia, Kabat or IMGT numbering systems.

[0850] In certain embodiments, the VH and / or VL of the antibody or antigen-binding fragment thereof comprises a framework region (FR) from a human immunoglobulin.

[0851] 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 both human 2Ig B7H3 and human 4Ig B7H3, and preferentially binds to human 4Ig B7H3. In certain embodiments, the antibody or antigen-binding fragment thereof binds to human 4Ig B7H3 but does not bind to human 2Ig B7H3.

[0852] In one aspect, the present 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).

[0853] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure comprise the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the IMGT numbering system:

[0854] (a) a heavy chain variable region (VH) comprising the following three CDRs: a HCDR1 having a sequence of SEQ ID NO: 10 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, a HCDR2 having a sequence of SEQ ID NO: 11 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, a HCDR3 having a sequence of SEQ ID NO: 12 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto; and / or,

[0855] a light chain variable region (VL) comprising the following three CDRs: LCDR1 having a sequence of SEQ ID NO: 20 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, LCDR2 having a sequence of GTF or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, LCDR3 having a sequence of SEQ ID NO: 22 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto;

[0856] (b) a heavy chain variable region (VH) comprising the following three CDRs: a HCDR1 having a sequence of SEQ ID NO: 30 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, a HCDR2 having a sequence of SEQ ID NO: 31 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, a HCDR3 having a sequence of SEQ ID NO: 32 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto; and / or,

[0857] a light chain variable region (VL) comprising the following three CDRs: LCDR1 having a sequence of SEQ ID NO: 40 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, LCDR2 having a sequence of GAS or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, LCDR3 having a sequence of SEQ ID NO: 42 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto;

[0858] (c) a heavy chain variable region (VH) comprising the following three CDRs: a HCDR1 having a sequence of SEQ ID NO: 10 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, a HCDR2 having a sequence of SEQ ID NO: 11 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, a HCDR3 having a sequence of SEQ ID NO: 12 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto; and / or,

[0859] a light chain variable region (VL) comprising the following three CDRs: LCDR1 having a sequence of SEQ ID NO: 40 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, LCDR2 having a sequence of GAS or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, LCDR3 having a sequence of SEQ ID NO: 42 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto;

[0860] or

[0861] (d) a heavy chain variable region (VH) comprising the following three CDRs: a HCDR1 having a sequence of SEQ ID NO: 30 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, a HCDR2 having a sequence of SEQ ID NO: 31 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, a HCDR3 having a sequence of SEQ ID NO: 32 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto; and / or,

[0862] A light chain variable region (VL) comprising the following three CDRs: LCDR1 having a sequence of SEQ ID NO: 20 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared thereto, LCDR2 having a sequence of GTF or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared thereto, LCDR3 having a sequence of SEQ ID NO: 22 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared thereto.

[0863] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure comprise the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the IMGT numbering system:

[0864] (a) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 with a sequence of SEQ ID NO: 10, HCDR2 with a sequence of SEQ ID NO: 11, and HCDR3 with a sequence of SEQ ID NO: 12; and / or,

[0865] a light chain variable region (VL) comprising the following three CDRs: LCDR1 of SEQ ID NO: 20, LCDR2 of GTF, and LCDR3 of SEQ ID NO: 22;

[0866] (b) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 with a sequence of SEQ ID NO: 10, HCDR2 with a sequence of SEQ ID NO: 11, and HCDR3 with a sequence of SEQ ID NO: 12; and / or,

[0867] a light chain variable region (VL) comprising the following three CDRs: LCDR1 of SEQ ID NO: 40, LCDR2 of GAS, and LCDR3 of SEQ ID NO: 42;

[0868] (c) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 with a sequence of SEQ ID NO: 30, HCDR2 with a sequence of SEQ ID NO: 31, and HCDR3 with a sequence of SEQ ID NO: 32; and / or,

[0869] a light chain variable region (VL) comprising the following three CDRs: LCDR1 of SEQ ID NO: 40, LCDR2 of GAS, and LCDR3 of SEQ ID NO: 42;

[0870] or

[0871] (d) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 with a sequence of SEQ ID NO: 30, HCDR2 with a sequence of SEQ ID NO: 31, and HCDR3 with a sequence of SEQ ID NO: 32; and / or,

[0872] The light chain variable region (VL) comprises the following three CDRs: LCDR1 with the sequence of SEQ ID NO: 20, LCDR2 with the sequence of GTF, and LCDR3 with the sequence of SEQ ID NO: 22.

[0873] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure comprise the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the Chothia numbering system:

[0874] (a) a heavy chain variable region (VH) comprising the following three CDRs: a HCDR1 having a sequence of SEQ ID NO: 4 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, a HCDR2 having a sequence of SEQ ID NO: 5 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, a HCDR3 having a sequence of SEQ ID NO: 6 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto; and / or,

[0875] a light chain variable region (VL) comprising the following three CDRs: LCDR1 having a sequence of SEQ ID NO: 14 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, LCDR2 having a sequence of SEQ ID NO: 15 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, LCDR3 having a sequence of SEQ ID NO: 16 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto;

[0876] (b) a heavy chain variable region (VH) comprising the following three CDRs: a HCDR1 having a sequence of SEQ ID NO: 4 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) therefrom, a HCDR2 having a sequence of SEQ ID NO: 5 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) therefrom, a HCDR3 having a sequence of SEQ ID NO: 6 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) therefrom; and / or,

[0877] a light chain variable region (VL) comprising the following three CDRs: LCDR1 having a sequence of SEQ ID NO: 34 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, LCDR2 having a sequence of SEQ ID NO: 35 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, LCDR3 having a sequence of SEQ ID NO: 36 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto;

[0878] (c) a heavy chain variable region (VH) comprising the following three CDRs: a HCDR1 having a sequence of SEQ ID NO: 24 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, a HCDR2 having a sequence of SEQ ID NO: 25 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, a HCDR3 having a sequence of SEQ ID NO: 26 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto; and / or,

[0879] a light chain variable region (VL) comprising the following three CDRs: LCDR1 having a sequence of SEQ ID NO: 34 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, LCDR2 having a sequence of SEQ ID NO: 35 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, LCDR3 having a sequence of SEQ ID NO: 36 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto;

[0880] or

[0881] (d) a heavy chain variable region (VH) comprising the following three CDRs: a HCDR1 having a sequence of SEQ ID NO: 24 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, a HCDR2 having a sequence of SEQ ID NO: 25 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, a HCDR3 having a sequence of SEQ ID NO: 26 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto; and / or,

[0882] A light chain variable region (VL) comprising the following three CDRs: LCDR1 having a sequence of SEQ ID NO: 14 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared thereto, LCDR2 having a sequence of SEQ ID NO: 15 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared thereto, LCDR3 having a sequence of SEQ ID NO: 16 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared thereto.

[0883] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure comprise the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the chothia numbering system:

[0884] (a) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 of SEQ ID NO: 4, HCDR2 of SEQ ID NO: 5, and HCDR3 of SEQ ID NO: 6; and / or,

[0885] a light chain variable region (VL) comprising the following three CDRs: LCDR1 of SEQ ID NO: 14, LCDR2 of SEQ ID NO: 15, and LCDR3 of SEQ ID NO: 16;

[0886] (b) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 of SEQ ID NO: 24, HCDR2 of SEQ ID NO: 25, and HCDR3 of SEQ ID NO: 26; and / or,

[0887] a light chain variable region (VL) comprising the following three CDRs: LCDR1 of SEQ ID NO: 34, LCDR2 of SEQ ID NO: 35, and LCDR3 of SEQ ID NO: 36;

[0888] (c) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 of SEQ ID NO: 4, HCDR2 of SEQ ID NO: 5, and HCDR3 of SEQ ID NO: 6; and / or,

[0889] a light chain variable region (VL) comprising the following three CDRs: LCDR1 of SEQ ID NO: 34, LCDR2 of SEQ ID NO: 35, and LCDR3 of SEQ ID NO: 36;

[0890] or

[0891] (d) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 of SEQ ID NO: 24, HCDR2 of SEQ ID NO: 25, and HCDR3 of SEQ ID NO: 26; and / or,

[0892] The light chain variable region (VL) comprises the following three CDRs: LCDR1 with the sequence of SEQ ID NO: 14, LCDR2 with the sequence of SEQ ID NO: 15, and LCDR3 with the sequence of SEQ ID NO: 16.

[0893] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure comprise the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the Kabat numbering system:

[0894] (a) a heavy chain variable region (VH) comprising the following three CDRs: a HCDR1 having a sequence of SEQ ID NO: 7 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) therefrom, a HCDR2 having a sequence of SEQ ID NO: 8 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) therefrom, a HCDR3 having a sequence of SEQ ID NO: 9 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) therefrom; and / or,

[0895] a light chain variable region (VL) comprising the following three CDRs: LCDR1 having a sequence of SEQ ID NO: 17 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, LCDR2 having a sequence of SEQ ID NO: 18 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, LCDR3 having a sequence of SEQ ID NO: 19 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto;

[0896] (b) a heavy chain variable region (VH) comprising the following three CDRs: a HCDR1 having a sequence of SEQ ID NO: 27 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, a HCDR2 having a sequence of SEQ ID NO: 28 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, a HCDR3 having a sequence of SEQ ID NO: 29 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto; and / or,

[0897] a light chain variable region (VL) comprising the following three CDRs: LCDR1 having a sequence of SEQ ID NO: 37 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared thereto, LCDR2 having a sequence of SEQ ID NO: 38 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared thereto, LCDR3 having a sequence of SEQ ID NO: 39 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared thereto;

[0898] (c) a heavy chain variable region (VH) comprising the following three CDRs: a HCDR1 having a sequence of SEQ ID NO: 7 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, a HCDR2 having a sequence of SEQ ID NO: 8 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, a HCDR3 having a sequence of SEQ ID NO: 9 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto; and / or,

[0899] a light chain variable region (VL) comprising the following three CDRs: LCDR1 having a sequence of SEQ ID NO: 37 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared thereto, LCDR2 having a sequence of SEQ ID NO: 38 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared thereto, LCDR3 having a sequence of SEQ ID NO: 39 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared thereto;

[0900] or

[0901] (d) a heavy chain variable region (VH) comprising the following three CDRs: a HCDR1 having a sequence of SEQ ID NO: 27 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, a HCDR2 having a sequence of SEQ ID NO: 28 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto, a HCDR3 having a sequence of SEQ ID NO: 29 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) thereto; and / or,

[0902] A light chain variable region (VL) comprising the following three CDRs: LCDR1 having a sequence of SEQ ID NO: 17 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared thereto, LCDR2 having a sequence of SEQ ID NO: 18 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared thereto, LCDR3 having a sequence of SEQ ID NO: 19 or a sequence having one or several amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared thereto.

[0903] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure comprise the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the Kabat numbering system:

[0904] (a) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 of SEQ ID NO: 7, HCDR2 of SEQ ID NO: 8, and HCDR3 of SEQ ID NO: 9; and / or,

[0905] a light chain variable region (VL) comprising the following three CDRs: LCDR1 of SEQ ID NO: 17, LCDR2 of SEQ ID NO: 18, and LCDR3 of SEQ ID NO: 19;

[0906] (b) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 of SEQ ID NO: 27, HCDR2 of SEQ ID NO: 28, and HCDR3 of SEQ ID NO: 29; and / or,

[0907] a light chain variable region (VL) comprising the following three CDRs: LCDR1 of SEQ ID NO: 37, LCDR2 of SEQ ID NO: 38, and LCDR3 of SEQ ID NO: 39;

[0908] (c) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 of SEQ ID NO: 7, HCDR2 of SEQ ID NO: 8, and HCDR3 of SEQ ID NO: 9; and / or,

[0909] a light chain variable region (VL) comprising the following three CDRs: LCDR1 of SEQ ID NO: 37, LCDR2 of SEQ ID NO: 38, and LCDR3 of SEQ ID NO: 39;

[0910] or

[0911] (d) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 of SEQ ID NO: 27, HCDR2 of SEQ ID NO: 28, and HCDR3 of SEQ ID NO: 29; and / or,

[0912] The light chain variable region (VL) comprises the following three CDRs: LCDR1 with the sequence of SEQ ID NO: 17, LCDR2 with the sequence of SEQ ID NO: 18, and LCDR3 with the sequence of SEQ ID NO: 19.

[0913] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure comprise the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein at least one CDR in the heavy chain variable region (VH) and / or light chain variable region (VL) contains a mutation compared to the CDR defined by IMGT, Chothia or Kabat, and the mutation is a substitution, deletion or addition of one or several amino acids or any combination thereof (e.g., substitution, deletion or addition of 1, 2 or 3 amino acids or any combination thereof).

[0914] Preferably, the substitutions described in the present disclosure are conservative substitutions.

[0915] In certain embodiments, the VH of the antibodies or antigen-binding fragments thereof of the present disclosure comprises a framework region (FR) derived from the heavy chain variable region (VH) of a human immunoglobulin, and / or the VL of the antibodies or antigen-binding fragments thereof comprises a framework region (FR) derived from the light chain variable region (VL) of a human immunoglobulin. Thus, in certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure are humanized. In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure are fully human.

[0916] In certain embodiments, the VH of the antibodies or antigen-binding fragments thereof of the present disclosure comprises a framework region (FR) derived from the heavy chain variable region (VH) of a human immunoglobulin, and / or the VL of the antibodies or antigen-binding fragments thereof comprises a framework region (FR) derived from the light chain variable region (VL) of a human immunoglobulin. Thus, in certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure are humanized. In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure are fully human.

[0917] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure comprise:

[0918] (a) a heavy chain framework region of a human immunoglobulin or a variant thereof, which has up to 20 conservative substitutions of amino acids (e.g., up to 20, up to 15, up to 10, or up to 5 conservative substitutions of amino acids; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 conservative substitutions of amino acids) compared to the amino acid sequence encoded by the germline antibody gene from which the variant is derived; and / or

[0919] (b) a light chain framework region of a human immunoglobulin or a variant thereof, which has up to 20 conservative substitutions of amino acids compared to the amino acid sequence encoded by the germline antibody gene from which the variant is derived (e.g., up to 20, up to 15, up to 10 or up to 5 conservative substitutions of amino acids; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 conservative substitutions of amino acids).

[0920] In certain embodiments, the degree of humanization of the antibodies or antigen-binding fragments 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%.

[0921] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure comprise:

[0922] (a) a heavy chain variable region (VH) comprising an amino acid sequence selected from the group consisting of:

[0923] (i) the sequence shown in SEQ ID NO: 3 or 23;

[0924] (ii) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence shown 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

[0925] (iii) 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;

[0926] and / or

[0927] (b) a light chain variable region (VL) comprising an amino acid sequence selected from the group consisting of:

[0928] (iv) the sequence shown in SEQ ID NO: 13 or 33;

[0929] (v) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence shown 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

[0930] (vi) 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: 13 or 33.

[0931] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure comprise the VH set forth in SEQ ID NO:3, and / or the VL set forth in SEQ ID NO:13.

[0932] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure comprise the VH set forth in SEQ ID NO:23, and / or, the VL set forth in SEQ ID NO:33.

[0933] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure comprise the VH set forth in SEQ ID NO: 3, and / or the VL set forth in SEQ ID NO: 33.

[0934] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure comprise the VH set forth in SEQ ID NO:23, and / or the VL set forth in SEQ ID NO:13.

[0935] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure comprise:

[0936] (a) VH of SEQ ID NO: 3 and VL of SEQ ID NO: 13;

[0937] (b) VH of SEQ ID NO: 23 and VL of SEQ ID NO: 33;

[0938] (c) VH of SEQ ID NO: 3 and VL of SEQ ID NO: 33;

[0939] (d) VH of SEQ ID NO: 23 and VL of SEQ ID NO: 13.

[0940] (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 described in any one of groups (a) to (f), respectively; or

[0941] (f) 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 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 any one of groups (a) to (d), respectively. Preferably, the substitutions are conservative substitutions.

[0942] In certain embodiments, the heavy chain of the antibodies or antigen-binding fragments thereof of the present disclosure comprises a heavy chain constant region (CH) of a human immunoglobulin or a variant thereof having up to 50 conservative substitutions of amino acids compared to the wild-type sequence from which it is derived (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 substitutions of amino acids; e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 conservative substitutions of amino acids). In certain embodiments, the light chain of the antibodies or antigen-binding fragments thereof of the present disclosure comprises a light chain constant region (CL) of a human immunoglobulin or a variant thereof having up to 50 conservative substitutions of amino acids compared to the wild-type sequence from which it is derived (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 substitutions of amino acids; e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 conservative substitutions of amino acids).

[0943] In some embodiments, the constant region is altered, for example, mutated, to modify the properties of the anti-B7H3 antibody molecule (e.g., to alter one or more of the following: Fc receptor binding, antibody glycosylation, the number of cysteine ​​residues, effector cell function, or complement function). Functional changes can be produced by replacing at least one amino acid residue in the antibody constant region with a different residue, for example, by altering the affinity of the antibody for an effector ligand (such as FcR or complement C1q), thereby altering (e.g., reducing) the effector function. The Fc region of an antibody mediates several important effector functions, such as ADCC, phagocytosis (ADCP), CDC, etc.

[0944] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure have 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 selected from, for example, the heavy chain constant regions of IgG1, IgG2, IgG3, and IgG4, more particularly selected from the heavy chain constant region of IgG1 (e.g., human IgG1). In some embodiments, the human IgG1 heavy chain constant region is as shown in SEQ ID NO: 43. In some embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure have 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 shown in SEQ ID NO: 44.

[0945] In some embodiments, the antibody or antigen-binding fragment thereof comprises the CH set forth in SEQ ID NO:43 or a variant thereof having 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; for example, 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.

[0946] In some embodiments, the antibody or antigen-binding fragment thereof comprises a light chain constant region or a 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) as set forth in SEQ ID NO: 44 or a variant thereof, wherein the variant has up to 20 conservative substitutions of amino acids compared to SEQ ID NO: 44 (e.g., up to 20, up to 15, up to 10, or up to 5 conservative substitutions of amino acids; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 conservative substitutions of amino acids), or 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 SEQ ID NO: 16;

[0947] 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.

[0948] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure comprise:

[0949] (a) a heavy chain comprising an amino acid sequence selected from the group consisting of:

[0950] (i) a sequence comprising the VH sequence shown in SEQ ID NO: 3 and the CH sequence shown in SEQ ID NO: 43;

[0951] (ii) a sequence having one or several amino acid substitutions, deletions or additions or any combination thereof compared to the sequence shown 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; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof); or

[0952] (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 shown in (i); and

[0953] (b) a light chain comprising an amino acid sequence selected from the group consisting of:

[0954] (iv) a sequence comprising the VL sequence shown in SEQ ID NO: 13 and the CL sequence shown in SEQ ID NO: 44;

[0955] (v) a sequence having one or several amino acid substitutions, deletions or additions or any combination thereof compared to the sequence shown in (iv) (e.g., at most 50, at most 45, at most 40, at most 35, at most 30, at most 25, at most 20, at most 15, at most 10 or at most 5 amino acid substitutions, deletions or additions or any combination thereof; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof); or

[0956] (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 shown in (iv).

[0957] In certain embodiments, the substitution described in (ii) or (v) is a conservative substitution.

[0958] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure comprise:

[0959] (a) a heavy chain comprising an amino acid sequence selected from the group consisting of:

[0960] (i) a sequence comprising the VH sequence shown in SEQ ID NO: 23 and the CH sequence shown in SEQ ID NO: 43;

[0961] (ii) a sequence having one or several amino acid substitutions, deletions or additions or any combination thereof compared to the sequence shown 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; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof); or

[0962] (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 shown in (i); and

[0963] (b) a light chain comprising an amino acid sequence selected from the group consisting of:

[0964] (iv) a sequence comprising the VL sequence shown in SEQ ID NO: 33 and the CL sequence shown in SEQ ID NO: 44;

[0965] (v) a sequence having one or several amino acid substitutions, deletions or additions or any combination thereof compared to the sequence shown in (iv) (e.g., at most 50, at most 45, at most 40, at most 35, at most 30, at most 25, at most 20, at most 15, at most 10 or at most 5 amino acid substitutions, deletions or additions or any combination thereof; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof); or

[0966] (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 shown in (iv).

[0967] In certain embodiments, the substitution described in (ii) or (v) is a conservative substitution.

[0968] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure comprise a heavy chain and a light chain,

[0969] The heavy chain comprises:

[0970] (i) the sequence shown in SEQ ID NO: 45;

[0971] (ii) a sequence having one or several amino acid substitutions, deletions or additions or any combination thereof compared to the sequence shown 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; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof); or

[0972] (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 shown in (i); and

[0973] The light chain comprises:

[0974] (iv) the sequence shown in SEQ ID NO: 46;

[0975] (v) a sequence having one or several amino acid substitutions, deletions or additions or any combination thereof compared to the sequence shown in (iv) (e.g., at most 50, at most 45, at most 40, at most 35, at most 30, at most 25, at most 20, at most 15, at most 10 or at most 5 amino acid substitutions, deletions or additions or any combination thereof; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof); or

[0976] (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 to the sequence set forth in (iv);

[0977] Preferably, the substitutions described in (ii) or (v) are conservative substitutions.

[0978] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present disclosure comprise a heavy chain and a light chain,

[0979] The heavy chain comprises:

[0980] (i) the sequence shown in SEQ ID NO: 47;

[0981] (ii) a sequence having one or several amino acid substitutions, deletions or additions or any combination thereof compared to the sequence shown 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; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof); or

[0982] (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 shown in (i); and

[0983] The light chain comprises:

[0984] (iv) the sequence shown in SEQ ID NO: 48;

[0985] (v) a sequence having one or several amino acid substitutions, deletions or additions or any combination thereof compared to the sequence shown in (iv) (e.g., at most 50, at most 45, at most 40, at most 35, at most 30, at most 25, at most 20, at most 15, at most 10 or at most 5 amino acid substitutions, deletions or additions or any combination thereof; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof); or

[0986] (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 to the sequence set forth in (iv);

[0987] Preferably, the substitutions described in (ii) or (v) are conservative substitutions.

[0988] In certain embodiments, the antibodies of the present disclosure are chimeric, humanized, or fully human antibodies. In certain embodiments, the antibodies of the present disclosure or their antigen-binding fragments are selected from the group consisting of scFv, Fab, Fab', (Fab')2, Fv fragments, disulfide-linked Fv (dsFv), and diabodies.

[0989] In certain embodiments, the antibodies of the present disclosure are scFvs. In certain embodiments, the scFvs of the present disclosure comprise:

[0990] (a) VH of SEQ ID NO: 3 and VL of SEQ ID NO: 13;

[0991] (b) VH of SEQ ID NO: 23 and VL of SEQ ID NO: 33;

[0992] (c) VH of SEQ ID NO: 3 and VL of SEQ ID NO: 33;

[0993] (d) VH of SEQ ID NO: 23 and VL of SEQ ID NO: 13.

[0994] (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 described in any one of groups (a) to (d), respectively; or

[0995] (f) 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 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 any one of groups (a) to (d), respectively. Preferably, the substitutions are conservative substitutions.

[0996] In certain embodiments, the antibodies of the present disclosure are scFvs. In certain embodiments, the scFvs of the present disclosure comprise:

[0997] (i) the sequence shown in SEQ ID NO: 1 or 2;

[0998] (ii) a sequence having one or several amino acid substitutions, deletions or additions or any combination thereof compared to the sequence shown 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; for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof); or

[0999] (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 shown in (i);

[1000] Preferably, the substitutions described in (ii) are conservative substitutions.

[1001] In certain embodiments, the antibodies of the disclosure are scFvs.

[1002] In certain embodiments, the scFv of the present disclosure comprises the sequence set forth in SEQ ID NO: 1 or 2.

[1003] Antibody derivatives

[1004] The antibodies or antigen-binding fragments thereof disclosed herein may be derivatized, for example, by being linked to another molecule (e.g., another polypeptide or protein). Generally, derivatization (e.g., labeling) of an antibody or antigen-binding fragment thereof will not adversely affect its binding to B7H3 (particularly human B7H3). Therefore, the antibodies or antigen-binding fragments thereof disclosed herein are also intended to include such derivatized forms. For example, an antibody or antigen-binding fragment thereof disclosed herein may be linked (by chemical coupling, genetic fusion, non-covalent linkage, or other means) to one or more other molecular groups, such as another antibody (e.g., to form a bispecific antibody), a detection reagent, a pharmaceutical agent, and / or a protein or polypeptide capable of mediating binding of the antibody or antigen-binding fragment thereof to another molecule (e.g., an avidin or polyhistidine tag).

[1005] One type of derivatized antibody (e.g., bispecific antibody) is produced by cross-linking two or more antibodies (of the same type or different types). Methods for obtaining bispecific antibodies are well known in the art, and examples thereof include, but are not limited to, chemical cross-linking, cell engineering (hybridoma method), or genetic engineering.

[1006] Another type of derivatized antibody is a labeled antibody. For example, the antibody of the present disclosure or its antigen-binding fragment can be connected to a detectable label. The detectable label described in the present disclosure can be any substance that can be detected by fluorescence, spectroscopy, photochemistry, biochemistry, immunology, electricity, optics or chemical means. Such labels are well known in the art, and examples thereof include, but are not limited to, enzymes (e.g., horseradish peroxidase, alkaline phosphatase, β-galactosidase, urease, glucose oxidase, etc.), radionuclides (e.g., 3H, 125I, 35S, 14C or 32P), 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)), acridinium ester compounds, magnetic beads (e.g., ), calorimetric labels such as colloidal gold or colored glass or plastic (e.g., polystyrene, polypropylene, latex, etc.) beads, and biotin for binding to avidin (e.g., streptavidin) modified with the above labels. Patents teaching the use of such labels include, but are not limited to, U.S. Patents 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). Detectable labels such as those described above can be detected by methods known in the art. For example, radioactive labels can be detected using photographic film or a scintillation counter, and fluorescent labels can be detected using a photodetector to detect emitted light. Enzyme labels 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, and calorimetric labels are detected by simple visualization of a colored label. In certain embodiments, such labels can be suitable for immunological detection (e.g., enzyme-linked immunosorbent assay, radioimmunoassay, fluorescent immunoassay, chemiluminescent immunoassay, etc.). In certain embodiments, the detectable labels described above can be linked to the antibodies or antigen-binding fragments thereof of the present disclosure via linkers of varying lengths to reduce potential steric hindrance.

[1007] In addition, the antibodies or antigen-binding fragments thereof of the present disclosure can also be derivatized with chemical groups, such as polyethylene glycol (PEG), methyl or ethyl groups, or glycosyl groups. These groups can be used to improve the biological properties of the antibodies, such as increasing serum half-life.

[1008] The antigen-binding fragments disclosed herein can be obtained by hydrolyzing intact antibody molecules (see Morimoto et al., J. Biochem. Biophys. Methods 24:107-117 (1992) and Brennan et al., Science 229:81 (1985)). Alternatively, these antigen-binding fragments can be produced directly from 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 directly isolated from recombinant host cell culture medium. Other techniques for preparing such antigen-binding fragments are well known to those of ordinary skill in the art.

[1009] The IgG isotype control antibodies disclosed herein are well known to those skilled in the art and can be purchased or prepared. For example, human anti-hen egg lysosomal IgG (anti-HEL, such as human IgG1, referred to as hIgG1) is derived from the variable region sequence of Fab F10.6.6 sequence in the study "Affinity maturation increases the stability and plasticity of the Fv domain of anti-protein antibodies" published by Acierno et al. (Acierno et al. J Mol Biol. 2007; 374(1): 130-46). The preparation method is as follows: Nanjing GenScript Biopharmaceuticals was commissioned to perform amino acid codon optimization and gene synthesis on the heavy and light chain genes (full sequence or variable region) of a human IgG antibody. Following the standard techniques described in the Molecular Cloning Manual (3rd Edition), standard molecular cloning techniques such as PCR, enzyme digestion, DNA gel recovery, ligation transformation, colony PCR, or enzyme digestion identification were used to subclone the heavy and light chain genes into mammalian expression systems for antibody heavy and light chain expression vectors, respectively. The heavy and light chain genes in the recombinant expression vectors were then sequenced and analyzed. After sequencing verification, large quantities of endotoxin-free expression plasmids were prepared and transiently co-transfected into HEK293 cells for recombinant antibody expression. After 7 days of culture, the cell culture fluid was collected and affinity purified using an rProtein A column (GE). The harvested antibody sample was then quality-assured using standard analytical techniques such as SDS-PAGE and SEC-HPLC.

[1010] In a ninth aspect, the present disclosure provides a multispecific antibody comprising the antibody or antigen-binding fragment thereof according to any one of the eighth aspects of the present disclosure, and another antibody or fragment thereof or antibody analog.

[1011] In certain embodiments, the multispecific antibody is a bispecific antibody, a trispecific antibody, or a tetraspecific antibody.

[1012] Therefore, 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, or a heavy chain variable region and / or a light chain variable region, or one or more CDRs thereof. According to codon degeneracy known in the art, in certain embodiments, the nucleotide sequence is replaceable according to codon degeneracy. In certain embodiments, the nucleotide sequence is codon-optimized.

[1013] In certain embodiments, the isolated nucleic acid molecules of the present disclosure comprise: (i) a first nucleic acid and a second nucleic acid encoding the heavy chain variable region and the light chain variable region of the antibody or antigen-binding fragment thereof of the present disclosure, respectively, or (ii) a first nucleic acid encoding the heavy chain variable region and the heavy chain constant region, and a second nucleic acid encoding the light chain variable region and the light chain constant region, respectively, or (iii) a first nucleic acid and a second nucleic acid encoding the heavy chain and the light chain of the antibody or antigen-binding fragment thereof of the present disclosure, respectively. In certain embodiments, the first nucleic acid and the second nucleic acid comprise a degenerate sequence or a substantially identical sequence to any of the first nucleic acid and the second nucleic acid in (i)-(iii) above. In certain embodiments, the degenerate sequence or substantially identical sequence refers to a sequence having at least about 85%, 90%, 95%, 99% or higher sequence identity or a sequence having one or more nucleotide substitutions, or a sequence that differs by no more than 3, 6, 15, 30 or 45 nucleotides compared to the nucleic acid molecules of (i)-(iii).

[1014] In an eleventh aspect, a vector (e.g., a cloning vector or an expression vector) is provided, comprising an isolated nucleic acid molecule of the present disclosure. In certain embodiments, the vector of the present disclosure is, for example, a plasmid, a cosmid, a phage, a lentivirus, etc. In certain embodiments, the vector is capable of expressing the antibody or antigen-binding fragment thereof of the present disclosure in a subject (e.g., a mammal, such as a human).

[1015] A twelfth aspect provides a host cell comprising an isolated nucleic acid molecule of the present disclosure or a vector of the present disclosure. The host cell can be a eukaryotic cell (e.g., a mammalian cell, an insect cell, a yeast cell) or a prokaryotic cell (e.g., an E. coli). Suitable eukaryotic cells include, but are not limited to, NSO 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 CHO (e.g., CHO-K1, CHO-S, CHO DXB11, CHO DG44).

[1016] In a thirteenth aspect, a method for preparing the antibody or antigen-binding fragment thereof, or multispecific antibody of the present disclosure is provided, comprising culturing the host cell of the present disclosure under conditions that allow 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.

[1017] In a fourteenth aspect, an antibody-drug conjugate is provided, wherein the antibody is the anti-B7H3 antibody described above, and is connected to a coupling moiety via a linker. The coupling moiety is selected from the group consisting of a detectable label, a radioisotope, a fluorescent substance, a luminescent substance, a colored substance, an enzyme, polyethylene glycol (PEG), a nuclide, a nucleic acid, a small molecule toxin, a polypeptide with binding activity, a protein, a receptor, a ligand, and other active substances that inhibit tumor cell growth or promote tumor cell apoptosis or necrosis.

[1018] In some embodiments, the antibody-drug conjugate comprises an anti-B7H3 antibody drug conjugate (B7H3-ADC) of the formula:

[1019] Ab-(LD)n,

[1020] in:

[1021] Ab is the antibody or antigen-binding fragment thereof of the eighth aspect, and;

[1022] D is a small molecule toxin drug moiety;

[1023] L is a bond or linking molecule that covalently links Ab and D;

[1024] n is an integer between 1-16 and represents the number of LDs covalently linked to the Ab.

[1025] In some embodiments, L of the antibody-drug conjugate comprises an amino acid residue or a short peptide consisting of 2-10 amino acid residues; the amino acid residue is selected from natural amino acid residues, non-natural amino acid residues, AA 1 the indicated amino acid residue or its stereoisomers;

[1026]

[1027] AA 1 Among the amino acid residues shown:

[1028] R a1 and R b Each independently selected from H,

[1029] Or, R a1 With R b Together with the carbon atoms they are commonly connected to form a 4-10 membered heterocyclic ring, wherein the heteroatoms in the 4-10 membered heterocyclic ring are selected from 1, 2, 3 or 4 of O, N and S; the 4-10 membered heterocyclic ring is optionally replaced by one or more R 0 replaced by;

[1030] r、r 1 、r 1a and r1b Each independently represents an integer from 0 to 20;

[1031] R m1 、R n1 、R m1a 、R n1a 、R m1b and R n1b Each independently is H, C 1-6 Alkyl, C 3-6 Cycloalkyl or -COOR x1 , where R x1 C 1-6 alkyl;

[1032] Or, R m1 and R n1 、R m1a and R n1a , and R m1b and R n1b Together with the nitrogen atom they are connected to, they form a 4-10 membered heterocyclic ring, wherein the heteroatoms in the 4-10 membered heterocyclic ring are selected from 1, 2, 3 or 4 of O, N and S; the 4-10 membered heterocyclic ring is optionally replaced by one or more R 0’ replaced by;

[1033] R z Selected from C 1-6 alkyl;

[1034] R 0 、R 0’ Each independently selected from C 1-6 Alkyl, C 3-6 Cycloalkyl, -NR m2 R n2 and optionally C 1-6 Alkyl-substituted 4-10 membered heterocyclic group; in the 4-10 membered heterocyclic ring, the heteroatoms are selected from 1, 2, 3 or 4 of O, N and S;

[1035] R m2 and R n2 Each independently selected from H and G 1-6 alkyl.

[1036] In some embodiments, AA 1 Among the amino acid residues shown, R a1 With R b Together with the carbon atoms they are connected to form a 5-6 membered heterocyclic ring, wherein the heteroatom in the 5-6 membered heterocyclic ring is N and the number of heteroatoms is 1 or 2; the 5-6 membered heterocyclic ring is optionally replaced by one or more R 0 The 5-6 membered heterocyclic ring is preferably a piperidine ring or a piperazine ring, and more preferably a piperidine ring, for example Carbon atom No. 1 is R a1 and R b carbon atoms that are linked together.

[1037] In some embodiments, r, r 1 、r 1a and r 1b Each independently represents 0, 1, 2, 3, 4 or 5; preferably, r, r 1 、r 1a and r 1b Each independently is 0 or 4; more preferably, r is 0, r 1 、r 1a and r 1b is 4; or r is 4, r 1 、r 1a and r 1b is 0.

[1038] In some embodiments, r, r 1 、r 1a and r 1b Not 0 at the same time.

[1039] In some embodiments, R m1 、R n1 、R m1a 、R n1a 、R m1b and R n1b Each independently is H, C 1-6 Alkyl or -COOR x1 , where R x1 C 1-6 Alkyl; preferably, each independently H, methyl, ethyl, n-propyl, -COOCH3, -COOCH2CH3, -COOCH2CH2CH3, -COOCH(CH3)2, -COOC(CH3)3 or -COOCH2CH2CH2CH3.

[1040] In some embodiments, R m1 and R n1 、R m1a and R n1a , and R m1b and R n1b Together with the nitrogen atom they are connected to form a 5-6 membered heterocyclic ring, wherein the heteroatom in the 5-6 membered heterocyclic ring is selected from 1 or 2 N atoms; the 5-6 membered heterocyclic ring is optionally replaced by one or more R 0’ substituted; the 5-6 membered heterocyclic ring is preferably a piperidine ring or a piperazine ring; further preferably Nitrogen atom No. 1 is R m1 and R n1 Commonly linked nitrogen atoms.

[1041] In some embodiments, R z Preferably C 1-6 Alkyl; more preferably methyl.

[1042] In some embodiments, R 0 and R 0’ Each independently is C 1-6 Alkyl, -NR m2 R n2 Or optionally C 1-6 Alkyl-substituted 5-6 membered heterocyclic group; in the 5-6 membered heterocyclic ring, the heteroatom is selected from 1 or 2 N atoms.

[1043] In some embodiments, R 0 Preferably C 1-6 Alkyl or C 1-6 Alkyl-substituted 5-6 membered heterocyclic group, wherein the 5-6 membered heterocyclic group is piperidinyl or piperazinyl; further preferably methyl or piperidinyl substituted by methyl; for example methyl or

[1044] In some embodiments, R 01’ Each independently preferably is C 1-6 Alkyl, -NR m2 R n2 Or optionally C 1-6 Alkyl-substituted 5-6 membered heterocyclic group; in the 5-6 membered heterocyclic ring, the heteroatom is 1 or 2 N; more preferably C 1-6 Alkyl or -NR m2 R n2 More preferably, methyl or -NR m2 R n2 , where R m2 and R n2 Each independently is preferably H or C 1-6 Alkyl, more preferably methyl.

[1045] In some embodiments, R 0’ Preferably C 1-6 Alkyl or -NR m2 R n2 ; R m2 and R n2 Each independently is preferably H or C 1-6 Alkyl (eg, methyl).

[1046] In some embodiments, AA 1 Among the amino acid residues shown, R a1 and Rb Either one is H, the other is Preferably, R a1 and R b Any one of them is H, and the other is selected from

[1047] In some embodiments, AA 1 The amino acid residues shown are preferably More preferably More preferably

[1048] Most preferably

[1049] In some embodiments, the antibody-drug conjugate consists of antibody-linker-drug, wherein the drug is selected from: microtubule blocking drugs, DNA damaging drugs, Bcl-xL inhibitors, etc.

[1050] In some embodiments, the antibody-drug conjugate consists of antibody-linker-drug, wherein the drug is selected from: Auristatins (e.g., MMAF, MMAE), maytansine derivatives (e.g., DM1, DM4), tubulysin, cryptocolistin, anti-mitotic inhibitors, pyrrole benzazepines and indolechlorobenzazepines, dukamycin, camptothecin, calicheamicin, etc.

[1051] In some embodiments, the antibody-drug conjugate consists of antibody-linker-drug, wherein the drug is selected from camptothecin (CPT) and its derivatives.

[1052] In some embodiments, the antibody-drug conjugate consists of antibody-linker-drug, wherein the drug is selected from topoisomerase I inhibitors.

[1053] In some embodiments, the antibody-drug conjugate consists of antibody-linker-drug, wherein the drug is selected from the group consisting of: SN-38, DXd.

[1054] In some embodiments, in the antibody-drug conjugate, D is a drug unit having Formula II:

[1055]

[1056] in,

[1057] R1 is H or F,

[1058] R2 is H or methyl;

[1059] Or R1 and R2 together with the carbon atom to which they are attached form

[1060] R3 is H;

[1061] R4 is selected from H, -(C1-C4 alkyl)-OH, -(C1-C4 alkenyl)-OH, -(C1-C4 alkyl)-NH2, or -(C1-C4 alkenyl)-NH2;

[1062] wherein L is linked via the hydroxyl or amine group present on D.

[1063] In some embodiments, L of the antibody-drug conjugate is selected from:

[1064]

[1065] Wherein, position 1 is connected to Ab, position 2 is connected to D; AA1 is as defined above.

[1066] It should also be noted that those skilled in the art will understand that L is linked to the sulfhydryl group contained in Ab (e.g., an antibody) after disulfide bond opening (e.g., reduction of disulfide bonds by the reducing agent TCEP can open disulfide bonds to form sulfhydryl groups -SH). In other words, the -S- between L and Ab is not a separate external sulfur atom. For example, in , the -S- is not a separate external sulfur atom, but rather the -S- formed by the sulfhydryl group contained in Tb after disulfide bond opening connecting with L.

[1067] In some preferred embodiments, the antibody-drug conjugate is selected from:

[1068]

[1069]

[1070]

[1071] wherein Ab is the B7H3 antibody 1D1-01 or 2E3-02, n is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, or 16, and q is preferably 1, 2, 3, 4, 5, 6, 7, or 8.

[1072] 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, wherein the ligand-drug conjugate has two or more q values, and the antibody-drug conjugate has two or more n values. The ligand-drug conjugate is a ligand-drug conjugate or antibody-drug conjugate (ADC) comprising a heterogeneous DAR distribution.

[1073] In some embodiments, when the ligand drug conjugates having one q value or n value account for the majority, the q value or n value is close to the DAR.

[1074] In some embodiments, the drug to antibody ratio (DAR) in the ligand drug conjugate is an integer or decimal selected from 1-10.

[1075] In some embodiments, the drug to antibody ratio (DAR) in the ligand drug conjugate is selected from the group consisting of 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.

[1076] In some embodiments, the LDC ADC comprises ADCs having a distribution of DARs of 1 to 8, for example, 1.5, 2, 4, 6, and 8 (i.e., 1.5, 2, 4, 6, and 8 drug loading species). Of note, degradation products may be produced such that the LDC may also contain DARs of 1, 3, 5, and 7. Furthermore, the ADCs in the LDC may also have a DAR greater than 8. The LDC is produced by reducing interchain disulfides followed by conjugation. In some embodiments, the LDC comprises both: a LDC having a DAR of 4 or less (i.e., a drug loading species of 4 or less) and a LDC having a DAR of 6 or greater (i.e., a drug loading species of 6 or greater).

[1077] In another aspect, the present disclosure provides a use of an antibody or antigen-binding fragment thereof in preparing 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: an antibody or antigen-binding fragment thereof, a nucleic acid, a vector, a host cell, a multispecific antibody, a conjugate, or a pharmaceutical composition described herein. Optionally, the diagnostic or therapeutic kit also includes instructions for use.

[1078] In the sixteenth aspect of the present disclosure, the present disclosure provides a pharmaceutical composition comprising substance A, and optionally one or more pharmaceutical excipients; 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, nucleic acid molecule, vector, host cell, multispecific antibody, and / or ligand-drug conjugate, 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.

[1079] In certain embodiments, the pharmaceutical compositions of the present disclosure comprise the antibodies or antigen-binding fragments thereof of the present disclosure, and a pharmaceutically acceptable carrier and / or excipient.

[1080] In certain embodiments, the pharmaceutical composition of the present disclosure comprises the 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 as described above.

[1081] In certain embodiments, the pharmaceutical compositions of the present disclosure comprise the multispecific antibodies of the present disclosure, and a pharmaceutically acceptable carrier and / or excipient.

[1082] In certain embodiments, pharmaceutical compositions of the present disclosure comprise a conjugate of the present disclosure and a pharmaceutically acceptable carrier and / or excipient.

[1083] In a seventeenth aspect of the present disclosure, the present disclosure provides the use of the aforementioned substance A or the aforementioned pharmaceutical composition in the preparation of a medicament for treating and / or preventing a disease associated with abnormal cell activity (e.g., a cancer disease). The substance A or the aforementioned pharmaceutical composition may be in a therapeutically effective amount.

[1084] In some embodiments, provided are uses of the antibodies or antigen-binding fragments thereof, nucleic acids, vectors, host cells, antibody-drug conjugates, or multispecific antibodies of the present disclosure in the preparation of drugs for modulating (inhibiting or blocking) the activity of B7H3.

[1085] In some embodiments, provided are uses of the antibodies or antigen-binding fragments thereof, nucleic acids, vectors, host cells, antibody-drug conjugates, or multispecific antibodies of the present disclosure in the preparation of medicaments for treating or preventing diseases associated with the activity of B7H3.

[1086] In some embodiments, provided are uses of the antibodies or antigen-binding fragments thereof, nucleic acids, vectors, host cells, antibody-drug conjugates, or multispecific antibodies of the present disclosure in the preparation of medicaments for treating or preventing tumors associated with B7H3 activity.

[1087] 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 carcinoma, epidermal cancer, non-Hodgkin's lymphoma, central nervous system tumor (e.g., glioma, glioblastoma multiforme, glioma, or sarcoma), prostate cancer, or thyroid cancer.

[1088] In some embodiments, the cancer disease is a cancer disease associated with Trop-2, Her 2, B7H3, Her 3, or EGFR.

[1089] In some embodiments, the cancer disease is a cancer disease associated with Trop-2 or Her 2.

[1090] In some embodiments, the cancer disease is a cancer disease associated with Trop-2.

[1091] In some embodiments, the cancer disease is a Her 3-associated cancer disease.

[1092] In some embodiments, the cancer disease is a cancer disease associated with EGFR.

[1093] In some embodiments, the cancer disease is a B7H3-associated cancer disease.

[1094] In some embodiments, the cancer disease is a solid tumor.

[1095] In some embodiments, the cancer disease is breast cancer or lung cancer (preferably non-small cell lung cancer).

[1096] In the seventeenth aspect of the present disclosure, the present disclosure provides the aforementioned substance A or the aforementioned pharmaceutical composition, which is used to treat and / or prevent diseases associated with abnormal cell activity (such as cancer diseases).

[1097] 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 carcinoma, epidermal cancer, non-Hodgkin's lymphoma, central nervous system tumor (e.g., glioma, glioblastoma multiforme, glioma, or sarcoma), prostate cancer, or thyroid cancer.

[1098] In some embodiments, the cancer disease is a cancer disease associated with Trop-2, Her 2, B7H3, Her 3, or EGFR.

[1099] In some embodiments, the cancer disease is a cancer disease associated with Trop-2 or Her 2.

[1100] In some embodiments, the cancer disease is a cancer disease associated with Trop-2.

[1101] In some embodiments, the cancer disease is a Her 3-associated cancer disease.

[1102] In some embodiments, the cancer disease is a cancer disease associated with EGFR.

[1103] In some embodiments, the cancer disease is a B7H3-associated cancer disease.

[1104] 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).

[1105] In the eighteenth aspect of the present disclosure, the present disclosure provides a method for preventing and / or treating diseases associated with abnormal cell activity (such as cancer diseases), which comprises: administering a preventive and / or therapeutically effective amount of the aforementioned substance A, or the aforementioned pharmaceutical composition, to an individual in need.

[1106] 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 carcinoma, epidermal cancer, non-Hodgkin's lymphoma, central nervous system tumor (e.g., glioma, glioblastoma multiforme, glioma, or sarcoma), prostate cancer, or thyroid cancer.

[1107] In some embodiments, the cancer disease is a cancer disease associated with Trop-2, Her 2, B7H3, Her 3, or EGFR.

[1108] In some embodiments, the cancer disease is a cancer disease associated with Trop-2 or Her 2.

[1109] In some embodiments, the cancer disease is a cancer disease associated with Trop-2.

[1110] In some embodiments, the cancer disease is a Her 3-associated cancer disease.

[1111] In some embodiments, the cancer disease is a cancer disease associated with EGFR.

[1112] In some embodiments, the cancer disease is a B7H3-associated cancer disease.

[1113] In some embodiments, the cancer disease is a solid tumor.

[1114] In some embodiments, the cancer disease is breast cancer or lung cancer (preferably non-small cell lung cancer).

[1115] In this disclosure, unless otherwise indicated, scientific and technical terms used herein have the meanings commonly understood by those skilled in the art. Furthermore, the laboratory procedures for cell culture, molecular genetics, nucleic acid chemistry, and immunology used herein are conventional procedures widely used in the relevant fields. To facilitate a better understanding of this disclosure, definitions and explanations of relevant terms are provided below.

[1116] As used herein, examples of the term "pharmaceutically acceptable salts" are organic acid addition salts formed with organic acids that form pharmaceutically acceptable anions, including but not limited to formates, acetates, propionates, benzoates, maleates, fumarates, succinates, tartrates, citrates, ascorbates, α-ketoglutarate, α-glycerophosphates, alkylsulfonates, or arylsulfonates; 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 hydrochlorides, hydrobromides, hydroiodides, nitrates, bicarbonates, carbonates, sulfates, or phosphates.

[1117] As used herein, the term "pharmaceutically acceptable carriers and / or excipients" refers to carriers and / or excipients that are pharmacologically and / or physiologically compatible with the subject and the active ingredient, which are well known in the art (see, for example, Remington's Pharmaceutical Sciences. Edited by Gennaro AR, 19th ed. Pennsylvania: Mack Publishing Company, 1995), and include, but are not limited to: pH adjusters, surfactants, adjuvants, ionic strength enhancers, diluents, agents for maintaining osmotic pressure, agents for delaying absorption, preservatives.

[1118] 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 affording a pharmaceutically acceptable anion.

[1119] For the purposes of this disclosure, pharmaceutical excipients refer to excipients and additives used in the production of pharmaceuticals and the preparation of prescriptions. These substances, in addition to the active ingredients, have been reasonably evaluated for safety and are included in pharmaceutical preparations. In addition to providing shape, acting as carriers, and improving stability, pharmaceutical excipients also have important functions such as solubilization, dissolution enhancement, and sustained-release control. They are important components that may affect the quality, safety, and efficacy of drugs. Based on their source, they can be categorized as natural, semi-synthetic, and fully synthetic. According to their functions and uses, they can be divided into: solvents, propellants, solubilizers, cosolvents, emulsifiers, colorants, adhesives, disintegrants, fillers, lubricants, wetting agents, osmotic pressure regulators, stabilizers, glidants, flavoring agents, preservatives, suspending agents, coating materials, fragrances, anti-adhesives, antioxidants, chelating agents, penetration enhancers, pH regulators, buffers, plasticizers, surfactants, foaming agents, defoaming agents, thickeners, inclusion agents, humectants, absorbents, diluents, flocculants and deflocculants, filter aids, release retardants, etc. According to their route of administration, they can be divided into oral, parenteral, mucosal, transdermal or topical administration, nasal or oral inhalation administration, and ocular administration, etc. The same pharmaceutical excipient can be used in pharmaceutical preparations for different routes of administration and have different functions and uses.

[1120] The pharmaceutical composition can be prepared into various suitable dosage forms according to the route of administration, such as tablets, capsules, granules, oral solutions, oral suspensions, oral emulsions, powders, tinctures, syrups, injections, suppositories, ointments, creams, pastes, ophthalmic preparations, pills, implants, aerosols, powder sprays, sprays, etc. The pharmaceutical composition or suitable dosage form can contain 0.01 mg to 1000 mg of the compound of the present disclosure or a pharmaceutically acceptable salt or conjugate thereof, preferably 0.1 mg to 800 mg, preferably 0.5-500 mg, preferably 0.5 to 350 mg, and particularly preferably 1-250 mg.

[1121] The pharmaceutical composition can be administered in the form of an injection, including an injection solution, sterile powder for injection, and concentrated solution for injection. Among them, the carriers and solvents that can be used include water, Ringer's solution, and isotonic sodium chloride solution. In addition, sterile fixed oils such as monoglycerides or diglycerides can also be used as solvents or suspending media.

[1122] As used herein, the term "treat" generally refers to obtaining a desired pharmacological and / or physiological effect. The effect may be prophylactic, in terms of completely or partially preventing a disease or its symptoms; and / or therapeutic, in terms of partially or completely stabilizing or curing a disease and / or causing side effects due to the disease. As used herein, "treat" encompasses any treatment of a disease in a patient, including: (a) preventing the onset of a disease or symptom in a patient who is susceptible to the disease or symptom but has not yet been diagnosed with the disease; (b) suppressing the symptoms of a disease, i.e., arresting its development; or (c) relieving the symptoms of a disease, i.e., causing regression of the disease or symptom.

[1123] In the present disclosure, the term "individual" includes humans and non-human animals. Exemplary human individuals include human individuals suffering from a disease (e.g., a disease described herein) (referred to as a patient) or normal individuals. The term "non-human animal" in the present disclosure includes all vertebrates, such as non-mammals (e.g., birds, amphibians, reptiles) and mammals, such as non-human primates, livestock and / or domesticated animals (e.g., sheep, dogs, cats, cows, pigs, etc.).

[1124] As used herein, the term "effective dose" refers to that amount of a compound which, when administered, will relieve to some extent one or more of the symptoms of the condition being treated.

[1125] In the present disclosure, the term "ligand drug conjugate" refers to a substance obtained by connecting a bioactive molecule (drug molecule) to a targeting portion. In some embodiments of the present disclosure, the bioactive molecule is connected to the targeting portion via a connector. The connector can be broken in a specific environment (e.g., hydrolases and / or low pH environments within a tumor) or under a specific action (e.g., the action of lysosomal proteases), thereby separating the bioactive molecule from the targeting portion. In some embodiments of the present disclosure, the connector comprises a cleavable or non-cleavable unit, such as a peptide or a disulfide bond. In some embodiments of the present disclosure, the bioactive molecule is directly connected to the targeting portion via a covalent bond, and the covalent bond can be broken under a specific environment or action, thereby separating the bioactive molecule from the targeting portion. In some embodiments of the present disclosure, the ligand drug conjugate comprises a targeting portion, a connector, and a fragment of a compound of formula II of the present disclosure.

[1126] In the present disclosure, the term "bioactive substance", "bioactive molecule" or "drug molecule" refers to a substance that inhibits or prevents cell function and / or causes cell death or destruction. In some embodiments of the present disclosure, the bioactive substance, bioactive molecule or drug molecule in the conjugate is a molecule with anti-tumor biological activity. For example: radioactive isotopes, such as At 211 , I 131 , I 125 、Y 90 、Re 186 、Re 188 、Sm 153 、Bi 212 、P 32 , Pb 212and radioactive isotopes of Lu; metal complexes, such as metal platinum complexes, metal gold complexes, oxaliplatin, etc.; glycopeptide antibiotics, such as bleomycin and bleomycin; DNA topoisomerase inhibitors, such as topoisomerase I inhibitors, camptothecin, hydroxycamptothecin, 9-aminocamptothecin, SN-38, irinotecan, topotecan, belotecan, rubitecan, topoisomerase II inhibitors, actinomycin D, doxorubicin, doxorubicin, duocarmycin, daunorubicin, mitoxantrone, podophyllotoxin, etoposide, etc.; drugs that interfere with DNA synthesis, such as methotrexate, 5-fluorouracil, cytarabine, gemcitabine, mercaptopurine, pentostatin, fludarabine, cladribine, nelarabine, etc.; drugs that act on structural proteins, such as tubulin inhibitors, vinca alkaloids, vincristine, vinblastine, paclitaxel, polyoxazine Citaxel, Cabazitaxel, etc.; tumor signaling pathway inhibitors, such as serine / threonine kinase inhibitors, tyrosine kinase inhibitors, aspartate kinase inhibitors or histidine kinase inhibitors; also including proteasome inhibitors, histone deacetylase inhibitors, tumor angiogenesis inhibitors, cell cycle protein inhibitors, maytansine derivatives, calicheamicin derivatives, auristatin derivatives, pyrrolobenzodiazepines (PBD) derivatives, melphalan, mitomycin C, chlorambucil, or other active substances that inhibit tumor cell growth, promote tumor cell apoptosis and necrosis; enzymes and fragments thereof, such as nucleolytic enzymes; antibiotics; toxins, such as small molecule toxins or enzymatically active toxins of bacterial, fungal, plant or animal origin, including fragments and / or variants thereof; growth inhibitors; drug moieties. The term "toxin" refers to a substance that can have a deleterious effect on cell growth or proliferation.

[1127] In this disclosure, the term "small molecule" refers to a small molecule drug with biological activity. The term "small molecule toxin" refers to a state in which a small molecule toxin has cell-damaging activity, i.e., a state in which a pathological change in a cell is caused in some form. Cell damage is not limited to direct damage, but includes all types of damage to the structure and function of the cell, such as DNA cleavage, base-dimer formation, chromosome cleavage, damage to the cell division machinery, and a decrease in the activity of various enzymatic enzymes.

[1128] In the present disclosure, the term "linker" refers to a fragment that connects a biologically active molecule (drug molecule) to a targeting moiety.

[1129] In this disclosure, the term "targeting moiety" refers to a portion of a conjugate that is capable of specifically binding to a target (or portion of a target) on the cell surface. Through the interaction of the targeting moiety with the target, the conjugate can be delivered to a specific cell population.

[1130] 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."

[1131] 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 having sulfhydryl groups, wherein the derivatization provides the antibody with a group or ability to react with a drug-linker conjugate. The sulfhydryl group (SH) can be derivatized by opening a disulfide bond (e.g., by reduction with the reducing agent TCEP).

[1132] As used herein, the terms "cancer" and "tumor" are used synonymously.

[1133] The term "gene" used herein includes not only DNA but also its mRNA, its cDNA, and its cRNA.

[1134] As used herein, the term "polynucleotide" is used with the same meaning as nucleic acid, and also includes DNA, RNA, probes, oligonucleotides, and primers.

[1135] The term "Tb" used herein is an abbreviation for "target binding", including antibodies or any molecules that bind to a target, and the term "Ab" is an abbreviation for "antibody" and is used indistinguishably from "antibody".

[1136] As used herein, the terms "polypeptide" and "protein" are used indiscriminately.

[1137] The term "cell" as used herein also includes cells in an animal individual and cultured cells.

[1138] The B7H3 involved in the anti-B7H3 antibodies described in the present disclosure may be conventional B7H3 in the art, such as soluble B7H3, membrane-form B7H3, etc., and also represents B7H3 variant 1 and / or B7H3 variant 2.

[1139] KD refers to the dissociation constant obtained from the ratio of Kd (the off-rate of a particular binding molecule-target protein interaction) to Ka (the on-rate of a particular binding molecule-target protein interaction) (or Kd / Ka, expressed in 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 using surface plasmon resonance, for example, a biosensor system such as the Biacore™ (GE Healthcare Life Sciences) system.

[1140] The term "B7H3 variant" refers to a polypeptide that has similar or identical functions to a B7H3 polypeptide, a B7H3 fragment, an anti-B7H3 antibody or an antibody fragment thereof, but does not necessarily comprise a similar or identical amino acid sequence of a B7H3 polypeptide, a B7H3 fragment, an anti-B7H3 antibody or a fragment thereof, or has a similar or identical structure of a B7H3 polypeptide, a B7H3 fragment, an anti-B7H3 antibody or a fragment thereof.

[1141] The term "Her3 variant" refers to a polypeptide that has similar or identical functions to a Her3 polypeptide, Her3 fragment, anti-Her3 antibody or antibody fragment thereof, but does not necessarily contain a similar or identical amino acid sequence of a Her3 polypeptide, Her3 fragment, anti-Her3 antibody or fragment thereof, or has a similar or identical structure of a Her3 polypeptide, Her3 fragment, anti-Her3 antibody or fragment thereof.

[1142] As used herein, the percent homology between two amino acid sequences is equivalent to the percent identity between the two sequences. The percent identity of a sequence between two sequences is a function of the number of identical positions shared by the sequences (i.e., % homology = number of identical positions / total number of positions x 100), taking into account the number of gaps and the length of each gap that need to be introduced for optimal alignment of the two sequences. Sequence comparisons and determination of percent identity between sequences can be performed using methods generally known in the art, and such sequence comparisons and determination of percent identity can be performed using mathematical algorithms. For example, the algorithm of Meyers and Miller, 1988 Comput. Appl. Biosci. 4: 11-17 (incorporated into the ALIGN program (version 2.0)) can be used to determine percent identity between amino acid sequences and / or between nucleotide sequences. In addition, the GAP program in the GCG software package available online from Accelrys (using its default parameters) can be used to determine percent identity between amino acid sequences or between nucleotide sequences. In one embodiment, the two sequences are of equal length.

[1143] 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 the B7H3 antibodies or antigen-binding fragments thereof disclosed herein can be determined using 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, and antibody homology modeling and protein docking prediction methods. As used herein, the phrase "antibodies that bind to the same epitope" refers to different antibodies that bind to a common epitope. If a second antibody binds to a portion of the peptide or a portion of the tertiary structure bound by the first antibody, it can be determined that the first and second antibodies bind to the same epitope.

[1144] In the present disclosure, the term "antibody" is interpreted in the broadest sense, including complete monoclonal antibodies, polyclonal antibodies, and multispecific antibodies (such as bispecific antibodies) formed by at least two complete antibodies, as long as they have the desired biological activity. In the present disclosure, "antibody" and "immunoglobulin" can be used interchangeably. As described herein, "antibody molecule" or "antibody" refers to immunoglobulin molecules and immunoactive parts of immunoglobulin molecules, i.e., molecules containing antigen binding sites that immunospecifically bind to antigens. Therefore, the term antibody broadly encompasses not only complete antibody molecules, but also fragments of the antibodies and variants (including derivatives) of the antibodies and antibody fragments. When "antibody molecule" or "antibody" is used in the same context as antigen-binding fragment, "antibody molecule" or "antibody" refers to complete antibody molecules or full-length antibodies. The term antibody molecule in this specification, for example, includes but is not limited to single-chain Fv (scFv), Fab fragments, Fab' fragments, F(ab')2, disulfide-linked Fv (sdFv), Fv, and complete antibodies or full-length antibodies. The term "single-chain Fv" or "scFv" refers to a polypeptide comprising the VL domain of an antibody connected to the VH domain of an antibody. For example, an antibody that immunospecifically binds to B7H3 may cross-react with other antigens. Preferably, an antibody that immunospecifically binds to B7H3 does not cross-react with other antigens. An antibody that immunospecifically binds to B7H3 can be identified, for example, by immunoassays or other methods known to those skilled in the art. An "intact" or "full-length" antibody refers to a protein comprising two heavy (H) chains and two light (L) chains interconnected by disulfide bonds, the protein comprising: (1) in the case of a heavy chain, a variable region (abbreviated herein as "VH") and a heavy chain constant region comprising three domains, CH1, CH2, and CH3; and (2) in the case of a light chain, a light chain variable region (abbreviated herein as "VL") and a light chain constant region comprising one domain, CL. The antibodies disclosed herein include, but are not limited to, monoclonal, multispecific, human or chimeric antibodies, single-chain antibodies, Fab fragments, F(ab') fragments, anti-idiotypic (anti-Id) antibodies (including, for example, anti-Id antibodies of the disclosed antibodies), and epitope-binding fragments of any of the above antibodies. The immunoglobulin molecules disclosed herein can be of any type (e.g., IgG, IgE, IgM, IgD, IgA, and IgY), class (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2), or subclass of immunoglobulin. Preferably, the antibodies disclosed herein comprise or consist of a VH domain, VHCDR (herein often referred to as HCDR), VL domain, or VLCDR (herein often referred to as LCDR) having any of the amino acid sequences described in the Sequence and Specific Information Tables, or a fragment or variant thereof.

[1145] In this disclosure, the term "monoclonal antibody" refers to an antibody derived from a population of substantially homogeneous antibodies, i.e., the individual antibodies comprising the population are identical except for possible naturally occurring mutations. Monoclonal antibodies have high specificity for a single determinant (epitope) of an antigen, whereas polyclonal antibodies, in contrast, comprise different antibodies directed against different determinants (epitopes). In addition to their specificity, monoclonal antibodies have the advantage of being synthesized without contamination by other antibodies. The modifier "monoclonal" herein indicates that the antibody is characterized by being derived from a substantially homogeneous population of antibodies and should not be construed as requiring production by a specific method.

[1146] In some embodiments of the present disclosure, monoclonal antibodies also specifically include chimeric antibodies, i.e., a portion of the heavy chain and / or light chain is identical or homologous to a certain type, class, or subclass of antibody, and the remaining portion is identical or homologous to another type, class, or subclass of antibody, as long as they have the desired biological activity (see, for example, US 4,816,567; and Morrison et al., 1984, PNAS, 81: 6851-6855). Chimeric antibodies that can be used in the present disclosure include primatized antibodies, which contain variable region antigen-binding sequences from non-human primates (e.g., monkeys, orangutans, etc.) and human constant region sequences.

[1147] The term "antigen-binding fragment" refers to a portion of an antibody, preferably an antigen-binding region or variable region. Examples of antibody fragments include Fab, Fab', F(ab')2, Fd, Fv, dAb and complementary determining region fragments, diabodies, linear antibodies and single-chain antibody molecules. The term "antigen-binding fragment" as used herein refers to a partial fragment of an antibody with antigen-binding activity, wherein the fragment has all or part of the function of an antibody, including but not limited to single-chain Fv (scFv), Fab, Fab', F(ab')2, disulfide-linked Fv (sdFv), Fv, di-scFv, etc. The term also includes Fab', which is a monovalent fragment of the variable region of an antibody obtained by treating F(ab')2 under reducing conditions. However, the term is not limited to these molecules, as long as the fragment has binding affinity to the antigen. In addition, these functional fragments include not only fragments obtained by treating the full-length molecule of the antibody protein with an appropriate enzyme, but also proteins produced in appropriate host cells using genetically modified antibody genes.

[1148] The term "Fab" as used herein 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, the Fab' disclosed herein also includes Fab' produced using genetically modified antibody genes.

[1149] As used herein, the term "scFv" refers to a single polypeptide chain comprising a VL and VH domain, wherein 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, Roseburg and Moore, eds., Springer-Verlag, New York, pp. 269-315 (1994)). Such scFv molecules may have the general structure: NH2-VL-linker-VH-COOH or NH2-VH-linker-VL-COOH. Suitable prior art linkers consist of repeated GGGGS amino acid sequences or variants thereof. For example, a linker having the amino acid sequence (GGGGS) 4 can be used, but variants thereof can also be used (Holliger et al. (1993), Proc. Natl. Acad. Sci. USA 90: 6444-6448). Other linkers useful in the present disclosure are described by Alfthan et al. (1995), Protein Eng. 8: 725-731, Choi et al. (2001), Eur. J. Immunol. 31: 94-106, Hu et al. (1996), Cancer Res. 56: 3055-3061, Kipriyanov et al. (1999), J. Mol. Biol. 293: 41-56, and Roovers et al. (2001), Cancer Immunol. In some cases, a disulfide bond may also be present between the VH and VL of the scFv. As used herein, the term "di-scFv" refers to an antibody fragment formed by linking two scFvs.

[1150] As used herein, the antibody or antigen-binding fragment thereof still has the activity of binding to the antigen even if it contains variants, amino acid substitutions, deletions or additions.

[1151] 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) through a coupling arm. The conjugate retains the activity of each antibody and thus has bifunctionality and bispecificity.

[1152] The term "multispecific antibody" includes, for example, trispecific antibodies, which are antibodies with three different antigen-binding specificities, and tetraspecific antibodies, which are antibodies with four different antigen-binding specificities.

[1153] 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), and a heavy chain constant region (CH1, CH2, and CH3). The constant region may be a native sequence (e.g., a native human constant region sequence) or an amino acid sequence variant thereof. An intact antibody preferably is an intact antibody having one or more effector functions.

[1154] The term "probody" refers to a modified antibody, including an antibody or an antibody fragment, that can specifically bind to its target and can be coupled to a masking group, wherein the masking group refers to a cleavage constant for the binding ability of the antibody or antibody fragment to its target that is at least 100 times, 1000 times, or 10,000 times greater than the cleavage constant for the binding ability of the antibody or antibody fragment to its target without the coupled masking group.

[1155] In the present disclosure, the "humanized" form of a non-human (e.g., mouse) antibody refers to a chimeric antibody that contains a minimal amount of non-human immunoglobulin sequence. Most humanized antibodies are those in which the hypervariable region residues of a human recipient immunoglobulin are replaced with 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, the framework region (FR) residues of the human immunoglobulin are also replaced with non-human residues. Moreover, the humanized antibody may also include residues that are not present in the recipient antibody or the donor antibody. These modifications are to further optimize the performance of the antibody. Humanized antibodies generally include at least one, typically two, variable regions in which all or nearly all of the hypervariable loops correspond to those of a non-human immunoglobulin, while the FRs are entirely or almost entirely human immunoglobulin sequences. The humanized antibody may also include at least a portion of an immunoglobulin constant region (Fc, typically a human immunoglobulin Fc). For details, see, for example, 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.

[1156] Intact antibodies can be divided into different "classes" based on the amino acid sequence of the constant region of their heavy chains. The five main classes are IgA, IgD, IgE, IgG, and IgM, and several of these classes are further divided into different "subclasses" (isotypes), such as IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2. The heavy chain constant regions of the different antibody classes are called α, β, ε, γ, and μ, respectively. The subunit structures and three-dimensional configurations of the different classes of immunoglobulins are well known in the art.

[1157] As used herein, the CDRs contained in the antibodies or antigen-binding fragments thereof of the present disclosure may be identified using various numbering systems known in the art. In certain embodiments, the CDRs contained in the antibodies or antigen-binding fragments thereof of the present disclosure are preferably identified using the Kabat, Chothia, AbM, or IMGT numbering systems. According to the IMGT numbering system, the corresponding numbers for the CDRs and FRs are as follows:

[1158]

[1159]

[1160] As used herein, the term "framework residue region" or "FR residues" refers to those amino acid residues in an antibody variable region other than the CDR residues as defined above.

[1161] As used herein, the term "germline antibody gene" refers to a gene encoding an immunoglobulin expressed by non-lymphocytes that has not undergone the genetic rearrangement and maturation process that results in the expression of a specific immunoglobulin. One advantage provided by various embodiments of the present disclosure stems from the recognition that the amino acid sequences encoded by germline antibody genes retain more of the important amino acid sequence structure characteristic of individual animal species than the amino acid sequences encoded by mature antibody genes. Therefore, when therapeutically applied to that species, the amino acid sequences are less likely to be recognized as foreign by that species.

[1162] As amino acid substitutions, in this specification, conservative amino acid substitutions are preferred. Conservative amino acid substitutions refer to substitutions that occur within a set of amino acids related to the amino acid side chain. Preferred amino acid sets are as follows: acidic set (aspartic acid and glutamic acid); basic set (lysine, arginine and histidine); non-polar set (alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine and tryptophan); and uncharged polar family (glycine, asparagine, glutamine, cysteine, serine, threonine and tyrosine). More preferred amino acid sets are as follows: aliphatic hydroxyl (serine and threonine); amide-containing set (asparagine and glutamine); aliphatic set (alanine, valine, leucine and isoleucine); and aromatic set (phenylalanine, tryptophan and tyrosine). Such amino acid substitutions are preferably carried out within a set that does not impair the performance of the substance having the original amino acid sequence.

[1163] 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) at the carboxyl terminus of the heavy chain of an antibody produced in cultured mammalian cells are deleted and the proline residue newly located 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.

[1164] The compilation of the twenty conventional amino acids involved herein follows conventional usage. See, for example, Immunology—A Synthesis (2nd Edition, E.S. Golub and D.R. Gren, Eds., Sinauer Associates, Sunderland, Mass. (1991)), which is incorporated herein by reference. 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 single-letter and three-letter abbreviations 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.

[1165] As used herein, the term "prevention" refers to a method implemented in order to prevent or delay the occurrence of a disease or illness or symptom (e.g., tumor and infectious disease) in a subject. As used herein, the term "treatment" refers to a method implemented in order to obtain a beneficial or desired clinical outcome. For the purposes of this disclosure, beneficial or desired clinical outcomes include, but are not limited to, alleviating symptoms, reducing the scope of the disease, stabilizing (i.e., no longer worsening) the state of the disease, delaying or slowing the development of the disease, improving or alleviating the state of the disease, and alleviating symptoms (whether partially or entirely), whether detectable or undetectable. In addition, "treatment" can also refer to, compared to the expected survival (if not receiving treatment), extending the survival period.

[1166] As used herein, the term "subject" refers to a mammal, such as a primate mammal, such as a non-human primate mammal or a human. In certain embodiments, the subject (e.g., human) has a tumor and an infectious disease, or is at risk of having the above-mentioned disease.

[1167] As used herein, the term "effective amount" refers to an amount sufficient to obtain or at least partially obtain the desired effect. For example, an effective amount for preventing a disease (e.g., tumors and infectious diseases) refers to an amount sufficient to prevent, stop, or delay the occurrence of a disease (e.g., tumors and infectious diseases); an effective amount for treating a disease refers to an amount sufficient to cure or at least partially stop the disease and its complications in a patient already suffering from the disease. Determining such an effective amount is well within the capabilities of those skilled in the art. For example, an amount effective for therapeutic use will depend on the severity of the disease to be treated, the overall state of the patient's own immune system, the patient's general condition such as age, weight and sex, the mode of administration of the drug, and other treatments administered simultaneously, etc.

[1168] As used herein, the term "effector function" refers to those biological activities attributable to the Fc region (a native sequence Fc region or an amino acid sequence variant Fc region) of an antibody, and varies with the antibody isotype.

[1169] The term "pharmaceutically acceptable" means that when the molecule itself, molecule fragment or composition is appropriately administered to an animal or human, it does not produce adverse, allergic or other untoward reactions. Specific examples of some substances that can serve as pharmaceutically acceptable carriers or components thereof include sugars (such as lactose), starch, cellulose and its derivatives, vegetable oils, gelatin, polyols (such as propylene glycol), alginic acid, etc.

[1170] 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 tumor cell line expressing B7H3 and measuring any changes in the cancer cells.

[1171] Although amino acid substitutions in antibodies are generally made with L-amino acids in the present disclosure, this is not limiting. In some embodiments, the antibody peptide chain may include one or more D-amino acids. Peptides containing D-amino acids are more stable and less susceptible to degradation in the oral cavity, intestinal tract, or plasma than peptides containing only L-amino acids.

[1172] The monoclonal antibodies used in the present disclosure can be produced by many methods. For example, the monoclonal antibodies used in the present disclosure can be obtained by the hybridoma method using cells from many species (including mice, hamsters, rats and humans) (see, for example, Kohler et al., 1975, Nature, 256:495), or by recombinant DNA technology (see, for example, US 4,816,567), or isolated from phage antibody libraries (see, for example, Clackson et al., 1991, Nature, 352:624-628; and Marks et al., 1991, Journal of Molecular Biology, 222:581-597).

[1173] Herein, unless otherwise expressly stated, the descriptions "each independently selected from" and "each independently selected from" used herein are interchangeable and should be understood in a broad sense. They can mean that in different groups, the specific options expressed by the same or different symbols do not affect each other, or that in the same group, the specific options expressed by the same or different symbols do not affect each other.

[1174] Herein, the term "direct bond" means that the groups on both sides are directly connected. For example, the compound shown in formula II If X is a direct bond, the structural formula is The remaining direct keys can be understood by referring to the above content.

[1175] Herein, the term "absence" means that the group does not exist. For example, the compound shown in formula II If W does not exist, the structural formula is

[1176] Compound represented by formula II In the equation, R1 and R2 form a carbon atom connected to them. The "dashed" bond indicates the position where the heterocycle is fused to the benzene ring, e.g., to form

[1177] Herein, the drug-linker conjugate shown in Formula III In the L4 When the numbers 1 and 2 represent the connection positions of L4 and the other groups, specifically, position 1 is connected to L3, position 2 is connected to the drug molecule, i.e. When connected, it forms The remaining reference numerals 1 and 2 can be understood by referring to the above content.

[1178] Herein, the compound shown in formula II In the formula, R3 and X together with the carbon atom to which they are attached form The dotted line indicates where the carbon ring is fused to the benzene ring and the pyridine ring to form

[1179] Herein, the definition of X is, for example, "X is selected from the group consisting of optionally substituted The substituent is selected from 1 or 2 C1-4 alkyl groups (such as methyl groups), and X can be, for example Other similar definitions of X can be understood with reference to the above content.

[1180] Herein, the definition of X is, for example, "X is selected from the group consisting of optionally substituted The substituent is selected from two C1-4 alkyl groups (such as methyl) and the carbon atoms to which they are simultaneously connected together to form a C3-6 cycloalkyl group (such as cyclopropyl). Then X can be, for example Other similar definitions of X can be understood with reference to the above content.

[1181] AA 1 The structure of the indicated amino acid residues If r is 0, it will be understood by those skilled in the art that AA 1 The structure of the amino acid residues shown will become

[1182] AA 1 The structure of the indicated amino acid residues In, if R a With R b Together with the carbon atoms to which they are attached, they form a 4-10 membered heterocyclic ring, which is optionally substituted by one or more R 0 substituted, wherein the term "the 4-10 membered heterocyclic ring is optionally substituted by one or more R 0 "substituted" means that the 4-10 membered heterocyclic ring may be unsubstituted or may be replaced by one or more R 0 substituted, and the plurality of R 0 In each R 0 The definitions of can be the same or different. Other similar definitions can be understood with reference to the above content.

[1183] 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, Lys-Ala-Asn, “the distal amino group of lysine (Lys) is optionally substituted with 1, 2 or 3 substituents selected from tert-butyloxycarbonyl, C1-6 alkyl (preferably methyl), and O” means that the distal amino group of Lys in each option of L3 is optionally substituted with 1, 2 or 3 substituents selected from tert-butyloxycarbonyl, C1-6 alkyl (preferably methyl), and O. Wherein, “the distal amino group of Lys” refers to the lysine residue The exposed amino group -NH2 in the lysine (Lys) is optionally substituted by one, two or three substituents selected from tert-butyloxycarbonyl, C1-6 alkyl (preferably methyl), and O. The expression "the distal amino group of the lysine (Lys) is optionally substituted by one, two or three substituents selected from tert-butyloxycarbonyl, C1-6 alkyl (preferably methyl), and O" means that the distal amino group of the lysine (Lys) may be unsubstituted or substituted by one, two or three substituents selected from tert-butyloxycarbonyl, C1-6 alkyl (preferably methyl), and O. For example, it may be substituted by one tert-butyloxycarbonyl, i.e., it becomes Alternatively, it can be substituted by two methyl groups, i.e. Or it can be replaced by 2 methyl groups and 1 O at the same time, which becomes It should be noted that "the distal amino group of the lysine (Lys) is substituted by O" means that the distal amino group of the lysine (Lys) is oxidized, i.e., becomes If the distal amino group is further substituted by two methyl groups, it becomes

[1184] Throughout this specification, substituents of the disclosed compounds are disclosed by group class or range. It is specifically noted that the disclosure includes every independent subcombination of the individual members of these group classes and ranges. For example, the term "C1-6 alkyl" specifically refers to the independently disclosed methyl, ethyl, C3 alkyl, C4 alkyl, C5 alkyl, and C6 alkyl.

[1185] As used herein, the term "C1-6 alkyl" refers to a linear or branched alkyl group containing 1 to 6 carbon atoms, including, for example, "C1-3 alkyl" or "C1-4 alkyl", methyl, ethyl, etc. Specific examples include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, pentyl, and hexyl.

[1186] As used herein, the term "C1-4 alkyl" refers to a linear or branched alkyl group containing 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, and tert-butyl.

[1187] In this article, the term “C 2-6 "Alkenyl" refers to a linear, branched or cyclic alkenyl group containing at least one double bond and having 2 to 6 carbon atoms, including, for example, "C 2-4 Examples include, but are not limited to, ethenyl, 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.

[1188] In this article, the term “C 2-6 "Alkynyl" refers to a straight or branched chain alkynyl group containing at least one triple bond and having 2 to 6 carbon atoms, including, for example, "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.

[1189] As used herein, the term "halogen" includes fluorine, chlorine, bromine, and iodine.

[1190] In this context, the term "3-6 membered cycloalkyl" or "C 3-6 The term "cycloalkyl" refers to a saturated cyclic alkyl group containing 3 to 6 carbon atoms, including cyclopropane (i.e., cyclopropyl), cyclobutane (i.e., cyclobutyl), cyclopentane (i.e., cyclopentyl), and cyclohexyl.

[1191] As used herein, the term "3-7 membered carbocycloalkyl" or "C 3-7 "Cycloalkyl" refers to a saturated cyclic alkyl group containing 3 to 7 carbon atoms, including cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and cycloheptyl.

[1192] As used herein, the term "C1-6 alkoxy" refers to an alkyl group as defined above that is attached to the parent molecular moiety via an oxygen atom. Specific examples include, but are not limited to, methoxy, ethoxy, propoxy, isopropoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, tert-butoxy, pentyloxy, hexyloxy, and the like.

[1193] In this article, the term “C 1-4 "Alkoxy" refers to an alkyl group, as defined above, attached to the parent molecular moiety through an oxygen atom. Specific examples include, but are not limited to, methoxy, ethoxy, propoxy, isopropoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, tert-butoxy, and the like.

[1194] In this article, the term "4-10 membered heterocyclic radical" refers to a cyclic group containing 4-10 ring atoms (wherein at least one ring atom is a heteroatom, such as a nitrogen atom, an oxygen atom or a sulfur atom). The term "4-6 membered heterocyclic radical" refers to a cyclic group containing 4-6 ring atoms (wherein at least one ring atom is a heteroatom, such as a nitrogen atom, an oxygen atom or a sulfur atom). Optionally, the ring atoms (such as carbon atoms, nitrogen atoms or sulfur atoms) in the cyclic structure can be oxoed. "4-8 membered heterocyclic radical" includes, for example, "4-8 membered nitrogen-containing heterocyclic radical", "4-8 membered oxygen-containing heterocyclic radical", "4-7 membered heterocyclic radical", "4-7 membered oxygen-containing heterocyclic radical", "4-7 membered heterocyclic radical", "4-6 membered heterocyclic radical", "5-7 membered heterocyclic radical", "5-6 membered heterocyclic radical", "5-6 membered nitrogen-containing heterocyclic radical", including but not limited to oxocyclobutane, pyrrolidinyl, tetrahydrofuranyl, piperidinyl, piperazinyl, tetrahydropyranyl, homopiperazinyl, etc.

[1195] As used herein, the term "4-10 membered heterocycle" refers to a ring containing 4-10 ring atoms (at least one of which is a heteroatom, such as a nitrogen atom, an oxygen atom, or a sulfur atom). The term "5-6 membered heterocycle" refers to a ring containing 5-6 ring atoms (at least one of which is a heteroatom, such as a nitrogen atom, an oxygen atom, or a sulfur atom), including but not limited to pyrrolidine, tetrahydrofuran, piperidine, piperazine, tetrahydropyran and the like.

[1196] As used herein, the term "aryl" refers to a monocyclic or polycyclic hydrocarbon group having aromatic properties, such as a 6-10 membered aryl group, a 5-8 membered aryl group, and the like. Specific examples include, but are not limited to, phenyl, naphthyl, anthracenyl, and phenanthrenyl. The "6-10 membered aryl" refers to an aryl group containing 6-10 ring atoms. The "C6-10 aryl" refers to an aryl group containing 6-10 carbon atoms.

[1197] In this article, term " heteroaryl " refers to the cyclic group with aromaticity, wherein at least one ring atom is a heteroatom, such as a nitrogen atom, an oxygen atom or a sulphur atom. Optionally, the ring atom (such as a carbon atom, a nitrogen atom or a sulphur atom) in the ring structure can be by oxo. Specific examples include but are not limited to 5-10 yuan heteroaryl, 5-6 yuan heteroaryl, 5-10 yuan nitrogen-containing heteroaryl, 6-10 yuan oxygen-containing heteroaryl, 6-8 yuan nitrogen-containing heteroaryl, 5-8 yuan oxygen-containing heteroaryl etc., 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, 1,3, 4-oxadiazolyl, pyridinyl, 2-pyridonyl, 4-pyridonyl, pyrimidinyl, 1,4-dioxadienyl, 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.

[1198] Bonds in structural formulae represented herein by a wavy line "~~" are intended to indicate that the structure represents either a cis or trans isomer, or a mixture of cis and trans isomers in any ratio.

[1199] The term "drug to antibody ratio" or "DAR" refers to the amount of drug, e.g., a small molecule toxin, attached to the antibody of an ADC. The DAR of an ADC can range from 1 to 16, but higher loadings (e.g., 20) are possible depending on the number of attachment sites on the antibody. The term DAR can be used when referring to the amount of drug loaded onto a single antibody, or alternatively, when referring to the average or mean DAR of a group of ADCs.

[1200] Advantageous Effects of the Invention

[1201] The present disclosure has obtained a novel class of antibody-bioactive molecule conjugates through extensive research to obtain bioactive drugs (bioactive molecules or payloads) with high activity and high cell permeability, linkers with high in vivo circulation stability and high chemical stability, enrichment of antibody-drug conjugates (ADCs) in the tumor microenvironment by regulating the physicochemical properties of the linker (such as alkalinity and lipophilicity), the unique in vivo enzymatic cleavage properties of the linker and the conjugation mode with the targeting moiety, combined with a large number of in vivo and in vitro drug efficacy screening and verification. The conjugates obtained by the above methods can achieve a variety of the following surprising technical effects:

[1202] The conjugate obtained by the above method has better solubility and excellent chemical stability. For example, the reversible Michael addition reaction caused by the maleimide linkage in traditional ADCs does not occur, so a high drug-antibody ratio can be obtained. In some embodiments, the DAR value of the conjugate can reach 6-8.

[1203] Having extremely high coupling efficiency. In some embodiments, the coupling efficiency can reach or exceed 90%;

[1204] Through extensive research, a class of linkers has been discovered that have high plasma stability but can be cleaved in the tumor microenvironment (both inside and outside tumor cells). Therefore, they can produce good anti-tumor effects in tumors with low or no antigen expression.

[1205] The conjugate (ADC) obtained by the above method improves the exposure of the entire ADC molecule to the relatively acidic tumor environment by adjusting the physicochemical properties of the linker and the entire ADC molecule. As a result, the ADC has better tumor tissue targeting, that is, the ability to be enriched in the tumor microenvironment, increases the ratio of the bioactive molecule concentration in the tumor to that in the blood, and reduces the mechanism-related toxicity of the ADC molecule (toxicity caused by the ADC binding to cell surface antigens in non-tumor tissues and internalization, also known as "on-target toxicity"), thereby having a higher therapeutic index;

[1206] The conjugate obtained according to the above method has high stability in the body circulation, reduces the shedding of drug molecules in non-target tissues, and reduces the "off-target" toxicity caused by the shedding of toxins in non-target tissues;

[1207] The bioactive molecule of the conjugate has higher anti-tumor cell activity and thus has an excellent bystander effect. The ADC can more effectively kill tumor cells with high antigen expression as well as tumor cells with low or no antigen expression in tumor tissues.

[1208] The toxin-linker disclosed herein can be combined with antibodies without endocytosis ability to form antibody-drug conjugates, utilizing its extracellular cleavage ability in the tumor microenvironment. Such antibody-drug conjugates still have high anti-tumor activity.

[1209] The toxin-linker disclosed herein, utilizing its extracellular cleavage ability and enrichment ability in the tumor microenvironment, can be combined with antibodies that lack endocytosis ability and antibodies that lack tumor extracellular antigen binding ability to form antibody-drug conjugates. Such antibody-drug conjugates still have high anti-tumor activity.

[1210] The present disclosure provides antibodies with a high degree of humanization or fully human antibodies, which can be safely administered to human subjects without eliciting an immunogenic response. The present disclosure also provides antibodies with a high ability to bind to both human and non-human B7H3, particularly antibodies that have cross-reactivity with B7H3 from non-human primates (e.g., cynomolgus macaques).

[1211] In summary, the linker, antibody and ADC of the present invention have significant clinical value. BRIEF DESCRIPTION OF THE DRAWINGS

[1212] Figure 1A ELISA binding detection of anti-B7H3 antibody 1D1-01 and human B7H3-4 IgG-His protein

[1213] Figure 1B ELISA binding detection of anti-B7H3 antibody 2E3-02 and human B7H3-4 IgG-His protein

[1214] Figure 2A ELISA binding detection of anti-B7H3 antibody 1D1-01 and monkey B7H3-4 IgG-His protein

[1215] Figure 2B ELISA binding detection of anti-B7H3 antibody 2E3-02 and monkey B7H3-4 IgG-His protein

[1216] Figure 3A Flow cytometry detection of anti-B7H3 antibody 1D1-01 binding to MCF-7 tumor cells

[1217] Figure 3B Flow cytometry detection of anti-B7H3 antibody 2E3-02 binding to MCF-7 tumor cells

[1218] Figure 4A Flow cytometry detection of anti-B7H3 antibody 1D1-01 binding to A549 tumor cells

[1219] Figure 4B Flow cytometry detection of anti-B7H3 antibody 2E3-02 binding to A549 tumor cells

[1220] Figure 5 Flow cytometry detection of anti-B7H3 antibody 2E3-02 binding to PC-3 tumor cells

[1221] Figure 6A Detection of specific binding of anti-B7H3 antibody 1D1-01 to B7H3 protein

[1222] Figure 6B Detection of specific binding of anti-B7H3 antibody 2E3-02 to B7H3 protein

[1223] Figure 7A Detection of endocytosis of candidate antibody 1D1-01 by tumor cells A549

[1224] Figure 7B Detection of endocytosis of candidate antibody 2E3-02 by tumor cells A549

[1225] Figure 7C Detection of endocytosis of candidate antibody 202-2-1 by tumor cells A549

[1226] Figure 8 SEC spectra before and after coupling

[1227] Figure 9: Test 1 of anti-B7H3-ADC inhibiting tumor growth in the NCI-HT29 xenograft tumor model

[1228] Figure 10: Anti-B7H3-ADC inhibition of tumor growth in the NCI-HT29 xenograft tumor model (Part 2)

[1229] Figure 11 Test of anti-B7H3-ADC inhibiting tumor growth in NCI-H358 xenograft tumor model

[1230] Figure 12 Test of anti-B7H3-ADC inhibiting tumor growth in esophageal squamous cell carcinoma PDX model

[1231] Figure 13: Anti-B7H3-ADC inhibition of tumor growth in prostate PDX model

[1232] Figure 14 Test of anti-Her3-ADC inhibiting tumor growth in NCI-H358 transplanted tumor model

[1233] Figure 15: Test 1 of anti-Her3-ADC inhibiting tumor growth in SW480 xenograft tumor model

[1234] Figure 16: Test 2 of anti-Her3-ADC inhibiting tumor growth in SW480 transplanted tumor model

[1235] Figure 17 Distribution ratio of A1.9 in plasma and tumor in HT29 model

[1236] Figure 18 shows the distribution ratio of ADC in plasma and tumor in the HT29 model.

[1237] Figure 19A Antibody-drug conjugate incubated in tumor homogenate and detected results

[1238] Figure 19B: Results of incubation of antibody-drug conjugates in muscle tissue homogenate DETAILED DESCRIPTION

[1239] The present disclosure will be further described below by way of specific embodiments, but this is not intended to limit the present disclosure. Those skilled in the art may make various modifications or improvements based on the teachings of the present disclosure without departing from the basic concept and scope of the present disclosure. Reagents or instruments used without indicating the manufacturer are conventional products that can be purchased commercially.

[1240] The abbreviations in this disclosure have the following meanings:

[1241]

[1242]

[1243] Sequence and its specific information:

[1244]

[1245]

[1246]

[1247]

[1248]

[1249]

[1250]

[1251]

[1252]

[1253] The present disclosure will now be described with reference to the following examples which are intended to illustrate the present disclosure rather than to limit it.

[1254] Unless otherwise specified, the molecular biology experimental methods and immunoassays used in the present disclosure are basically carried out with reference to the methods described in J. Sambrook et al., Molecular Cloning: A Laboratory Manual, 2nd edition, Cold Spring Harbor Laboratory Press, 1989, and FM Ausubel et al., Molecular Biology: A Compendium of Laboratory Manuals, 3rd edition, John Wiley & Sons, Inc., 1995. It will be appreciated by those skilled in the art that the embodiments describe the present disclosure by way of example and are not intended to limit the scope of protection claimed in the present disclosure.

[1255] Preparation plan

[1256] The structures of the compounds described in the following examples were determined by nuclear magnetic resonance ( 1 H NMR) or mass spectrometry (MS).

[1257] Nuclear magnetic resonance (NMR) 1 H NMR was measured using a Bruker 400 MHz nuclear magnetic resonance instrument; the measurement solvent was deuterated methanol (CD3OD), deuterated chloroform (CDCl3), or hexadeuterated dimethyl sulfoxide (DMSO-d6); and the internal standard was tetramethylsilane (TMS).

[1258] The abbreviations used in the nuclear magnetic resonance (NMR) spectra in the examples are shown below.

[1259] 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.

[1260] The mass spectrometry (MS) was performed using an Agilent (ESI) mass spectrometer, model Agilent 6120B.

[1261] 1. Synthesis of Bioactive Molecules and Intermediates Used in the Synthesis of Drug-Linker Compounds

[1262] A. Synthesis of Bioactive Molecules

[1263] 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)

[1264]

[1265] Step 1: 4-Bromo-1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazole (4.6 g) was dissolved in anhydrous tetrahydrofuran (50 ml). The above solution was cooled to -78°C with dry ice-acetone under nitrogen protection, and then n-butyl lithium (12 ml, 2M) 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 was complete, the reaction mixture was naturally warmed to room temperature and stirred for 5 hours. The reaction mixture was quenched with methanol (3 ml) and ethyl acetate (200 ml) was added. The solution was washed with water (100 ml x 3). The organic phase was dried and the organic solvent was removed. The target product (2-amino-4-fluoro-5-methylphenyl)(1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazol-4-yl)methanone was separated by silica gel column chromatography. ESI-MS (m / z): 304 [M+H] + .

[1266] 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 for 4 hours. The mixture was dissolved in ethyl acetate (300 ml), the organic phase was washed with water (100 ml x 2), dried, and the organic solvent 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] + .

[1267] Example A1.2: Synthesis of (S)-7-ethyl-7-hydroxy-14-(1H-pyrazol-4-yl)-10,13-dihydro-11H-[1,3]dioxo[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (A1.2)

[1268]

[1269] Using the same method and reaction conditions as in Example A1.1, substituting methyl 6-aminobenzo[d][1,3]dioxolane-5-carboxylate for methyl 2-amino-4-fluoro-5-methylbenzoate, 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) was obtained. ESI-MS (m / z): 459 [M+H]. + .

[1270] Example A1.3: Synthesis of (S)-14-(3-aminophenyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolano[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (A1.3)

[1271]

[1272] Compound (A1.3-A) (1.4 g), compound (A1.3-B) (2.2 g), Pd2(DBA)3 (300 mg), tricyclohexylphosphine (300 mg), and potassium acetate (1.1 g) were added sequentially to a mixed solvent of dioxane (30 ml) and water (5 ml). The mixture was heated and stirred at 100°C under nitrogen for 12 hours. After cooling, ethyl acetate (200 ml) was added, the mixture was washed with water (100 ml), dried, and separated by silica gel column chromatography to obtain the target product, (S)-14-(3-aminophenyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolano[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (A1.3). ESI-MS (m / z): 484 [M+H] + .

[1273] Example A1.4: Synthesis of (S)-14-(4-aminophenyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolano[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (A1.4)

[1274]

[1275] Compound (A1.4-A) (1.4 g), compound (A1.4-B) (2.2 g), Pd2(DBA)3 (300 mg), tricyclohexylphosphine (300 mg), and potassium acetate (1.1 g) were added sequentially to a mixed solvent of dioxane (30 ml) and water (5 ml). The mixture was heated and stirred at 100°C under nitrogen for 12 hours. After cooling, ethyl acetate (200 ml) was added, the mixture was washed with water (100 ml), dried, and separated by silica gel column chromatography to obtain the target product (S)-14-(4-aminophenyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolano[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (A1.4).

[1276] ESI-MS (m / z): 484 [M+H] + ;

[1277] 1H 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).

[1278] Example A1.5: Synthesis of (S)-14-(3-aminopropyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolano[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (A1.5)

[1279]

[1280] Step 1: Synthesis of (S)-14-(3-chloropropyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolano[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione

[1281]

[1282] Under ice-cooling conditions, to a 75% sulfuric acid solution (5 mL) of compound (S) 7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxetine[4,5-g]pyrano[3′,4′:6,7]indeno[1,2-b]quinoline-8,11(7H)-dione (A1.5-A, 500 mg) were added ferrous sulfate heptahydrate (570 mg dissolved in 1 mL of water) and 4,4-dimethoxychlorobutane (3.89 g). The reaction mixture was stirred for three minutes, after which hydrogen peroxide (29%, 2.5 mL) was added dropwise. The reaction mixture was stirred at 0°C for 5 minutes, then warmed to room temperature and stirred for 3 hours. The reaction solution was diluted with water (50 mL) and extracted with ethyl acetate (80 mL × 2). The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to give a crude product. The crude product was further purified using a C18 column (acetonitrile / 0.05% formic acid aqueous solution: 5%-60%) to give the title compound (yellow solid, 400 mg, yield: 67%).

[1283] LCMS (ESI) [M+H] + :468.9;

[1284] 1H 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).

[1285] Step 2: Synthesis of (S)-14-(3-azidopropyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolano[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione

[1286]

[1287] To a solution (3 mL) of compound (S)-14-(3-chloropropyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolane-[4,5-g]pyrano[3′,4′:6,7]indolizine-[1,2-b]quinoline-8,11(7H)-dione (200 mg) in N,N-dimethylformamide was added sodium azide (284 mg), and the reaction solution was stirred at 100°C for 1 hour. Water (30 mL) was added to the reaction solution, and the mixture 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]dioxolane-[4,5-g]pyrano[3′,4′:6,7]indolizine-[1,2-b]quinoline-8,11(7H)-dione (yellow solid, 180 mg, yield: 88%).

[1288] LCMS (ESI) [M+H] + :476;

[1289] 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).

[1290] Step 3: Synthesis of (S)-14-(3-aminopropyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolano[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione

[1291]

[1292] To a mixture of (S)-14-(3-azidopropyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolano[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (130 mg, 0.274 mmol) in tetrahydrofuran (3 mL) and water (1 mL) was added triphenylphosphine (108 mg, 0.411 mmol) 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, which was acidified with 2N hydrochloric acid (3 mL) and extracted with ethyl acetate (10 mL). The aqueous phase was purified by 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]dioxolane-[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%).

[1293] LCMS (ESI) [M+H] + :449.9;

[1294] 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).

[1295] 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]dioxolano[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)-2-hydroxyacetamide (A1.6)

[1296]

[1297] Step 1: Preparation of (S)-2-(ethyl(2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxol[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)amino)-2-oxoethyl acetate (A1.6-B):

[1298]

[1299] Compound (S)-7-ethyl-14-(2-(ethylamino)ethyl)-7-hydroxy-10,13-dihydro-11H-[1,3]dioxetine[4,5-g]pyrano[3′,4′:6,7]indolizine[1,2-b]quinoline-8,11(7H)-dione (A1.6-A, 50 mg), acetoxyacetyl chloride (73 mg), and triethylamine (50 mg) were added sequentially to dichloromethane (5 mL) and stirred at room temperature for 1 hour. The reaction solution was evaporated to remove the solvent to obtain 75 mg of the crude oily compound.

[1300] LCMS (ESI) [M+H] + :564.2.

[1301] Step 2: Synthesis of (S)-N-ethyl-N-(2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxolano[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)-2-hydroxyacetamide (A1.6)

[1302]

[1303] The crude compound (S)-2-(ethyl(2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxol[4,5-g]pyrano[3′,4′:6,7]indolizine[1,2-b]quinolin-14-yl)ethyl)amino)-2-oxoethyl acetate (75 mg, 0.13 mmol) was dissolved in a mixed solution of concentrated hydrochloric acid and anhydrous ethanol (volume ratio of 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% trifluoroacetic acid in water, 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]dioxolano[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)-2-hydroxyacetamide (A1.6) (15 mg, yield: 26%) as a white solid.

[1304] LCMS (ESI) [M+H] + :522.0;

[1305] 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).

[1306] 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]dioxolano[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)-2-hydroxyacetamide (A1.7)

[1307]

[1308] Step 1: Preparation of compound methylbenzyl (3-(6-nitrobenzo[d][1,3]dioxetyl-5-yl)-3-oxopropyl)carbamate

[1309]

[1310] 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 reacted in a sealed container at 100°C for 16 hours. The reaction mixture 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 resulting aqueous phase was adjusted to pH 9 with sodium bicarbonate, and then benzyl chloroformate (513 mg, 3.0 mmol) was added and allowed to react 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 give a crude product. The crude product was separated and purified by C18 (acetonitrile / water containing 0.05% formic acid = 5-95%) to give the target compound (A1.7-B, 180 mg, yield 23%).

[1311] LCMS (ESI) [M+H] + :387.1.

[1312] 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).

[1313] Step 2: Preparation of compound benzyl (3-(6-aminobenzo [d] [1,3] dioxin-5-yl) -3-oxypropyl) (methyl) carbamate (A1.7-C)

[1314]

[1315] Compound A1.7-B, methylbenzyl(3-(6-nitrobenz[d][1,3]dioxetyl-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). Iron powder (130 mg) was then added to the reaction mixture, and the reaction was stirred at 80°C for 2 hours. The reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to a solid. The crude product was separated and purified by TLC (petroleum ether:ethyl acetate = 2 / 1) to obtain the target compound A1.7-C (76 mg).

[1316] LCMS (ESI) [M+H] + :357.0.

[1317] Step 3: Preparation of benzyl (S)-(2-(7-ethyl-7-hydroxy-8,11-dioxy-7,8,11,13-tetrahydro-10H-[1,3]dioxetane[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)(methyl)carbamate (A1.7-D)

[1318]

[1319] At room temperature, 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). The solution was clarified and mixed, then concentrated under reduced pressure and evacuated to a vacuum using an oil pump. The reaction was incubated at 120°C under vacuum for 2 hours. The reaction solution was cooled to room temperature, water (30 mL) was added, and the mixture was extracted with dichloromethane (30 mL x 3). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain the target compound A1.7-D (50 mg).

[1320] LCMS (ESI) [M+H] + =584.0.

[1321] Step 4: Preparation of (S) 7-ethyl-7-hydroxy-14-(2-(methylamino)ethyl)-10,13-dihydro-11H-[1,3]dioxetine[4,5-g]pyrano[3′,4′:6,7]indolizine[1,2-b]quinoline-8,11(7H)-dione (A1.7-E)

[1322]

[1323] Compound A1.7-D benzyl (S)-(2-(7-ethyl-7-hydroxy-8,11-dioxy-7,8,11,13-tetrahydro-10H-[1,3]dioxetane[4,5-g]pyrano[3′,4′:6,7]indolizine[1,2-b]quinolin-14-yl)ethyl)(methyl)carbamate (50 mg) was dissolved in dichloromethane (5 mL) at room temperature, and trimethylsilyl iodide (51 mg, 0.26 mmol) was added at 0°C and reacted for 3 hours. The reaction solution was concentrated to remove the solvent and purified by HPLC (acetonitrile / water containing 0.05% formic acid) to give compound A1.7-E (15 mg) as a brown solid.

[1324] LCMS (ESI) [M+H] +:450.0.

[1325] Step 5: Preparation of compound (S)-2-((2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxol[4,5-g]pyrano[3′,4′:6,7]indolizin[1,2-b]quinolin-14-yl)ethyl)(methyl)amino)-2-oxoethyl acetate (A1.7-F)

[1326]

[1327] Compound A1.7-E(S)7-ethyl-7-hydroxy-14-(2-(methylamino)ethyl)-10,13-dihydro-11H-[1,3]dioxetine[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). Acetoxyacetyl chloride (20 mg) was added and the mixture was reacted at 0°C for 1 hour. The reaction solution was concentrated to remove the solvent to obtain a crude product. The resulting solid was used directly in the next step without further purification.

[1328] LCMS (ESI) [M5+H] + =550.1.

[1329] Step 6: Preparation of compound (S)-N-(2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxetane[4,5-g]pyrano[3′,4′:6,7]indolizine[1,2-b]quinolin-14-yl)ethyl)-2-hydroxy-N-methylacetamide (A1.7)

[1330]

[1331] Compound A1.7-F(S)-2-((2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxol[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-14-yl)(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 purified by high performance liquid chromatography (acetonitrile / water containing 0.05% formic acid) to give compound A1.7 (1.6 mg) as a white solid.

[1332] LCMS (ESI) [M+H] + :508.2.

[1333] Example A1.8: Synthesis of (S)-N-isopropyl-N-(2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxolano[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)-2-hydroxyacetamide (A1.8)

[1334]

[1335] Step 1: Preparation of isopropylbenzyl (3-(6-nitrobenzo[d][1,3]dioxin-5-yl)-3-oxopropyl)carbamate (A1.8-B)

[1336]

[1337] To a solution of compound (A1.8-A) 3-(isopropylamino)-1-(6-nitrobenzo[d][1,3]dioxin-5-yl)propan-1-one (900 mg) in dichloromethane (10 mL) were added triethylamine (1.8 g) and benzyloxyphosgene (734 mg, 4.3 mmol) in sequence, and the mixture was allowed to react at room temperature for 2 hours. The reaction mixture 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 obtain a crude product. The crude product was separated and purified by column chromatography (petroleum ether:ethyl acetate = 3 / 1) to obtain the target compound (A1.8-B) (600 mg).

[1338] LCMS (ESI) [M+H] + :414.9;

[1339] 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).

[1340] Step 2: Preparation of compound benzyl (3-(6-aminobenzo [d] [1,3] dioxin-5-yl) -3-oxopropyl) (isopropyl) carbamate (A1.8-C)

[1341]

[1342] Compound (A1.8-B) isopropylbenzyl (3-(6-nitrobenz[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). Iron powder (135 mg) was then added to the reaction mixture, and the reaction was stirred at room temperature for 2 hours. The reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to a solid. The crude product was separated and purified by column chromatography (petroleum ether:ethyl acetate = 3 / 1) to obtain the target compound (A1.8-C) (65 mg).

[1343] LCMS (ESI) [M+H] + :395.2.

[1344] Step 3: Preparation of (S)7-ethyl-7-hydroxy-14-(2-(isopropylamino)ethyl)-10,13-dihydro-11H-[1,3]dioxetine[4,5-g]pyrano[3′,4′:6,7]indolizine[1,2-b]quinoline-8,11(7H)-dione (A1.8-D)

[1345]

[1346] 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) and p-toluenesulfonic acid (33 mg, 0.17 mmol) were dissolved in dichloromethane (10 mL) solution. After the solution was clarified and mixed, it was concentrated under reduced pressure and evacuated to vacuum with an oil pump. The reaction was carried out under vacuum at 120 ° C for 2 hours. The reaction solution was cooled to room temperature, and water (50 mL) was added. The mixture was extracted with dichloromethane (30 mL × 3). The combined organic phases were dried over anhydrous sodium sulfate and filtered. The filtrate was concentrated under reduced pressure. The crude product was separated and purified by column chromatography (dichloromethane / methanol = 10 / 1) to obtain the target compound (A1.8-D) (40 mg).

[1347] LCMS (ESI) [M+H] + :478.0;

[1348] 1H NMR (400MHz, DMSO-d6) δ8.46 (s, 1H), 7.67 (s, 1H), 7.57 (s, 1H), 7.27 (s, 1H), 6.52 (s, 1H), 6.33 (s, 2H), 5.44 (s, 2H), 5.35 (s, 2H), 3.41 (br s, 2H), 3.23 (br s, 3H), 1.94-1.78 (m, 2H), 1.25 (d, J=6.4Hz, 6H), 0.88 (t, J=7.3Hz, 3H).

[1349] Step 4: Preparation of compound ((S)-2-((2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxa[4,5-g]pyrano[3′,4′:6,7]indolizin[1,2-b]quinolin-14-yl)ethyl)(isopropyl)amino)-2-oxoacetic acid ethyl ester (A1.8-E)

[1350]

[1351] To a dichloromethane solution (2 mL) of compound (S)-7-ethyl-7-hydroxy-14-(2-(isopropylamino)ethyl)-10,13-dihydro-11H-[1,3]dioxacyclo[4,5-g]pyrano[3′,4′:6,7]indolizine[1,2-b]quinoline-8,11(7H)-dione (A1.8-D) (40 mg) was added triethylamine (35 mg) and 2-chloro-2-oxoethyl acetate (57 mg) under ice-cooling. The mixture was reacted at zero degrees for 30 minutes. The reaction solution was concentrated under reduced pressure to afford compound A1.8-E, which was used directly in the next step.

[1352] LCMS (ESI) [M+H] + :578.0.

[1353] Step 5: Preparation of compound (S)-N-(2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxo[4,5-g]pyrano[3′,4′:6,7]indolizine[1,2-b]quinolin-14-yl)ethyl)-2-hydroxy-N-isopropylacetamide (A1.8)

[1354]

[1355] To a solution (3 mL) of compound (S)-ethyl 2-((2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxa[4,5-g]pyrano[3′,4′:6,7]indolizin[1,2-b]quinolin-14-yl)ethyl)(isopropyl)amino)-2-oxoacetate (A1.8-E, 50 mg) in ethanol was added concentrated hydrochloric acid (0.5 mL), and the mixture was reacted at 70°C for 1 hour. The reaction solution was concentrated to obtain a crude product. The product was then purified by HPLC (acetonitrile / 0.05% formic acid in water) to obtain the title compound (A1.8, 3 mg).

[1356] LCMS (ESI) [M+H] + :464.0;

[1357] 1 H NMR (400MHz, DMSO-d6) δ7.99 (s, 1H), 7.53 (s, 1H), 7.26 (s, 1H), 6.31 (s, 2H), 5.43 (s, 2H), 5.36 (s, 2H), 4.22 (s, 2H), 3.99-3.94 ( m, 1H), 3.44 (dd, J=16.7, 7.8Hz, 2H), 3.33-3.21 (m, 2H), 1.95-1.79 (m, 2H), 1.19 (dd, J=16.4, 5.8Hz, 6H), 0.88 (t, J=7.2Hz, 3H).

[1358] Example A1.9: Synthesis of (S)-7-ethyl-7-hydroxy-14-(3-hydroxypropyl)-10,13-dihydro-11H-[1,3]dioxolano[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (A1.9)

[1359]

[1360] Compound (S)-7-ethyl-7-hydroxy-14-(3-chloropropyl)-10,13-dihydro-11H-[1,3]dioxolano[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (100 mg, 0.213 mmol) was dissolved in 10% sulfuric acid (5 mL) and reacted at 110°C for 48 hours. Saturated sodium bicarbonate (30 mL) was added to the reaction solution, and the mixture was extracted with dichloromethane (10 mL x 5), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. Purification by HPLC (acetonitrile / water containing 0.05% formic acid) gave (S)-7-ethyl-7-hydroxy-14-(3-hydroxypropyl)-10,13-dihydro-11H-[1,3]dioxolane-[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (A1.9, 1.78 mg).

[1361] LCMS (ESI) [M+H] + :451.0;

[1362] 1 H NMR (400MHz, DMSO-d6) δ7.63 (s, 1H), 7.50 (s, 1H), 7.24 (s, 1H), 6.48 (s, 1H), 6.28 (s, 2H), 5.47- 5.37(m, 2H), 5.32-5.19(m, 2H), 3.51-3.46(m, 2H), 3.17-3.13(m, 2H), 1.92-1.76(m, 4H), 0.90- 0.84(m,3H).

[1363] Example A1.10: Synthesis of (S)-4-ethyl-8-fluoro-4-hydroxy-11-(3-hydroxypropyl)-9-methyl-1,12-dihydro-14H-pyrano[3′,4′:6,7]indolizino[1,2-b]quinoline-3,14(4H)-dione (A1.10)

[1364]

[1365] Step 1:

[1366] To a solution of compound A1.10-A (10 g) in 1,2-dichloroethane (200 mL) at 0°C, 1 mol / L boron trichloride (96 mL) and 4-chlorobutyronitrile (9.9 g) were added dropwise. The mixture was stirred at 80°C for 2 hours. The reaction mixture was cooled to room temperature, 2 mol / L hydrochloric acid (90 mL) was added, and the mixture was refluxed at 80°C with stirring for 0.5 hours. The reaction mixture was cooled to room temperature, diluted with a small amount of water, and extracted with dichloromethane (200 mL x 3). The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product. The crude product was separated and purified by column chromatography (petroleum ether:ethyl acetate = 10 / 1) to obtain the target compound A1.10-B (4 g).

[1367] LCMS (ESI) [M+H] + :230.0.

[1368] Step 2:

[1369] To compound A1.10-B (50 mg) was added (S)-4-ethyl-4-hydroxy-7,8-dihydro-1H-pyrano[3,4-f]indolizine-3,6,10(4H)-trione (35 mg) and p-tol...

Claims

1. The ligand-drug conjugate shown in Formula XV, Or a stereoisomer of the ligand-drug conjugate, its prodrug, its pharmaceutically acceptable salt, or its pharmaceutically acceptable solvate. in: Tb is the ligand or target portion that binds to the target. q represents the drug-ligand coupling ratio; D represents a bioactive molecular fragment; L1 is an extension unit; L2 either does not exist or is a connection unit; L3 is selected from amino acid residues or short peptides composed of 2-10 amino acid residues; L4 may or may not exist; if L4 exists, L4 is selected from... Bit 1 is connected to L3, and bit 2 is connected to D.

2. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate as claimed in claim 1, characterized in that, It satisfies one or more of the following conditions: (1) Tb is an antibody or its antigen-binding fragment; (2) q is selected from any value between 0.1 and 16.0; (3) D is a molecular fragment with anti-tumor bioactivity; (4) L1 is selected from: Each Z is independently selected from direct bond, carbon-carbon triple bond, carbon-carbon double bond, C6-10 aryl, 5-10 heteroaryl, amide group, sulfonamide group, imine group and CF2; Rx and Ry are each independently selected from H and C1-4 alkyl groups; 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 between 0 and 20; Position 1 is connected to Tb via an S atom, and position 2 is connected to L2 or L3; (5) L2 either does not exist or exists. If L2 exists, L2 is selected from: y1, y2, y3 and y4 are each independently selected from any integer between 0 and 20, with 1 bit connected to L1 and 2 bits connected to L3; (6) L3 is selected from amino acid residues or short peptides composed of 2-10 amino acid residues; the amino acid residues are selected from natural amino acid residues, non-natural amino acid residues, or amino acids. 1 The indicated amino acid residue or its stereoisomer; AA 1 The structures of the amino acid residues shown are as follows: Where: R a R b Each is independently selected from H, And R a R b Not both H; Or, R a With R b Together with the carbon atoms they are linked to, they form 4-10 membered heterocycles, which are optionally separated by one or more R atoms. 0 Replaced; r、r 1 Each integer is independently selected from any integer between 0 and 20; R m1 R n1 Each is independently selected from H, C1-6 alkyl, C3-6 cycloalkyl, and -COOR. x1 ; R x1 Selected from C1-6 alkyl groups; Or, R m1 With R n1 Together with the nitrogen atoms bonded to them, they form 4-10 membered heterocycles, which are optionally separated by one or more R atoms. 0’ Replaced; R z Selected from C1-6 alkyl groups; R 0 R 0’ Each is independently selected from C1-6 alkyl, C3-6 cycloalkyl, -NR m2 R n2 And optional 4-10 membered heterocyclic groups substituted with C1-6 alkyl groups; R m2 R n2 Each is independently selected from H and C1-6 alkyl groups; (7) L4 either does not exist or exists. If L4 exists, L4 is selected from... Bit 1 is connected to L3, and bit 2 is connected to D.

3. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate as described in claim 2, characterized in that, It satisfies one or more of the following conditions: (1) Tb is an antibody or its antigen-binding fragment, which satisfies one or more of the following conditions: (i) The antibody or its antigen-binding fragment includes Fab, Fab′, F(ab′)2, Fd, Fv, dAb, complementarity-determining region fragment, non-human antibody, humanized antibody, chimeric antibody, fully human antibody, proantibody, monoclonal antibody, bispecific antibody or multispecific antibody; (ii) Tb is a monoclonal antibody or its antigen-binding fragment; (iii) Tb is an antibody or its antigen-binding fragment that has endocytosis, non-endocytosis, or weak endocytosis; (iv) Tb is an antibody or its antigen-binding fragment that has the activity of binding free antigens in tumor tissue and / or antigens on the surface of tumor cells; (v)Tb is an antibody or its antigen-binding fragment that does not have the activity of binding to free antigens in tumor tissue and / or antigens on the surface of tumor cells; (2) q is selected from any value between 0.1, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 and ; (3) In the bioactive molecular fragment, the bioactive molecule is a DNA topoisomerase inhibitor or a microtubule protein inhibitor. (4) L1 is selected from Each Z is independently selected from direct bond, carbon-carbon triple bond, carbon-carbon double bond, C6-10 aryl, 5-10 heteroaryl, and amide group; 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; each y2 is independently selected from any integer between 0 and 15; each y3 is independently selected from 1, 2, and 3; each y4 is independently selected from 0 and 1; position 1 is connected to Tb through an S atom, and position 2 is connected to L2 or L3; (5) L2 either does not exist or exists. If L2 exists, L2 is selected from... y1 is selected from any integer between 1 and 6, each y2 is independently selected from any integer between 0 and 10, each y3 is independently selected from 1 and 2, each y4 is independently selected from 0 and 1, 1 bit is connected to L1, and 2 bits are connected to L3; (6) L3 is selected from amino acid residues Val, D-Val, Cit, Phe, Lys, Lys(Ac), Leu, Gly, Ala, Asn, Asp, Arg, and AA. 1 Or selected from 2-10 of Val, Cit, Phe, Lys, D-Val, Leu, Gly, Ala, Asn, Asp, and AA 1 Short peptides composed of amino acid residues; (7)AA 1 Among the amino acid residues, R a R b In the given information, one of them is H, and the other is selected from... Or, AA 1 Among the amino acid residues, R a With R b Together with the carbon atoms they are bonded to, they form R 0 Substituted 5-6 membered heterocycles; (8)AA 1 Among the amino acid residues, r, r 1 Each is independently selected from 0, 1, 2, 3, 4, and 5; (9)AA 1 Among the amino acid residues, R m1 R n1 Each is independently selected from H, methyl, ethyl, n-propyl, n-butyl, -COOCH3, -COOCH2CH3, -COOCH2CH2CH3, -COOCH(CH3)2, -COOC(CH3)3 and -COOCH2CH2CH2CH3; or, R m1 With R n1 Together with the nitrogen atoms they are bonded to, they form R 0’ Substituted 5-6 membered heterocycles; (10)AA 1 Among the amino acid residues, R z It is methyl; (11)AA 1 Among the amino acid residues, R 0 R 0’ Each is independently selected from C1-6 alkyl groups, -NR m2 R n2 And optional 5-6 membered heterocyclic groups substituted with C1-6 alkyl groups; (12) L4 either does not exist or exists. If L4 exists, L4 is selected from... Bit 1 is connected to L3, and bit 2 is connected to D.

4. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate as described in claim 3, characterized in that, It satisfies one or more of the following conditions: (1) Tb is anti-B7H3 antibody or its antigen-binding fragment, anti-Trop-2 antibody or its antigen-binding fragment, anti-Her 2 antibody or its antigen-binding fragment, anti-Her 3 antibody or its antigen-binding fragment, anti-EGFR antibody or its antigen-binding fragment, or IgG1 isotype antibody against chicken lysin antibody; (2) q is selected from any value between 0.1, 1, 2, 3, 4, 5, 6, 7, 8 and ; (3) When the bioactive molecule in the bioactive molecular fragment is a DNA topoisomerase inhibitor, the DNA topoisomerase inhibitor is a camptothecin-type bioactive molecule. (4) When the bioactive molecule in the bioactive molecular fragment is a tubulin inhibitor, the tubulin inhibitor is an MMAF-type tubulin inhibitor or an MMAE-type tubulin inhibitor. (5) In L1, each Z is independently selected from direct bond, carbon-carbon triple bond and carbon-carbon double bond; (6) In L1, y1 is 4, 5 or 6; (7) In L1, each y2 is independently selected from any integer between 6 and 10; (8) L2 either does not exist or exists. If L2 exists, L2 is selected from... Bit 1 is connected to L1, and bit 2 is connected to L3; (9) L3 is selected from 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 , AA 1 -Gly, Ala-Ala-Ala, Ala-Ala-Asn, Ala-Ala-Asp, Val-AA 1 -Gly, Ala-AA 1 -Gly, Gly-AA 1 -Gly, Lys-Ala-Ala-Asn, Lys-Ala-Ala-Asp, Gly-Phe-Gly, Gly-Gly-Phe-Gly, 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; (10)AA 1 Of the amino acid residues shown, R a R b In the given information, one of them is H, and the other is selected from... Or, R a With R b Together with the carbon atoms they are bonded to, they form R 0 The substituted 5-6 member heterocycle is replaced by R 0 Substituted piperidine ring or piperazine ring; (11)AA 1 Among the amino acid residues, r, r 1 Each is independently selected from 0 and 4; (12)AA 1 Among the amino acid residues, R m1 R n1 Each is independently selected from H, C1-6 alkyl, C3-6 cycloalkyl, and tert-butoxycarbonyl; or, R m1 With R n1 Together with the nitrogen atoms they are bonded to, they form R 0’ Substituted piperidine ring or piperazine ring; (13)AA 1 Among the amino acid residues, R 0 The group is selected from C1-6 alkyl groups and 5-6 membered heterocyclic groups substituted with C1-6 alkyl groups, wherein the 5-6 membered heterocyclic group is selected from piperidinyl and piperazineyl; (14)AA 1 Among the amino acid residues, R 0’ Selected from C1-6 alkyl and -NR m2 R n2 ; (15)AA 1 Among the amino acid residues, R m2 R n2 It is methyl; (16) L4 either does not exist or exists. If L4 exists, then L4 is... Bit 1 is connected to L3, and bit 2 is connected to D.

5. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate as described in any one of claims 1-4, characterized in that, It satisfies one or more of the following conditions: (1) q is selected from any value between 2, 3, 4, 5, 6, 7, 8 and; (2) Tb can be any of the following schemes: (i) Tb is an antibody against Her2 or its antigen-binding fragment, wherein the antibody against Her2 is anbenitamab, coprelotamab, disitamab, gancotamab, margetuximab, pertuzumab, timigutuzumab, zanidatamab, Trastuzumab, Pertuzumab or its antigen-binding fragment. (ii) Tb is an antibody against Trop-2 or its antigen-binding fragment, wherein the antibody against Trop-2 is datopotamab, sacituzumab or its antigen-binding fragment; or (iii) Tb is an anti-EGFR antibody or its antigen-binding fragment, wherein the anti-EGFR antibody is demupitamab, depatuxizumab, futuximab, imgatuzumab, lapatuximab, losatuxizumab, matuzumab, modotuximab, necitumumab, nimotuzumab, panitumumab, pimurutamab, serclutamab, tomouzotuximab, zalutumumab, Cetuximab or its antigen-binding fragment; or (iv) Tb is an anti-B7H3 antibody or its antigen-binding fragment, wherein the anti-B7H3 antibody is enoblituzumab, mirzotamab, omburtamab, 1D1-01, 2E3-02 antibody or its antigen-binding fragment; or (v) Tb is an anti-B7H3 antibody or its antigen-binding fragment, wherein the antigen-binding fragment of the anti-B7H3 antibody contains a complementarity-determining region (CDR), wherein the CDR is defined according to the Kabat numbering system: (a) HCDR1 with sequence SEQ ID NO: 7, HCDR2 with sequence SEQ ID NO: 8, and HCDR3 with sequence SEQ ID NO: 9; and / or, LCDR1 with sequence SEQ ID NO: 17, LCDR2 with sequence SEQ ID NO: 18, and LCDR3 with sequence SEQ ID NO: 19; (b) HCDR1 of SEQ ID NO: 27, HCDR2 of SEQ ID NO: 28, and HCDR3 of SEQ ID NO: 29; and / or, LCDR1 with sequence SEQ ID NO: 37, LCDR2 with sequence SEQ ID NO: 38, and LCDR3 with sequence SEQ ID NO: 39; (c) HCDR1 with sequence SEQ ID NO: 7, HCDR2 with sequence SEQ ID NO: 8, and HCDR3 with sequence SEQ ID NO: 9; and / or, LCDR1 with sequence SEQ ID NO: 37, LCDR2 with sequence SEQ ID NO: 38, and LCDR3 with sequence SEQ ID NO: 39; Or (d) the sequence is HCDR1 of SEQ ID NO: 27, the sequence is HCDR2 of SEQ ID NO: 28, and the sequence is HCDR3 of SEQ ID NO: 29; and / or, LCDR1 with sequence SEQ ID NO: 17, LCDR2 with sequence SEQ ID NO: 18, and LCDR3 with sequence SEQ ID NO: 19; or (vi) Tb is an anti-Her3 antibody or its antigen-binding fragment, wherein the anti-Her3 antibody is barecetamab, duligotuzumab, elgemtumab, lumretuzumab, patritumab, seribantumab, 202-2-1 or its antigen-binding fragment; or (vii) Tb is an anti-Her3 antibody or its antigen-binding fragment, wherein the anti-Her3 antibody or its antigen-binding fragment contains a complementarity-determining region (CDR), wherein the CDR is defined according to the Kabat numbering system: The sequences are HCDR1 (SEQ ID NO: 53), HCDR2 (SEQ ID NO: 54), and HCDR3 (SEQ ID NO: 55); and / or, LCDR1 with sequence SEQ ID NO: 59, LCDR2 with sequence SEQ ID NO: 60, and LCDR3 with sequence SEQ ID NO: 21; (viii) Antibodies or antigen-binding fragments thereof that have tumor cell endocytosis activity and binding activity to free antigens in tumor tissue and / or tumor cell surface antigens; (ix) Antibodies or antigen-binding fragments of tumor cells with weak or no endocytic activity and which have binding activity to free antigens in tumor tissue and / or antigens on the surface of tumor cells. (x) Tumor cells have weak or no endocytic activity, and there are no antibodies or antigen-binding fragments of them that bind to free antigens in tumor tissue or antigens on the surface of tumor cells; for example, Tb is an anti-chicken lysozyme human IgG1 isotype antibody. (xi) Antibodies or antigen-binding fragments of non-endocytic antigens; for example, ALCAM / CD166; (3) L1 is selected from m is selected from 2, 3 and 4, y1 is selected from any integer between 1 and 6, each y2 is independently selected from any integer between 0 and 10, each y3 is independently selected from 1 or 2, position 1 is connected to Tb through the S atom, and position 2 is connected to L2 or L3. (4) L2 either does not exist or exists. If L2 exists, L2 is selected from... Bit 1 is connected to L1, and bit 2 is connected to L3; (5) L3 is selected from AA 1 、AA 1 -Gly, Val-Cit, Val-AA 1 -Gly, AA 1 -Ala-Asn and Gly-Gly-Phe-Gly; (6)AA 1 Of the amino acid residues shown, R a With R b Together with the carbon atoms they are bonded to, they form R 0 The substituted 5-6 member heterocycle is replaced by R 0 Substituted piperidine ring; (7)AA 1 Among the amino acid residues, r, r 1 In the middle, one is 0 and the other is 4; (8)AA 1 Among the amino acid residues, R m1 R n1 Each is independently selected from H and C1-6 alkyl groups; or, R m1 With R n1 Together with the nitrogen atoms they are bonded to, they form R 0’ Substituted piperidine ring; (10)AA 1 Among the amino acid residues, R 0 Selected from methyl, ethyl and methyl-substituted 5-6 membered heterocyclic groups, wherein the 5-6 membered heterocyclic group is piperidinyl; (11)AA 1 Among the amino acid residues, R 0’ Selected from methyl and -NR m2 R n2 ; (12) When the bioactive molecule in the bioactive molecular fragment is a DNA topoisomerase inhibitor and the DNA topoisomerase inhibitor is a camptothecin-type bioactive molecule, the camptothecin-type bioactive molecule is camptothecin, DXD, camptothecin with substituent modification or DXD with substituent modification.

6. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate as claimed in any one of claims 1-4, characterized in that, It satisfies one or more of the following conditions: (1) q is selected from any value between 3, 4, 5, 6, 7, 8 and ; (2) Tb can be any of the following schemes: (vi) Tb is an anti-B7H3 antibody or its antigen-binding fragment, wherein the anti-B7H3 antibody or its antigen-binding fragment contains a complementarity-determining region (CDR), wherein the CDR is defined according to the Kabat numbering system: (a) HCDR1 with sequence SEQ ID NO: 7, HCDR2 with sequence SEQ ID NO: 8, and HCDR3 with sequence SEQ ID NO: 9; and / or, LCDR1 with sequence SEQ ID NO: 17, LCDR2 with sequence SEQ ID NO: 18, and LCDR3 with sequence SEQ ID NO: 19; (b) HCDR1 of SEQ ID NO: 27, HCDR2 of SEQ ID NO: 28, and HCDR3 of SEQ ID NO: 29; and / or, LCDR1 with sequence SEQ ID NO: 37, LCDR2 with sequence SEQ ID NO: 38, and LCDR3 with sequence SEQ ID NO: 39; (c) HCDR1 with sequence SEQ ID NO: 7, HCDR2 with sequence SEQ ID NO: 8, and HCDR3 with sequence SEQ ID NO: 9; and / or, LCDR1 with sequence SEQ ID NO: 37, LCDR2 with sequence SEQ ID NO: 38, and LCDR3 with sequence SEQ ID NO: 39; Or (d) the sequence is HCDR1 of SEQ ID NO: 27, the sequence is HCDR2 of SEQ ID NO: 28, and the sequence is HCDR3 of SEQ ID NO: 29; and / or, LCDR1 with sequence SEQ ID NO: 17, LCDR2 with sequence SEQ ID NO: 18, and LCDR3 with sequence SEQ ID NO: 19; Furthermore, the anti-B7H3 antibody or its antigen-binding fragment comprises a VH containing one set of heavy chain CDRs from (a) to (d) and a VL containing one set of light chain CDRs from (a) to (d); (ii) Tb is an anti-Her3 antibody or its antigen-binding fragment. The anti-Her3 antibody or its antigen-binding fragment contains a complementarity-determining region (CDR), wherein the CDR is defined according to the Kabat numbering system: The sequences are HCDR1 (SEQ ID NO: 53), HCDR2 (SEQ ID NO: 54), and HCDR3 (SEQ ID NO: 55); and / or, LCDR1 with sequence SEQ ID NO: 59, LCDR2 with sequence SEQ ID NO: 60, and LCDR3 with sequence SEQ ID NO: 21; The anti-Her3 antibody or its antigen-binding fragment comprises VH of the above-mentioned heavy chain CDRs and VL containing the above-mentioned light chain CDRs; (3) L1 is selected from Position 1 is connected to Tb via an S atom, and position 2 is connected to L2 or L3; (4) L2 either does not exist or exists. If L2 exists, L2 is selected from... Bit 1 is connected to L1, and bit 2 is connected to L3; (5) L3 is selected from AA 1 Val-AA 1 -Gly; (6)AA 1 Among the amino acid residues, R 0 Selected from methyl, ethyl and 7. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate as described in any one of claims 1-4, characterized in that, It satisfies one or more of the following conditions: (1) q is selected from any value between 4, 5, 6, 7, 8 and ; (2) Tb can be any of the following schemes: (i) Tb is an anti-B7H3 antibody or its antigen-binding fragment, wherein the anti-B7H3 antibody or its antigen-binding fragment comprises: VH as shown in SEQ ID NO: 3 or 23, and / or VL as shown in SEQ ID NO: 13 or 33; (ii) Tb is an anti-Her3 antibody or its antigen-binding fragment, wherein the anti-Her3 antibody or its antigen-binding fragment comprises: VH as shown in SEQ ID NO: 49, and / or VL as shown in SEQ ID NO: 50; (3) L1 is selected from Position 1 is connected to Tb via an S atom, and position 2 is connected to L2 or L3; (4) L2 does not exist or (5) L3 is selected from Val-AA 1 -Gly; (6)AA 1 Of the amino acid residues shown, R a With R b Together with the carbon atoms bonded to them, they form Carbon atom number 1 is related to R a and R b Carbon atoms linked together; (7) r、r 1 In the middle, r is 4, r 1 When R is 0, m1 R n1 Each is independently selected from H and C1-6 alkyl groups; r is 0, r 1 When R is 4, m1 R n1 Each is independently selected from C1-6 alkyl groups, or R m1 With R n1 Together with the nitrogen atoms bonded to them, they form Carbon atom number 1 is related to R a and R b Carbon atoms that are connected together.

8. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate as claimed in claim 7, characterized in that, (1) q is selected from any value between 6, 7, 8 and ; (2) Tb can be any of the following schemes: (i) Tb is an anti-B7H3 antibody or its antigen-binding fragment, wherein the anti-B7H3 antibody or its antigen-binding fragment comprises: VH as shown in SEQ ID NO: 3 or 23, and / or VL as shown in SEQ ID NO: 13 or 33; Further includes: (a) The heavy chain constant region (CH) of human immunoglobulins or a variant thereof; and / or (b) The light chain constant region (CL) of human immunoglobulins or a variant thereof. (ii) Tb is an anti-Her3 antibody or its antigen-binding fragment, wherein the anti-Her3 antibody or its antigen-binding fragment comprises: VH as shown in SEQ ID NO: 49, and / or VL as shown in SEQ ID NO: 50; Further includes: (a) The heavy chain constant region (CH) of human immunoglobulins or a variant thereof; and / or (b) The light chain constant region (CL) of human immunoglobulins or a variant thereof; (3) r、r 1 In the middle, r is 4, r 1 When R is 0, m1 R n1 Each is independently selected from H and C1-6 alkyl groups; r is 0, r 1 When R is 4, m1 R n1 Each is independently selected from C2-6 alkyl groups; (4) L1 is selected from Position 1 is connected to Tb via an S atom, and position 2 is connected to L2 or L3; for example, L1 is selected from... Position 1 is connected to Tb via an S atom, and position 2 is connected to L2 or L3.

9. The ligand-drug conjugate as claimed in claim 8, or a stereoisomer of the ligand-drug conjugate, its prodrug, its pharmaceutically acceptable salt, or its pharmaceutically acceptable solvate, characterized in that, (1) Tb can be any of the following schemes: (i) Tb is an anti-B7H3 antibody or its antigen-binding fragment, wherein the anti-B7H3 antibody or its antigen-binding fragment comprises: VH as shown in SEQ ID NO: 3, CH as shown in SEQ ID NO: 43 or a variant thereof, and / or VL as shown in SEQ ID NO: 13, CL as shown in SEQ ID NO: 44 or a variant thereof; VH as shown in SEQ ID NO: 23, CH as shown in SEQ ID NO: 43 or a variant thereof, and / or VL as shown in SEQ ID NO: 33, CL as shown in SEQ ID NO: 44 or a variant thereof; Preferably, the B7H3 antibody or its antigen-binding fragment comprises: The heavy chain shown in SEQ ID NO: 45, and / or the light chain shown in SEQ ID NO: 46; The heavy chain shown in SEQ ID NO: 47, and / or the light chain shown in SEQ ID NO: 48; (ii) Tb is an anti-Her3 antibody or its antigen-binding fragment, wherein the anti-Her3 antibody or its antigen-binding fragment comprises: VH as shown in SEQ ID NO: 49, CH as shown in SEQ ID NO: 43 or a variant thereof, and / or VL as shown in SEQ ID NO: 50, CL as shown in SEQ ID NO: 44 or a variant thereof; Preferably, the anti-Her3 antibody or its antigen-binding fragment comprises: The heavy chain shown in SEQ ID NO: 51, and / or the light chain shown in SEQ ID NO: 52; (2)R m1 R n1 Each is independently selected from C1-6 alkyl groups, wherein the C1-6 alkyl groups are selected from methyl, ethyl, and n-propyl; or R m1 R n1 Each is independently selected from C2-6 alkyl groups, wherein the C2-6 alkyl groups are selected from ethyl and n-propyl.

10. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate as claimed in any one of claims 1-9, characterized in that, It satisfies one or more of the following conditions: (1) When Tb is an antibody or its antigen-binding fragment, and the antibody or its antigen-binding fragment includes a single-chain antibody, the single-chain antibody is scFv; (2) Tb is an antibody or its antigen-binding fragment with endocytic activity; (3)AA 1 The amino acid residues shown are selected from Preferably, AA 1 The amino acid residues shown are selected from More preferably, AA 1 The amino acid residues shown are selected from The most preferred option is AA. 1 The amino acid residues shown are selected from 11. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate as claimed in any one of claims 1-10, characterized in that, L3 is selected from X - Selected from halide ions, carboxylate ions, sulfate ions, hydrogen sulfate ions and OH- ions - One bit is connected to L1 or L2, and two bits are connected to L4 or D; Preferably, L3 is selected from X - Selected from halide ions, carboxylate ions, sulfate ions, hydrogen sulfate ions and OH- ions - One bit is connected to L1 or L2, and two bits are connected to L4 or D; More preferably, L3 is selected from X - Selected from halide ions, carboxylate ions, sulfate ions, hydrogen sulfate ions and OH- ions - One bit is connected to L1 or L2, and two bits are connected to L4 or D; Preferably, L3 is selected from One bit is connected to L1 or L2, and two bits are connected to L4 or D; Most preferably, L3 is selected from One bit is connected to L1 or L2, and two bits are connected to L4 or D; For example The structure is selected from the following structural fragments:

12. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate as claimed in any one of claims 1-11, characterized in that, in The structures are selected from the following: Of these, bit 1 is connected to Tb, and bit 2 is connected to D.

13. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate as claimed in any one of claims 1-12, characterized in that, in, The ligand-drug conjugate has the structure shown in Formula I: Wherein: Tb, L1, L2, L3, L4 and q are defined as described in any one of claims 1-12; 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 atom attached thereto form a 5-7 membered carbon ring or a 5-7 membered heterocycle, wherein the heterocycle contains 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 atom attached thereto, form a 5-7 membered carbon ring or a 5-7 membered heterocycle, said heterocycle containing one or more O, S, N, carbonyl, sulfoxide, or sulfone groups or any combination thereof, or... R3 and R2 together with the carbon atom attached thereto form a 5-7 membered carbon ring or a 5-7 membered heterocycle, wherein the heterocycle contains one or more O, S, N, carbonyl, sulfoxide or sulfone groups or any combination thereof; W may or may not exist. If W exists, W is selected from -O-, -S-, -NR4-, One bit is connected to X, and two bits are connected to L4 or L3; X is selected from the direct bond, optionally substituted -O-(CH2). n3 -, -NR4-(CH2) n3 -、-S-(CH2) n3 - Carbonyl-(CH2) n3 -, -SO2-(CH2) n3 -、-(CH2) n1 -、 C3-6 cycloalkyl, C6-10 aryl, 5-10 heteroaryl, and 4-10 heterocyclic groups, with the 1-position connected to the parent ring and the 2-position connected to W or L4; the substituents are selected from one or more C1-4 alkyl, C3-6 cycloalkyl, or multiple C1-4 alkyl groups and carbon atoms simultaneously connected to them to form C3-6 cycloalkyl groups. Each M is independently selected from direct bonds and -CR 5a R 5b -; R4, R5, R 5a R 5b R6 and 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 between 0 and 6; Position 1 is attached to the camptothecin nucleus, and position 2 is attached to W or L4.

14. The ligand-drug conjugate as claimed in claim 13, or a stereoisomer of the ligand-drug conjugate, its prodrug, its pharmaceutically acceptable salt, or its pharmaceutically acceptable solvate, characterized in that, It satisfies one or more of the following conditions: (1) R1 and R2 are each independently selected from H, halogen, C1-4 alkyl; or, R1 and R2 together with the carbon atom attached thereto form a 5-6 membered heterocycle, the heterocycle containing one, two or three O, S or N or any combination thereof; (2) R3 is selected from H, C1-4 alkyl; or, R3 and X together with the carbon atom attached thereto form a 5-6 membered carbon ring; (3) W either does not exist or exists. When W exists, W is selected from -O-, -S-, -NR4-, One bit is connected to X, and two bits are connected to L4 or L3; (4) X is selected from the substituted -(CH2) group. n1 -、 C6-10 aryl, 5-10 heteroaryl, and 4-10 heterocyclic groups, with the parent ring attached at position 1 and W or L4 attached at position 2; the substituents are selected from one or two C1-4 alkyl groups, or two C1-4 alkyl groups and carbon atoms attached to them to form C3-6 cycloalkyl groups. (5) R4 and R5 are each independently selected from H, C1-4 alkyl and C3-6 cycloalkyl; (6)R 5a R 5b Each is independently selected from H and C1-4 alkyl groups; (7) Each R7 is independently selected from H and C1-4 alkyl groups; (8) n is selected from 1, 2 and 3; (9) n1 is selected from 1, 2, 3 and 4; (10) n2 is 1; (10) n3 is 0.

15. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate as claimed in claim 13 or 14, characterized in that, It satisfies one or more of the following conditions: (1) R1 is selected from H and halogens, and R2 is selected from H and C1-4 alkyl; or, R1 and R2 and the carbon atom attached thereto form The dashed lines indicate the positions where the heterocyclic ring and the benzene ring are fused; (2) R3 is H; or, R3 and X together with the carbon atom attached to them form The dashed lines indicate the positions where the carbon ring is fused with the benzene ring and the pyridine ring; (3) W either does not exist or exists. When W exists, W is selected from -O-, -S-, -NR4-, One bit is connected to X, and two bits are connected to L4 or L3; (4) Each R4 is independently selected from H, C1-4 alkyl and C3-6 cycloalkyl, and R5 is H; (5)R 5a R 5b Each is independently selected from H and methyl; (6) R7 is H; (7) n is 1.

16. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate as claimed in any one of claims 13-15, characterized in that, It satisfies one or more of the following conditions: (1) R1 is H or F, and R2 is H or methyl; or, R1 and R2 together with the carbon atom attached to them form The dashed lines indicate the positions where the heterocyclic ring and the benzene ring are fused; (2) W is selected from -O-, -NR4- and One bit is connected to X, and two bits are connected to L4 or L3; (3) Each R4 is independently selected from H, methyl, ethyl, n-propyl, isopropyl, tert-butyl and cyclopropyl, and R5 is H; (4) X is selected from any alternative substitution. The 1-position is connected to the parent ring, and the 2-position is connected to W or L4; the substituent is selected from one or two C1-4 alkyl groups, or two C1-4 alkyl groups and carbon atoms connected to them to form C3-6 cycloalkyl groups; the C1-4 alkyl group is such as methyl; the C3-6 cycloalkyl group is such as cyclopropyl.

17. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate as claimed in any one of claims 13-16, characterized in that, (1) W is selected from -O- and -NR4-; (2) X is selected from Position 1 is connected to the mother ring, and position 2 is connected to W or L4.

18. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate as claimed in any one of claims 13-17, characterized in that, When W does not exist, X is selected from... Bit 1 is connected to the parent ring, and bit 2 is connected to L4; when W exists, X is selected from... Position 1 is connected to the mother ring, and position 2 is connected to W.

19. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate as claimed in any one of claims 13-18, characterized in that, The structures are selected from the following: One bit is connected to L4; when L4 does not exist, one bit is connected to L3.

20. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate as claimed in any one of claims 13-19, characterized in that, The ligand-drug conjugate has the structure of formula I-1, formula I-2 or formula I-3: Wherein, Tb, L1, L2, L3, L4, X, R1, R2, R3, R4 and q are as defined in any one of claims 13-19; Wherein, Tb, L1, L2, L3, L4, X, R1, R2, R3 and q are as defined in any one of claims 13-19; Wherein, Tb, L1, L2, L3, L4, X, R1, R2, R3, R4, R5, n, and q are as defined in any one of claims 13-19.

21. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate as claimed in any one of claims 13-20, characterized in that, The ligand-drug conjugate has the structure of formula I-1A, I-1B, I-2A, I-2B, I-3A, or I-3B: Wherein, Tb, L2, L3, L4, X, R1, R2, R3, R4 and q are as defined in any one of claims 13-19; Wherein, Tb, L2, L3, L4, X, R1, R2, R3 and q are as defined in any one of claims 13-19; Wherein, Tb, L2, L3, L4, X, R1, R2, R3, R4 and q are as defined in any one of claims 13-19.

22. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate according to any one of claims 13-21, characterized in that, in, The ligand-drug conjugate has the structure of formula IA or formula IB: Among them, Tb, X, R1, R2, R3, R a R b and q as defined in any one of claims 13-19; Among them, Tb, X, R1, R2, R3, R a R b And q as defined in any one of claims 13-19.

23. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate according to any one of claims 1-22, characterized in that, in, The ligand-drug conjugates are selected from the following:

24. The ligand-drug conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt, or pharmaceutically acceptable solvate according to any one of claims 1-23, characterized in that, in, The ligand-drug conjugate is selected from: Wherein, Tb1 is an anti-B7H3 antibody or its antigen-binding fragment, such as enoblituzumab, mirzotamab, omburtamab, 1D1-01, 2E3-02 antibody; q is selected from any value between 0.1 and 16.0, preferably any value between 2 and 8, more preferably q is 2, 4, 6 or 8; Wherein, Tb2 is an anti-Trop-2 antibody or its antigen-binding fragment, such as datopotamab or sacituzumab; q is selected from any value between 0.1 and 16.0, preferably any value between 2 and 8, and more preferably, q is 2, 4, 6 or 8; Wherein, Tb3 is an anti-Her2 antibody or its antigen-binding fragment, such as anbenitamab, coprelotamab, disitamab, gancotamab, margetuximab, pertuzumab, timigutuzumab, zanidatamab, or trastuzumab; q is selected from any value between 0.1 and 16.0, preferably any value between 2 and 8, and more preferably, q is 2, 4, 6, or 8; Wherein, Tb4 is an anti-Her3 antibody or its antigen-binding fragment, such as barecetamab, duligotuzumab, elgemtumab, lumretuzumab, patritumab, seribantumab, and the antibody shown in IMGT / mAb-DB ID: 546; q is selected from any value between 0.1 and 16.0, preferably any value between 2 and 8, and more preferably, q is 2, 4, 6, or 8; Wherein, Tb5 is an anti-EGFR antibody or its antigen-binding fragment, such as demupitamab, depatuxizumab, futuximab, imgatuzumab, lapatuximab, losatuxizumab, matuzumab, modotuximab, necitumumab, nimotuzumab, panitumumab, pimurutamab, serclutamab, tomuzotuximab, zalutumumab, Cetuximab; q is selected from any value between 0.1 and 16.0, preferably any value between 2 and 8, more preferably q is 2, 4, 6 or 8; Wherein, Tb6 is an antibody that has no tumor cell endocytic activity and no tumor cell surface antigen binding activity, or an antibody that has the activity of binding non-endocytic (e.g., ALCAM / CD166) antigens, such as: IgG isotype antibody that does not have the corresponding cell surface antigen in the human body, anti-CD166 antibody; more specifically, anti-chicken lysozyme human IgG1 isotype antibody; q is selected from any value between 0.1 and 16.0, preferably any value between 2 and 8, more preferably q is 2, 4, 6 or 8; Wherein, Tb7 is an antibody with weak or no endocytic activity of tumor cells, but with activity of binding to tumor cell surface antigens; for example, the antibody has the ability to bind to B7H3, Her3, GD-2, Trop-2, EGFR, CD19, CD30, GPNMB, CD20, CD79b and BCMA antigens but does not have endocytic activity; q is selected from any value between 0.1 and 16.0, preferably any value between 2 and 8, more preferably q is 2, 4, 6 or 8; Wherein, Tb8 is an antibody with tumor cell endocytosis activity and tumor cell surface antigen binding activity, such as 1D1-01, 2E3-02 antibody, sacituzumab, pertuzumab, Trastuzumab or Cetuximab antibody; q is selected from any value between 0.1 and 16.0, preferably any value between 2 and 8, more preferably q is 2, 4, 6 or 8.

25. A linker in a ligand-drug conjugate, comprising the following fragment: in, The 2nd position is linked to a bioactive molecular fragment; The definitions of L3 and L4 are as described in any one of claims 1-24; Preferably, its structure is as follows: Among them, position 1 is linked to the ligand or target portion that binds to the target, and position 2 is linked to the bioactive molecular fragment. The definitions of L3 and L4 are as described in any one of claims 1-24; Preferably, the definitions of the target-binding ligand or targeting portion and the bioactive molecular fragment are as described in any one of claims 1-24, specifically Tb and D.

26. The compound represented by Formula II: Or a stereoisomer of the compound, its prodrug, its pharmaceutically acceptable salt, its pharmaceutically acceptable solvate, or its ligand-drug conjugate. in, The definitions of R1, R2, R3 and X are as described in any one of claims 13-24; W may or may not exist. If W exists, W is selected from -OH, -SH, -NHR4. 1 bit is connected to X; Preferably, W is absent or present; when W is present, W is selected from -OH, -SH, -NHR4, ... 1 bit is connected to X; Furthermore, when W does not exist, X is connected to H; wherein, when R1 and R2 are both H, and X is -(CH2). n1 When n1 is 1, 2, 3, or 4, W is not -OH or -NHR4; and the compound shown in Formula II does not contain 27. The compound of claim 26, wherein, The compounds shown in Formula II are selected from the following:

28. The drug-linked conjugate shown in Formula III, Or a stereoisomer of the drug linker conjugate, its prodrug, its pharmaceutically acceptable salt, or its pharmaceutically acceptable solvate. in: The definitions of R1, R2, R3, X, L1, L2, L3, and L4 are as described in any one of claims 1-24; the definition of W is as described in any one of claims 13-24; bit 1 of L1 is connected to Lg; Lg is a leaving group, selected from halogens, sulfones, and tertiary amine salts (Me3N). + Et3N + ), diazonium salts, -OMs, MeSO2- and CF3SO3-; Preferably, Lg is selected from F, Cl, and MeSO2-; the tertiary amine salt is selected from Me3N. + and Et3N + ; More preferably, Lg is selected from F and MeSO2-.

29. The drug linker conjugate of claim 28, or a stereoisomer of the drug linker conjugate, its prodrug, its pharmaceutically acceptable salt, or its pharmaceutically acceptable solvate, characterized in that, It has the structure of formula III-(1), formula III-(2), or formula III-(3): Wherein, L1, L2, L3, L4, X, R1, R2, R3, R4 and Lg are as defined in claim 28; Wherein, L1, L2, L3, L4, X, R1, R2, R3, and Lg are as defined in claim 28. L1, L2, L3, L4, X, R1, R2, R3, R4, R5, n, and Lg are as defined in claim 28.

30. The drug linker conjugate of claim 28 or 29, or a stereoisomer of the drug linker conjugate, its prodrug, its pharmaceutically acceptable salt, or its pharmaceutically acceptable solvate, characterized in that, It has the structure of formula III-(1A), III-(1B), III-(2A), III-(2B), III-(3A) or III-(3B); The structure of formula III-(1A) or III-(1B): Wherein, L2, L3, L4, X, R1, R2, R3, R4 and Lg are as defined in claim 28 or 29; The structure of formula III-(2A) or III-(2B) is as follows: Wherein, L2, L3, L4, X, R1, R2, R3 and Lg are as defined in claim 28 or 29; The structure of formula III-(3A) or III-(3B) is as follows: L2, L3, L4, X, R1, R2, R3, R4 and Lg are as defined in claim 28 or 29.

31. The drug linker conjugate or its stereoisomer, prodrug, pharmaceutically acceptable salt thereof, or pharmaceutically acceptable solvate thereof, as described in any one of claims 28-30, characterized in that... in, The drug linker conjugate has a structure of formula III-A or formula III-B: Among them, Lg, X, R1, R2, R3, R a R b and q as defined in any one of claims 28-30; Among them, Lg, X, R1, R2, R3, R a R b And q as defined in any one of claims 28-30.

32. The drug linker conjugate or stereoisomer of said drug linker conjugate, its prodrug, its pharmaceutically acceptable salt or its pharmaceutically acceptable solvate, as described in any one of claims 28-31, characterized in that, in, The drug conjugate is selected from the following:

33. A method for preparing the ligand-drug conjugate according to any one of claims 28-32, comprising: Tb conjugate with the drug linker shown in Formula III The coupling reaction is carried out under suitable solvents and conditions; in: L1, L2, L3, L4, and Tb are as defined in any one of claims 1-24; R1, R2, R3, X, and W are as defined in claims 13-24; Lg is defined as in any one of claims 28-32.

34. The method of claim 33, wherein, The method includes conjugating Tb with the drug linker conjugate shown in Formula III. The steps involved in forming CS bonds through a coupling reaction under suitable solvents and conditions; Preferably, the molar ratio of Tb to the drug linker conjugate is 1:(1-20); Preferably, the coupling reaction is carried out in water and / or an organic solvent; preferably, the organic solvent is selected from N,N-dimethylformamide, dimethyl sulfoxide, N-methylpyrrolidone, nitrile (e.g., acetonitrile), alcohol (e.g., methanol, ethanol) or any combination thereof; the nitrile may be acetonitrile, and the alcohol may be methanol or ethanol; Preferably, the method further includes a step of purifying the coupling product; preferably, the coupling product is purified by a chromatography method; preferably, the chromatography method includes one or more of ion exchange chromatography, hydrophobic chromatography, reversed-phase chromatography, or affinity chromatography.

35. An antibody or antigen-binding fragment thereof that binds to B7H3, said antibody or antigen-binding fragment comprising a complementarity-determining region (CDR) as follows: (a) HCDR1 or a variant thereof, HCDR2 or a variant thereof, and HCDR3 or a variant thereof contained in the heavy chain variable region VH shown in SEQ ID NO: 3 or 23; and / or (b) LCDR1 or a variant thereof, LCDR2 or a variant thereof, and LCDR3 or a variant thereof contained in the variable region VL of the light chain shown in SEQ ID NO: 13 or 33; Preferably, the variant of the sequence is a CDR with one or more amino acid substitutions, deletions, or additions compared to its source CDR, such as a CDR with one, two, or three amino acid substitutions, deletions, or additions; preferably, the substitutions are conservative substitutions.

36. The antibody or antigen-binding fragment thereof according to claim 35, wherein, The antibody or its antigen-binding fragment comprises: (1) VH and / or VL, where defined according to the IMGT numbering system: (a) The VH comprises: HCDR1 with the sequence SEQ ID NO: 10, HCDR2 with the sequence SEQ ID NO: 11, and HCDR3 with the sequence SEQ ID NO: 12; and / or, The VL includes: LCDR1 with the sequence SEQ ID NO: 20, LCDR2 with the sequence GTF, and LCDR3 with the sequence SEQ ID NO: 22; (b) The VH comprises: HCDR1 with the sequence SEQ ID NO: 10, HCDR2 with the sequence SEQ ID NO: 11, and HCDR3 with the sequence SEQ ID NO: 12; and / or, The VL includes: LCDR1 with the sequence SEQ ID NO: 40, LCDR2 with the sequence GAS, and LCDR3 with the sequence SEQ ID NO: 42; (c) The VH comprises: HCDR1 with the sequence SEQ ID NO: 30, HCDR2 with the sequence SEQ ID NO: 31, and HCDR3 with the sequence SEQ ID NO: 32; and / or, The VL includes: LCDR1 with the sequence SEQ ID NO: 40, LCDR2 with the sequence GAS, and LCDR3 with the sequence SEQ ID NO: 42; or (d) The VH comprises: HCDR1 with the sequence SEQ ID NO: 30, HCDR2 with the sequence SEQ ID NO: 31, and HCDR3 with the sequence SEQ ID NO: 32; and / or, The VL includes: LCDR1 with the sequence SEQ ID NO: 20, LCDR2 with the sequence GTF, and LCDR3 with the sequence SEQ ID NO: 22; (2) VH and / or VL, where defined according to the chothia numbering system: (a) The VH comprises: HCDR1 with the sequence SEQ ID NO: 4, HCDR2 with the sequence SEQ ID NO: 5, and HCDR3 with the sequence SEQ ID NO: 6; and / or, The VL includes: LCDR1 with the sequence SEQ ID NO: 14, LCDR2 with the sequence SEQ ID NO: 15, and LCDR3 with the sequence SEQ ID NO: 16; (b) The VH comprises: HCDR1 with the sequence SEQ ID NO: 24, HCDR2 with the sequence SEQ ID NO: 25, and HCDR3 with the sequence SEQ ID NO: 26; and / or, The VL includes: LCDR1 with the sequence SEQ ID NO: 34, LCDR2 with the sequence SEQ ID NO: 35, and LCDR3 with the sequence SEQ ID NO: 36; (c) The VH comprises: HCDR1 with the sequence SEQ ID NO: 4, HCDR2 with the sequence SEQ ID NO: 5, and HCDR3 with the sequence SEQ ID NO: 6; and / or, The VL includes: LCDR1 with the sequence SEQ ID NO: 34, LCDR2 with the sequence SEQ ID NO: 35, and LCDR3 with the sequence SEQ ID NO: 36; or (d) The VH comprises: HCDR1 with the sequence SEQ ID NO: 24, HCDR2 with the sequence SEQ ID NO: 25, and HCDR3 with the sequence SEQ ID NO: 26; and / or, The VL includes: LCDR1 with the sequence SEQ ID NO: 14, LCDR2 with the sequence SEQ ID NO: 15, and LCDR3 with the sequence SEQ ID NO: 16; (3) VH and / or VL, where defined according to the kabat numbering system: (a) The VH comprises: HCDR1 with the sequence SEQ ID NO: 7, HCDR2 with the sequence SEQ ID NO: 8, and HCDR3 with the sequence SEQ ID NO: 9; and / or, The VL includes: LCDR1 with the sequence SEQ ID NO: 17, LCDR2 with the sequence SEQ ID NO: 18, and LCDR3 with the sequence SEQ ID NO: 19; (b) The VH comprises: HCDR1 with the sequence SEQ ID NO: 27, HCDR2 with the sequence SEQ ID NO: 28, and HCDR3 with the sequence SEQ ID NO: 29; and / or, The VL includes: LCDR1 with the sequence SEQ ID NO: 37, LCDR2 with the sequence SEQ ID NO: 38, and LCDR3 with the sequence SEQ ID NO: 39; (c) The VH comprises: HCDR1 with the sequence SEQ ID NO: 7, HCDR2 with the sequence SEQ ID NO: 8, and HCDR3 with the sequence SEQ ID NO: 9; and / or, The VL includes: LCDR1 with the sequence SEQ ID NO: 37, LCDR2 with the sequence SEQ ID NO: 38, and LCDR3 with the sequence SEQ ID NO: 39; or (d) The VH comprises: HCDR1 with the sequence SEQ ID NO: 27, HCDR2 with the sequence SEQ ID NO: 28, and HCDR3 with the sequence SEQ ID NO: 29; and / or, The VL includes: LCDR1 with the sequence SEQ ID NO: 17, LCDR2 with the sequence SEQ ID NO: 18, and LCDR3 with the sequence SEQ ID NO: 19; Optionally, the antibody or its antigen-binding fragment disclosed herein comprises a heavy chain variable region VH and / or a light chain variable region VL, wherein, compared with the CDRs defined by IMGT, chothia, or kabat, at least one CDR in the heavy chain variable region VH and / or the light chain variable region VL contains a mutation, said mutation being a substitution, deletion, or addition of one or more amino acids or any combination thereof, for example, a substitution, deletion, or addition of one, two, or three amino acids or any combination thereof; preferably, said substitution is a conserved substitution; More preferably, the antibody or its antigen-binding fragment binds to human B7-H3 and / or monkey B7-H3.

37. The antibody or antigen-binding fragment thereof according to claim 35 or 36, wherein, The antibody or its antigen-binding fragment comprises: (a) VH as shown in SEQ ID NO: 3 or 23, and / or VL as shown in any one of SEQ ID NO: 13 or 33; (b) a VH 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 any VH in (a); and / or, a VL 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 any VL in (a); or (c) A VH having, compared to any VH in (a), one or more amino acid substitutions, deletions, or additions, or any combination thereof, such as 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions, or additions, or any combination thereof; and / or, a VL having, compared to any VL in (a), one or more amino acid substitutions, deletions, or additions, or any combination thereof, such as 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions, or additions, or any combination thereof; preferably, the substitution is a conservative substitution.

38. The antibody or antigen-binding fragment thereof according to any one of claims 35-37, wherein, The antibody or its antigen-binding fragment comprises: (a) VH of the sequence shown in SEQ ID NO: 3 and VL of the sequence shown in SEQ ID NO: 13; (b) VH of the sequence shown in SEQ ID NO: 23 and VL of the sequence shown in SEQ ID NO: 33; (c) VH of the sequence shown in SEQ ID NO: 3 and VL of the sequence shown in SEQ ID NO: 33; (d) VH of the sequence shown in SEQ ID NO: 23 and VL of the sequence shown in SEQ ID NO: 13; (e) VH and VL, compared with VH and VL in any of groups (a) to (d), wherein VH 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; and / or, wherein VL 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; or (f) VH and VL, compared with VH and VL in any of groups (a) to (d), wherein VH has one or more amino acid substitutions, deletions or additions or any combination thereof, such as 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid substitutions, deletions or additions or any combination thereof; and / or, wherein VL has one or more amino acid substitutions, deletions or additions or any combination thereof, such as 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.

39. The antibody or antigen-binding fragment thereof according to any one of claims 35-38, wherein, The antibody or its antigen-binding fragment is a chimeric antibody, a humanized antibody, or a fully human antibody; Optionally, the antibody or its antigen-binding fragment is selected from scFv, Fab, Fab', (Fab')2, Fv fragment, disulfide-linked Fv (dsFv), diabody, and multispecific antibody; Optionally, the scFv includes: (i) The sequence shown in SEQ ID NO: 1 or 2; (ii) A sequence having, compared to the sequence shown in (i), one or more amino acid substitutions, deletions, or additions, or any combination thereof (e.g., substitutions, deletions, or additions of up to 50, 45, 35, 25, 15, 10, or 5 amino acids, or any combination thereof; e.g., substitutions, deletions, or additions of 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids, 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 with the sequence shown in (i); Preferably, the substitution described in (ii) is a conservative substitution.

40. The antibody or antigen-binding fragment thereof according to any one of claims 35-39, wherein, The antibody or its antigen-binding fragment further comprises: (a) the heavy chain constant region CH of human immunoglobulins or a variant thereof; and / or (b) The light chain constant region CL of human immunoglobulins or a variant thereof. The variant has at least 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with its derived wild-type sequence; or, the variant has one or more amino acid substitutions, deletions, or additions, or any combination thereof, such as up to 50, 45, 40, 35, 30, 25, 20, 15, 10, or 5 amino acid substitutions, deletions, or additions, or any combination thereof, such as 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; Preferably, the heavy chain constant region is an IgG heavy chain constant region, such as an IgG1, IgG2, IgG3 or IgG4 heavy chain constant region; and / or, the light chain constant region is a κ or λ light chain constant region; More preferably, the antibody or its antigen-binding fragment contains a human IgG1 heavy chain constant region; and / or the antibody or its antigen-binding fragment contains a human κ light chain constant region.

41. The antibody or antigen-binding fragment thereof according to claim 40, wherein, The heavy chain constant region includes CH as shown in SEQ ID NO: 43 or a variant thereof, the variant having at most 20 conserved substitutions compared to SEQ ID NO: 43, such as at most 20, at most 15, at most 10, or at most 5 conserved substitutions, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 conserved substitutions; or 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 SEQ ID NO: 43; and / or The light chain constant region includes the CL shown in SEQ ID NO: 44 or a variant thereof, the variant having at most 20 conserved substitutions of amino acids compared to SEQ ID NO: 44, such as at most 20, at most 15, at most 10 or at most 5 conserved substitutions of amino acids, such as 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 conserved substitutions of amino acids, or 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 SEQ ID NO: 44; Preferably, the antibody or its antigen-binding fragment comprises the heavy chain constant region CH shown in SEQ ID NO: 43 and the light chain constant region CL shown in SEQ ID NO:

44.

42. The antibody or antigen-binding fragment thereof according to any one of claims 35-41, wherein, The antibodies include: (a) The heavy chain comprising VH as shown in SEQ ID NO: 3 and CH as shown in SEQ ID NO: 43, and the light chain comprising VL as shown in SEQ ID NO: 13 and CL as shown in SEQ ID NO: 44; preferably comprising the heavy chain as shown in SEQ ID NO: 45 and the light chain as shown in SEQ ID NO: 46; (b) The heavy chain comprising VH as shown in SEQ ID NO: 23 and CH as shown in SEQ ID NO: 43, and the light chain comprising VL as shown in SEQ ID NO: 33 and CL as shown in SEQ ID NO: 44; preferably comprising the heavy chain as shown in SEQ ID NO: 47 and the light chain as shown in SEQ ID NO:

48.

43. A multispecific antibody comprising the antibody or antigen-binding fragment thereof as described in any one of claims 35-42, and additional antibodies or fragments thereof or antibody analogs; Preferably, the multispecific antibody is a bispecific antibody, a trispecific antibody, or a tetraspecific antibody.

44. An isolated nucleic acid molecule encoding an antibody or antigen-binding fragment thereof as described in any one of claims 35-42, or a multispecific antibody as described in claim 43.

45. A vector comprising the isolated nucleic acid molecule of claim 44; preferably, the vector is a cloning vector or an expression vector.

46. ​​A host cell comprising the isolated nucleic acid molecule of claim 44 or the vector of claim 45.

47. A method for preparing the antibody or antigen-binding fragment thereof according to any one of claims 35-42, or the multispecific antibody according to claim 43, comprising culturing the host cell according to claim 46 under conditions allowing expression of the antibody or antigen-binding fragment thereof, and recovering the antibody or antigen-binding fragment thereof, or the multispecific antibody, from the cultured host cell culture.

48. An antibody-drug conjugate, wherein the antibody is the antibody of any one of claims 35-42 or its antigen-binding fragment, or the multispecific antibody of claim 43, which is connected to a conjugation portion via a linker, said conjugation portion being selected from: detectable markers, radioisotopes, fluorescent substances, luminescent substances, colored substances, enzymes, polyethylene glycol, radionuclides, nucleic acids, small molecule toxins, polypeptides with binding activity, proteins, receptors, ligands, and other active substances that inhibit tumor cell growth, promote tumor cell apoptosis or necrosis.

49. A ligand-drug conjugate comprising the ligand-drug conjugate of any one of claims 1-28, wherein the ligand-drug conjugate has two or more q values; Optionally, the drug-to-antibody ratio (DAR) in the ligand-drug conjugate is selected from an integer or decimal number between 1 and 10; Preferably, the drug-to-antibody ratio (DAR) in the ligand-drug conjugate is selected from 1.5-2.5, 3.5-4.5, 5.5-6.5, and 7.5-8.5; Preferably, the drug-to-antibody ratio (DAR) in the ligand-drug conjugate is selected from about 2.0, 4.0, 6.0, and 8.0; Preferably, the drug-to-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.

50. A pharmaceutical composition comprising substance A and optionally one or more pharmaceutical excipients, wherein substance A is a ligand-drug conjugate as claimed in any one of claims 1-24, or a stereoisomer of said ligand-drug conjugate, a prodrug thereof, a pharmaceutically acceptable salt thereof or a pharmaceutically acceptable solvate thereof, or a compound as claimed in claim 26 or 27, or a drug linker conjugate as claimed in any one of claims 28-32, or an antibody or an antigen-binding fragment thereof as claimed in any one of claims 35-42, or a multispecific antibody as claimed in claim 43, or a nucleic acid molecule as claimed in claim 44, or a carrier as claimed in claim 45, or a host cell as claimed in claim 46, or a ligand-drug conjugate as claimed in claim 49; preferably further comprising a pharmaceutically acceptable carrier and / or excipient.

51. Use of substance A or the pharmaceutical composition of claim 50 in the preparation of a medicament for treating and / or preventing diseases associated with abnormal cell activity; wherein substance A is a ligand-drug conjugate as described in any one of claims 1-24, or a stereoisomer of said ligand-drug conjugate, its prodrug, its pharmaceutically acceptable salt or its pharmaceutically acceptable solvate, or a compound as described in claim 26 or 27, or a drug linker conjugate as described in any one of claims 28-32, or an antibody or its antigen-binding fragment as described in any one of claims 35-42, or a multispecific antibody as described in claim 43, or a nucleic acid molecule as described in claim 44, or a carrier as described in claim 45, or a host cell as described in claim 46, or a ligand-drug conjugate as described in claim 49; the disease associated with abnormal cell activity may be cancer; Preferably, the cancer is selected from esophageal cancer, brain tumor, lung cancer, 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, rectal cancer, colorectal cancer, liver cancer, kidney cancer, urothelial carcinoma, epidermal cancer, non-Hodgkin's lymphoma, central nervous system tumors, 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 is selected from colon cancer, colorectal cancer, colon adenocarcinoma, lung cancer, breast cancer, prostate cancer, and esophageal squamous cell carcinoma; More preferably, the cancer is a cancer related to B7H3; More preferably, the cancer is a Her3-related cancer; More preferably, the cancer is an EGFR-related cancer; More preferably, the cancer is a cancer associated with Trop-2 or Her 2; Most preferably, the cancer is breast cancer or lung cancer is non-small cell lung cancer.

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