BCMA Monoclonal Antibodies and Antibody-Drug Conjugates

Abstract

The present disclosure relates to antibody drug conjugates (ADCs) comprising an antibody or monoclonal antibody against B-cell maturation antigen (BCMA), or an antigen-binding fragment thereof, conjugated to a cytotoxin. The BCMA antibodies and their ADCs are useful for the detection of BCMA as well as the treatment of multiple myeloma cells, B-cell or plasma cell mediated diseases, and immune disorders. Additionally, pharmaceutical compositions and methods of medical treatment are disclosed.

Description

[Technical field]

[0001] This application claims the benefit of PCT / CN2021 / 128453, filed November 3, 2021, which is incorporated herein by reference.

[0002] Sequence Listing Reference This application contains an electronic sequence listing in the file named FE00688PCT-Sequence listing xml, created on Oct. 10, 2022, and containing 40 KB, which is incorporated herein by reference. [Background technology]

[0003] B cell maturation antigen (BCMA, CD269) is a member of the tumor necrosis factor (TNF) receptor superfamily (Non-Patent Document 1). BCMA binds to two different ligands, B cell activating factor (BAFF; also known as BlyS, TALL-1, TNFSF13B, and THANK) and proliferation-inducing ligand (APRIL, TNFSF13) (Non-Patent Documents 2 and 3). The BCMA ligand further binds to two TNF receptors, transmembrane activator, calcium modulator, cyclophilin ligand interactor (TACI), and BAFF receptor (BAFF-R, also called BR3) (Non-Patent Document 4). Thus, BCMA is known primarily for its functional activity in mediating the survival of plasma cells that maintain long-term humoral immunity.

[0004] BCMA expression has been associated with many cancers, autoimmune diseases, and infectious diseases (Non-Patent Document 5). BCMA protein is highly expressed on the surface of plasma cells derived from multiple myeloma patients (Non-Patent Documents 6-8). Therefore, BCMA has been widely studied as a therapeutic target for multiple myeloma and autoimmune diseases in the past decade (Non-Patent Documents 9 and 10).

[0005] Multiple myeloma (MM) is a frequent hematological malignancy occurring worldwide (Non-Patent Document 11). It is a hematological malignancy characterized by proliferation of plasma cells with or without production of monoclonal immunoglobulins. Management of MM patients usually begins with induction / neoadjuvant therapy (including chemotherapy, radiation, surgery, and biophosphonates), usually followed by proteasome inhibitors (PIs) including dexamethasone, immunomodulatory agents (IMIDs), followed by autologous (hematopoietic) stem cell transplantation (ASCT) in eligible patients. In the past 5 years, the US FDA has approved three monoclonal antibodies (daratumumab, isatuximab, and elotuzumab), an exporter-1 inhibitor (selinexor), an anti-BCMA antibody-drug conjugate (belantamab mafodotin), and two BCMA chimeric antigen receptor (CAR) T-cell therapies (idecabtagenevicleucel and ciltacabtageneautoleucel) for the treatment of multiple myeloma. Numerous clinical trials are underway with novel targets and constructs, including bispecific antibodies targeting BCMA, GPRC5D, and FCRH5 in multiple myeloma. With incremental improvements in treatment regimens, the survival of patients with multiple myeloma (MM) has been significantly extended. Nevertheless, MM remains a nightmare with poor prognosis, and the disease itself remains incurable (Non-Patent Documents 12 and 13). And the median overall survival of patients who progress after multiple lines of treatment, including the use of monoclonal antibodies, is only 8.6 months. Although the newly approved BCMA CAR-T cell therapy has demonstrated an overall efficacy of over 80% in clinical studies, production from the patient's own cells has its own limitations and challenges, resulting in very expensive treatment costs. In the case of the first-in-class BCMA ADC, belantamab mafodotin, the overall response rate for MM was only 32%, and more than 60% of patients with this ADC reported unique side effects of some ocular toxicities that required dose adjustments, dose delays, and treatment discontinuation (Non-Patent Document 14). Therefore, there remains an urgent need for more MM treatment options in terms of feasibility, safety, and promising efficacy, especially therapies that overcome the resistance of existing drugs. [Prior art documents]

Non-licensed literature

[0006]

Non-licensed literature 1

Non-licensed Document 2

Non-licensed Document 4

Non-licensed Document 5

Non-licensed Document 6

Non-licensed Document 7

Non-licensed literature 9

Non-licensed literature 10

[0007] In this application, we disclose BCMA antibodies and antibody-drug conjugates (ADCs) that are directed against B-cell maturation antigen (BCMA) and comprise a BCMA monoclonal antibody or a BCMA antigen-binding fragment thereof conjugated to a cytotoxic agent. The BCMA antibodies and their ADCs are useful for the detection of BCMA as well as the treatment and diagnosis of various cancers, autoimmune diseases, and infectious diseases. In the present invention, the techniques of specific conjugation methods (PCT / CN2021 / 128453) are still applied to construct these BCMA ADCs. Furthermore, pharmaceutical compositions, screening, and medical treatment methods are disclosed. [Means for solving the problem]

[0008] The present invention provides antigen binding proteins that specifically bind to BCMA (CD269), such as antibodies that specifically bind to BCMA and inhibit binding of BAFF and / or APRIL to the BCMA receptor. The present invention also provides antigen binding proteins that specifically bind to BCMA and inhibit binding of BAFF and / or APRIL to BCMA and can be internalized. The BCMA monoclonal antibody comprises (a) a heavy chain variable region comprising a complementarity determining region 1 (HCDR1) amino acid sequence of SEQ ID NO:1, a HCDR2 amino acid sequence of SEQ ID NO:2, and a HCDR3 of SEQ ID NO:3; and (b) a light chain variable region comprising a LCDR1 amino acid sequence of SEQ ID NO:4, a LCDR2 amino acid sequence of SEQ ID NO:5, and a LCDR3 amino acid sequence of SEQ ID NO:6.

[0009] The present invention also provides an antibody drug conjugate (ADC) comprising a monoclonal antibody or antigen-binding fragment thereof against B-cell maturation antigen (BCMA) conjugated to a cytotoxic agent. In a further embodiment, the antigen binding protein is conjugated to a toxin, such as a tubulysin analog, a PBD dimer, or an auristatin analog.

[0010] Furthermore, the present invention provides compositions comprising the above-described antibody-drug conjugates and a pharma- ceutically acceptable carrier, and methods of killing multiple myeloma cells that express BCMA (including multiple myeloma stem cells) by contacting the multiple myeloma cells with the ADC.

[0011] In another aspect of the invention, there is provided a method of treating a human patient suffering from a B-cell related disorder or disease, such as an antibody-mediated disease or a plasma cell-mediated disease, or a plasma cell malignancy such as multiple myeloma (MM), comprising administering to said patient a therapeutically effective amount of an antigen-binding antibody and / or ADC thereof described herein.

[0012] In a further aspect of the invention, there is provided a method of treating a human patient suffering from rheumatoid arthritis, psoriasis, type 1 diabetes, or multiple sclerosis, the method comprising administering to said patient a therapeutically effective amount of an antigen binding protein and / or ADC described herein. [Brief description of the drawings]

[0013] [Figure 1] Binding affinity of hybridoma antibody BCMA-A2-6H4-5D2 and positive control antibody J6M0 to recombinantly expressed TrxA-BCMA is shown. [Diagram 2] Binding affinity of hybridoma antibody BCMA-A2-6H4-5D2, chimeric antibody c5D2, humanized antibody hu5D2, and positive control antibody J6M0 to recombinantly expressed TrxA-BCMA is shown. [Diagram 3] 1 shows the binding affinity of humanized antibody hu5D2, hu5D2-binding ADCs, and an isotype control antibody to the endogenous BCMA-expressing cell line NCI-H929. [Figure 4A] 1 shows killing of BCMA-overexpressing RPMI-8226 cell lines by the antibody drug conjugate hu5D2-tubulysin B analog conjugate (C-390). [Figure 4B] 1 shows killing of the cell line NCI-H929 by the antibody drug conjugate hu5D2-Tublysin B analog conjugate (C-390) compared to ADC J6M0-Tublysin B analog conjugate (C-390), naked hu5D2 antibody, unconjugated tubulysin B analog (Compound 390), and paclitaxel. [Figure 4C] 1 shows killing of the cell line MM.1S by the antibody drug conjugate hu5D2-Tublysin B analog conjugate (C-390) compared to ADC J6M0-Tublysin B analog conjugate (C-390), naked hu5D2 antibody, unconjugated tubulysin B analog (Compound 390), and paclitaxel. [Figure 4D] 1 shows killing of the BCMA negative expressing cell line Jurkat by the antibody drug conjugate hu5D2-Tublysin B analog conjugate (C-390) compared to ADC J6M0-Tublysin B analog conjugate (C-390), naked hu5D2 antibody, unconjugated tubulysin B analog (Compound 390), and paclitaxel. [Figure 4E]1 shows killing of the BCMA expressing cell line U266B1 by BCMA antibody (hu5D2)-drug conjugates: C-221, C-202, C-88, C-326, C-30 compared to paclitaxel. [Diagram 5] Figure 1 shows MS / MS daughter ion or product ion spectra of glycopeptides of BCMA antibodies: (a): non-glycosylated peptide; (b): Man5 containing glycopeptide; (c): G0F-GlcNAc containing glycopeptide; (d): G0 containing glycopeptide; (e): G0F containing glycopeptide; (f): G1 containing glycopeptide; (g): G1F containing glycopeptide; (h): G2F containing glycopeptide. [Figure 6] Mid-level characterization of BCMA-Tublysin B analog ADC (C-390) after N-deglycosylation and reduction: (a) rpHPLC chromatograms of the ADC fragments obtained after deglycosylation and DTT reduction: light chains (LC) with 0 or 1 drug molecule attached (L0 and L1), (b) heavy chains with 0, 1, 2, or 3 drug molecules attached (H0, H1, H2, and H3). [Figure 7] Percentage of drug-loaded peptides in BCMA ADC (C-390). (a): LC peptide with 0 or 1 drug molecule attached [GEC] (D0 and D1). (b): HC peptide with 0 or 1 drug molecule attached at the arm region [SCDK] (D0 and D1). (c): HC peptide with 0, 1, or 2 drug molecules attached at the hinge region [THTCPPCPAPELLGGPSVFLFPPKPK] (D0, D1, and D2). [Figure 8] MS / MS daughter or product ion spectra of drug-loaded peptides of BCMA-ADC(C-390): (a): heavy chain [SC(223)DK] + 1 drug; (b): heavy chain [THTC(229)PPCPAPELLGGPSVFLFPPKPK] + 1 drug; (c): heavy chain [THTCPPC(232)PAPELLGGPSVFLFPPKPK] + 1 drug; (d): heavy chain [THTC(229)PPC(232)PAPELLGGPSVFLFPPKPK] + 2 drugs; (e): light chain [GEC(219)] + 1 drug. [Figure 9]Changes in tumor volume of BCMA(hu5D2) ADCs (C-68a, C-115, C-192, C-202, C-221, C-290, C-306, C-385, C-390, C-399, C-402, C-417, DAR is shown in Table 7.) at a series of single doses (3 mg / kg) using a human multiple myeloma NCI-H929 cell xenograft mouse model, compared with belantamab mcMMAF and PBS buffer (control). This figure shows that all 13 conjugates have antitumor activity, and the order of antitumor activity is C-385 < C-306 < C-290 < C-68a < C-115 < BCMA-mcMMAF < C-192 < C-202 < C-399 < C-390 < C-417 < C-402 < C-221. [Figure 10] Changes in tumor volume of hu5D2-ADCs at a series of single doses (5 mg / kg) using a human multiple myeloma JJN-3 cell xenograft mouse model, compared with belantamab mcMMAF and PBS buffer (control). This figure shows that all 9 conjugates have antitumor activity, and the order of antitumor activity is paclitaxel < C-385 < belantamab mcMMAF < C-195 < C-137 < C-181b < C-126 < C-83 < C-277 < C-258. [Figure 11] Changes in tumor volume of hu5D2-ADCs at a series of single doses (2 mg / kg) using a human multiple myeloma NCI-H929 cell xenograft mouse model, compared with belantamab mcMMAF and PBS buffer (control). This figure shows that all 7 conjugates have antitumor activity, and the order of antitumor activity is belantamab mcMMAF < C-406 < C-396 < C-399 < C-400 < C-221b < C-402. [Figure 12] Shows the general synthesis of components of the bis-conjugate. [Figure 13] Shows the synthesis of camptothecin analogs containing bis-conjugates. [Figure 14] Shows the synthesis of camptothecin analogs containing bis-conjugates. [Figure 15] Shows the synthesis of camptothecin analogs containing bis-conjugates. [Figure 16] The general synthesis of the components of the bis-linker is shown. [Figure 17] 1 shows the synthesis of camptothecin analogs containing bisconjugated linkers. [Figure 18] 1 shows the synthesis of camptothecin analogs containing bisconjugated linkers. [Figure 19] 1 shows the synthesis of camptothecin analogs containing bisconjugated linkers. [Figure 20] The general synthesis of the components of the bis-linker is shown. [Figure 21] 1 shows the synthesis of camptothecin analogs containing bisconjugated linkers. [Figure 22] 1 shows the synthesis of tubulysin B analogs containing bisconjugated linkers. [Diagram 23] 1 shows the synthesis of tubulysin B analogs containing bisconjugated linkers. [Figure 24] 1 shows the synthesis of tubulysin B analogs containing bisconjugated linkers. [Diagram 25] 1 shows the synthesis of tubulysin B analogs containing bisconjugated linkers. [Figure 26] 1 shows the synthesis of tubulysin B analogs containing bisconjugated linkers. [Figure 27] 1 shows the synthesis of tubulysin B analogs containing bisconjugated linkers. [Figure 28] 1 shows the synthesis of components of a tubulysin B analogue. [Figure 29] 1 shows the synthesis of tubulysin B analogs containing bisconjugated linkers. [Diagram 30] 1 shows the synthesis of tubulysin B analogs containing bisconjugated linkers. [Diagram 31] 1 shows the synthesis of tubulysin B analogs containing bisconjugated linkers. [Diagram 32] 1 shows the synthesis of tubulysin B analogs containing bisconjugated linkers. [Diagram 33] 1 shows the synthesis of tubulysin B analogs containing bisconjugated linkers. [Diagram 34] 1 shows the synthesis of tubulysin B analogs containing bisconjugated linkers. [Diagram 35] 1 shows the synthesis of tubulysin B analogs containing bisconjugated linkers. [Diagram 36] 1 shows the synthesis of camptothecin analogs and amanitin analog components containing bis-conjugated linkers. [Figure 37] 1 shows the synthesis of amanitin analogs containing bisconjugated linkers. [Figure 38] 1 shows the synthesis of amanitin analogs containing bisconjugated linkers. [Figure 39] 1 shows the synthesis of amanitin analogs containing bisconjugated linkers. [Diagram 40] 1 shows the synthesis of amanitin analogs containing bisconjugated linkers. [Diagram 41] 1 shows the synthesis of a tubulysin B analog containing a linker. [Diagram 42] 1 shows the synthesis of a tubulysin B analog containing a linker. [Diagram 43] 1 shows the synthesis of a tubulysin B analog containing a linker. [Diagram 44] 1 shows the synthesis of a tubulysin B analog containing a linker. [Diagram 45] 1 shows the synthesis of a tubulysin B analog containing a linker. [Figure 46] 1 shows the synthesis of a tubulysin B analog containing a linker. [Figure 47] 1 shows the synthesis of a tubulysin B analog containing a linker. [Figure 48] 1 shows the synthesis of a tubulysin B analog containing a linker. [Figure 49] 1 shows the synthesis of a tubulysin B analog containing a linker. [Figure 50] 1 shows the synthesis of a tubulysin B analog containing a linker. [Figure 51] 1 shows the synthesis of a tubulysin B analog containing a linker. [Figure 52] 1 shows the synthesis of a tubulysin B analog containing a linker. [Diagram 53] 1 shows the synthesis of a tubulysin B analog containing a linker. [Figure 54]1 shows the synthesis of a tubulysin B analog containing a linker. [Figure 55] 1 shows the synthesis of a tubulysin B analog containing a linker. [Figure 56] 1 shows the synthesis of a tubulysin B analog containing a linker and a PBD analog component. [Figure 57] The synthesis of PBD analogs containing linkers is shown. [Figure 58] The synthesis of PBD analogs containing linkers is shown. [Figure 59] The synthesis of components of a PBD analog containing a linker and a tubulysin B analog containing a linker is shown. [Figure 60] 1 shows the synthesis of a tubulysin B analog containing a linker. [Figure 61] 1 shows the synthesis of a tubulysin B analog containing a linker. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0014] definition

[0015] "Alkyl" refers to an aliphatic hydrocarbon group or monovalent group derived from an alkane by removing one or two hydrogen atoms from a carbon atom. 1 ~C 8It may be linear or branched having 1 to 8 carbon atoms. "Branched" refers to a linear alkyl group to which one or more low C alkyl groups, such as methyl, ethyl, or propyl groups, are bonded. Specific examples of the alkyl group include methyl, ethyl, n-propyl, i-propyl, n-butyl, t-butyl, n-pentyl, 3-pentyl, octyl, nonyl, decyl, cyclopentyl, cyclohexyl, 2,2-dimethylbutyl, 2,3-dimethylbutyl, 2,2-dimethylpentyl, 2,3-dimethylpentyl, 3,3-dimethylpentyl, 2,3,4-trimethylpentyl, 3-methylhexyl, 2,2-dimethylhexyl, 2,4-dimethylhexyl, 2,5-dimethylhexyl, 3,5-dimethylhexyl, 2,4-dimethylpentyl, 2-methylheptyl, 3-methylheptyl, n-heptyl, isoheptyl, n-octyl, and isooctyl. 1 ~C 8 An alkyl group may be unsubstituted or substituted with one or more substituents, including but not limited to the following substituents: 1 -C 8 Alkyl, -O-(C 1 ~C 8 Alkyl), -aryl, -C(O)R', -OC(O)R', -C(O)OR', -C(O)NH 2 , -C(O)NHR', -C(O)N(R') 2 , -NHC(O)R', -SR', -S(O) 2 R', -S(O)R', -OH, -halogen, -N 3 , -NH 2 , -NH(R'), -N(R') 2 and -CN, wherein each R' is independently C 1 ~C 8 It is selected from alkyl and aryl.

[0016] "Halogen" refers to fluorine, chlorine, bromine, or iodine atoms, with fluorine and chlorine atoms being preferred.

[0017] "Heteroalkyl" refers to a C alkyl group in which 1 to 4 carbon atoms are independently replaced by a heteroatom selected from the group consisting of O, S, and N. 2 ~C 8 It means alkyl.

[0018] "Carbocycle" refers to a monocyclic or bicyclic saturated or unsaturated ring having 3 to 8 carbon atoms or 7 to 13 carbon atoms. Monocyclic carbocycles have 3 to 6, more typically 5 or 6 ring atoms. Bicyclic carbocycles have 7 to 12 ring atoms arranged as a [4,5], [5,5], [5,6], or [6,6] bicycle, or 9 to 10 ring atoms arranged as a [5,6] or [6,6] bicycle. Representative C 3 ~C 8 Carbocyclic rings (C 3 ~C 8 Carbocycles include, but are not limited to, -cyclopropyl, -cyclobutyl, -cyclopentyl, -cyclopentadienyl, -cyclohexyl, -cyclohexenyl, -1,3-cyclohexadienyl, -1,4-cyclohexadienyl, -cycloheptyl, -1,3-cycloheptadienyl, -1,3,5-cycloheptatrienyl, -cyclooctyl, and -cyclooctadienyl.

[0019] C 3 ~C 8 Carbocycle (C 3 ~C 8 A carbocycle refers to a saturated or unsaturated non-aromatic hydrocarbon carbocyclic compound having 3, 4, 5, 6, 7, or 8 carbon atoms. 3 ~C 8 A carbocycle may be unsubstituted or substituted with one or more substituents, including, but not limited to, -C 1 ~C 8 Alkyl, -O-(C 1 ~C 8 Alkyl), -aryl, -C(O)R', -OC(O)R', -C(O)OR', -C(O)NH 2 , -C(O)NHR', -C(O)N(R') 2, -NHC(O)R', -SR', -S(O)R', -S(O) 2 R', -OH, -halogen, -N 3 , -NH 2 , -NH(R'), -N(R') 2 and -CN, where each R' is independently C 1 ~C 8 It is selected from alkyl and aryl.

[0020] "Alkenyl" refers to an aliphatic hydrocarbon group having a carbon-carbon double bond, which may be straight or branched, having 2 to 8 carbon atoms in the chain. Alkenyl groups include, for example, ethenyl, propenyl, n-butenyl, i-butenyl, 3-methylbut-2-enyl, n-pentenyl, hexylenyl, heptenyl, and octenyl.

[0021] "Alkynyl" refers to an aliphatic hydrocarbon group having a carbon-carbon triple bond and may be straight or branched having 2 to 8 carbon atoms in the chain. Alkynyl groups include, for example, ethynyl, propynyl, n-butynyl, 2-butynyl, 3-methylbutynyl, 5-pentynyl, n-pentynyl, hexynyl, heptynyl, and octynyl.

[0022] "Alkylene" refers to a saturated, straight or branched chain or cyclic hydrocarbon group of 1 to 18 carbon atoms having two monovalent radical centers derived by the removal of two hydrogen atoms from the same or two different carbon atoms of a parent alkane. Exemplary alkylene groups include methylene (-CH 2 -), 1,2-ethyl (-CH 2 CH 2 -), 1,3-propyl (-CH 2 CH 2 CH 2 -), 1,4-butyl (-CH 2 CH 2 CH 2 CH 2 -) and the like, but are not limited to these.

[0023] "Alkenylene" refers to an unsaturated, straight or branched chain or cyclic hydrocarbon group of 2 to 18 carbon atoms having two monovalent radical centers derived by the removal of two hydrogen atoms from the same or two different carbon atoms of a parent alkene. Typical alkenylene groups include, but are not limited to, 1,2-ethylene (-CH=CH-).

[0024] "Alkynylene" refers to an unsaturated, straight or branched chain or cyclic hydrocarbon group of 2 to 18 carbon atoms having two monovalent radical centers derived by the removal of two hydrogen atoms from the same or two different carbon atoms of a parent alkyne. Typical alkynylene groups include, but are not limited to, acetylene, propargyl, and 4-pentynyl.

[0025] "Aryl" or Ar refers to an aromatic or heteroaromatic group consisting of one or several rings, containing 3 to 14 carbon atoms, preferably 6 to 10 carbon atoms. The term "heteroaromatic group" refers to an aromatic group in which one or several carbons, preferably 1, 2, 3 or 4 carbon atoms, on the aromatic group are replaced by O, N, Si, Se, P or S, preferably O, S and N. The term aryl or Ar also refers to an aromatic group in which one or several H atoms are independently replaced by -R', -halogen, -OR', -SR', -NR'R'', -N=NR', -N=R', -NR'R'', -NO 2 , -S(O)R', -S(O) 2 R', -S(O) 2 OR', -OS(O) 2 It also refers to being replaced by -OR', -PR'R'', -P(O)R'R'', -P(OR')(OR'', -P(O)(OR')(OR'', or -OP(O)(OR')(OR''), where R', R'' are independently H, alkyl, alkenyl, alkynyl, heteroalkyl, aryl, arylalkyl, carbonyl, or a pharmaceutical salt.

[0026] "Heterocycle" refers to a ring system in which one to four ring carbon atoms are independently replaced with a heteroatom from the group O, N, S, Se, B, Si, and P. Preferred heteroatoms are O, N, and S. Heterocycles are described in The Handbook of Chemistry and Physics, 78th Edition, CRC Press, Inc., 1997-1998, p. 225-226, the disclosure of which is incorporated herein by reference. Preferred non-aromatic heterocycles include, but are not limited to, epoxy, aziridinyl, thiiranyl, pyrrolidinyl, pyrazolidinyl, imidazolidinyl, oxiranyl, tetrahydrofuranyl, dioxolanyl, tetrahydropyranyl, dioxanyl, dioxolanyl, piperidyl, piperazinyl, morpholinyl, pyranyl, imidazolinyl, pyrrolinyl, pyrazolinyl, thiazolidinyl, tetrahydropyranyl, dihydropyranyl, tetrahydropyridyl, dihydropyridyl, tetrahydropyrimidinyl, dihydrothiopyranyl, azepanyl, as well as fused systems resulting from condensation with a phenyl group.

[0027] The term "heteroaryl" or aromatic heterocycle refers to a 3- to 14-membered, preferably 5- to 10-membered, aromatic hetero, monocyclic, bicyclic, or polycyclic ring. Examples include pyrrolyl, pyridyl, pyrazolyl, thienyl, pyrimidinyl, pyrazinyl, tetrazolyl, indolyl, quinolinyl, purinyl, imidazolyl, thienyl, thiazolyl, benzothiazolyl, furanyl, benzofuranyl, 1,2,4-thiadiazolyl, isothiazolyl, triazolyl, tetrazolyl, isoquinolyl, benzothienyl, isobenzofuryl, pyrazolyl, carbazolyl, benzimidazolyl, isoxazolyl, pyridyl-N-oxide, and fused systems resulting from condensation with a phenyl group.

[0028] "Alkyl", "cycloalkyl", "alkenyl", "alkynyl", "aryl", "heteroaryl", "heterocyclic", and the like, also refer to the corresponding "alkylene", "cycloalkylene", "alkenylene", "alkynylene", "arylene", "heteroarylene", "heterocyclene", and the like, formed by the removal of two hydrogen atoms.

[0029] "Arylalkyl" refers to an alkyl group consisting of a carbon atom, typically a terminal or sp 3 It refers to an acyclic alkyl group in which one of the hydrogen atoms bonded to a carbon atom is replaced with an aryl group. Typical arylalkyl groups include, but are not limited to, benzyl, 2-phenylethan-1-yl, 2-phenylethen-1-yl, naphthylmethyl, 2-naphthylethan-1-yl, 2-naphthylethene-1-yl, naphthobenzyl, 2-naphthophenylethan-1-yl, and the like.

[0030] "Heteroarylalkyl" refers to a heteroaryl group consisting of a carbon atom, typically a terminal or sp 3 It refers to an acyclic alkyl group in which one of the hydrogen atoms bonded to a carbon atom is replaced with a heteroaryl group. Typical heteroarylalkyl groups include, but are not limited to, 2-benzimidazolylmethyl, 2-furylethyl, and the like.

[0031] Examples of "hydroxy protecting groups" include, but are not limited to, methoxymethyl ether, 2-methoxyethoxymethyl ether, tetrahydropyranyl ether, benzyl ether, p-methoxybenzyl ether, trimethylsilyl ether, triethylsilyl ether, triisopropylsilyl ether, t-butyldimethylsilyl ether, triphenylmethylsilyl ether, acetate, substituted acetate, pivaloate, benzoate, methanesulfonate, and p-toluenesulfonate.

[0032] "Leaving group" refers to a functional group that can be replaced by another functional group. Such leaving groups are well known in the art and include, for example, but are not limited to, halides (e.g., chloride, bromide, and iodide), methanesulfonyl (mesyl), p-toluenesulfonyl (tosyl), trifluoromethylsulfonyl (triflate), and trifluoromethylsulfonate. The leaving group is preferably selected from nitrophenol; N-hydroxysuccinimide (NHS); phenol; dinitrophenol; pentafluorophenol; tetrafluorophenol; difluorophenol; monofluorophenol; pentachlorophenol; triflate; imidazole; dichlorophenol; tetrachlorophenol; 1-hydroxybenzotriazole; tosylate; mesylate; 2-ethyl-5-phenylisoxazolium-3'-sulfonate, anhydrides formed with themselves or with other anhydrides (e.g., acetic anhydride, formic anhydride); or intermediates generated by condensation reagents for peptide coupling reactions or Mitsunobu reactions.

[0033] The following abbreviations may be used herein and have the definitions given below: Boc, tert-butoxycarbonyl; BroP, bromotrispyrrolidinophosphonium hexafluorophosphate; CDI, 1,1'-carbonyldiimidazole; DCC, dicyclohexylcarbodiimide; DCE, 1,2-dichloroethane; DCM, dichloromethane; DIAD, diisopropyl azodicarboxylate; DIBAL-H, diisobutylaluminum hydride; DIPEA, diisopropylethylamine; DEPC, diethylphosphoroanidinium; DMA, N,N-dimethylacetamide; DMAP, 4-(N,N-dimethylamino)pyridine; DMF, N,N-dimethylformamide; DMSO, dimethylsulfoxide; DTT, dithiothreitol. tol; EDC, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride; ESI-MS, electrospray mass spectrometry; HATU, O-(7-azabenzotriazol-1-yl)-N,N,N'-N'-tetramethyluronium hexafluorophosphate; HOBt, 1-hydroxybenzotriazole; HPLC, high pressure liquid chromatography; NHS, N-hydroxysuccinimide; MMP, 4-methylmorpholine; PAB, p-aminobenzyl; PBS, phosphate buffered saline (pH 7.0-7.5); PEG, polyethylene glycol; SEC, size exclusion chromatography; TCEP, tris(2-carboxyethyl)phosphine; TFA, trifluoroacetic acid; THF, tetrahydrofuran; Val, valine.

[0034] The "amino acid(s)" may be natural and / or unnatural amino acids, preferably α-amino acids. Natural amino acids are those encoded by the genetic code: alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tyrosine, tryptophan, and valine. Unnatural amino acids are derived forms of protein-forming amino acids, including, for example, hydroxyproline, lanthionine, 2-aminoisobutyric acid, dehydroalanine, γ-aminobutyric acid (neurotransmitter), ornithine, citrulline, β-alanine (3-aminopropanoic acid), γ-carboxyglutamate, selenocysteine ​​(present in most eukaryotes, but not directly encoded by DNA), pyrrolysine (only found in some archaea and one bacterium), N-formylmethionine (often the first amino acid in proteins in bacteria, mitochondria, and chloroplasts), 5-hydroxytryptophan, L-dihydroxyphenylalanine, triiodothyronine, L-3,4-dihydroxyphenylalanine (DOPA), and O-phosphoserine. The term amino acid also includes amino acid analogs and mimetics. Analogs have the same general H group as the natural amino acids, except that the R group is not one found in the natural amino acids. 2 N(R)CHCO 2An amino acid mimetic is a compound having an H structure. Examples of analogues include homoserine, norleucine, methionine-sulfoxide, and methionine methylsulfonium. Preferably, an amino acid mimetic is a compound that has a structure different from the general chemical structure of an α-amino acid, but functions similarly thereto. The term "unnatural amino acid" is intended to represent the "D" stereochemical form, whereas natural amino acids are in the "L" form. When 1 to 8 amino acids are used in the present application, it is preferred that the amino acid sequence is a cleavage recognition sequence for a protease. Many cleavage recognition sequences are known in the art, see, e.g., Matayoshi et al. Science 247: 954 (1990); Dunn et al. Meth. Enzymol. 241: 254 (1994); Seidah et al. Meth. Enzymol. 244: 175 (1994); Thornberry, Meth. Enzymol. 244: 615 (1994); Weber et al. Meth. Enzymol. 244: 595 (1994); Smith et al. Meth. Enzymol. 244: 412 (1994); and Bouvier et al. Meth. Enzymol. 248: 614 (1995); the disclosures of which are incorporated herein by reference. In particular, the sequence is selected from the group consisting of Val-Cit, Ala-Val, Ala-Ala, Val-Val, Val-Ala-Val, Lys-Lys, Ala-Asn-Val, Val-Leu-Lys, Cit-Cit, Val-Lys, Ala-Ala-Asn, Lys, Cit, Ser, and Glu.

[0035] "Pharmaceutically" or "pharmacologically acceptable" means that the corresponding compound or compound composition does not produce harmful, allergic or other adverse reactions when administered to animals or humans in a suitable manner.

[0036] "Pharmaceutically acceptable solvate" or "solvate" refers to an association of one or more solvent molecules with a disclosed compound. Examples of solvents that form pharmacologically acceptable solvates include, but are not limited to, water, isopropanol, ethanol, methanol, DMSO, ethyl acetate, acetic acid, and ethanolamine.

[0037] Pharmaceutically acceptable auxiliary materials include all carriers, diluents, auxiliaries or excipients, such as preservatives, antioxidants, fillers, disintegrants, wetting agents, emulsifiers, suspending agents, solvents, dispersion media, coating agents, antibacterial agents, antifungal agents, isotonic agents, absorption retardants, etc. In the pharmaceutical field, it is a common practice to add these auxiliary materials to active drug components. It can be said that it is reasonable to add auxiliary materials to drug components, except when the auxiliary materials are incompatible with the active drug components. In order to obtain good results, active auxiliary materials may be added to drug components.

[0038] In the present invention, the term "pharmaceutical salt" refers to the salt derivative of the compound of the present invention. Through appropriate modification, the compound of the present invention can be formed into a corresponding acid salt or alkali salt. The pharmaceutical salt includes commonly used non-toxic salts or quaternary ammonium salts, which can be prepared with the compound of the present invention and a corresponding non-toxic inorganic or organic acid. For example, inorganic acids include hydrochloric acid, hydrobromic acid, sulfuric acid, aminosulfonic acid, phosphoric acid, and nitric acid, and organic acids include acetic acid, propionic acid, succinic acid, tartaric acid, citric acid, methanesulfonic acid, benzenesulfonic acid, glucuronic acid, glutamic acid, benzoic acid, salicylic acid, toluenesulfonic acid, oxalic acid, fumaric acid, and lactic acid, which can be used to form pharmaceutical acceptable salts. Other salts include ammonium salts such as trometamol, meglumine, and pyrrole ethanol, and metal salts such as sodium, potassium, calcium, zinc, and magnesium.

[0039] In the present invention, pharmaceutical salts can be prepared from parent compounds containing acidic or basic residues by conventional chemical methods. Generally, these salts can be obtained by reacting the free acid or free base forms of these compounds with a stoichiometric amount of an appropriate base or acid in water, an organic solvent, or a mixture of the two. Generally, ether, ethyl acetate, ethanol, isopropanol, or acetonitrile are preferred as non-aqueous reaction solvents. A list of suitable salts can be found in "Remington's Pharmaceutical Sciences", 17th Edition, Mack Publishing Company, Easton, PA, 1985, page 1418, the disclosure of which is incorporated by reference.

[0040] "Administering" or "Administration" refers to any manner in which a pharmaceutical or other agent is transferred, delivered, introduced, or conveyed to a subject. Such manners include oral administration, topical contact, intravenous, intraperitoneal, intramuscular, intralesional, nasal, subcutaneous, or cavity administration. Also contemplated by the present invention is the use of a device or instrument in administering an agent. Such devices may utilize active or passive transport and may be slow or fast release delivery devices.

[0041] The abbreviations and chemical names of the biological buffers are as follows: ACES (N-(2-acetamido)-2-aminoethanesulfonic acid) is used for buffering at pH 6.1 to 7.5 (pKa = 6.88). ADA (N-(2-acetamido)iminodiacetic acid, N-(carbamoylmethyl)iminodiacetic acid) is useful for buffering at pH 6.0-7.2 (pKa=6.65). AMPD (2-amino-2-methyl-1,3-propanediol) is useful for buffering at pH 7.8 to 9.7. AMPSO (N-(1,1-dimethyl-2-hydroxyethyl)-3-amino-2-hydroxypropanesulfonic acid). BES (N,N-bis(2-hydroxyethyl)-2-aminoethanesulfonic acid). Bicine (N,N-bis(2-hydroxyethyl)glycine), bis(2-hydroxyethyl)amino-tris(hydroxymethyl)methane) is used as a buffer at pH 5.8-7.2 (pKa=8.35). BisTris (Bis-(2-hydroxyethyl)amino-tris(hydroxymethyl)methane). BisTrisBisTrispropane (1,3-Bis[tris(hydroxymethyl)methylamino]propane). DIPSO (N,N-bis(2-hydroxyethyl)-3-amino-2-hydroxypropanesulfonic acid) is used for buffering at pH 7.0 to 8.2. Gly-Gly (diglycine, glycyl-glycine) is used for buffering at pH 7.5-8.9 (pKa=8.30). HEBPS (N-(2-hydroxyethyl)piperazine-N'-(4-butanesulfonic acid) is a homologue of HEPES and EPPS with a higher pKa (pKa = 8.30) and is used for buffering at pH 7.6-9.0. HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid; 2-morpholinoethanesulfonic acid; 2-(4-morpholino)ethanesulfonic acid; 2-(N-morpholino)ethanesulfonic acid; morpholine-4-ethanesulfonic acid hydrate) is widely used for buffering at pH 6.8-8.2 and has a pKa of 7.45-7.65 at 20 °C. HEPPS or EPPS (3-[4-(2-hydroxyethyl)-1-piperazinyl]propanesulfonic acid hydrate, 4-(2-hydroxyethyl)piperazine-1-(2-hydroxypropanesulfonic acid) hydrate) is used as a buffer at pH 7.3-8.7 (pKa=8.00 / piperazine ring). HEPPSO (4-(2-hydroxyethyl)piperazine-1-(2-hydroxypropanesulfonic acid) hydrate). MES (2-(N-morpholino)ethanesulfonic acid, monohydrate) is used as a buffer at pH 5.2-7.1 (pKa: 6.16). MOBS (4-morpholinebutanesulfonic acid; 3-(N-morpholino)butanesulfonic acid hemi-sodium salt) is a homologue of MES and MOPS with a higher pKa / , and is used in buffer solutions with a pH range of 6.9 to 8.3 (pKa: 7.6). MOPS (4-morpholinepropanesulfonic acid sodium salt). MOPSO (β-hydroxy-4-morpholinepropanesulfonic acid, 3-morpholino-2-hydroxypropanesulfonic acid). PIPES (piperazine-1,4-bis(2-ethanesulfonic acid) is used to buffer at pH 6.1-7.5 (pKa=6.80). POPSO (Piperazine-1,4-bis(2-hydroxypropanesulfonic acid) dihydrate). TAPS ([(2-hydroxy-1,1-bis(hydroxymethyl)ethyl)amino]-1-propanesulfonic acid). TAPSO (2-hydroxy-3-[tris(hydroxymethyl)methylamino]-1-propanesulfonic acid). TES (2-[(2-hydroxy-1,1-bis(hydroxymethyl)ethyl)amino]ethanesulfonic acid). Tricine (piperazine-N,N'-bis[2-hydroxypropanesulfonic acid]) is used to buffer at pH 7.4-8.8 (pKa: 8.16).

[0042] The term "antibody" is used herein in the broadest sense to encompass a variety of antibody structures, including, but not limited to, monoclonal antibodies, polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies), antibody fragments and fusion proteins comprising antibodies that exhibit the desired antigen-binding activity, and other modified configurations of immunoglobulin molecules that contain antigen recognition sites. Antibodies include antibodies of any class, such as IgG, IgA, or IgM (or subclasses thereof), and an antibody need not be of a particular class. Depending on the amino acid sequence of the constant region of the antibody's heavy chain, immunoglobulins can be assigned to various classes. There are five major classes of immunoglobulins: IgA, IgD, IgE, IgG, and IgM, and several of these can be further divided into subclasses (isotypes), e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2. The heavy chain constant regions corresponding to the various classes of immunoglobulins are called alpha, delta, epsilon, gamma, and mu, respectively. The subunit structures and three-dimensional configurations of the various classes of immunoglobulins are well known. "Antibody fragment" refers to a molecule other than an intact antibody that contains a portion of an intact antibody and binds to the antigen to which the intact antibody binds. Examples of antibody fragments include, but are not limited to, Fv, Fab, Fab', Fab-SH, F(ab') 2Humanized antibodies include: diabodies, linear antibodies, single chain antibody molecules (e.g., scFv), and multispecific antibody fragments formed from antibody fragment(s). A "humanized" antibody refers to a chimeric antibody comprising amino acid residues from a non-human HVR and amino acid residues from a human FR. In certain embodiments, a humanized antibody comprises substantially all of at least one, and typically two, variable domains, with all or substantially all of the HVRs (e.g., CDRs) corresponding to those of a non-human antibody, and with all or substantially all of the HVRs (e.g., CDRs) corresponding to the variable domains of the non-human antibody. Substantially all of the FRs correspond to the FRs of a human antibody. A humanized antibody may optionally comprise at least a portion of an antibody constant region derived from a human antibody. A "humanized form" of an antibody, e.g., a non-human antibody, refers to an antibody that has undergone humanization. The term "variable region" or "variable domain" refers to the domain of an antibody heavy or light chain that is involved in binding the antibody to an antigen. The heavy and light chain variable domains (VH and VL, respectively) of natural antibodies generally have a similar structure, with each domain containing four conserved framework regions (FR) and three hypervariable regions (HVR) (see, e.g., Kindt et al., Kuby Immunology, 6 th ed., WH Freeman and Co., page 91 (2007). A single VH or VL domain may be sufficient to confer antigen-binding specificity. Furthermore, antibodies that bind to a specific antigen can be isolated using the VH or VL domain from an antibody that binds the antigen to screen libraries of complementary VH or VL domains, respectively. See, e.g., Portolano et al., J. Immunol.150: 880-887 (1993); Clarkson et al., Nature 352: 624-628 (1991).

[0043] The term "monoclonal antibody" as used herein refers to an antibody obtained from a population of substantially homogeneous antibodies, i.e., the individual antibodies constituting the population are identical except for naturally occurring mutations that may be present in minor amounts. Monoclonal antibodies are highly specific, acting specifically on a single antigenic site. The modifier "monoclonal" indicates that the antibody is obtained from a population of substantially homogeneous antibodies, and should not be construed as requiring production of the antibody by any particular method. For example, monoclonal antibodies used in accordance with the present invention can be made by the hybridoma method first described by Kohler and Milstein, Nature 256:495, 1975, or by recombinant DNA methods such as those described in U.S. Pat. No. 4,816,567. Monoclonal antibodies can also be isolated from phage libraries generated using the techniques described in, for example, McCafferty et al., Nature 348:552-554, 1990.

[0044] As used herein, a "humanized" antibody refers to a form of a non-human (e.g., murine) antibody that is a chimeric immunoglobulin, immunoglobulin chain, or fragment thereof (such as Fv, Fab, Fab', F(ab')2, or other antigen binding subsequence of the antibody) that contains minimal sequence derived from the non-human immunoglobulin. Preferably, humanized antibodies are human immunoglobulins (recipient antibody) in which residues from a complementarity determining region (CDR) of the recipient are replaced by residues from a CDR of a non-human species (donor antibody) such as mouse, rat, or rabbit having the desired specificity, affinity, and capacity. In some cases, Fv framework region (FR) residues of the human immunoglobulin are replaced by corresponding non-human residues. Furthermore, humanized antibodies may comprise residues that are found neither in the recipient antibody nor in the imported CDR or framework sequences, but are included to further refine and optimize antibody performance. Generally, a humanized antibody comprises substantially all of at least one, typically two, variable domains, in which all or substantially all of the CDR regions correspond to those of a non-human immunoglobulin and all or substantially all of the FR regions are of the consensus sequence of a human immunoglobulin. The humanized antibody will optimally also comprise at least a portion of an immunoglobulin constant region or domain (Fc), typically of a human immunoglobulin. Antibodies with modified Fc regions are preferred, as described in WO 99 / 58572. Another form of humanized antibody has one or more CDRs (CDR L1, CDR L2, CDR L3, CDR H1, CDR H2, or CDR H3) that are "derived from" one or more CDRs of the original antibody, also referred to as one or more CDRs.

[0045] As used herein, "human antibody" refers to an antibody having an amino acid sequence that corresponds to that of an antibody produced by a human and / or an antibody made using any of the techniques for making human antibodies disclosed herein or known to those of skill in the art. This definition of a human antibody includes antibodies that comprise at least one human heavy chain polypeptide or at least one human light chain polypeptide. One such example is an antibody that comprises a mouse light chain and a human heavy chain polypeptide. Human antibodies can be produced using a variety of techniques known in the art. In one embodiment, human antibodies are selected from phage libraries that express human antibodies (Vaughan et al., Nature Biotechnology, 14:309-314, 1996; Sheets et al., Proc. Natl. Acad. Sci (USA) 95:6157-6162, 1998; Hoogenboom and Winter, J. Mol. Biol., 227:381, 1991; Marks et al., J. Mol. Biol., 222:581, 1991). Human antibodies can also be produced by immunization of animals transgenic for human immunoglobulin loci in place of endogenous loci, e.g., mice in which the endogenous immunoglobulin genes have been partially or completely inactivated. This approach is described in U.S. Patent Nos. 5,545,807; 5,545,806; 5,569,825; 5,625,126; and 5,633,425. Alternatively, human antibodies can be prepared by immortalizing human B lymphocytes that produce antibodies against the target antigen (such B lymphocytes can be recovered from an individual or from single cell cloning of cDNA or immunized in vitro). See, for example, Cole et al. Monoclonal Antibodies and Cancer Therapy, Alan R. Liss, p. 77, 1985; Boerner et al., J. Immunol., 147 (1):86-95, 1991; and U.S. Patent No. 5,750,373.

[0046] The term "chimeric antibody" is intended to refer to an antibody in which the variable region sequences are derived from one species and the constant region sequences are derived from another species, e.g., the variable region sequences are derived from a murine antibody and the constant region sequences are derived from a human antibody.

[0047] The terms "polypeptide", "oligopeptide", "peptide", and "protein" are used interchangeably herein and refer to a chain of amino acids of any length, preferably relatively short (e.g., 10-100 amino acids). The chain may be linear or branched, may contain modified amino acids, and / or may be interrupted by non-amino acids. The term also includes amino acid chains that have been modified, either naturally or by intervention, for example, by formation of disulfide bonds, glycosylation, lipidation, acetylation, phosphorylation, or other manipulations or modifications, such as conjugation with a labeling component. Also included in this definition are polypeptides that contain, for example, analogs of one or more amino acids (including, for example, unnatural amino acids, etc.), as well as other modifications known in the art. It is understood that polypeptides can exist as single chains or linked chains.

[0048] A "monovalent antibody" contains one antigen binding site per molecule (e.g., IgG or Fab). In some cases, a monovalent antibody can have one or more antigen binding sites, but the binding sites are from different antigens.

[0049] A "monospecific antibody" contains two identical antigen-binding sites per molecule, such as two binding sites that bind to the same epitope on an antigen (e.g., IgG). Thus, they compete to bind one type of antigen molecule. Most naturally occurring antibodies are monospecific. In some cases, a monospecific antibody may be a monovalent antibody (e.g., Fab).

[0050] A "bivalent antibody" contains two antigen-binding sites per molecule (e.g., IgG). In some cases, the two binding sites may have the same antigen specificity. However, a bivalent antibody may be bispecific.

[0051] "Bispecific" or "dual-specific" is a hybrid antibody that has two different antigen-binding sites. The two antigen-binding sites of a bispecific antibody bind to two different epitopes located on the same or different protein targets.

[0052] A "bifunctional" antibody is one that has the same antigen-binding site (ie, the same amino acid sequence) in both arms, but each arm can recognize two different antigens.

[0053] A "heteromultimer," "heteromultimeric complex," or "heteromultimeric polypeptide" is a molecule that comprises at least a first polypeptide and a second polypeptide, where the second polypeptide differs in amino acid sequence from the first polypeptide at at least one amino acid residue. A heteromultimer may comprise a "heterodimer" formed by the first and second polypeptides, or may comprise a higher order tertiary structure that is formed when polypeptides are present in addition to the first and second polypeptides.

[0054] A "heterodimer," "heterodimeric protein," "heterodimeric complex," or "heterodimeric polypeptide" is a molecule comprising a first peptide and a second peptide, where the amino acid sequence of the second peptide differs from the amino acid sequence of the first peptide by at least one amino acid residue.

[0055] In this patent application, the terms "hinge region", "hinge sequence" and variants thereof include the meanings well known in the art, e.g., as set forth in Janeway et al., ImmunoBiology: the immune system in health and disease, (Elsevier Science Ltd., NY) (4th ed., 1999); Bloom et al., Protein Science (1997), 6:407-415; Humphreys et al., J. Immunol. Methods (1997), 209:193-202.

[0056] In this patent application, "immunoglobulin-like hinge region", "immunoglobulin-like hinge sequence" and variants thereof refer to the hinge region and hinge sequence of an immunoglobulin-like or antibody-like molecule (e.g., an immunoadhesin). In some embodiments, the immunoglobulin-like hinge region can come from or be derived from any of the IgG1, IgG2, IgG3, or IgG4 subtypes, or IgA, IgE, IgD, or IgM, including chimeric forms thereof, e.g., chimeric IgG1 / 2 hinge regions.

[0057] In this patent application, the term "immune effector cell" or "effector cell" refers to a cell within the natural cellular repertoire of the human immune system that can be activated to affect the viability of a target cell, including the ability of the cell to survive, proliferate, and / or interact with other cells.

[0058] The antibodies of the present invention can be prepared by techniques well known in the art, such as recombinant techniques, phage display techniques, synthetic techniques, or a combination of these techniques with other techniques known in the art (see, e.g., Jayasena, SD, Clin Chem., 45: 1628-50, 1999 and Fellouse, FA, et al., J. Mol. Biol., 373(4): 924-40, 2007).

[0059] In this patent application, the term "cytotoxic agent" refers to a substance that inhibits or prevents cellular function and / or causes cell death or destruction. Cytotoxic agents include radioisotopes (e.g., At211, I131, I125, Y90, In111, Re186, Re188, Sm153, Bi212, P32, Pb212, Zr89, F18, and radioisotopes of Lu, such as Lu177); chemotherapeutic agents or drugs (e.g., tubulysin, maytansine, auristatin, DNA minor groove binders (such as PBD dimers), ducarmysin, topoisomerase inhibitors, RNA polymerase inhibitors, DNA alkylating agents, melanoma inhibitors, erythropoietin ... growth inhibitors; enzymes and fragments thereof, such as nucleases; antibiotics; toxins, such as small molecule toxins or enzymatically active toxins of bacterial, fungal, plant, or animal origin (including fragments and / or variants thereof); and various anti-tumor or anti-cancer agents disclosed throughout this application.

[0060] "Linker" refers to a chemical moiety that contains a covalent bond or chain of atoms that covalently attaches an antibody to a drug moiety. In various embodiments, the linker includes a divalent radical, such as alkyldiyl, aryldiyl, heteroaryldiyl, -(CR 2 ) n O(CR 2 ) n moieties such as -, repeating units of alkyloxy (e.g., polyethyleneoxy, PEG, polymethyleneoxy), and alkylamino (e.g., polyethyleneamino); diacid esters and amides include succinate, succinamide, diglycolate, malonate, and caproamide. In various embodiments, the linker can include one or more amino acid residues such as valine, phenylalanine, lysine, and homolysine.

[0061] In the present specification and claims, the terms "comprise", "comprising", "including" and "includes" are used to indicate the presence of features, integers, components, or steps, but do not exclude the presence or coexistence of one or more other features, integers, components, steps, or groups thereof. The novel conjugates disclosed herein use bridge linkers. Some suitable linkers and their synthetic examples are described herein in embodiments 1-468.

[0062] BCMA antibodies and their antibody-drug conjugates

[0063] The present invention provides monoclonal antibodies that specifically bind to BCMA (CD269). Unless otherwise specified, BCMA refers to human BCMA. Exemplary human nucleic acid and amino acid sequences are provided by SEQ ID NOs: 1 and 2. Unless otherwise clear from the context, reference to BCMA refers to at least the extracellular domain of the protein (approximately residues 1-54 of SEQ ID NO: 7) and sometimes the entire protein.

[0064] The invention provides a method for treating a medical disorder in a human subject, the medical disorder being associated with the presence of pathogenic B cells that express B-cell maturation antigen (BCMA), the method comprising administering to the human subject an isolated monoclonal antibody or antigen-binding fragment thereof that binds BCMA (CD269).

[0065] The BCMA antibodies of the present invention (e.g., hu5D2) are humanized monoclonal antibodies that specifically bind to human BCMA, as described in the Examples. The 5D2 antibody was produced by hybridoma BCMA-A2-6H4-5D2. The deposit at the China Center for Typical Culture Collection (CCTCC) was made under the Budapest Treaty on June 23, 2022. The CCTCC is located at Wuhan University, Wuhan, Hubei Province, China, zip code 430000. The CCTCC deposit was assigned the accession number CCTCC C2022188. The Hu5D2 antibody inhibits binding to both BCMA and its ligands (APRIL and BAFF). The Hu5D2 antibody induces ADCC upon binding to human IgG1 and induces signaling through the Fcγ receptor. The Hu5D2 antibody can also be incorporated into antibody-drug conjugates to deliver the conjugated drug to the interior of cells expressing BCMA.

[0066] The Hu5D2 antibody is another humanized monoclonal antibody that specifically binds to human BCMA, inhibits binding to its ligands, and can deliver the bound drug to the interior of cells expressing BCMA.

[0067] The present invention provides antigen binding proteins capable of binding and internalizing membrane-bound targets. In a further embodiment, an immunoconjugate is provided comprising the antigen binding protein of the present invention and a cytotoxic agent. In a further embodiment, the antigen binding protein has ADCC effector function, e.g., the antigen binding protein has enhanced ADCC effector function.

[0068] In one such embodiment, there is provided an antigen binding protein or fragment thereof that specifically binds to BCMA, e.g. which specifically binds to human BCMA (hBCMA) and inhibits binding of BAFF and / or APRIL to the BCMA receptor.

[0069] In a further embodiment, the antigen binding protein or fragment of the invention specifically binds to BCMA and inhibits binding of BAFF and / or APRIL to BCMA, wherein the antigen binding protein or fragment thereof has the ability to bind to FcγRIIIA and mediate FcgRIIIA mediated effector, or has enhanced Fc.γRIIIA mediated effector function. In one embodiment of the invention provided herein, the antigen binding protein is capable of internalization.

[0070] In one aspect of the invention there is provided an antigen binding protein according to the invention as described herein which binds to non-membrane bound BCMA, such as serum BCMA.

[0071] In one aspect of the invention there is provided an antigen binding protein as described herein, wherein the antigen binding protein comprises a CDRH3 of SEQ ID NO:3 or a variant of SEQ ID NO:3.

[0072] In a further aspect of the invention there is provided an antigen binding protein as described herein further comprising one or more of the following: CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and / or CDRL3 of SEQ ID NO: 6, and variants thereof.

[0073] The antigen binding proteins of the invention may comprise the heavy and light chain variable regions of the invention, which may be formatted into the structure of a natural antibody or a functional fragment or equivalent thereof. Thus, the antigen binding proteins of the invention may comprise the VH regions of the invention formatted into a full-length antibody, a (Fab')2 fragment, a Fab fragment, or equivalents thereof (scFV, dimer, trimer, or tetramer, tandem antibody, etc.) when combined with an appropriate light chain. The antibody may be IgG1, IgG2, IgG3, or IgG4; or IgM, IgA, IgE, or IgD, or modified variants thereof. The constant domain of the antibody heavy chain may be selected accordingly. The light chain constant domain may be a kappa or lambda constant domain. Furthermore, the antigen binding proteins may comprise all classes of modifications, such as IgG dimers, Fc variants that no longer bind Fc receptors or mediate C1q binding. The antigen binding protein may be a chimeric antibody of the type described in WO86 / 001533, which comprises an antigen binding region and a non-immunoglobulin region.

[0074] The constant region is selected depending on the required function, for example IgG1 exhibits lytic capability through binding to complement and / or mediates ADCC (antibody dependent cellular cytotoxicity).

[0075] The antigen binding proteins of the present invention are derived from a murine antibody having the variable regions set forth in SEQ ID NO: 10 and 11, or a non-murine equivalent thereof, such as a rat, human, chimeric or humanized variant. For example, the antigen binding proteins of the present invention are derived from an antibody having the variable heavy chain sequence set forth in SEQ ID NO: 10 and / or the variable light chain sequence set forth in SEQ ID NO: 11.

[0076] In one aspect of the invention there is provided an antigen binding protein comprising an isolated heavy chain variable domain selected from any one of SEQ ID NOs: 8, 10 or 13.

[0077] In another aspect of the invention there is provided an antigen binding protein comprising an isolated light chain variable domain selected from any one of SEQ ID NOs: 9, 11 or 15.

[0078] In a further aspect of the invention there is provided an antigen binding protein comprising an isolated heavy chain variable domain selected from any one of SEQ ID NOs: 8, 10 or 13 and an isolated light chain variable domain selected from any one of SEQ ID NOs: 9, 11 or 15.

[0079] In one aspect, the antigen binding protein of the present invention is provided as an antigen binding protein comprising a heavy chain variable region encoded by SEQ ID NO:20 or 22 and a light chain variable region encoded by SEQ ID NO:21 or 23.

[0080] In one aspect, a polynucleotide encoding the isolated variable heavy chain is provided, wherein said polynucleotide comprises SEQ ID NO: 28, 29, or 30.

[0081] In one aspect, a polynucleotide encoding the isolated variable light chain is provided, wherein said polynucleotide comprises SEQ ID NO: 31, 32 or 33.

[0082] In a further aspect, the antigen binding protein may comprise any one of the variable heavy chains described herein in combination with any of the light chains described herein.

[0083] In one aspect the antigen binding protein is an antibody or antigen binding fragment thereof comprising one or more CDRs according to the invention as described herein or any one or both of the heavy or light chain variable domains of the invention as described herein. In one embodiment the antigen binding protein binds to primate BCMA. In one such embodiment the antigen binding protein further binds to non-human primate BCMA, such as cynomolgus BCMA.

[0084] In another embodiment, the antigen binding protein is selected from the group consisting of a dAb, Fab, Fab', F(ab'), Fv, diabody, triabody, tetrabody, miniantibody, and minibody.

[0085] In one aspect of the invention, the antigen binding protein is a humanized or chimeric antibody, in a further aspect the antibody is humanized. In one aspect the antibody is a monoclonal antibody.

[0086] In another aspect, the antigen binding protein binds to human BCMA with high affinity, for example as measured by Biacore, the antigen binding protein binds to human BCMA with an affinity of 20 nM or less, or an affinity of 15 nM or less, or an affinity of 5 nM or less, or an affinity of 1000 pM or less, or an affinity of 500 pM or less, or an affinity of 400 pM or less, or an affinity of 300 pM or less, or for example with an affinity of about 120 pM. In a further embodiment, the antigen binding protein binds to human BCMA with between about 100 pM and about 500 pM, or between about 100 pM and about 400 pM, or between about 100 pM and about 300 pM, as measured by Biacore. In one embodiment of the invention, the antigen binding protein binds to BCMA with an affinity of less than 150 pM.

[0087] In one such embodiment, this is measured by Biacore, for example as described in Example 4.

[0088] In another aspect, the antigen binding protein binds to human BCMA and neutralises binding of the ligands BAFF and / or APRIL to the BCMA receptor in a cell neutralisation assay, wherein the antigen binding protein has a 1050 of about 1 nM to about 500 nM, or about 1 nM to about 100 nM, or about 1 nM to about 50 nM, or about 1 nM to about 25 nM, or about 5 nM to about 15 nM. In a further embodiment of the invention, the antigen binding protein binds to BCMA and neutralises BCMA in a cell neutralisation assay, wherein the antigen binding protein has a 1050 of about 10 nM.

[0089] Antigen binding proteins, such as antibodies of the present invention, can be produced by transfection of a host cell with an expression vector comprising the coding sequence of an antigen binding protein of the present invention. The expression vector or recombinant plasmid is produced by placing the coding sequence of the antigen binding protein in operative association with conventional regulatory control sequences capable of controlling the replication and expression in and / or secretion from the host cell. Regulatory sequences include promoter sequences, such as the CMV promoter, and signal sequences that may be derived from other known antibodies. Similarly, a second expression vector can be produced with a DNA sequence encoding a complementary antigen binding protein light or heavy chain. In certain embodiments, this second expression vector is identical to the first expression vector, except as far as the coding sequences and selection markers are concerned, in order to ensure as far as possible that each polypeptide chain is functionally expressed. Alternatively, the heavy and light chain coding sequences of the antigen binding protein may be present on a single vector.

[0090] The selected host cell is co-transfected with both the first and second vectors (or simply transfected with a single vector) by conventional techniques to generate a transfected host cell of the invention containing both recombinant or synthetic light and heavy chains. The transfected cells are then cultured by conventional techniques to produce the engineered antigen binding protein of the invention. Antigen binding proteins containing both recombinant heavy and / or light chains are screened from the culture by a suitable assay, such as ELISA or RIA. Similar conventional techniques can be used to construct other antigen binding proteins.

[0091] A vector suitable for the cloning and subcloning steps used in constructing the method and composition of the present invention can be selected by those skilled in the art. For example, conventional pUC series cloning vectors can be used. One vector, pUC19, is commercially available from suppliers such as AmershamBioscience (Buckinghamshire, UK) or GenScript (Nanjing, China). Furthermore, any vector that can be easily replicated, has abundant cloning sites and selectable genes (e.g., antibiotic resistance), and can be easily manipulated can be used for cloning. Therefore, the selection of the cloning vector is not a limiting factor in the present invention.

[0092] The expression vector may also be characterized by a gene suitable for amplifying the expression of the heterologous DNA sequence, for example, the mammalian dihydrofolate reductase gene (DHFR). Other vector sequences include polyA signal sequences, such as bovine growth hormone (BGH) and the beta globin promoter sequence (beta globulo). The expression vectors useful herein may be synthesized by techniques well known to those skilled in the art.

[0093] The components of such vectors, such as replicons, selection genes, enhancers, promoters, signal sequences, etc., can be obtained from commercial or natural sources or synthesized by known procedures for use in directing the expression and / or secretion of recombinant DNA products in a selected host. Other suitable expression vectors, of which a large variety are known in the art for mammalian, bacterial, insect, yeast, and fungal expression, can also be selected for this purpose.

[0094] The present invention also encompasses cell lines transfected with recombinant plasmids containing the coding sequences of the antigen binding proteins of the present invention. Host cells useful for the cloning and other manipulations of these cloning vectors are also conventional. However, cells from various strains of E. coli can be used for replicating the cloning vectors and other steps in the construction of the antigen binding proteins of the present invention.

[0095] Suitable host cells or cell lines for expression of the antigen binding proteins of the invention include mammalian cells such as NS0, Sp2 / 0, CHO (e.g. DG44), COS, HEK, fibroblasts (e.g. 3T3), and myeloma cells. For example, the antigen binding proteins of the invention may be expressed in CHO or myeloma cells. Human cells may also be used, allowing the molecules to be modified with human glycosylation patterns.

[0096] Alternatively, other eukaryotic cell lines may be used. The selection of suitable mammalian host cells and methods for transformation, culture, amplification, screening, and product production and purification are known in the art. See, for example, Sambrook et al., (1989). Molecular cloning: a laboratory manual, 2 nd See ed. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY.

[0097] Bacterial cells may prove useful as suitable host cells for the expression of recombinant Fabs or other embodiments of the invention (see, e.g., Pluckthun, A., Immunol. Rev., 130:151-188 (1992)). However, because proteins expressed in bacterial cells tend to be in unfolded or improperly folded or non-glycosylated forms, recombinant Fabs produced in bacterial cells must be screened for retention of antigen-binding ability. If the molecule expressed by the bacterial cell is produced in a properly folded form, the bacterial cell will be a desirable host, or in another embodiment, the molecule may be expressed in a bacterial host and then refolded. For example, in the field of biotechnology, various strains of E. coli are well known host cells used for expression. Various strains of Bacillus subtilis, Streptomyces, other bacilli, etc. may also be used in this method.

[0098] If desired, strains of yeast cells known to those skilled in the art can also be used as host cells, as can insect cells such as Drosophila and Lepidoptera, and viral expression systems. See, for example, Miller et al., Genetic Engineering, 8:277-298, Plenum Press (1986) and McGuire, S et a., Trends Genet. (2004) 20, 384-391, and references cited therein.

[0099] The general methods by which vectors can be constructed, the transfection methods required to produce the host cells of the invention, and the culture methods required to produce the antigen binding proteins of the invention from such host cells can all be conventional techniques. Typically, the culture methods of the invention are serum-free culture methods, usually by culturing the cells in suspension without serum. Similarly, the antigen binding proteins of the invention can be purified from the cell culture contents after production according to standard procedures in the art, including ammonium precipitation, affinity columns, column chromatography, gel electrophoresis, and the like. Such techniques are within the scope of the skilled artisan and do not limit the invention. For example, the preparation of modified antibodies is described in WO99 / 058679 and WO96 / 016990. Yet another method of expression of antigen binding proteins can utilize expression in transgenic animals, as described in US Pat. No. 4,873,316. ​​This relates to an expression system using an animal casein promoter, which when transgenically incorporated into a mammal, enables the female to produce the desired recombinant protein in her milk.

[0100] In a further embodiment of the invention there is provided a method of producing an antibody of the invention comprising culturing a host cell transformed or transfected with a vector encoding the light chain and / or the heavy chain of an antibody of the invention and recovering the antibody produced thereby.

[0101] According to the invention there is provided a method for producing an anti-BCMA antibody of the invention which binds to and neutralises the activity of human BCMA, comprising the steps of providing a first vector encoding an antibody heavy chain, providing a second vector encoding an antibody light chain, transforming a mammalian host cell (e.g. CHO) with said first and second vectors, culturing the host cell in step (c) under conditions conducive to secretion of the antibody from the host cell into the culture medium, and recovering the secreted antibody in step (d).

[0102] The antibodies expressed in the desired manner are tested for in vitro activity using a suitable assay.Currently, a conventional ELISA assay format is used to evaluate the qualitative and quantitative binding of antibodies to BCMA.In addition, other in vitro assays can be used to verify neutralizing effects before subsequent human clinical studies are performed to evaluate the persistence of antibodies in the body despite normal clearance mechanisms.

[0103] The dosage and duration of treatment relate to the relative duration of the molecules of the invention (antibodies and antibody-drug conjugates) in human circulation and can be adjusted by those skilled in the art depending on the condition being treated and the general health of the patient. It is anticipated that repeated dosing (e.g., once a week, or once every two weeks, or once every three weeks, or once every four weeks) over an extended period (e.g., 4-6 months) may be required to achieve maximum therapeutic efficacy.

[0104] In one embodiment of the present invention, a recombinantly transformed, transfected or transduced host cell is provided comprising at least one expression cassette, for example the expression cassette comprises a polynucleotide encoding a heavy chain of an antigen binding protein according to the invention as described herein, further comprising a polynucleotide encoding a light chain of an antigen binding protein according to the invention as described herein, or there are two expression cassettes, a first cassette encoding a light chain and a second cassette encoding a heavy chain. For example, in one embodiment, a first expression cassette comprises a polynucleotide encoding a heavy chain of an antigen binding protein comprising a constant region or an antigen binding fragment thereof linked to a constant region according to the invention as described herein, and further comprising a second expression cassette comprising a polynucleotide encoding a light chain of an antigen binding protein comprising a constant region or an antigen binding fragment thereof linked to a constant region according to the invention as described herein, for example the first expression cassette comprises a polynucleotide encoding a heavy chain selected from SEQ ID NO: 18 or 25 and the second expression cassette comprises a polynucleotide encoding a light chain selected from SEQ ID NO: 19 or 27.

[0105] In another embodiment of the present invention, a stably transformed host cell is provided comprising a vector comprising one or more expression cassettes encoding a heavy and / or light chain of an antibody comprising a constant region or an antigen-binding fragment thereof that binds to a constant region as described herein. For example, such a host cell may comprise a first vector encoding a light chain and a second vector encoding a heavy chain, e.g., the first vector encoding a heavy chain selected from SEQ ID NO: 18 or 25, and the second vector encoding a light chain, e.g., the light chain of SEQ ID NO: 19 or 27. In one such example, the first vector encoding a heavy chain selected from SEQ ID NO: 18, and the second vector encoding a light chain, e.g., the light chain of SEQ ID NO: 19.

[0106] In another embodiment of the invention there is provided a host cell according to the invention as described herein, wherein the cell is a eukaryotic cell, for example the cell is mammalian. Examples of such cell lines include CHO or NS0.

[0107] In another embodiment of the invention, there is provided a method for producing an antibody comprising a constant region or an antigen-binding fragment thereof linked to a constant region according to the invention as described herein, the method comprising culturing a host cell in a medium, e.g. a serum-free medium.

[0108] In another embodiment of the invention, there is provided a method according to the invention described herein, wherein the antibody is further purified to at least 95% or more (e.g., 98% or more) relative to the antibody comprising serum-free medium.

[0109] In yet another embodiment, a pharmaceutical composition is provided comprising the antigen binding protein and a pharma- ceutically acceptable carrier.

[0110] In another embodiment of the present invention, a kit of parts is provided comprising a composition according to the present invention as described herein together with instructions for use.

[0111] The mode of administration of the therapeutic agent of the present invention can be any suitable route that delivers the therapeutic agent to the host. The antigen binding proteins and pharmaceutical compositions of the present invention are particularly useful for parenteral administration, i.e., subcutaneous (sc), intrathecal, intraperitoneal, intramuscular (im), or intravenous (iv). In one such embodiment, the antigen binding protein of the present invention is administered intravenously or subcutaneously.

[0112] The therapeutic agent of the present invention may be prepared as a pharmaceutical composition containing an effective amount of the antigen binding protein of the present invention as an active ingredient in a pharma- ceutically acceptable carrier. In one embodiment, the prophylactic agent of the present invention is an aqueous suspension or solution containing the antigen binding protein in a ready-to-inject form. In one embodiment, the suspension or solution is buffered at physiological pH. In one embodiment, a composition for parenteral administration will contain a solution of the antigen binding protein of the present invention or a cocktail thereof dissolved in a pharma- ceutically acceptable carrier. In one embodiment, the carrier is an aqueous carrier. A variety of aqueous carriers may be used, e.g., 0.9% saline, 0.3% glycine, and the like. These solutions may be made sterile and generally free of particulate matter. These solutions may be sterilized by conventional, well-known sterilization techniques (e.g., filtration). The composition may contain pharma- ceutical acceptable auxiliary substances, such as pH adjusting and buffering agents, as necessary to approximate physiological conditions. The concentration of the antigen binding protein of the present invention in such pharmaceutical preparations may vary over a wide range, i.e., from less than about 0.5%. Typically, this will be at least about 1% by weight to as much as about 15 or 20% by weight, selected primarily based on fluid volumes, viscosities, and the like, in accordance with the particular mode of administration selected.

[0113] Thus, a pharmaceutical composition of the invention for intravenous infusion can be made up to contain about 250 ml of sterile Ringer's solution and about 1 to about 30 or 5 mg to about 25 mg of an antigen-binding protein of the invention per ml of Ringer's solution. Actual methods for preparing parenterally administrable compositions are well known or will be apparent to those skilled in the art and are described in further detail, for example, in Remington's Pharmaceutical Science, 15. sup. th ed., Mack Publishing Company, PA, USA. For the preparation of intravenously administrable antigen-binding protein formulations of the present invention, see Parkins D. and Lasmar U. "The formulation of Biopharmaceutical products", Pharm. Sci. Tech. Today, 3 (2000) 129-137; Wang, W "Instability, stabilisation and formulation of liquid protein pharmaceuticals", Int. J. Pharm 185 (1999) 129-188; Jorgensen, L. et al, "Recent trends in stabilising peptides and proteins in pharmaceutical formulation - considerations in the choice of excipients" Expert Opin Drug Deliv. 6 (2009) 1219-1230; Akers, MJ "Excipient-Drug interactions in Parenteral Formulations", J. Pharm Sci 91 (2002) 2283-2300; Imamura, K et al "Effects of types of sugar on stabilization of Protein in the dried state", J Pharm Sci 92 (2003) 266-274; Izutsu, Kkojima, S."Excipient crystallinity and its protein-structure-stabilizing effect during freeze-drying", J. Pharm. Pharmacol, 54 (2002) 1033-1039; Johnson, R, et al "Mannitol-sucrose mixtures--versatile formulations for protein lyophilization", J. Pharm. Sci, 91 (2002) 914-922; Kerwin B. “Polysorbates 20 and 80 used in the formulation of protein biotherapeutics: structure and degradation pathways” J. Pharm Sci. 97 (2008) 2924-2935; Ha, E., et al "Peroxide formation in polysorbate 80 and protein stability", J. Pharm Sci, 91 (2002), 2252-2264, and He, F., et al, “Effect of sugar molecules on the viscosity of high concentration monoclonal antibody See, “Scientific Solutions” Pharm Res. 28 (2011) 1552-1560, the entire contents of which are incorporated herein by reference and the reader is specifically referred to.

[0114] In one embodiment, the antibody of the present invention, when in a pharmaceutical formulation, is present in a unit dose form. The appropriate therapeutically effective dose will be readily determined by one of skill in the art. The appropriate dose can be calculated according to the patient's body weight, for example, the appropriate dose may be in the range of about 0.1 to about 200 mg / kg, such as about 1 to about 20 mg / kg, such as about 10 to about 20 mg / kg, or for example, about 1 to about 15 mg / kg, such as about 5 to about 15 mg / kg. To effectively treat conditions such as multiple myeloma, SLE, or IPT in humans, the appropriate dose may be in the range of about 0.1 to about 2000 mg, such as about 0.1 to about 500 mg, such as about 500 mg, such as about 0.1 mg to about 150 mg, or about 0.1 to about 80 mg, or about 0.1 to about 60 mg, or about 0.1 to about 40 mg, or for example, about 1 to about 100 mg, or about 1 to about 50 mg. The antigen binding proteins of the present invention may be administered parenterally, for example subcutaneously, intravenously or intramuscularly. Such doses may, if necessary, be repeated at appropriate time intervals appropriately selected by the physician.

[0115] The antigen binding proteins described herein can be lyophilized for storage and reconstituted in a suitable carrier prior to use, a technique that has been shown to be effective with conventional immunoglobulins and peroxide and reconstitution techniques known in the art.

[0116] In another aspect of the invention there is provided an antigen binding protein as described herein for use in medicine.

[0117] In one aspect of the invention there is provided an antigen binding protein according to the invention as described herein for use in the treatment of rheumatoid arthritis, type 1 diabetes, multiple sclerosis or psoriasis, said method comprising the step of administering to said patient a therapeutically effective amount of an antigen binding protein as described herein.

[0118] In one embodiment of the invention there is provided a method for treating cancer in a human comprising administering to said human an antigen binding protein that specifically binds to BCMA, optionally wherein the antigen binding protein is part of an immune complex.

[0119] In another aspect of the invention, the present invention is directed to the treatment of multiple myeloma (MM), lymphocytic leukemia (CLL), non-secretory multiple myeloma, smoldering multiple myeloma, monoclonal gammopathy of undetermined significance (MGUS), solitary plasmacytoma (bone, extramedullary), lymphoplasmacytic lymphoma (LPL), Waldenstrom's macroglobulinemia, plasma cell leukemia, primary amyloidosis (AL), heavy chain disease, systemic lupus erythematosus (SLE), POEMS syndrome / osteosclerotic myeloma, types I and II cryoglobulinemia, light chain deposition disease, Goodpasture's syndrome, idiopathic thrombocytopenic purpura (I thrombocytopenic purpura), and / or other inflammatory diseases. There is provided an antibody according to the invention as described herein for use in the treatment of a B cell or plasma cell mediated disease, or an antibody mediated disease or disorder selected from acute glomerulonephritis, pemphigus and pemphigoid disorders, epidermolysis bullosa acquisita, or any non-Hodgkin's lymphoma B cell leukemia or Hodgkin's lymphoma (HL) with BCMA expression, or any disease in which the patient expresses neutralizing antibodies to recombinant protein replacement therapy, wherein the method comprises administering to said patient a therapeutically effective amount of an antigen binding protein as described herein.

[0120] B cell disorders can be categorized as defects in B cell development / immunoglobulin production (immunodeficiency) and excessive / uncontrolled proliferation (lymphoma, leukemia). As used herein, B cell disorders refers to both types of disease, and methods are provided for treating B cell disorders using antigen binding proteins.

[0121] In certain embodiments, the disease or disorder is selected from the group consisting of multiple myeloma (MM), chronic lymphocytic leukemia (CLL), solitary plasmacytoma (bone, extramedullary), Waldenstrom's macroglobulinemia.

[0122] In one aspect of the invention, the disease is multiple myeloma, smoldering multiple myeloma (SMM), or solitary plasmacytoma (bone, extramedullary).

[0123] In one aspect of the invention, the disease is multiple myeloma.

[0124] In one aspect of the invention, the disease is systemic lupus erythematosus (SLE).

[0125] In one aspect of the invention, the disease is idiopathic thrombocytopenic purpura (ITP).

[0126] There is also provided the use of the antigen binding proteins described herein in the manufacture of a medicament for the treatment of the diseases and disorders described herein.

[0127] For example, in one aspect of the invention there is provided the use of an antigen binding protein as described herein for use in the treatment or prevention of diseases and disorders that respond to modulation (such as inhibition or or blocking) of the interaction between BCMA and the ligands BAFF and APRIL.

[0128] In another aspect of the invention there is provided the use of an antigen binding protein as described herein for use in the treatment or prophylaxis of an antibody-mediated or plasma cell-mediated disease or disorder selected from rheumatoid arthritis, type 1 diabetes, multiple sclerosis, rheumatoid arthritis, or psoriasis.

[0129] In another aspect of the invention, the present invention is directed to the treatment of multiple myeloma (MM), chronic lymphocytic leukemia (CLL), monoclonal gammopathy of undetermined significance (MGUS), smoldering multiple myeloma (SMM), isolated plasmacytoma (bone, extramedullary), Waldenstrom's macroglobulinemia, primary amyloidosis (AL), heavy chain disease, systemic lupus erythematosus (SLE), POEMS syndrome / osteosclerotic myeloma, types I and II cryoglobulinemia, light chain deposition disease, Goodpastures syndrome, idiopathic hematologic malignancies, and idiopathic hematologic malignancies. There is provided use of an antibody as described herein for use in the treatment or prevention of a plasma cell mediated disease or disorder selected from thrombocytopenic purpura (ITP), acute glomerulonephritis, pemphigus and pemphigoid disorders and epidermolysis bullosa acquisita, non-Hodgkin's lymphomas and leukemias with BCMA expression, or any disease in which patients express neutralizing antibodies to recombinant protein replacement therapy, wherein the method comprises administering to said patient a therapeutically effective amount of an antigen binding protein as described herein.

[0130] In one aspect, the present invention provides a pharmaceutical composition comprising the antibody of the present invention or a functional fragment thereof and a pharma- ceutically acceptable carrier for the treatment or prevention of rheumatoid arthritis, type 1 diabetes, multiple sclerosis or psoriasis, or an antibody-mediated or plasma cell-mediated disease or disorder, wherein the antibody-mediated or plasma cell-mediated disease or disorder is selected from multiple myeloma (MM), chronic lymphocytic leukemia (CLL), monoclonal gammopathy of undetermined significance (MGUS), smoldering multiple myeloma (SMM), solitary plasmacytoma (bone, extramedullary), Waldenstrom's macroglobulinemia, primary cell-mediated disease or disorder. The antigen binding protein is selected from amyloidosis (AL), heavy chain disease, systemic lupus erythematosus (SLE), POEMS syndrome / osteosclerotic myeloma, types I and II cryoglobulinemia, light chain deposition disease, Goodpastures syndrome, idiopathic thrombocytopenic purpura (ITP), acute glomerulonephritis, pemphigus and pemphigoid disorders epidermolysis bullosa acquisita, any non-Hodgkin's lymphoma and leukemia with BCMA expression, or any disease in which a patient expresses neutralizing antibodies to recombinant protein replacement therapy, wherein the method comprises administering to the patient a therapeutically effective amount of an antigen binding protein as described herein.

[0131] In another embodiment of the invention there is provided a method of treating a human patient suffering from rheumatoid arthritis, type 1 diabetes, multiple sclerosis or psoriasis, or an antibody mediated or plasma cell mediated disorder or disease, comprising administering a therapeutically effective amount of an antigen binding protein according to the invention as described herein, for example a method of treating a human patient suffering from an antibody mediated or plasma cell mediated disease or disorder selected from: In another aspect of the invention there is provided an antigen binding protein according to the invention as described herein for use in the treatment of an antibody mediated or plasma cell mediated disease or disorder selected from multiple myeloma (MM), chronic lymphocytic leukemia (CLL), monoclonal gammopathy of undetermined significance (MGUS). ), smoldering multiple myeloma (SMM), isolated plasmacytoma (bone, extramedullary), Waldenstrom's macroglobulinemia, primary amyloidosis (AL), heavy chain disease, systemic lupus erythematosus (SLE), POEMS syndrome / osteosclerotic myeloma, types I and II cryoglobulinemia, mild linked deposition disease, Goodpastures syndrome, idiopathic thrombocytopenic purpura (ITP), acute glomerulonephritis, pemphigus and pemphigoid disorders and epidermolysis bullosa acquisita, non-Hodgkin's lymphomas and leukemias with BCMA expression, or diseases in which the patient produces neutralizing antibodies to recombinant protein replacement therapy, wherein said method comprises the step of administering a pharmaceutical composition comprising an antigen binding protein according to the invention in combination with a pharma- ceutical acceptable carrier.

[0132] In a further embodiment, a method of treating a human patient suffering from multiple myeloma (MM) is provided.

[0133] BCMA antibodies are useful for any therapy in which targeting of BCMA is desired, such as adoptive cell transfer (ACT), bispecific T cell engagers (BiTEs), and nanoparticles. In one embodiment, the present disclosure provides a chimeric antigen receptor (CAR) comprising an antigen binding domain of a BCMA monoclonal antibody described herein linked to a T cell activation moiety. A "chimeric antigen receptor (CAR)" is an artificially constructed hybrid protein or polypeptide that comprises an antigen binding domain of an antibody (e.g., a single chain variable fragment (scFv)) linked to a T cell signaling moiety or a T cell activation moiety. CAR structures have evolved over the past two decades, most commonly incorporating single-chain variable fragments (scFv) derived from monoclonal antibodies (mAbs) and signaling motifs from TCR chains (referred to as "first generation" CARs (see, e.g., Okur, FV, Brenner, MK, Methods Mol. Biol., 651: 319-45 (2010); and Lee et al., Clin. Cancer. Res., 18(10): 2780-2790 (2012))).Recently, second and third generation CARs have been developed that incorporate one ("second generation") or two ("third generation") costimulatory activation motifs from, for example, CD28, 4-1BB (CD137), and / or CD134 (OX-40) that enhance proliferation, cytotoxicity, and persistence in vivo (e.g., Finney et al., J. Immunol., 172: 104-13 (2004); Altvater et al, Clin Exp Immunol. 144(3): 447-57 (2006); Chu et al., J Transl Med. 20(1): 240 (2022); Maher et al., Nat Biotechnol., 20:70-5 (2002); Milone et al., Mol Ther., 17: 1453-64 (2009); See SafarzadehKozani et al, Biomark res. 10(1): 24(2022); Xu et al, Blood Sci. 1(2): 156-160 (2019) and Qian et al., Front Immunol. 13: 841425 (2022)).

[0134] The antigen-binding domain of the CAR may comprise a whole monoclonal antibody or a monoclonal antibody fragment, as described herein. In one embodiment, the antigen-binding domain of the CAR may comprise a single chain Fv (scFv) fragment of an anti-BCMA monoclonal antibody. Methods for generating chimeric antigen receptors and CARs are further described, for example, in Ohmine K, and Uchibori R. Int J Hematol. 115(6): 799-810 (2022); Ding L, et al, Stem Cell Investig. 8:1 (2021); Riviere, I. and M. Sadelain, Mol. Ther., 25(5): 1117-1124 (2017); Chan LY et al., Biomedicines. 10(4): 804 (2022); Davila, ML and M. Sadelain, Int. J. Hematol., 104(1): 6-17 (2016); and Mohty et al, Leukemia. 33(12): 2767-2778 (2019).

[0135] The term "antibody-drug conjugate (ADC)" refers to a compound comprising a monoclonal antibody (mAb) linked via a chemical linker to a cytotoxic agent (typically a small molecule drug with high systemic toxicity). An ADC can be represented by the formula: [ka]

[0136] In the formula, D, D 1 , and D 2 L, L are small molecule cytotoxins or functional small molecules commonly referred to as payloads. 1 , and L 2is a conjugate. In some embodiments, the ADC may include a small molecule cytotoxin that has been chemically modified to include a conjugate, or the conjugate is part of a payload called a traceless conjugate. Conjugates are generally used to attach cytotoxins to antibodies or antigen-binding fragments thereof. Upon binding to a target antigen on the cell surface, the ADC is internalized and transported to lysosomes, where the cytotoxin is released either by proteolysis of the cleavable conjugate (e.g., by cathepsin B found in lysosomes) or by proteolysis of the antibody if it is attached to the cytotoxin via a non-cleavable linkage. The cytotoxin then travels from the lysosome to the cytosol or nucleus, where it binds to the target depending on the mechanism of action.

[0137] The antibody-drug conjugate may comprise a whole antibody or an antibody fragment. A whole antibody is usually composed of four polypeptides: two identical copies of a heavy (H) chain polypeptide and two identical copies of a light (L) chain polypeptide. Each heavy chain contains one N-terminal variable (VH) region and three C-terminal constant (CH1, CH2, CH3) regions, and each light chain contains one N-terminal variable (VL) region and one C-terminal constant (CL). The variable regions of each pair of light and heavy chains form the antigen-binding site of the antibody. The VH and VL regions have the same general structure, and each region contains four framework regions, whose sequences are relatively conserved. The framework regions are connected by three complementarity determining regions (CDRs). The three CDRs, known as CDR1, CDR2, and CDR3, form the "hypervariable region" of the antibody that is responsible for antigen binding.

[0138] ADCs may include antigen-binding fragments of antibodies. The terms "antibody fragment," "antigen-binding fragment," "functional fragment of an antibody," and "antigen-binding portion" are used interchangeably herein and refer to one or more fragments or portions of an antibody that retain the ability to specifically bind to an antigen. An antibody fragment may include, for example, one or more CDRs, a variable region (or a portion thereof), a constant region (or a portion thereof), or a combination thereof. Examples of antibody fragments include, but are not limited to: (i) a Fab fragment, which is a monovalent fragment consisting of the VL, VH, CL, and CH1 domains; (ii) an F(ab')2 fragment, which is a bivalent fragment containing two Fab fragments linked by a disulfide bridge at the hinge region; (iii) an Fv fragment consisting of the VL and VH domains of a single arm of an antibody; and (iv) a single chain Fv (scFv). These are monovalent molecules consisting of two domains of an Fv fragment (i.e., VL and VH) linked by a synthetic linker that allows the two domains to be synthesized as a single polypeptide chain (see, e.g., Kabat EA, Wu TT., J Immunol. 1991, 147(5): 1709-19); and (v) diabodies, which are dimers of polypeptide chains, each of which is connected to a VL by a peptide linker that is too short to allow pairing between the VH and VL on the same polypeptide chain, promoting pairing between complementary domains on different VH-VL polypeptide chains to generate a dimeric molecule with two functional antigen-binding sites (see, e.g., Hudson P. J, Kortt A. A, J Immunol Methods. 1999, 231(1-2): 177-89; Holliger P, Winter G. Cancer Immunol Immunother. 1997, 231(1-2): 177-89). 45(3-4):128-30).

[0139] In one embodiment, the antibody-drug conjugate described herein comprises a monoclonal antibody or antigen-binding fragment thereof against B-cell maturation antigen (BCMA, also known as CD269). The monoclonal antibody or antigen-binding fragment thereof may comprise (a) a heavy chain variable region comprising a complementarity determining region 1 (HCDR1) amino acid sequence of SEQ ID NO: 1, a HCDR2 amino acid sequence of SEQ ID NO: 2, and a HCDR3 amino acid sequence of SEQ ID NO: 3, and (b) a light chain variable region comprising a complementarity determining region 1 (LCDR1) amino acid sequence of SEQ ID NO: 4, a LCDR2 amino acid sequence of SEQ ID NO: 5, and a LCDR3 amino acid sequence of SEQ ID NO: 6. In another embodiment, the monoclonal antibody comprises a heavy chain variable region comprising an amino acid sequence of SEQ ID NO: 8 and / or a light chain variable region comprising an amino acid sequence of SEQ ID NO: 9.

[0140] The monoclonal antibody or antigen-binding fragment thereof against BCMA may comprise any suitable binding affinity for BCMA or an epitope thereof. The term "affinity" refers to the equilibrium constant for the reversible binding of two agents and is expressed as the dissociation constant (K D The affinity of an antibody or antigen-binding fragment thereof for an antigen or epitope of interest can be measured using any method known in the art. Such methods include, for example, fluorescence activated cell sorting (FACS), surface plasmon resonance (e.g., Biacore TM , ProteOn TM ), biolayer interferometry (BLI, e.g., Octet), kinetic exclusion assays (e.g., KinExA TM ), separable beads (e.g., magnetic beads), antigen panning, and / or ELISA (see, e.g., JR Crowther, Methods Mol Biol. 2000, 149:III-IV, 1-413). It is known in the art that the binding affinity of a particular antibody varies depending on the method used to analyze the binding affinity.

[0141] The affinity of the binding agent for the ligand, such as the affinity of an antibody for an epitope, can be, for example, from about 1 picomolar (pM) to about 1 micromolar (1 μM) (e.g., about 1 picomolar (pM) to about 1 nanomolar (nM), or about 1 nM to about 1 micromolar (μM)). In one embodiment, the monoclonal antibody or antigen-binding fragment thereof can bind BCMA with a Kd of 100 nanomolar or less (e.g., 100 nM, about 90 nM, about 80 nM, about 70 nM, about 60 nM, about 50 nM, about 40 nM, about 30 nM, about 20 nM, or about 10 nM, or a range defined by any two of the foregoing values).

[0142] In another embodiment, the monoclonal antibody may bind to BCMA with a Kd of 10 nanomolar or less (e.g., about 9 nM, about 8 nM, about 7 nM, about 6 nM, about 5 nM, about 4 nM, about 3 nM, about 2 nM, about 1 nM, about 0.9 nM, about 0.8 nM, about 0.7 nM, about 0.6 nM, about 0.5 nM, about 0.4 nM, about 0.3 nM, about 0.2 nM, about 0.1 nM, about 0.05 nM, about 0.02 nM, about 0.01 nM, about 0.001 nM, or a range defined by any two of the foregoing values).

[0143] In another embodiment, the monoclonal antibody may bind to BCMA with a Kd of 200 pM or less (e.g., about 190 pM, about 175 pM, about 150 pM, about 125 pM, about 110 pM, about 100 pM, about 90 pM, about 80 pM, about 70 pM, about 60 pM, about 50 pM, about 40 pM, about 30 pM, about 25 pM, about 20 pM, about 15 pM, about 10 pM, about 5 pM, about 1 pM, or a range defined by any two of the foregoing values).

[0144] In one embodiment, the affinity of the BCMA antibody, or antigen-binding fragment thereof, for monomeric BCMA as measured by surface plasmon resonance (SPR) is about 90 nM, about 80 nM, about 70 nM, about 60 nM, about 50 nM, about 40 nM, about 30 nM, or a range defined by any two of the foregoing values, e.g., about 50 nM to about 70 nM, about 55 nM to about 65 nM, or about 58 nM to about 62 nM.

[0145] In one embodiment, the affinity of the BCMA antibody, or antigen-binding fragment thereof, for membrane bound BCMA as measured by FACS is 10 nanomolar or less (e.g., about 9 nM, about 8 nM, about 7 nM, about 6 nM, about 5 nM, about 4 nM, about 3 nM, about 2 nM, about 1 nM, about 0.9 nM, about 0.8 nM, about 0.7 nM, about 0.6 nM, about 0.5 nM, about 0.4 nM, about 0.3 nM, about 0.2 nM, about 0.1 nM, about 0.05 nM, about 0.02 nM, about 0.01 nM, about 0.001 nM, or a range defined by any two of the foregoing values).

[0146] The antigen-binding portion or fragment of a monoclonal antibody can be of any size, so long as the portion binds to BCMA. In this regard, an antigen-binding portion or fragment of a monoclonal antibody against BCMA (also referred to herein as an "anti-BCMA monoclonal antibody") desirably comprises about 5-25 amino acids (e.g., about 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 35, or a range defined by any two of the foregoing values).

[0147] In one embodiment, the antibody-drug conjugate comprises a variable region of an anti-BCMA monoclonal antibody. In this regard, the ADC may comprise a light chain variable region, a heavy chain variable region, or both a light chain variable region and a heavy chain variable region of an anti-BCMA monoclonal antibody. Preferably, the ADC comprises a light chain variable region and a heavy chain variable region of an anti-BCMA monoclonal antibody. In one embodiment, the monoclonal antibody of the ADC described herein comprises (a) a heavy chain variable region comprising the complementarity determining region 1 (HCDR1) amino acid sequence TSFLIHW (SEQ ID NO: 1), the HCDR2 amino acid sequence FIIPGNGGTKYNQKFQ (SEQ ID NO: 2), and the HCDR3 amino acid sequence YDGSFEGYFDV (SEQ ID NO: 3), and (b) a light chain variable region comprising the complementarity determining region 1 (LCDR1) amino acid sequence SSQSLVHSDGNTYLH (SEQ ID NO: 4), the LCDR2 amino acid sequence KVSNRDS (SEQ ID NO: 5), and the LCDR3 amino acid sequence SQSTHWPWT (SEQ ID NO: 6). In another embodiment, the monoclonal antibodies of the ADCs described herein may comprise a heavy chain variable region comprising the amino acid sequence of SEQ ID NO:8 and / or a light chain variable region comprising the amino acid sequence of SEQ ID NO:9.

[0148] The BCMA monoclonal antibody or antigen-binding fragment thereof may be conjugated to a cytotoxin using any suitable method known in the art, including site-specific or non-site-specific conjugation methods. Conventional antibody conjugation strategies typically rely on randomly (i.e., non-specifically) conjugating a payload to an antibody, antigen-binding fragment thereof, via lysine or cysteine. Thus, in some embodiments, an antibody or antigen-binding fragment thereof is randomly conjugated to a cytotoxic agent, for example, by partial reduction of the antibody or antibody fragment, followed by reaction with a desired agent, with or without a linking moiety. For example, the antibody or antigen-binding fragment thereof may be reduced using dithiothreitol (DTT), TCEP, thiolethenol, or a similar reducing agent. The cytotoxic agent, with or without a linking moiety, may then be added in molar excess to the reduced antibody or antibody fragment in the presence of dimethylsulfoxide (DMSO) or DMA. After conjugation, an excess of free cysteine ​​may be added to quench unreacted agent. The cytotoxic agent, with or without a linking moiety bearing an amino-reactive group, a phenol-reactive group, or other reactive group (e.g., NHS, PFP), can be added directly to the antibody or antibody fragment in molar excess in the presence of DMSO or DMA to form the conjugate. The reaction mixture can then be purified by chromatography or buffer exchange into phosphate buffered saline (PBS).

[0149] The terms "cytotoxin" and "cytotoxic agent" refer to any molecule that inhibits or prevents the function of a cell and / or causes destruction of a cell (cell death) and / or exerts an anti-proliferative effect. The cytotoxin or cytotoxic agent of an ADC is also referred to in the art as the "payload" of the ADC. Many classes of cytotoxic agents are known in the art to have potential utility in ADC molecules and can be used in the ADCs described herein. Such classes of cytotoxic agents include, for example, anti-microtubule agents (e.g., tubulysins, auristatins, and maytansinoids), DNA minor groove binders (e.g., pyrrolobenzodiazepines (PBDs) or indolinobenzodiazepines (IGNs)), RNA polymerase II inhibitors (e.g., amatoxins), DNA topoisomerase I inhibitors (e.g., camptothecins), and DNA alkylating agents (e.g., duocarmycins, CC-1065, pyrrolobenzodiazepine or indolinobenzodiazepine pseudodimers). Particular cytotoxic agents that may be used in the ADCs described herein include, but are not limited to, for example, tubulysins, amanitins, auristatins, calicheamicins, camptothecins, daunomycins, doxorubicins, duocarmycins, dolastatins, enediynes, lexitropsins, taxanes, puromycins, maytansinoids, vinca alkaloids, and pyrrolobenzodiazepines (PBDs). More specifically, the cytotoxic agent may be, for example, tubulysin, auristatin (AFP, MMAF, MMAE, AEB, AEVB, E), paclitaxel, docetaxel, CC-1065 (ducarmycin, DC1, DC4, CBI dimer), camptothecin (SN-38, topotecan), morpholino-doxorubicin, rhizoxin, cyanomorpholino-doxorubicin, dolastatin-10, echinomycin, combretastatin, calicheamicin, maytansine (DM1, DM4, DM21), vinblastine, methotrexate, netropsin, or a derivative or analog thereof. Cytotoxins suitable for use in ADCs are also described, for example, in International Patent Application Publication No. PCT / CN2021 / 128453.

[0150] Generally, a chemotherapeutic agent or functional compound may also be conjugated to the BCMA antibody of the invention. The chemotherapeutic agent or functional compound is selected from the group consisting of:

[0151] (1) a chemotherapeutic agent selected from the group consisting of: a) alkylating agents selected from the group consisting of: nitrogen mustards: chlorambucil, chlornaphazine, cyclophosphamide, dacarbazine, estramustine, ifosfamide, mechlorethamine, mechlorethamine oxide hydrochloride, mannomustine, mitobronitol, melphalan, mitolactol, pipobroman, nobembitine, phenesterine, prednimustine, thiotepa, trofosfamide, uracil mustard; CC-1065 and synthetic analogs of adozelesin, carzelesin and bizelesin; duocarmycin and its synthetic analogs KW-2189 and CBI-TMI; benzodiazepine dimers or pyrrolobenzodiazepine (PBD) dimers, tomaymycin dimers, indolinobenzodiazepine dimers. nitrosourea compounds including carmustine, lomustine, chlorozotocin, fotemustine, nimustine, ranimustine; alkylsulfonates including busulfan, treosulfan, improsulfan, and piposulfan; triazenes or dacarbazines; platinum-containing compounds including carboplatin, cisplatin, and oxaliplatin; aziridines, benzodopa, carboquone, meturedopa, and uredopa; ethylenimines and methylamelanamines including altretamine, triethylenemelamine, triethylenephosphoramide, triethylenethiophosphoramine, and trimethylolomelamine; b) plant alkaloids selected from the group consisting of: vinca alkaloids including vincristine, vinblastine, vindesine, vinorelbine, navelbine; taxoids including paclitaxel and docetaxel and analogs thereof; maytansinoids including DM1, DM2, DM3, DM4, DM5, DM6, DM7, maytansine, ansamitocins and analogs thereof; cryptophycins including the cryptophycin 1 and cryptophycin 8 groups; epothilones, eleutherobin, discodermolide, bryostatins, dolostatins, auristatins, tubulysins, cephalostatins; pancratistatin; sarcodictiin; spongistatin; c) DNA topoisomerase inhibitors selected from: epipodophyllins, including 9-aminocamptothecin, camptothecin, crisnatol, daunomycin, etoposide, etoposide phosphate, irinotecan, mitoxantrone, novantrone, retinoic acid (retinols), teniposide, topotecan, 9-nitrocamptothecin, or RFS 2000; and mitomycins, and their analogs; d) Antimetabolites selected from: {[Antifolates: (DHFR inhibitors: including methotrexate, trimetrexate, denopterin, pteropterin, aminopterin (4-aminopteroic acid), or other folate analogs); IMP dehydrogenase inhibitors (including mycophenolic acid, tiazofurin, ribavirin, EICAR); Ribonucleotide reductase inhibitors (including hydroxyurea, deferoxamine)]; [Pyrimidine analogs : uracil analogues (including ancitabine, azacitidine, 6-azauridine, capecitabine, carmofur, cytarabine, dideoxyuridine, doxifluridine, enocitabine, 5-fluorouracil, floxuridine, raltitrexed); cytosine analogues (including cytarabine, cytosine arabinoside, fludarabine); purine analogues (including azathioprine, fludarabine, mercaptopurine, thiamiprine, thioguanine); folic acid supplements such as folinic acid}, and nicotinamide phosphate ribonuclease (NAMPT) inhibitors; e) Hormonal therapy agents selected from the following: Receptor antagonists: [Antiestrogen: (including megestrol, raloxifene, tamoxifen); LHRH agonists: (including goserelin, leuprolide acetate); Antiandrogens: (bicalutamide, flutamide, calsterone, dromostanolone propionate, epithiostanol, goserelin, leuprolide, mepitiostane, nilutamide, testolactone, trilostane, and other similar antineoplastic agents] drogen inhibitors.)];Retinoids / deltoids:[Vitamin D3 analogs:(including CB1093, EB1089, KH1060, cholecalciferol, ergocalciferol);Photodynamic therapy agents:(including verteporfin, phthalocyanines, photosensitizer Pc4, demethoxyhypocrelin A);Cytokines:(including interferon alpha, interferon gamma, tumor necrosis factor (TNF), TNF domain-containing human proteins)]}; f) kinase inhibitors selected from the following: BIBW2992 (anti-EGFR / Erb2), imatinib, gefitinib, pegaptanib, sorafenib, dasatinib, sunitinib, erlotinib, nilotinib, lapatinib, axitinib, pazopanib, vandetanib, E7080 (anti-VEGFR2), mubritinib, ponatinib, bafetinib, bosutinib, cabozantinib, vismodegib, iniparib, ruxolitinib, CYT387, axitinib, tivozanib, sorafenib, bevacizumab, cetuximab, trastuzumab, ranibizumab, panitumumab, ispinesib; g) a poly(ADP-ribose) polymerase (PARP) inhibitor selected from the group consisting of olaparib, niraparib, iniparib, talazoparib, veliparib, CEP9722 (Cephalon), E7016 (Eisai), BGB-290 (Beigene), or 3-aminobenzamide; h) enediyne antibiotics (selected from the calicheamicins, calicheamicin gamma 1, delta 1, alpha 1, or beta 1; dynemicins including dynemicin A and deoxydynemicin; esperamicin, kedarcidin, C-1027, maduropeptin, or neocarzinostatin chromophores and related chromoprotein enediyne antibiotic chromophores), aclacinomycins, actinomycin, anthramycin, azaserine, bleomycins, cactinomycin, carabicin, carminomycin, carzinophilin; chromomycins, dactinomycin, daunomycin, Antibiotics selected from rubicin, detorubicin, 6-diazo-5-oxo-L-norleucine, doxorubicin, morpholino-doxorubicin, cyanomorpholino-doxorubicin, 2-pyrrolino-doxorubicin and deoxydoxorubicin, epirubicin, eribulin, esorubicin, idarubicin, marcelomycin, nitomycins, mycophenolic acid, nogalamycin, olivomycins, peplomycin, potofilomycin, puromycin, queramycin, rodorubicin, streptonigrin, streptozocin, tubercidin, ubenimex, zinostatin, zorubicin; i) Polyketides (acetogenins), bullatacin and bullatacinone; gemcitabine, epoxomicins and carfilzomib, bortezomib, thalidomide, lenalidomide, pomalidomide, tosedostat, zybrestat, PLX4032, STA-9090, Stimuvax, allovectin-7, zygeva, probenge, elvoi, isoprenylation inhibitors and lovastatin, dopaminergic neurotoxins and 1-methyl-4-phenyl Pyridine ion, cell cycle inhibitors (including staurosporine), actinomycins (including actinomycin D and dactinomycin), amanitins, bleomycins (including bleomycin A2, bleomycin B2, and peplomycin), anthracyclines (including daunorubicin, doxorubicin (adriamycin), idarubicin, epirubicin, pirarubicin, and zorubicin), mitoxantrone, MDR inhibitors or verapamil, Ca 2+ATP inhibitors or thapsigargin, histone deacetylase inhibitors (including vorinostat, romidepsin, panobinostat, valproic acid, mocetinostat (MGCD0103), belinostat, PCI-24781, entinostat, SB939, resminostat, gibinostat, AR-42, CUDC-101, sulforaphane, trichostatin A); thapsigargin, celecoxib, glitazones, epigallocatechin gallate, disulfiram, salinosporamide A, antiadrenal drugs (selected from the group consisting of aminoglutethimide, mitotane, trilostanca); aceglatone; aldophosphamide glicoside; aminolevulinic acid; amsacrine; arabinoside, bestravsil; Bisantrene;Edatrexate;Defofamine;Demecolcine;Diaziquone;Elfornitine (DFMO), Elfomitine;Elliptinium acetate;Etocuruside, Gallium nitrate, Gacytosine, Hydroxyurea;Ibandronate, Lentinan;Lonidamine;Mitoguazone;Mopidamol;Nitracrine;Pentostatin;Phenamet;Pirarubicin;Podophyllic acid;2-Ethylhydrazide;Procarbazine;PSK (registered trademark);Razoxane;Rhizoxin;Sizofiran;Spirogermanium;Tenuazonic acid;Triaziquone;2,2',2''-Trichlorotriethylamine;Tricothecenes (including T2 toxin, verrucarin A, roridin A, and anguidine);Urethanes, siRNAs, antisense drugs;

[0152] (2) Anti-autoimmune disease drugs: cyclosporine, cyclosporine A, aminocaproic acid, azathioprine, bromocriptine, chlorambucil, chloroquine, cyclophosphamide, corticoids (including amcinonide, betamethasone, budesonide, hydrocortisone, flunisolide, fluticasone propionate, fluocortolone danazol, dexamethasone, triamcinolone acetonide, and beclomethasone dipropionate), DHEA, etanercept, hydroxychloroquine, infliximab, meloxicam, methotrexate, mofetil, mycophenolate, prednisone, sirolimus, and tacrolimus;

[0153] (3) Anti-infectives, including but not limited to: a) Aminoglycosides: amikacin, astromycin, gentamicin (netilmicin, sisomicin, isepamicin), hygromycin B, kanamycin (amikacin, arbekacin, bekanamycin, dibekacin, tobramycin), neomycin (framycin, paromomycin, ribostamycin), netilmicin, spectinomycin, streptomycin, tobramycin, verdamycin; b) Amphenicols: azidamphenicol, chloramphenicol, florfenicol, thiamphenicol; c) Ansamycins: geldanamycin, herbimycin; d) Carbapenems: biapenem, doripenem, ertapenem, imipenem, cilastatin, meropenem, panipenem; e) Cephalosporins: carbacephems (loracarbef), cephacetrile, cefaclor, cephradine, cefadroxil, cephalonium, cephaloridine, cephalothin or cephalothin, cephalexin, cephaloglycin, cefamandole, cephapirin, cefatrizine, cefazaflur, cephazedone, cefazolin, cefbuperazone, cefcapene, cefdaloxime, cefepime, cefminox, cefoxitin, cefprozil, cefroxadine, ceftezole, cefuroxime, cefixime, cefdinir, cefditoren, cefepime, fetameth, cefmenoxime, cefodizime, cefonicid, cefoperazone, ceforanide, cefotaxime, cefotiam, cefozopran, cephalexin, cefpimizole, cefpiramide, cefpirome, cefpodoxime, cefprozil, cefquinome, cefsulodin, ceftazidime, cefteram, ceftibuten, ceftiolene, ceftizoxime, ceftobiprol, ceftriaxone, cefuroxime, cefuzonam, cephamycins (including cefoxitin, cefotetan, cefmetazole), oxacephems (flomoxef, latamoxef); f) Glycopeptides: bleomycin, vancomycin (including oritavancin, telavancin), teicoplanin (dalbavancin), ramoplanin; g) Glycylcyclines: Tigecycline; h) β-lactamase inhibitors: penams (sulbactam, tazobactam), clavams (clavulanic acid); i) Lincosamides: clindamycin, lincomycin; j) Lipopeptides: daptomycin, A54145, calcium-dependent antibiotics (CDA); k) Macrolides: azithromycin, cethromycin, clarithromycin, dirithromycin, erythromycin, flurithromycin, josamycin, ketolides (telithromycin, cethromycin), midecamycin, miocamycin, oleandomycin, rifamycins (rifampicin, rifampin, rifabutin, rifapentine), rokitamycin, roxithromycin, spectinomycin, spiramycin, tacrolimus (FK506), troleandomycin, telithromycin; l) Monobactams: aztreonam, tigemonam; m) Oxazolidinones: Linezolid; n) Penicillins: amoxicillin, ampicillin, pivampicillin, hetacillin, bacampicillin, methampicillin, talampicillin, azidocillin, azlocillin, benzylpenicillin, benzathine benzylpenicillin, benzathine phenoxymethylpenicillin, clometocillin, procaine benzylpenicillin, carbenicillin (carindacillin), cloxacillin, dicloxacillin, epicillin, flucloxacillin, mecillinam (pivmecillinam), mezlocillin, methicillin, nafcillin, oxacillin, penamecillin, penicillin, phenethicillin, phenoxymethylpenicillin, piperacillin, propicillin, sulbenicillin, temocillin, ticarcillin; o) Polypeptides: bacitracin, colistin, polymyxin B; p) Quinolones: alatrofloxacin, balofloxacin, ciprofloxacin, clinafloxacin, danofloxacin, difloxacin, enoxacin, enrofloxacin, floxin, garenoxacin, gatifloxacin, gemifloxacin, grepafloxacin, canotrovafloxacin, levofloxacin, lomefloxacin, marbofloxacin, moxifloxacin, nadifloxacin, norfloxacin, orbifloxacin, ofloxacin, pefloxacin, trovafloxacin, grepafloxacin, sitafloxacin, sparfloxacin, temafloxacin, tosufloxacin, trovafloxacin; q) Streptogramins: Pristinamycin, quinupristin / dalfopristin; r) Sulfonamides: mafenide, prontosil, sulfacetamide, sulfamethizole, sulfanilamide, sulfasalazine, sulfisoxazole, trimethoprim, trimethoprim-sulfamethoxazole (co-trimoxazole); s) Steroidal antibacterial agents: fusidic acid, etc.; t) Tetracyclines: doxycycline, chlortetracycline, clomocycline, demeclocycline, lymecycline, meclocycline, methacycline, minocycline, oxytetracycline, penimepicycline, rolitetracycline, tetracycline, glycylcyclines (tigecycline, etc.); u) Other antibiotics: annonacin, arsphenamine, bactoprenol inhibitors (bacitracin), DADAL / AR inhibitors (cycloserine), dictyostatin, discodermolide, eleutherobin, epothilone, ethambutol, etoposide, faropenem, fusidic acid, furazolidone, isoniazid, laulimalide, metronidazole, mupirocin, mycolactone, NAM synthesis inhibitors (fosfomycin), nitrofurantoin, paclitaxel, platensimycin, pyrazinamide, quinupristin / dalfopristin, rifampicin (rifampin), tazobactam tinidazole, uvaricin;

[0154] (4)Antiviral drugs: a) Entry / fusion inhibitors: aplaviroc, maraviroc, vicriviroc, gp41 (enfuvirtide), PRO140, CD4 (ibalizumab); b) Integrase inhibitors: Raltegravir, Elvitegravir, Globoidonan A; c) Maturation inhibitors: Bevirimat, Vivecon; d) Neuraminidase inhibitors: oseltamivir, zanamivir, peramivir; e) Nucleosides and Nucleotides: Abacavir, Acyclovir, Adefovir, Amdoxovir, Apricitabine, Brivudine, Cidofovir, Clevudine, Dexerbucitabine, Didanosine (DDI), Elvucitabine, Emtricitabine (FTC), Entecavir, Famciclovir, Fluorouracil (5-FU), 3'-Fluoro substituted 2',3'-deoxynucleoside analogues (e.g., 3'-Fluoro-2',3'-dideoxythymidine (FLT) and 3'-Fluoro-2',3'-deoxythymidine (FLT)). dideoxyguanosine (FLG), fomivirsen, ganciclovir, idoxuridine, lamivudine (3TC), L-nucleosides (e.g., β-L-thymidine and β-L-2'-deoxycytidine), penciclovir, rasvir, ribavirin, stampidine, stavudine set (d4T), taribavirin (viramidine), telbivudine, tenofovir, trifluridine, valacyclovir, valganciclovir, zalcitabine (ddC), zidovudine (AZT); f) Non-nucleosides: amantadine, ateviridine, capravirine, diarylpyrimidines (etravirine, rilpivirine), delavirdine, docosanol, emivirine, efavirenz, foscarnet (phosphorylformate), imiquimod, interferon alpha, loviride, rhodnosine, methisazone, nevirapine, NOV-205, peginterferon alpha, podophyllotoxin, rifampicin, rimantadine, resiquimod (R-848), tromantadine; g) Protease inhibitors: amprenavir, atazanavir, boceprevir, darunavir, fosamprenavir, indinavir, lopinavir, nelfinavir, pleconaril, ritonavir, saquinavir, telaprevir (VX-950), tipranavir; h) Other antiviral drugs: Abzyme, Arbidol, Calanolide A, Seragenin, Cyanovirin-N, Diarylpyrimidines, Epigallocatechin gallate (EGCG), Foscarnet, Griffithsin, Taribavirin (Viramidine), Hydroxyurea, KP-1461, Miltefosine, Pleconaril, Portmanteau inhibitors, Ribavirin, Seliciclib;

[0155] (5) 3 H, 11 C. 14 C. 18 F, 32 P, 35 S, 64 Cu, 68 Ga, 86 Y, 99 Tc, 111 In, 123 I, 124 I, 125 I, 131 I, 133 Xe, 177 Lu, 211 At, or 213 The radioisotope may be selected from the group consisting of Bi (radionuclide).

[0156] (6) Chromophore molecules capable of absorbing UV light, fluorescent light, IR light, near-IR light, or visible light; xanthophores, erythrophores, iridophores, leucophores, melanophores, and cerebrophores; a class or subclass of fluorescent molecules that are fluorescent chemicals that re-emit light with light; a class or subclass of visual light-sensitive molecules; a class or subclass of luminescent molecules; a class or subclass of luminescent molecules; and a class or subclass of luciferin compounds; a non-protein organic fluorophore selected from the following: xanthene derivatives (fluorescein, rhodamine, Oregon Green, eosin ... cyanine derivatives (including cyanine, indocarbocyanine, oxacarbocyanine, thiacarbocyanine, and merocyanine); squalene derivatives and ring-substituted squaraines (including Seta, SeTau, and Square dyes); naphthalene derivatives (including dansyl and prodan derivatives); coumarin derivatives; oxadiazole derivatives (including pyridyloxazole, nitrobenzoxadiazole, and benzoxadiazole); anthracene derivatives (including anthraquinones including DRAQ5, DRAQ7, and CyTRAK Orange); pyrene derivatives (cascade blue); oxazine derivatives (including Nile red, Nile blue, cresyl violet, oxazine 170); acridine derivatives (including proflavine, acridine orange, acridine yellow); arylmethine derivatives (including auramine, crystal violet, malachite green). Tetrapyrrole derivatives (including porphines, phthalocyanines, bilirubin); fluorophore analogs and derivatives including: CF dyes, DRAQ and CyTRAK probes, BODIPY, Alexa Fluor, DyLight Fluor, Atto and Trancy, FluoProbes, Abberior dyes, DY and MegaStokes dyes, SulfoCy dyes, HiLyte Fluor, Seta, SeTau and Square dyes, Quasar and Cal Flour dyes, SureLight dyes (APC, RPEPerCP, phycobilisomes), APC, APCXL, RPE, BPE, allophycocyanin (APC), aminocoumarins, APC-Cy7 conjugates, BODIPY-FL, Cascade Blue, Cy2, Cy3, Cy3.5, Cy3B, Cy5, Cy5.5, Cy7, Fluorescein, FluorX, Hydroxycoumarin, Lissamine Rhodamine B, Lucifer Yellow, Methoxycoumarin, NBD, Pacific Blue, Pacific Orange, PE-Cy5 conjugate, PE-Cy7 conjugate, PerCP, R-Phycoerythrin (PE), Red613, Seta-555-Azide, Seta-555-DBCO, Seta-555-NHS, Seta-580-NHS, Seta-680-NHS, Seta-780-NHS, Seta-APC-780, Seta-PerCP-680, Seta-R-PE-670, SeTau380-NHS, SeTau405-Maleimide, SeTau405-NHS, SeTau425-NHS, SeTau647-NHS, Texas Red, TRITC, TruRed, X-Rhodamine, 7-AAD (7-aminoactinomycin D, CG selective), Acridine Orange, Chromomycin A3, CyTRAK Orange (Red color excitation dark), DAPI, DRAQ5, DRAQ7, ethidium bromide, Hoechst 33258, Hoechst 33342, LDS751, mithramycin, propidium iodide (PI), SYTOX blue, SYTOX green, SYTOX orange, thiazole orange, TO-PRO: cyanine monomer, TOTO-1, TO-PRO-1, TOTO-3, TO-PRO-3, YOseta-1, YOYO-1; fluorescent dyes including the following compounds or their derivatives: DCFH (2'7' dichlorodihydro-fluorescein, oxidized form), DHR (dihydrorhodamine 123, oxidized form, photocatalytic oxidation), Fluo-3 (AM ester, pH>6), Fluo-4 (AM ester, pH7.2), Indo-1 (AM ester, low / high calcium (Ca. 2+)), SNARF (pH6 / 9), Allophycocyanin (APC), AmCyan1 (tetramer, Clontech), AsRed2 (tetramer, Clontech), Azami Green (monomer), Azurite, B-Phycoerythrin (BPE), Cerulean, CyPet, DsRed monomer (Clontech), DsRed2 ("RFP"), EBFP, EBFP2, ECFP, EGFP (weak dimer), Emerald (weak dimer), EYFP (weak dimer), GFP (S65A mutant), GFP (S65C mutant), GFP (S65L mutant), GFP (S65T mutant), GFP (Y66F mutant), GFP (Y66H mutant), GFP (Y66W mutant), GFP uv , HcRed1, J-Red, Katyusha, Kusabira Orange (monomer, MBL), mCFP, mCherry, mCitrine, Midriishi Cyan (dimer, MBL), mKate (TagFP635, monomer), mKeima-Red (monomer), mKO, mOrange, mPlum, mRaspberry, mRFP1 (monomer), mStrawberry, mTFP1, mTurquoise2, P3 (phycobilisome complex), Peridinin Chlorophyll (PerCP), R-Phycoerythrin (RPE), T-Sapphire, TagCFP (dimer), TagGFP (dimer), TagRFP (dimer), TagYFP (dimer), tdTomato (tandem dimer), Topaz, TurboFP602 (dimer), TurboFP635 (dimer), TurboGFP (dimer), TurboRFP (dimer), TurboYFP635 (dimer), Venus, native GFP, YPet, Zs Green 1 (tetramer), Zs Yellow 1 (tetramer), and their derivatives;

[0157] (7) Cell receptor agonists or cell stimulating molecules that can be selected from the following: folic acid derivatives; glutamic acid urea derivatives; somatostatin and its analogs (selected from the group consisting of octreotide (sandostatin) and lanreotide (somatilin); aromatic sulfonamides; pituitary adenylate cyclase activating peptide (PACAP) (PAC1); vasoactive intestinal peptide (VIP / PACAP) (VPAC1, VPAC2); melanocyte stimulating hormone (α-MSH); cholecystokinin (CCK) / gastrin receptor agonists, bombesin (Pyr-Gln-Arg-Leu-Gly-Asn-Gln-Trp-Ala-Val-Gly-His-Leu-Met-NH 2 neurotensin receptor ligands (NTR1, NTR2, NTR3), substance P (NK1 receptor) ligands; neuropeptide Y (Y1-Y6); homing peptides including RGD (Arg-Gly-Asp), NGR (Asn-Gly-Arg), dimeric and multimeric cyclic RGD peptides (cRGDfV), TAASGVRSMH, and LTLRWVGLMS (chondroitin sulfate proteoglycan NG2 receptor ligand), and F3 peptide; cell membrane permeable peptides. peptide hormones, luteinizing hormone releasing hormone (LHRH) agonists and antagonists, and gonadotropin releasing hormone (GnRH) agonists, which act by targeting follicle stimulating hormone (FSH) and luteinizing hormone (LH) as well as testosterone production, including buserelin (Pyr-His-Trp-Ser-Tyr-D-Ser(OtBu)-Leu-Arg-Pro-NHEt), gonadorelin (Pyr-His-Trp-Ser-Tyr-Gly-Leu-Arg-Pro-Gly-NH 2 ), goserelin (Pyr-His-Trp-Ser-)Tyr-D-Ser(OtBu)-Leu-Arg-Pro-AzGly-NH 2), histrelin (Pyr-His-Trp-Ser-Tyr-D-His(N-benzyl)-Leu-Arg-Pro-NHEt), leuprolide (Pyr-His-Trp-Ser-Tyr-D-Leu-Leu-Arg-Pro-NHEt), nafarelin (Pyr-His-Trp-Ser-Tyr-2Nal-Leu-Arg-Pro-Gly-NH 2 ), Triptorelin (Pyr-His-Trp-Ser-Tyr-D-Trp-Leu-Arg-Pro-Gly-NH 2 ), nafarelin, deslorelin, abarelix (Ac-D-2Nal-D-4-chloroPhe-D-3-(3-pyridyl)Ala-Ser-(N-Me)Tyr-D-Asn-Leu-isopropylLys-Pro-DAla-NH 2 ), Cetrorelix (Ac-D-2Nal-D-4-chloroPhe-D-3-(3-pyridyl)Ala-Ser-Tyr-D-Cit-Leu-Arg-Pro-D-Ala-NH 2 ), degarelix (Ac-D-2Nal-D-4-chloroPhe-D-3-(3-pyridyl)Ala-Ser-4-aminoPhe(L-hydroorotyl)-D-4-aminoPhe(carbamoyl)-Leu-isopropylLys-Pro-D-Ala-NH 2 ), and ganirelix (Ac-D-2Nal-D-4-chloroPhe-D-3-(3-pyridyl)Ala-Ser-Tyr-D-(N9,N10-diethyl)-homoArg-Leu-(N9,N10-diethyl)-homoArg-Pro-D-Ala-NH 2 ); pattern recognition receptors (PRRs) selected from Toll-like receptor (TLR) ligands, C-type lectins, and Nodlike receptor (NLR) ligands; calcitonin receptor agonists; integrin receptors and their receptor subtypes (α V β 1 , α V β 3 , α V β 5 , α V β 6 , α 6 β 4 , α 7 β1 , α L β 2 , α IIb β 3 agonists (selected from the group consisting of GRGDSPK, cyclo(RGDfV)(L1) and its derivatives [cyclo(-N(Me)R-GDfV), cyclo(R-Sar-DfV), cyclo(RG-N(Me)D-fV), cyclo(RGD-N(Me)fV), cyclo(RGDf-N(Me)V-) (silengitide)]; anticalins (derivatives of lipocalin); adnectins (tenth FN3 (fibronectin)); designed ankyrin repeat proteins (DARPins); avimers; EGF receptor or VEGF receptor agonists;

[0158] (8) A pharma- ceutically acceptable salt, acid, derivative, hydrate or hydrated salt of any of the above drugs; or a crystal structure, or an optical isomer, racemate, diastereomer or enantiomer of any of the above drugs.

[0159] In another embodiment, drug D can be a polyalkylene glycol used to extend the half-life of a cell-binding antibody or antibody-like protein molecule when administered to a mammal. Polyalkylene glycols include, but are not limited to, polyalkylene glycols selected from poly(ethylene glycol) (PEG), poly(propylene glycol), and copolymers of ethylene oxide and propylene oxide; particularly preferred are PEGs, and more particularly preferred are monofunctionally activated hydroxyPEGs (e.g., hydroxyPEGs activated at one end, including hydroxyPEG-monocarboxylic acids, hydroxyPEG-monoaldehydes, hydroxyPEG-monoamines, hydroxyPEG-monohydrazides, hydroxyPEG-monocarbazides, hydroxyPEG-monoiodoacetamides, hydroxyPEG-monomaleimides, hydroxyPEG-orthopyridine disulfides, hydroxyPEG-monoximes, hydroxyPEG-monophenylcarbonates, hydroxyPEG-monophenylglyoxals, hydroxyPEG-monothiazolidine-2-thiones, hydroxyPEG-monothioesters, hydroxyPEG-monothiols, hydroxyPEG-monotriazines, and hydroxyPEG-monovinylsulfones).

[0160] In certain such embodiments, the polyalkylene glycol has a molecular weight of about 10 Daltons to about 200 kDa, preferably about 88 Da to about 40 kDa; two branches each having a molecular weight of about 88 Da to about 40 kDa; and more preferably, two branches each having a molecular weight of about 88 Da to about 20 kDa. In a particular embodiment, the polyalkylene glycol is poly(ethylene) glycol and has a molecular weight of about 10 kDa; about 20 kDa, or about 40 kDa. In certain embodiments, the PEG is PEG 10 kDa (linear or branched), PEG 20 kDa (linear or branched), or PEG 40 kDa (linear or branched). Many U.S. patents, such as U.S. Patent Nos. 5,428,128; 5,621,039; 5,622,986; 5,643,575; 5,728,560; 5,730,990; 5,738,846; 5,811,076; 5,824,701; 5,840,900; 5,880,131; 5,900,402; 5,902,588; 5,919,455; 5,951,974; 5,965,119; 5,965,566; 5,969,040; 5,981,709; 6,011,042; 6,042,822; Nos. 6,113,906; 6,127,355; 6,132,713; 6,177,087; and 6,180,095 disclose the preparation of linear or branched "non-antigenic" PEG polymers and their derivatives or conjugates.

[0161] In yet another embodiment, D is more preferably a potent cytotoxic agent, such as tubulysin and its analogs, maytansine and its analogs, taxanes and its analogs, CC-1065 and its analogs, daunorubicin or doxorubicin and its analogs, amatoxin and its analogs, benzodiazepine dimers (dimers of pyrrolobenzodiazepines (PBD), tomaymycin, anthramycin, indolinobenzodiazepines, imidazobenzothiadiazepines, or oxazolidinobenzodiazepines) and their analogs, calicheamicin, and enediyne antibiotic analogs, actinomycin and its analogs, azaserine and its analogs, bleomycin and its analogs, epirubicin and its analogs, tamoxifen and its analogs, idarubicin and its analogs, dolastatins and their analogs, auristatins (including monomethylauristatin (MMAE), MMAF, auristatin PYE, auristatin TP, auristatin 2-AQ, 6-AQ, EB (AEB), and EFP (AEFP)) and their analogs, combretastatins, duocarmycins and their analogs, and its analogues, camptothecin and its analogues, geldanamycin and its analogues, methotrexate and its analogues, thiotepa and its analogues, vindesine and its analogues, vincristine and its analogues, hemiasterin and its analogues, nazmamide and its analogues, spliceostatin, pladienolide, microginin and its analogues, radiosumin and its analogues, alterobactin and its analogues, microsclerodermin and its analogues, theonellamide and its analogues, The therapeutic agent is selected from esperamicin and its analogs, PNU-159682 and its analogs, protein kinase inhibitors, MEK inhibitors, KSP inhibitors, nicotinamide phosphoribosyltransferase (NAMPT) inhibitors, immunotoxins, specific cell receptor agonists, cell stimulatory or intracellular signaling molecules, one or more of DNA, RNA, mRNA, small interfering RNA (siRNA), microRNA (miRNA), and PIWI-interacting RNA (piRNA), as well as stereoisomers, equivalents, analogs, or derivatives of the above-mentioned drugs.

[0162] Tubulysin and its analogs are well known in the art and can be isolated from natural sources according to known methods or prepared synthetically according to known methods (e.g., Balasubramanian, R., et al. J. Med. Chem., 2009, 52, 238-40; Wipf, P., et al. Org. Lett., 2004, 6, 4057-60; Pando, O., et al. J. Am. Chem. Soc., 2011, 133, 7692-5; Reddy, JA, et al. Mol. Pharmaceutics, 2009, 6, 1518-25; Raghavan, B., et al. J. Med. Chem., 2008, 51, 1530-33; Patterson, AW, et al. J. Org. Chem., 2008, 73, 4362-9; Pando, O., et al. Org. Lett., 2009, 11 (24), 5567-9; Wipf, P., et al. Org. Lett., 2007, 9 (8), 1605-7; Friestad, GK, Org. Lett., 2004, 6, 3249-52; Peltier, HM et al., J. Am. Chem. Soc., 2006, 128, 16018-9; Chandrasekhar, S., et al. J. Org. Chem., 2009, 74, 9531-4; Liu, Y., et al. Mol. Pharmaceutics, 2012, 9, 168-75; Friestad, GK, et al Org. Lett., 2009, 11, 1095-8; Kubicek, K., et al., Angew Chem Int Ed Engl, 2010.49: 4809-12; Chai, Y., et al., Chem Biol, 2010, 17 : 296-309; Ullrich, A., et al.,Angew Chem Int Ed Engl, 2009, 48, 4422-5; Sani, M., et al., Angew Chem Int Ed Engl, 2007, 46, 3526-9. Domling, A., et al., Angew Chem Int Ed Engl, 2006, 45, 7235-9; Patent Applications: Zanda, M., et al., Canadian Patent Application CA 2710693 (2011); Chai, Y., et al., European Patent Application 2174947 (2010), WO 2010034724; Leamon, C., et al., WO2010033733, WO2009002993; Ellman, J., et al., PCT WO2009134279; WO 2009012958, US Applications 20110263650, 20110021568; Matschiner, G., et al., WO2009095447; Vlahov, I., et al., WO2009055562, WO2008112873; Low, P., et al., WO2009026177; Richter W., WO2008138561; Kjems, J., et al.,WO 2008125116; Davis, M., et al., WO2008076333; Diener, J., et al., U.S. Patent Application No. 20070041901, WO2006096754; Matschiner, G., et al., WO2006056464; Vaghefi, F., et al., WO2006033913; Doemling, A., German Patent Application DE102004030227, WO2004005327, WO2004005326, WO2004005269; Stanton, M., et al, US Patent No. 20040249130; Hoefle, G., et al, Ger. OffenDE10254439, DE10241152, DE10008089; Leung, D., et al. al, WO2002077036; Reichenbach, H., et al, Ger. Offen. DE19638870; Wolfgang, R., US20120129779; Chen, H. Preferred structures of tubulysin for conjugation with cell-binding molecules by the methods of the present patent application are described in patent application PCT / IB2012 / 053554.

[0163] A tubulysin analog having the following formula (IV): or a pharma- ceutically acceptable salt, hydrate, or hydrated salt; or a polymorphic crystal structure; or an optical isomer, racemate, diastereomer, or enantiomer thereof: [ka]

[0164] During the ceremony, [ka] is independent, L 1 and / or L 2 The binding site is the site that binds to the two [ka] L 1 and L 2 If it binds to both, R 1 and R 2 , or Z 2 and Z 3 is preferably a double bond.

[0165] R 1 , R 2 , R 3 , and R 4 are independent of each other, H, C 1 ~C 8 Alkyl;C 2 ~C 8 Heteroalkyl or heterocyclic ring; C 3 ~C 8Ar is aryl, Ar-alkyl, cycloalkyl, alkylcycloalkyl, heterocycloalkyl, heteroalkylcycloalkyl, carbocycle, or alkylcarbonyl; or R 1 R 2 , R 1 R 3 , R 2 R 3 , R 3 R 4 , R 5 R 6 , R 11 R 12 , or R 13 R 14 forms a 3- to 7-membered carbocyclic, cycloalkyl, heterocyclic, heterocycloalkyl, aromatic, or heteroaromatic ring system; L 1 Or L 2 When independently or simultaneously bound to R 1 and R 2 may be absent independently; Y 1 is N or CH.

[0166] In the formula, R 5 , R 6 , R 8 , R 10 , and R 11 are independently H or C 1 ~C 4 It is alkyl or heteroalkyl.

[0167] In the formula, R 7 are independent, H, R 14 , -R 14 C(=O)X 1 R 15 ; or -R 14 X 1 R 15 X 1 are O, S, SS, NH, and CH 2 , or NR 14 It is.

[0168] In the formula, R 9 are H, OH, -O-, =O, -OR 14 , -OC(=O)R 14 , -OC(=O)NHR 14-, -OC(=O)R 14 SSR 15 -, OP(=O)(OR 14 )-, -OC(=O)NR 14 R 15 ,OP(=O)(OR 14 ), OR 14 OP(=O)(OR 15 ) is selected.

[0169] R 11 are independent, H, R 14 , -R 14 C(=O)R 15 , -R 14 C(=O)X 2 R 15 Wherein, X 2 is -O-, -S-, -NH-, or -N(R 14 )-.

[0170] R 12 -COOH, -COSH, -CONH 2 , CONHNH 2 ,CONHNHR 15 , -CONH(R 15 ), -COOR 15 , -R 15 COR 16 , -R 15 COOR 16 , -R 15 C(O)NH 2 , -R 15 C(O)NHR 16 , -COSR 15 , R 15 S(=O) 2 R 16 , -R 15 P(=O)(OR 17 ) 2 , -R 15 OP(=O)(OR 17 ) 2 , -COOCH 2 OP(=O)(OR 17 ) 2 , -COX 2 SO 2 R 17 , -COOR 15 X 2R 16 , tetrazole, imidazole, or triazole; X 2 -O-, -S-, -NH-, -N(R 15 )-, -OR 15 -, -SR 15 -, CH 2 , or -NHR 15 - is;;R 12 L 1 Or L 2 When binding to R 12 -C(O)O-, -C(O)NH-, -C(=O)NHS(O) 2 R 15 - or -C(=O)N(R 15 )-.

[0171] R 13 and R 14 is independent, C 1 ~C 8 Alkyl, heteroalkyl: C 2 ~C 8 Alkenyl, alkynyl, heteroalkyl, heterocycloalkyl;C 3 ~C 8 Aryl, Ar-alkyl.

[0172] Z 2 and Z 3 are independently H, O, S, NH, N(R 15 ), NHNH, -OH, -SH, -NH 2 , NH, NHNH2, -NH(R 15 ), -OR 15 , CO, -COX2, -COX2R 16 , R 17 , F, Cl, Br, I, SR 16 , N.R. 16 R 17 , N=NR 16 , N=R 16 , NO2, SOR 16 R 17 , SO2R 16 , SO3R 16 , OSO3R 16 , P.R. 16 R 17 , POR 16 R17 , PO2R 16 R 17 , OP(O)(OR 17 )2, OCH2OP(O)(OR 17 )2, O.C.(O)R 17 ,OC(O)OP(O)(OR 17 )2, PO(OR 16 )(OR 17 ), OP(O)(OR 17 )OP(O)(OR 17 )2, OC(O)NHR 17 ;-O-(C 4 ~C 12 Glycoside), -N-(C 4 ~C 12 Glycoside);C 1 ~C 8 Alkyl, heteroalkyl;C 2 ~C 8 Alkenyl, alkynyl, heteroalkyl, heterocycloalkyl;C 3 ~C 8 Aryl, Ar-alkyl, carbocyclic, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl, or an ester, ether, or amide of 2 to 8 carbon atoms; or a peptide containing 1 to 8 amino acids (NH(Aa) 1~8 or CO(Aa) 1~8 (N-terminus or C-terminus (Aa) r , r=1 to 8 identical or different amino acid sequences), or the formula (OCH 2 CH 2 ) p or (OCH 2 CH(CH 3 )) p (wherein p is an integer from 0 to about 1000), or a combination of the above groups; X 2 are O, S, SS, NH, and CH. 2 , O.H., S.H., and N.H. 2 , C.H.R. 15 , or NR 15 It is.

[0173] R 15 , R 16 , and R 17are independent of each other, H, C 1 ~C 8 Alkyl, heteroalkyl;C 2 ~C 8 Alkenyl, alkynyl, heteroalkyl, heterocycloalkyl;C 3 ~C 8 Aryl, Ar-alkyl, carbocycle, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl, alkylcarbonyl, or Na + , K + , Cs + , Li + , Ca 2+ , Mg + , Zn 2+ , N + (R 1 )(R 2 )(R 3 )(R 4 ) or HN + (C 2 H 5 OH) 3 It's salt.

[0174] Y 1 and Y 2 is independently N or CH; q is 0 or 1; when q=0, Y 3 does not exist, and Y 4 , Y 5 , Y 6 , and Y 7 are independently CH, N, NH, O, S, or N(R1), so that Y 2 , Y 4 , Y 5 , Y 6 , and Y 7 forms heteroaromatic rings of furan, pyrrole, thiophene, thiazole, oxazole, imidazole, pyrazole, triazole, tetrazole, thiadiazole; when q=1, Y 3 , Y 4 , Y 5 , Y 6 , and Y 7 are independently CH or N, so Y 2 , Y 3 , Y 4 , Y 5 , Y6 , and Y 7 form the aromatic rings of benzene, pyridine, pyridazine, pyrimidine, pyrazine, triazine, tetrazine, and pentazine.

[0175] In particular, tubulysin analogs have the structure shown below: [ka] TIFF2024541059000006.tif231170TIFF2024541059000007.tif246170TIFF2024541059000008.tif251170 TIFF2024541059000009.tif224170TIFF2024541059000010.tif246170TIFF2024541059000011.tif252170 TIFF2024541059000012.tif252170TIFF2024541059000013.tif227170TIFF2024541059000014.tif222170 TIFF2024541059000015.tif251170TIFF2024541059000016.tif241170TIFF2024541059000017.tif121170

[0176] In the formula, R 20 is H;C 1 ~C 8 linear or branched alkyl or heteroalkyl; 2 ~C 8 Straight-chain or branched alkenyl, alkynyl, alkylcycloalkyl, heterocycloalkyl; 3 ~C 8 Ar-alkyl, heterocyclic, carbocyclic, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl; carbonate (-C(O)OR 17 ), carbamates (-C(O)NR 17 R 18 or a carboxylate, ester, ether, or amide having 1 to 8 carbon atoms; or an amino acid having 1 to 8 carbon atoms; or a compound of the formula (OCH2 CH 2 ) p or (OCH 2 CH(CH 3 )) p where p is an integer from 0 to about 1000; or R 20 is absent and the oxygen atom forms a ketone, or a combination of the above;

[0177] Z 2 and Z 3 are independently H, OH, and NH 2 , O, NH, COOH, COO, C(O), C(O), C(O)NH, C(O)NH 2 , R 18 , O.C.H. 2 OP(O)(OR 18 ) 2 ,OC(O)OP(O)(OR 18 ) 2 , OPO(OR 18 ) 2 , NHPO(OR 18 ) 2 , OP(O)(OR 18 )OP(O)(OR 18 ) 2 , O.C.(O)R 18 , O.C.(O)N.H.R. 18 , O.S.O. 2 (OR 18 ), O-(C 4 ~C 12 -glycoside), C 1 ~C 8 linear or branched alkyl or heteroalkyl; 2 ~C 8 Straight-chain or branched alkenyl, alkynyl, alkylcycloalkyl, heterocycloalkyl; 3 ~C 8 Ar-alkyl, heterocyclic, carbocyclic, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl; carbonate (-C(O)OR 17 ), carbamates (-C(O)NR 17 R 18 ).

[0178] R 17 and R 18 are independently H, linear or branched alkyl or heteroalkyl, C 2 ~C 8 Straight-chain or branched alkenyl, alkynyl, alkylcycloalkyl, heterocycloalkyl; 3 ~C 8 Ar-alkyl, heterocyclic, carbocyclic, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl; carbonate (-C(O)OR 17 ), carbamates (-C(O)NR 17 R 18 ).

[0179] R 19 are H, OH, and NH. 2 , O.S.O. 2 (OR 18 ), XCH 2 OP(O)(OR 18 ) 2 , XPO(OR 18 ) 2 , XC(O)OP(O)(OR 18 ) 2 , XC(O)R 18 , XC(O)NHR 18 , C 1 ~C 8 Alkyl or carboxylate; C 2 ~C 8 Alkenyl, alkynyl, alkylcycloalkyl, heterocycloalkyl;C 3 ~C 8 aryl or alkylcarbonyl; or a pharmaceutical salt.

[0180] X is O, S, NH, NHNH, or CH 2 It is.

[0181] R 7 is the same as defined above;

[0182] In the formula, the binding site in formulae (IV-01) to (IV-79) [ka] is the same as the binding site described in formula (IV).

[0183] Calicheamicin and related enediyne antibiotics are described in: Nicolaou, KC et al, Science 1992, 256, 1172-1178; Proc. Natl. Acad. Sci USA. 1993, 90, 5881-8), US Patent Nos. 4, 970, 198; 5, 053, 394; 5, 108, 912; 5, 264, 586; 5, 384, 412; 5, 606, 040; 5, 712, 374; 5, 714, 586; 5, 739, 116; 5, 770, 701; 5, 770, 710; 5, 773, 001; 877, 296; 6, 015, 562; 6, 124, 310; 8, 153, 768. Exemplary enediynes include, but are not limited to, calicheamicin, esperamicin, uncialamicin, dynemicin, and derivatives thereof. The structure of calicheamicin is preferably of the following formula: or an isotope of a chemical element, or a pharma- ceutically acceptable salt, hydrate, or hydrated salt, or a polymorphic crystal structure, or an optical isomer, racemate, diastereomer, or enantiomer thereof. [ka]

[0184] Geldanamycin is a benzoquinone ansamycin antibiotic that binds to Hsp90 (heat shock protein 90) and is used as an antitumor drug. Exemplary geldanamycins include, but are not limited to, 17-AAG (17-N-allylamino-17-demethoxygeldanamycin) and 17-DMAG (17-dimethylaminoethylamino-17-demethoxygeldanamycin).

[0185] Maytansine or its derivatives, maytansinoids, inhibit cell proliferation by inhibiting microtubule formation during mitosis through inhibition of tubulin polymerization. See Remillard et al., Science 189:1002-1005 (1975). Exemplary maytansine and maytansinoids include, but are not limited to, mertansine (DM1, DM4), maytansinol and its derivatives, and ansamitocins. Maytansinoids are disclosed in U.S. Pat. Nos. 4,256, 746, 4,361, 650, 4,307, 016, 4,294, 757, 4,294, 757, 4,371, 533, 4,424, 219, 4,331, 598, 4,450, 254, 4,364, 866, 4,313, 946, 4,315, 929, 4,362, 663, 4,322, 348, 4,371, 533, 4,424, 219, 5,208, 020, 5,416, 064, 5, 208, 020; 5, 416, 064; 6, 333.410; 6, 441, 163; 6, 716, 821, 7, 276, 497, 7, 301, 019, 7, 303, 749, 7, 368, 565, 7, 411, 063, 7, 851, 432, and 8, 163, 888. The structure of the maytansinoid is preferably represented by the following formula:

[0186] [ka]

[0187] Camptothecins (CPTs) and their derivatives are topoisomerase inhibitors that cause DNA damage that prevents DNA religation and thus triggers apoptosis and have been described in the following publications: Shang, XF et al, Med Res Rev. 2018, 38(3) 775-828; Botella, P. and Rivero-Buceta, EJ Control Release 2017, 247: 28-54; Martino, E. et al, Bioorg Med Chem Lett. 2017, 27(4):701-707; Lu, A., et al., Acta Pharmacol Sin 2007, 28(2): 307-314. Camptothecins include SN-38, topotecan, irinotecan (CPT-11), ciratecan (DB-67, AR-67), cositecan (BNP-1350), etirinotecan, exatecan, lurtotecan, gimatecan (ST1481), belotecan (CKD-602), rubitecan, and several others (Shang, XF et al, Med Res Rev. 2018, 38(3):775-828).So far, three CPT analogs, topotecan, irinotecan, and belotecan, have been approved and used in cancer chemotherapy (Palakurthi, S., Expert Opin Drug Deliv. 2015;12(12):1911-21; Shang, XF et al, Med Res Rev. 2018, 38(3):775-828), and both SN-38 and exatecan have been successfully used as payloads for ADC conjugates in clinical trials (Ocean, AJ et al, Cancer. 2017, 123(19): 3843-3854; Starodub, AN, et al, Clin Cancer Res. 2015, 21(17): 3870-8; Cardillo, TM, et al, Bioconjug Chem. 2015, 26(5): 919-31; Ogitani, Y . et al, Bioorg Med Chem Lett. 2016, 26(20): 5069-5072; Takekawa, N. et al, Int J Cancer. 2017 Oct 15;141(8):1682-1689. U.S. Patents 7, 591, 994; 7, 999, 083, 8, 080, 250, 8, 268, 317; U.S. Patent Applications 20130090458, 20140099258, 20150297748, 20160279259).

[0188] Camptothecin (CPT) has the structure shown in the following formula, or one or more isotopes of chemical elements, or a pharma- ceutically acceptable salt, hydrate, or hydrated salt, or a polymorphic crystal structure of these compounds, or an optical isomer, racemate, diastereomer, or enantiomer: [ka]

[0189] In the formula, R 1 , R 2 , and R 4 are independently H, F, Cl, Br, CN, and NO2 , C 1 ~C 8 Alkyl;OC 1 ~C 8 Alkyl;NH-C 1 ~C 8 Alkyl;C 2 ~C 8 Heteroalkyl, alkylcycloalkyl, heterocycloalkyl;C 3 ~C 8 aryl, Ar-alkyl, heterocycle, carbocycle, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl; or an ester, ether, amide, carbonate, urea, or carbamate having 2 to 8 carbon atoms; R 3 H, OH, NH 2 , C 1 ~C 8 Alkyl;OC 1 ~C 8 Alkyl;NH-C 1 ~C 8 Alkyl;C 2 ~C 8 heteroalkyl, alkylcycloalkyl, heterocycloalkyl; or an ester, ether, amide, carbonate, urea, or carbamate having 2 to 8 carbon atoms; or R 1 R 2 , R 2 R 3 , and R 3 R 4 independently form a 5- to 7-membered carbocyclic, heterocyclic, heterocycloalkyl, aromatic, or heteroaromatic ring system.

[0190] [ka] is an internal site to which L1 or L2 can bind.

[0191] The structure of camptothecin is preferably the following formula, or one or more isotopes of the chemical elements, or a pharma- ceutically acceptable salt, hydrate, or hydrated salt, or a polymorphic crystal structure of these compounds, or an optical isomer, racemate, diastereomer, or enantiomer: [ka] TIFF2024541059000024.tif246170TIFF2024541059000025.tif196170

[0192] During the ceremony, [ka] L 1 Or L 2 It is the binding site for;

[0193] P 1 are H, OH, and NH. 2 , COOH, C(O)NH 2 , O.C.H. 2 OP(O)(OR 18 ) 2 ,OC(O)OP(O)(OR 18 ) 2 , OPO(OR 18 ) 2 , NHPO(OR 18 ) 2 , O.C.(O)R 18 , OP(O)(OR 18 )OP(O)(OR 18 ) 2 , O.C.(O)N.H.R. 18 , O C (O) N (C 2 H 4 ) 2 NCH 3 , O.S.O. 2 (OR 18 ), O-(C 4 -C 12 -glycosides), OC(O)N(C 2 H 4 ) 2 CH 2 N(C2 H 4 ) 2 CH 3 , O-(C 1 ~C 8 (linear or branched alkyl), C 1 ~C 8 linear or branched alkyl or heteroalkyl; 2 ~C 8 Straight or branched alkenyl, alkynyl, alkylcycloalkyl, heterocycloalkyl; 3 ~C 8 Aryl, Ar-alkyl, heterocyclic, carbocyclic, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl; carbonate (-C(O)OR 17 ), carbamates (-C(O)NR 17 R 18 );R 17 and R 18 are independently H, linear or branched alkyl or heteroalkyl; C 2 ~C 8 Straight or branched alkenyl, alkynyl, alkylcycloalkyl, heterocycloalkyl; 3 ~C 8 Linear or branched aryl, Ar-alkyl, heterocyclic, carbocyclic, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl; carbonate (-C(O)OR 17 ), carbamates (-C(O)NR 17 R 18 ).

[0194] Combretastatins are naturally occurring phenols with intratumoral vascular disrupting properties. Exemplary combretastatins and their derivatives include, but are not limited to, combretastatin A-4 (CA-4), CA4-βGal, CA-4PD, CA4-NPs, and ombrabulin:

[0195] [ka]

[0196] Taxanes, including the cytotoxic natural product paclitaxel (taxol), the semisynthetic derivative docetaxel (taxotere), and their analogs suitable for conjugation are described in K C. Nicolaou et al., J. Am. Chem. Soc. 117, 2409-20, (1995); Ojima et al, J. Med. Chem. 39:3889-3896 (1996); 40:267-78 (1997); 45, 5620-3 (2002); Ojima et al., Proc. Natl. Acad. Sci., 96:4256-61 (1999); Kim et al., Bull. Korean Chem. Soc., 20, 1389-90 (1999); Miller, et al. J. Med. Chem., 47, 4802-5 (2004); US Patent Nos. 5, 475, 011 5, 728, 849, 5, 811, 452; 6, 340, 701; 6, 372, 738; 6, 391, 913, 6.436, 931; 6, 589, 979; 6, 596, 757; 6, 706, 708; 7, 008, 942; 7, 186, 851; 7, 217, 819; 7, 276, 499; 7, 598, 290; and 7, 667, 054. The structure of the taxane is preferably of the formula:

[0197] [ka]

[0198] During the ceremony, [ka] L 1 Or L 2 Ar and Ar' are independently aryl or heteroaryl.

[0199] Anthracyclines are mammalian DNA topoisomerase II inhibitors that can stabilize enzyme-DNA complexes in which DNA strands are cleaved and are covalently linked to antibody-like proteins. These anticancer drugs have played an important role in the treatment of many forms of solid tumors and acute leukemias for the past several decades. However, anthracyclines cause cardiovascular morbidity and mortality (Sagi, JC, et al, Pharmacogenomics. 2016, 17(9), 1075-87; McGowan, JV, et al, Cardiovasc Drugs Ther. 2017, 31(1), 63-75). Therefore, to increase the specific activity of such molecules while reducing cardiotoxicity, researchers have actively used conjugation of anthracyclines with cell-binding antibodies or antibody-like protein molecules as a general approach to improve the therapeutic index of these drugs (Mollaev, M. et al, Int J Pharm. 2018 Dec 29. pii: S0378-5173(18) 30991-8; Rossin, R., et al, Bioconjug Chem. 2016, 27(7):1697-706; Dal Corso, A,. et al, J Control Release. 2017, 264:211-218). Exemplary anthracyclines include, but are not limited to, daunorubicin, doxorubicin (i.e., adriamycin), epirubicin, idarubicin, valrubicin, and mitoxantrone. The structure of the anthracycline used in the present application is preferably represented by the following formula:

[0200] [ka] TIFF2024541059000031.tif223170

[0201] Vinca alkaloids are a set of antimitotic and antimicrotubule alkaloid agents that act by inhibiting the ability of cancer cells to divide. Vinca alkaloids include vinblastine, vincristine, vindesine, leurosine, vinorelbine, catharanthine, vindoline, vincaminol, vineridine, minovincine, methoxyminovincine, minovincine, vincadifformine, desoxyvincaminol, vincamazine, vincamine, vinpocetine, and vinbrunin. The structure of the vinca alkaloids is preferably vinblastine, vincristine, which has the following formula:

[0202] [ka] TIFF2024541059000033.tif145170

[0203] Dolastatins and their peptide analogs and derivatives, the auristatins, are highly potent antimitotic agents that have been shown to have anticancer and antifungal activity (see, e.g., U.S. Patent No. 5,663,149 and Pettit et al., Antimicrob. Agents Chemother. 42:2961-2965, 1998). Exemplary dolastatins and auristatins include, but are not limited to, dolastatin 10, auristatin E (AE), auristatin EB (AEB), auristatin EFP (AEFP), MMAD (monomethylauristatin D or monomethyldolastatin 10), MMAF (monomethylauristatin F or N-methylvaline-valine-dolaisoloiin-dolaproine-phenylalanine), MMAE (monomethylauristatin E or N-methylvaline-valine-dolaisoloiin-dolaproine-norephedrine), 5-benzoylvaleric acid-AE ester (AEVB), auristatin F phenylenediamine (AFP), and other novel auristatins. Auristatins are described in Int. J. Oncol. 15: 367-72 (1999); Molecular Cancer Therapeutics, vol. 3, No. 8, pp. 921-32 (2004); U.S. Application Nos. 11134826, 20060074008, 2006022925.U.S. Patent Nos. 4414205, 4753894, 4764368, 4816444, 4879278, 4943628, 4978744, 5122368, 5165923, 5169774, 5286637, 5410024, 5521284, 5530097, 5554725, 5585089, 5599902, 5629197, 5635483, 5654399, 5663149, 5665860, 5708146, 5714586, 5741892, 5767236, 5767237, 5780588, 5821337, 5840699, 5965537, 6004934, 6033876, 6034065, 6048720, 6054297, 6054561, 6124431, 6143721, 6162930, 6214345, 6239104, 6323315, Listed in 6342219, 6342221, 6407213, 6569834, 6620911, 6639055, 6884869, 6913748, 7090843, 7091186, 7097840, 7098305, 7098308, 7498298, 7375078, 7462352, 7553816, 7659241, 7662387, 7745394, 7754681, 7829531, 7837980, 7837995, 7902338. The structure of the auristatin analog is preferably of the following formulas (Ih-01), (Ih-02), (Ih-03), (Ih-04), (Ih-05), (Ih-06), (Ih-07), (Ih-08), (Ih-09), (Ih-10), and (Ih-11), or one or more isotopes of chemical elements, or a pharma- ceutically acceptable salt, hydrate, or hydrated salt, or a polymorphic crystal structure of these compounds, or an optical isomer, racemate, diastereomer, or enantiomer:

[0204] [ka] JPEG2024541059000035.jpg254170

[0205] In the formula, R 1 , R 2 , R 3 , R4 , and R 5 are independent, H;C 1 ~C 8 or a peptide containing 1 to 8 amino acids of the formula (OCH 2 CH 2 ) p or (OCH 2 CH(CH 3 )) p (wherein p is an integer from 1 to about 5000); 1 R 2 , R 2 R 3 , R 1 R 3 , or R 3 R 4 may be taken together to form a 3-8 membered ring of an alkyl, aryl, heteroaryl, heteroalkyl, or alkylcycloalkyl group.

[0206] Y 1 and Y 2 is the connection site (independently binds to L1 and / or L2) [ka] and independently O, NH, NHNH, or NR 5 , S, C(O)O, C(O)NH, OC(O)NH, OC(O)O, NHC(O)NH, NHC(O)S, OC(O)N(R 1 ), N(R 1 )C(O)N(R 2 ), C(O)NHNHC(O), and C(O)NR 1 or the connection site [ka] If not bonded to OH, NH 2, NHNH 2 , NHR 5 , SH, C(O)OH, C(O)NH 2 , OC(O)NH 2 , OC(O)OH, NHC(O)NH 2 , NHC(O)SH, OC(O)NH(R 1 ), N(R 1 )(C(O)NH(R 2 ), C(O)NHNHC(O)OH, and C(O)NHR 1 is.

[0207] R 12 is OH, NH 2 , NHR 1 , NHNH 2 , NHNHCOOH, O-R 1 -COOH, NH-R 1 -COOH, NH-(Aa) n COOH, O(CH 2 CH 2 O) p CH 2 CH 2 OH, O(CH 2 CH 2 O) p CH 2 CH 2 NH 2 , NH(CH 2 CH 2 O) p CH 2 CH 2 NH 2 , NR 1 R 1’ , NHOH, NHOR 1 , O(CH 2 CH 2 O) p CH 2 CH 2 , NH(CH 2 CH 2 O) p CH 2 CH 2 , NH-Ar-COOH, NH-Ar-NH 2 , O(CH 2 CH 2 O) p CH 2 CH2 NHSO 3 H、NH(CH 2 CH 2 O) p CH 2 CH 2 NHSO 3 H、R 1 -NHSO 3 H、NH-R 1 -NHSO 3 H、O(CH 2 CH 2 O) p CH 2 CH 2 NHPO 3 H 2 NH(CH 2 CH 2 O) p CH 2 CH 2 NHPO 3 H 2 OR 1 、R 1 -NHPO 3 H 2 、R 1 -OPO 3 H 2 、O(CH 2 CH 2 O) p CH 2 CH 2 OPO 3 H 2 OR 1 -NHPO 3 H 2 、NH-R 1 -NHPO 3 H 2 NH(CH 2 CH 2 NH) p CH 2 CH 2 NH 2 NH(CH 2 CH 2 S) p CH 2 CH 2 NH 2 NH(CH 2 CH 2 NH) p CH 2 CH 2OH, NH(CH 2 CH 2 S) p CH 2 CH 2 OH, NH-R 1 -NH 2 , or NH(CH 2 CH 2 O) p CH 2 CH 2 NHPO 3 H 2 where Aa is 1 to 8 identical or different amino acids; p is 1 to 5000.

[0208] R 1 , R 2 , R 3 , R 4 , R 5 , R 5’ , Z 1 , Z 2 , and n are as defined above.

[0209] Hemiasterlin and its analogs (such as HTI-286) bind to tubulin, disrupt normal microtubule dynamics, and depolymerize microtubules in stoichiometric amounts. The structure of maytansinoids is preferably of the formula:

[0210] [ka]

[0211] In the formula, R 1 , R 2 , R 3 , R 4 , and R 5 are independent, H;C 1 ~C 8 linear or branched alkyl, aryl, heteroaryl, heteroalkyl, alkylcycloalkyl, ester, ether, amide, amine, heterocycloalkyl, or acyloxylamine; or a peptide containing 1 to 8 amino acids, or a peptide of the formula (OCH 2 CH 2 ) p Or (OCH2 CH(CH 3 )) p (wherein p is an integer of 1 to about 5000); 2 R 3 can form a 3- to 8-membered ring of an alkyl, aryl, heteroaryl, heteroalkyl, or alkylcycloalkyl group.

[0212] Eribulin binds primarily to a small number of high affinity sites at the plus ends of pre-existing microtubules and has both cytotoxic and non-cytotoxic modes of action. Its cytotoxic effects are related to its antimitotic activity, inducing apoptosis of cancer cells after prolonged and irreversible mitotic inhibition (Kuznetsov, G. et al, Cancer Research. 2004, 64 (16): 5760-6.; Towle, M. J et al, Cancer Research, 2010, 71 (2): 496-505). Preclinical studies in human breast cancer models have shown that in addition to its cytotoxic, antimitotic-based mechanisms, eribulin also exerts complex effects on the biology of surviving cancer cells and residual tumors that appear to be independent of its antimitotic effects. Eribulin has been approved by the US FDA for the treatment of metastatic breast cancer that has received at least two chemotherapy regimens for advanced disease, including both anthracycline and taxane chemotherapy, and for the treatment of liposarcoma (a type of soft tissue sarcoma) that cannot be removed by surgery (unresectable) or has progressed (metastatic). Eribulin has been used as the payload in ADC conjugates (US20170252458). The structure of eribulin is preferably of the following formula Eb01:

[0213] [ka]

[0214] Nicotinamide phosphoribosyltransferase inhibitors (NAMPT) could be interesting ADC payloads due to their unique mechanism of potent activity (Sampath D, et al, Pharmacol Ther 2015; 151, 16-31). NAMPT regulates intracellular levels of nicotinamide adenine dinucleotide (NAD). NAD functions as an essential redox cofactor that supports energy and anabolic metabolism. NAD plays several important roles in metabolism; it acts as a coenzyme in redox reactions, as a donor of the ADP-ribose moiety in ADP-ribosylation reactions, as a precursor of the second messenger molecule cyclic ADP-ribose, as well as a substrate for bacterial DNA ligase and NAD + In addition to these metabolic functions, they also act as a group of enzymes called sirtuins that use NAD to remove acetyl groups from proteins. +It appears as an adenine nucleotide and is released from cells spontaneously or by regulated mechanisms (Smyth L. M, et al, J. Biol. Chem. 2004, 279 (47), 48893-903; Billington R. A, et al, Mol Med. 2006, 12, 324-7) and therefore may have important extracellular roles (Billington R. A, et al, Mol Med. 2006, 12, 324-7). In the presence of inhibitors of NAMPT, NAD levels fall below those required for metabolism, creating an energy crisis and leading to cell death. Thus far, clinical NAMPT inhibitor candidates FK-866, CHS-828, and GMX-1777 have progressed into clinical trials, but all encountered dose-limiting toxicity before achieving objective responses (Holen K., et al, Invest New Drugs 2008, 26, 45-51; Hovstadius, P., et al, Clin Cancer Res 2002, 8, 2843-50; Pishvaian, MJ, et al, J Clin Oncol 2009, 27, 3581). Thus, the use of ADCs for targeted delivery of NAMPT inhibitors may avoid systemic toxicity and achieve a broader therapeutic index. The structure of the NAMPT inhibitor is preferably of the following formula: NP01, NP02, NP03, NP04, NP05, NP06, NP07, NP08, and NP09, or one or more isotopes of chemical elements, or a pharma- ceutically acceptable salt, hydrate, or hydrated salt, or a polymorphic crystal structure of these compounds, or an optical isomer, racemate, diastereomer, or enantiomer:

[0215] [ka]

[0216] During the ceremony, [ka] is the same as above;X 5 is F, Cl, Br, I, OH, OR 1 , R 1 , OPO 3 H 2 , O.S.O. 3 H, N.H.R. 1 , O.C.O.R. 1 , N.H.C.O.R. 1 It is.

[0217] Benzodiazepine dimers and their analogs (e.g., pyrrolobenzodiazepine (PBD) or (tomaymycin) dimers, indolinobenzodiazepine (IGN) dimers, imidazobenzothiadiazepine dimers, or oxazolidinobenzodiazepine dimers) are antitumor agents that contain one or more imine functional groups that bind to double-stranded DNA, or their equivalents. PBD and IGN molecules are based on the natural product athramycin and interact with DNA in a sequence-selective manner, with a preference for purine-guanine-purine sequences. Benzodiazepine dimers that are preferred in the present invention are exemplified by the following U.S. Patents: 8,153,627; 8,034,808; 7,834,005; 7,741,319; 7,704,924; 7,691,848; 7,678,787; 7,612,062; 7,608,615; 7,557,099; 7,528,128; 7,528,126; 7,511,032; 7,429,658; 7,407,951; 7,326,700; 7,312,210; 7,265,105; 7,202,239; 7,189,710; 7,173,026; 7,109,193; 7,067,511; 7,064,120; 7,056,913; 7,049,311; 7,022,699; 7,015,215; 6,979,684; 6,951,853; 6,884,799; 6,800,622; 6, 747, 144; 6,660,856; 6, 608, 192; 6,562,806; 6, 977, 254; 6,951,853; 6,909,006; 6, 344, 451; 5,880,122; 4, 935, 362; 4,764,616; 4,761,412; 4,723,007; 4,723,003; 4,683,230; 4,663,453; 4,508,647; 4, 464, 467; 4,427,587; 4,000,304; U.S. Patent Application Publication Nos. 20100203007, 20100316656, 20030195196.Exemplary structures of antibody-benzodiazepine dimer conjugates are shown below as PB01, PB02, PB03, PB04, PB05, PB06, PB07, PB08, PB09, PB10, PB11, PB12, PB13, PB14, PB15, PB16, PB17, PB18, PB19, and PB20.

[0218] [ka] TIFF2024541059000043.tif252170TIFF2024541059000044.tif236170 TIFF2024541059000045.tif41170 or isotopes of one or more chemical elements, or pharma- ceutically acceptable salts, hydrates, or hydrated salts, or polymorphic crystal structures of these compounds, or optical isomers, racemates, diastereomers, or enantiomers.

[0219] In the formula, X 1 , X 2 , Y 1 , Y 2 , Z 1 , Z 2 , and n are defined as above. Preferably, X 1 , X 2 , Y 1 , and Y 2 are independently O, N, NH, NHNH, and NR 5 , S, C(O)O, C(O)NH, OC(O)NH, OC(O)O, NHC(O)NH, NHC(O)S, OC(O)N(R 1 ), N(R 1 )C(O)N(R 1 ), CH, C(O)NHNHC(O), and C(O)NR 1 It is.

[0220] R 1 , R 2 , R 3 , R 1’ , R 2’ , and R 3’are independently: H; F; Cl; = O; = S; OH; SH; C 1 -C 8 Linear or branched alkyl, aryl, alkenyl, heteroaryl, heteroalkyl, alkylcycloalkyl, ester (COOR 5 Or -OC(O)R 5 ), Ester (OR 5 ), amide (CONR 5 ), carbamates (OCONR 5 ), amines (NHR 5 , N.R. 5 R 5’ ), heterocycloalkyl, or acyloxylamine (-C(O)NHOH, -ONHC(O)R 5 or a peptide comprising 1 to 20 natural or unnatural amino acids, or a peptide of the formula (OCH 2 CH 2 ) p Or (OCH 2 CH(CH 3 )) p where p is an integer from 1 to about 1000; 1 R 2 , R 2 R 3 , R 1 R 3 , R 1’ R 2’ , R 2’ R 3’ , or R 1’ R 3’ can independently form a 3- to 8-membered ring of an alkyl, aryl, heteroaryl, heteroalkyl, or alkylcycloalkyl group.

[0221] X 3 and Y 3 are independently N, NH, CH, or CR 1 or X 3 and Y 3 Any one of may be omitted.

[0222] R 1 and R 2 is C 1 ~C 8linear or branched alkyl, heteroalkyl; 3 ~C 8 aryl, heteroaryl, alkylcycloalkyl, acylosyl, alkylaryl, alkylaryloxyl, alkylarylamino, alkylarylthiol; or 1 to 6 identical or different amino acid / peptide sequences (Ar)r (r=1-6).

[0223] R 4 , R 5 , R 5’ , R 6 , R 12 , and R 12’ are independently H, OH, and NH 2 , NH(CH 3 ), N.H.N.H. 2 , COOH, SH, OZ 3 , S.Z. 3 , F, Cl, or C 1 -C 8 It is a straight or branched alkyl, aryl, heteroaryl, heteroalkyl, alkylcycloalkyl, or acyloxylamine.

[0224] Z 3 H,OP(O)(OM 1 )(OM 2 ), O.C.H. 2 OP(O)(OM 1 )(OM 2 ), O.S.O. 3 M 1 , or O-glycosides (glucosides, galactosides, mannosides, glucuronosides / glucuronides, allosides, fructosides, etc.), NH-glycosides, S-glycosides, or CH 2 -Glycoside; M 1 and M 2 are independently H, Na, K, Ca, Mg, and NH 4 , N.R. 1 R 2 R 3 is;

[0225] X 6 is CH, N, P(O)NH, P(O)NR 1 , CHC(O)NH, C3 ~C 8 Aryl, heteroaryl, alkylcycloalkyl, acyloxy, alkylaryl, alkylaryloxy, alkylarylamino, or Aa (an amino acid, preferably selected from Lys, Phe, Asp, Glu, Ser, Thr, His, Cys, Tyr, Trp, Gln, Asn, Arg).

[0226] [ka] has the same definition as above.

[0227] CC-1065 analogs and duocarmycin analogs are also preferably used in the conjugates of the present invention. Examples of CC-1065 analogs and duocarmycin analogs, as well as their synthesis, are described in Warpehoski, et al, J. Med. Chem. 31:590 603 (1988); D. Boger et al.,J. Org. Chem; 66; 6654-61, 2001; U.S. Patent Nos.: 4169888, 4391904, 4671958, 4816567, 4912227, 4923990, 4952394, 4975278, 4978757, 4994578, 5037993, 5070092, 5084468, 5101038, 5117006, 5137877, 5138059, 5147786, 5187186, 5223409, 5225539, 5288514, 5324483, 5332740, 5332837, 5334528, 5403484, 5427908, 5475092, 5495009, 5530101, 5545806, 5547667, 5569825, 5571698, 5573922, 558071 7, 5585089, 5585499, 5587161, 5595499, 5606017, 5622929, 5625126, 5629430, 5633425, 5641780, 5660829, 5661016, 5686237, 5693762, 5703080, 5712374, 5714586, 5739116, 5739350, 5770429, 5773001, 5773435, 5786377 5786486, 5789650, 5814318, 5846545, 5874299, 5877296, 5877397, 5885793, 5939598, 5962216, 5969108, 5985908, 6060608, 6066742 , 6075181, 6103236, 6114598, 6130237, 6132722, 6143901, 6150584, 6162963, 6172197, 6180370, 6194612, 6214345, 6262271, 6281 354, 6310209, 6329497, 6342480, 6486326, 6512101, 6521404, 6534660, 6544731, 6548530, 6555313, 6555693 ,6566336,6,586,618,6593081,6630579,6756397,6759509,6762179,6884869,6897034,6946455,7049316,7087600,7091186,7115573,7129261,7214663,7223837,7304032,7329507,7,Nos. 329,760, 7,388,026, 7,655,660, 7,655,661, 7,906,545, and 8,012,978. Exemplary structures of antibody-CC-1065 analog conjugates via the linkers of this patent are shown below in CC01, CC02, CC03, CC04, CC05, CC06, and CC07.

[0228] [ka]

[0229] During the ceremony, X 1 , X 2 , Y 1 , and Y 2 is the connection site [ka] and independently O, NH, NHNH, or NR 5 , S, C(O)O, C(O)NH, OC(O)NH, OC(O)O, NHC(O)NH, NHC(O)S, OC(O)N(R 1 ), N(R 1 )C(O)N(R 2 ), C(O)NHNHC(O), and C(O)NR 1 or the connection site [ka] If not bonded to OH, NH 2 , N.H.N.H. 2 , N.H.R. 1 , SH, C(O)OH, C(O)NH 2 , O-C(O)NH 2 , OC(O)OH, NHC(O)NH 2 , NHC(O)SH, OC(O)NH(R 1 ), N(R 1 )C(O)NH(R 2 ), C(O)NHNHC(O)OH, and C(O)NHR 1 It is.

[0230] Z 3 H,PO(OM 1 )(OM 2 ), SO 3 M 1 , C.H. 2 PO(OM 1 )(OM 2 ), C.H. 3 N(CH 2 CH 2 ) 2 NC(O)-, O(CH 2 CH 2 ) 2 NC(O)-, R 1 , or a glycoside; R 1 , R 2 , R 3 , M 1 , M 2 , and n are as defined above.

[0231] Amatoxins and their analogues are a subgroup that includes at least 10 toxic compounds that are found primarily in several genera of poisonous mushrooms, most notably Amanita phalloides, and several other mushrooms, and are preferred for use in the conjugates of this patent. These ten amatoxins, namely α-amanitin, β-amanitin, γ-amanitin, ε-amanitin, amanulin, amanuric acid, amaninamide, and amanine, proamanitin, are rigid bicyclic peptides synthesized as precursors of proteins containing 35 amino acids, which are finally cleaved by eight amino acid prolyl oligopeptidases (Litten, W. 1975 Scientific American232 (3): 90-101; HE Hallen, et al 2007 Proc. Nat. Aca. Sci. USA 104, 19097-101; K. Baumann, et al, 1993 Biochemistry 32 (15): 4043-50; Karlson-Stiber C, Persson H. 2003, Toxicon 42 (4): 339-49; Horgen, PA et al. 1978 Arch. Microbio. 118 (3): 317-9). Amatoxins kill cells by inhibiting RNA polymerase II (Pol II) and blocking gene transcription and protein synthesis (Brodner, OG and Wieland, T. 1976 Biochemistry, 15(16): 3480-4; Fiume, L., Curr Probl Clin Biochem, 1977, 7: 23-8; Karlson-Stiber C, Persson H. 2003, Toxicon 42(4): 339-49; Chafin, DR , Guo, H. & Price, DH 1995 J. Biol. Chem. 270 (32): 19114-19; Wieland (1983) Int. J. Pept. Protein Res. 22(3): 257-76.). Amatoxins were collected from Amanita phalloides (Yocum, RR1978 Biochemistry 17(18): 3786-9; Zhang, P. et al, 2005, FEMS Microbiol. Lett.252(2), 223-8), or by fermentation with Basidiomycete (Muraoka, S. and Shinozawa T., 2000 J. Biosci. Bioeng. 89(1): 73-6), or by fermentation with A. Fissa (Guo, XW, et al, 2006 Wei Sheng Wu Xue Bao 46(3): 373-8), or by culturing A. cholera mushroom (WO / 1990 / 009799, JP11137291). However, the yields of these isolations and fermentations are rather low (less than 5 mg / l of culture). In the past 30 years, several methods for the preparation of amatoxins and their analogues have been reported (WE Savige, A. Fontana, Chem. Commun. 1976, 600-1; Zanotti, G., et al, Int J Pept Protein Res. 1981. 18(2): 162-8; Wieland, T., et al, Eur. J. Biochem. 1981, 117, 161-4; PA Bartlett, et al, Tetrahedron Lett. 1982, 23, 619-22; Zanotti, G., et al., Biochim Biophys Acta, 1986. 870(3): 454-62; Zanotti, G., et al., Int. J. Peptide Protein Res. 1987, 30, 323-9; Zanotti, G., et al. al., Int. J. Peptide Protein Res. 1987, 30, 450-9; Zanotti, G., et al., Int J Pept Protein Res, 1988. 32(1): 9-20; G. Zanotti, T. et al, Int. J. Peptide Protein Res. 1989, 34, 222-8; Zanotti, G., et al. al., Int J Pept Protein Res, 1990. 35(3): 263-70; Mullersman, JE and JF Preston, 3rd, Int J Pept Protein Res, 1991. 37(6): 544-51; Mullersman, JE, et al, Int J Pept Protein Res, 1991. 38(5): 409-16; Zanotti, G., et al, Int J Pept Protein Res, 1992. 40(6): 551-8; Schmitt, W. et al, J. Am. Chem. Soc. 1996, 118, 4380-7; Anderson, M. O., et al, J. Org. Chem., 2005, 70(12): 4578-84; JP May, et al, J. Org. Chem. 2005, 70, 8424-30; F. Brueckner, P. Cramer, Nat. Struct. Mol. Biol. 2008, 15, 811-8; JP May, DM Perrin, Chem. Eur. J. 2008, 14, 3404-9; JP May, et al, Chem. Eur. J. 2008, 14, 3410-17; Q. Wang, et al, Eur. J. Org. Chem. 2002, 834-9; May, J.P. and D.M. Perrin, Biopolymers, 2007. 88(5): 714-24; May, J.P., et al., Chemistry, 2008. 14(11): 3410-7; S. De Lamo Marin, et al, Eur. J. Org. Chem. 2010, 3985-9; Pousse, G., et al., Org Lett, 2010. 12(16): 3582-5; Luo, H., et al., Chem Biol, 2014. 21(12): 1610-7; Zhao, L., et al., Chembiochem, 2015.16(10): 1420-5), most of which are partial synthetic methods. Due to their extremely potent potency and unique cytotoxic mechanism, amatoxins have been used as conjugate payloads (Fiume, L., Lancet, 1969. 2 (7625): 853-4; Barbanti-Brodano, G. and L. Fiume, Nat New Biol, 1973. 243(130): 281-3; Bonetti, E., M. et al, Arch Toxicol, 1976. 35(1): p. 69-73; Davis, MT, Preston, JF Science 1981, 213, 1385-1388; Preston, JF, et al, Arch Biochem Biophys, 1981. 209(1): 63-71; H. Faulstich, et al, Biochemistry 1981, 20, 6498-504; Barak, LS, et al., Proc Natl Acad Sci USA, 1981. 78(5): 3034-8; Faulstich, H. and L. Fiume, Methods Enzymol, 1985. 112: 225-37; Zhelev, Z., A. et al, Toxicon, 1987. 25(9): 981-7; Khalacheva, K., et al. al, Eksp Med Morfol, 1990. 29(3): 26-30; U. Bermbach, H. Faulstich, Biochemistry 1990, 29, 6839-45; Mullersman, JE and JF Preston, Int. J. Peptide Protein Res. 1991, 37, 544-51; Mullersman, JE and JF Preston, Biochem Cell Biol, 1991. 69(7): 418-27; J. Anderl, H. Echner, H. Faulstich, Beilstein J. Org. Chem. 2012, 8, 2072-84; Moldenhauer, G., et al, J. Natl. Cancer Inst. 2012, 104, 622-34; A. Moshnikova, et al; Biochemistry 2013, 52, 1171-8; Zhao, L., et al., Chembiochem, 2015. 16(10): 1420-5; Zhou, B., et al., Biosens Bioelectron, 2015. 68: 189-96; WO2014 / 043403, US20150218220, EP 1661584). We have been studying the conjugation of amatoxins for some time. The structures of the amatoxins applicable to the present invention are preferably the following Am01, Am02 and Am03, or isotopic substitutions of one or more chemical elements, or pharma- ceutically acceptable salts, hydrates or hydrated salts, or polycrystalline structures of these compounds, or optical isomers, racemates, diastereomers or enantiomers:

[0232] [ka]

[0233] During the ceremony, X 1 and Y 1 are independently O, NH, NHNH, and NR 5 , S, C(O)O, C(O)NH, OC(O)NH, OC(O)O, NHC(O)NH, NHC(O)S, OC(O)N(R 1 ), N(R 1 )C(O)N(R 1 ), C.H. 2 , CHNH, CH 2 O, C(O)NHNHC(O), and C(O)NR 1 It is.

[0234] R 7 , R 8 , and R 9 are independently H, OH, and OR 1 , N.H. 2 , N.H.R. 1 , C 1 ~C6 Alkyl, or lacking.

[0235] Y 2 , O, O 2 , N.R. 1 , NH, or absent.

[0236] R 10 is CH 2 , O, NH, NR 1 , NHC(O), NHC(O)NH, NHC(O)O, OC(O)O, C(O), OC(O), OC(O)(NR 1 ), (NR 1 )C(O)(NR 1 ), C(O)R 1 , or lacking.

[0237] R 11 OH, NH 2 , N.H.R. 1 , N.H.N.H. 2 , NHNHCOOH, OR 1 -COOH, NH-R 1 -COOH, NH-(Aa) r COOH, O(CH 2 CH 2 O) p CH 2 CH 2 OH, O(CH 2 CH 2 O) p CH 2 CH 2 NH 2 , NH(CH 2 CH 2 O) p CH 2 CH 2 NH 2 , N.R. 1 R 2 , O(CH 2 CH 2 O) p CH 2 CH 2 -COOH, NH(CH 2 CH 2 O) p CH 2 CH 2COOH, NH-Ar-COOH, NH-Ar-NH 2 , O(CH 2 CH 2 O) p CH 2 CH 2 -NHSO 3 H, NH(CH 2 CH 2 O) p CH 2 CH 2 NHSO 3 H, R 1 -NHSO 3 H, NH-R 1 -NHSO 3 H, O(CH 2 CH 2 O) p -CH 2 CH 2 NHPO 3 H 2 , NH(CH 2 CH 2 O) p CH 2 CH 2 NHPO 3 H 2 , OR 1 , R 1 -NHPO 3 H 2 , R 1 -OPO 3 H 2 , O(CH 2 CH 2 O) p CH 2 CH 2 OPO 3 H 2 , OR 1 -NHPO 3 H 2 , N.H.-R. 1 -NHPO 3 H 2 , or NH(CH 2 CH 2 O) p CH 2 -CH 2 NHPO 3 H 2 where, (Aa) r is 1 to 8 amino acids.

[0238] n and m 1 are independently 1 to 20.

[0239] p is 1 to 5000.

[0240] R 1 , R 2 , Ar, and [ka] is the same as the definition above.

[0241] Spliceostatins and pladienolides are antitumor compounds that inhibit splicing and interact with the spliceosome SF3b. ​​Examples of spliceostatins include, but are not limited to, spliceostatin A, FR901464, and (2S,3Z)-5-{[(2R,3R,5S,6S)-6-{(2E,4E)-5-[(3R,4R,5R,7S)-7-(2-hydrazinyl-2-oxoethyl)-4-hydroxy-1,6-dioxaspiro[2.5]oct-5-yl]-3-methylpenta-2,4-dien-1-yl-1}-2,5-dimethyltetrahydro-2H-pyran-3-yl]amino}-5-oxopent-3-en-2-yl acetate, which has the core structure of Sp-01:

[0242] [ka]

[0243] Examples of pladienolides include, but are not limited to, pladienolide B, pladienolide D, and E7107.

[0244] The addition of phosphate groups (PO 4Protein kinase inhibitors that block the action of enzymes that add a 5'-amino acid to a protein can modulate protein function. They can be used to treat diseases by modulating cell function to overcome overactivity of protein kinases (including mutant or overexpressed kinases) in cancer or other disease factors. The structures of protein kinase inhibitors are preferably those having the following formulae PK01 to PK40: adavosertib, afatinib, axitinib, bafetinib, bosutinib, cobimetinib, crizotinib, cabozantinib, dasatinib, entrectinib, erdafitinib, erlotinib, erlotinib, fostamatinib, gefitinib, ibrutinib, imatinib, lapatinib, lenvatinib, mubritinib, nissert ... Lotinib, Pazopanib, Pegaptanib, Ponatinib, Revastinib, Regorafenib, Ruxolitinib, Sorafenib, Sunitinib, SU6656, Tofacitinib, Vandetanib, Vemurafenib, Entrectinib, Palbociclib, Ribociclib, Abemaciclib, Dacomitinib, Neratinib, Rociletinib (CO-1686), Osimertinib, AZD3759, Nazartinib (EGF816):

[0245] [ka] TIFF2024541059000054.tif252170TIFF2024541059000055.tif241170 TIFF2024541059000056.tif236170TIFF2024541059000057.tif254170 TIFF2024541059000058.tif236170

[0246] In the formula, Z 5 and Z 5’ are independently O, NH, NHNH, and NR 5 , S, C(O)O, C(O)NH, OC(O)NH, OC(O)O, NHC(O)O, NHC(O)NH, NHC(O)S, OC(O)N(R 1 ), N(R 1 )C(O)N(R 2), C(O)NHNHC(O), and C(O)NR 1 is selected from.

[0247] MEK inhibitors inhibit the mitogen-activated protein kinases MEK1 and / or MEK2, which are often overactivated in some cancers. MEK inhibitors are used specifically to treat BRAF-mutated melanoma, KRAS / BRAF-mutated colorectal cancer, breast cancer, and non-small cell lung cancer (NSCLC). MEK inhibitors are selected from PD0325901, selumetinib (AZD6244), cobimetinib (XL518), refametinib, trametinib (GSK1120212), pimasertib, binimetinib (MEK162), AZD8330, RO4987655, RO5126766, WX-554, E6201, GDC-0623, PD-325901, and TAK-733. Preferred MEK inhibitors are selected from trametinib (GSK1120212), cobimetinib (XL518), binimetinib (MEK162), selumetinib, which have the following formula:

[0248] [ka]

[0249] In the formula, Z 5 are O, NH, NHNH, and NR 5 , S, C(O)O, C(O)NH, OC(O)NH, OC(O)O, NHC(O)O, NHC(O)NH, NHC(O)S, OC(O)N(R 1 ), N(R 1 )C(O)N(R 2 ), C(O)NHNHC(O), and C(O)NR 1 is selected from.

[0250] The protease inhibitor used as the payload of the conjugate is preferably selected from carfilzomib, clindamycin, retapamulin, and indibulin, which have the following structures:

[0251] [ka]

[0252] Immunotoxins are generally macromolecular drugs that are cytotoxic proteins derived from bacterial or plant proteins, such as diphtheria toxin (DT), cholera toxin (CT), trichosanthin (TCS), dianthin, Pseudomonas exotoxin A (ETA), erythrogenic toxin, diphtheria toxin, AB toxin, type III exotoxin, and the like. They may also be highly toxic bacterial pore-forming protoxins that require proteolytic treatment for activation. Examples of such protoxins include proerolysin and its genetically modified form, topsalicin. Topsalicin is a modified recombinant protein designed to be selectively activated by an enzyme present in the prostate, causing localized cell death and tissue destruction without damaging adjacent tissues and nerves. The immunotoxins of the present invention are preferably conjugated by the methods of the present invention to an amino acid having a free amino, thiol, or carboxyl group, more preferably the N-terminal amino acid.

[0253] Additionally, cell receptor agonists, cell stimulatory molecules, or intracellular signaling molecules can be conjugated as drug D by the methods of the present invention.

[0254] The cell-binding ligands and receptor agonists are selected from the following: folic acid derivatives; glutamic acid urea derivatives; somatostatin and its analogs (selected from the group consisting of octreotide (Sandostatin) and lanreotide (Somatilin); aromatic sulfonamides; pituitary adenylate cyclase activating peptide (PACAP) (PAC1); vasoactive intestinal peptide (VIP / PACAP) (VPAC1, VPAC2); melanocyte stimulating hormone (α-MSH); cholecystokinin (CCK) / gastrin receptor agonists, bombesin (Pyr-Gln-Arg-Leu-Gly-Asn-Gln-Trp-Ala-Val-Gly-His-Leu-Met-NH 2neurotensin receptor ligands (NTR1, NTR2, NTR3), substance P (NK1 receptor) ligands; neuropeptide Y (Y1-Y6); homing peptides including RGD (Arg-Gly-Asp), NGR (Asn-Gly-Arg), dimeric and multimeric cyclic RGD peptides (cRGDfV), TAASGVRSMH, and LTLRWVGLMS (chondroitin sulfate proteoglycan NG2 receptor ligand), and F3 peptide; cell membrane permeable peptides. peptide hormones, luteinizing hormone releasing hormone (LHRH) agonists and antagonists, and gonadotropin releasing hormone (GnRH) agonists, which act by targeting follicle stimulating hormone (FSH) and luteinizing hormone (LH) as well as testosterone production, including buserelin (Pyr-His-Trp-Ser-Tyr-D-Ser(OtBu)-Leu-Arg-Pro-NHEt), gonadorelin (Pyr-His-Trp-Ser-Tyr-Gly-Leu-Arg-Pro-Gly-NH 2 ), goserelin (Pyr-His-Trp-Ser-)Tyr-D-Ser(OtBu)-Leu-Arg-Pro-AzGly-NH 2 ), histrelin (Pyr-His-Trp-Ser-Tyr-D-His(N-benzyl)-Leu-Arg-Pro-NHEt), leuprolide (Pyr-His-Trp-Ser-Tyr-D-Leu-Leu-Arg-Pro-NHEt), nafarelin (Pyr-His-Trp-Ser-Tyr-2Nal-Leu-Arg-Pro-Gly-NH 2 ), Triptorelin (Pyr-His-Trp-Ser-Tyr-D-Trp-Leu-Arg-Pro-Gly-NH 2 ), nafarelin, deslorelin, abarelix (Ac-D-2Nal-D-4-chloroPhe-D-3-(3-pyridyl)Ala-Ser-(N-Me)Tyr-D-Asn-Leu-isopropylLys-Pro-DAla-NH 2), Cetrorelix (Ac-D-2Nal-D-4-chloroPhe-D-3-(3-pyridyl)Ala-Ser-Tyr-D-Cit-Leu-Arg-Pro-D-Ala-NH 2 ), degarelix (Ac-D-2Nal-D-4-chloroPhe-D-3-(3-pyridyl)Ala-Ser-4-aminoPhe(L-hydroorotyl)-D-4-aminoPhe(carbamoyl)-Leu-isopropylLys-Pro-D-Ala-NH 2 ), and ganirelix (Ac-D-2Nal-D-4-chloroPhe-D-3-(3-pyridyl)Ala-Ser-Tyr-D-(N9,N10-diethyl)-homoArg-Leu-(N9,N10-diethyl)-homoArg-Pro-D-Ala-NH 2 ); pattern recognition receptors (PRRs) selected from Toll-like receptor (TLR) ligands, C-type lectins, and Nodlike receptor (NLR) ligands; calcitonin receptor agonists; integrin receptors and their receptor subtypes (α V β 1 , α V β 3 , α V β 5 , α V β 6 , α 6 β 4 , α 7 β 1 , α L β 2 , α IIb β 3 agonists (selected from the group consisting of GRGDSPK, cyclo(RGDfV)(L1) and its derivatives [cyclo(-N(Me)R-GDfV), cyclo(R-Sar-DfV), cyclo(RG-N(Me)D-fV), cyclo(RGD-N(Me)fV), cyclo(RGDf-N(Me)V-) (silengitide)]; anticalins (derivatives of lipocalin); adnectins (tenth FN3 (fibronectin)); designed ankyrin repeat proteins (DARPins); avimers; EGF receptor or VEGF receptor agonists;

[0255] The cell binding molecule / ligand or cell receptor agonist is selected from the following: LB01 (folic acid), LB02 (PMSA ligand), LB03 (PMSA ligand), LB04 (PMSA ligand), LB05 (somatostatin), LB06 (somatostatin), LB07 (octreotide, somatostatin analog), LB08 (ramareotide, somatostatin analog), LB09 (vapreotide (sambar), somatostatin analog), LB10 (CAIX ligand), LB11 (CAIX ligand), LB12 (gastrin releasing peptide receptor (GRPr), MBA), LB13 (luteinizing hormone releasing hormone (LH-RH) ligand and GnRH), LB14 (luteinizing hormone releasing hormone (LH-RH) and GnRH ligand), LB15 (GnRH antagonist, abarelix), LB16 (cobalamin, vitamin B12 analog), LB17 (cobalamin, vitamin B12 analog), LB18 (α v β 3 for integrin receptors), LB19 (heterobivalent peptide ligand for VEGF receptors), LB20 (neuromedin B), LB21 (bombesin for G protein-coupled receptors), LB22 (TLR for Toll-like receptors), 2 ), LB23 (for androgen receptor), LB24 (α vCilengitide / cyclo(-RGDfV-) for integrin receptors, LB23 (Fludrocortisone), LB25 (Rifabutin analog), LB26 (Rifabutin analog), LB27 (Rifabutin analog), LB28 (Fludrocortisone), LB29 (Dexamethasone), LB30 (Fluticasone propionate), LB31 (Beclomethasone dipropionate), LB32 (Triamcinolone acetonide), LB33 (Prednisone), LB34 (Prednisolone), LB35 (Methylprednisolone), LB36 (Betamethasone), LB37 (Irinotecan analog), LB3 8 (crizotinib analog), LB39 (bortezomib analog), LB40 (carfilzomib analog), LB41 (carfilzomib analog), LB42 (leuprolide analog), LB43 (triptorelin analog), LB44 (clindamycin), LB45 (liraglutide analog), LB46 (semaglutide analog), LB47 (retapamulin analog), LB48 (indibulin analog), LB49 (vinblastine analog), LB50 (lixisenatide analog), LB51 (osimertinib analog), LB52 (nucleoside analog), LB53 (erlotinib analog), or LB54 (lapatinib analog);

[0256] [ka] TIFF2024541059000062.tif237170TIFF2024541059000063.tif248170TIFF202 4541059000064.tif236170TIFF2024541059000065.tif246170TIFF2024541059 000066.tif248170TIFF2024541059000067.tif250170TIFF2024541059000068. tif237170TIFF2024541059000069.tif232170TIFF2024541059000070.tif61170

[0257] In the formula, X 4 and Y 1are independently O, NH, NHNH, and NR 1 , S, C(O)O, C(O)NH, OC(O)NH, OC(O)O, NHC(O)NH, NHC(O)S, OC(O)N(R 1 ), N(R 1 )C(O)N(R 1 ), C.H. 2 , C(O)NHNHC(O), and C(O)NR 1 It is.

[0258] In another embodiment, one or more of DNA, RNA, mRNA, small interfering RNA (siRNA), microRNA (miRNA), and PIWI-interacting RNA (piRNA) are conjugated as drugs by the methods of the present invention:

[0259] [ka]

[0260] During the ceremony, [ka] is the attachment site for the side chain linker of this patent. [ka] is a single-stranded or double-stranded DNA, RNA, mRNA, siRNA, miRNA, or piRNA; X 1 and Y is independently O, NH, NHNH, NR 1 , S, C(O)O, C(O)NH, OC(O)NH, OC(O)O, NHC(O)NH, NHC(O)S, OC(O)N(R 1 ), N(R 1 )C(O)N(R 1 ), C.H. 2 , C(O)NHNHC(O), and C(O)NR 1 It is.

[0261] L1 and L 2 are the same or different, and each independently represents O, NH, S, NHNH, N(R 3 ), N(R 3 )N(R 3 '), or the formula (OCH 2 CH 2 ) p OR 3 , (OCH 2 CH(CH 3 )) p OR 3 , NH(CH 2 CH 2 O) p OR 3 , NH(CH 2 CH(CH 3 )O)pR3 p OR 3 , N[(CH 2 CH 2 O) p R 3 ][(CH 2 CH 2 O) p’ R 3’ , (OCH 2 CH 2 ) p COOR 3 , or C.H. 2 CH 2 (OCH 2 CH 2 ) p COOR 3 (wherein p and p' are independently an integer selected from 0 to about 1000), or a combination thereof; 1 -C 8 Alkyl;C 2 -C 8 Heteroalkyl, alkylcycloalkyl, heterocycloalkyl;C 3 -C 8 aryl, Ar-alkyl, heterocycle, carbocycle, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl; 3 and R 3’ are independent of each other, H, C 1 -C 8 Alkyl;C2 -C 8 or an ester, ether, or amide having from 1 to 8 carbon atoms; or one to eight natural or unnatural amino acids as described in the definitions; or a group of the formula (OCH 2 CH 2 ) p Or (OCH 2 CH(CH 3 )) p where p is an integer from 0 to about 1000, or a combination thereof.

[0262] L 1 Or L 2 may contain self-immolative or non-self-immolative moieties, peptidyl units, hydrazone bonds, disulfide, ester, oxime, amide, or thioether bonds, where the self-immolative units include aromatic compounds electronically similar to para-aminobenzylcarbamoyl (PAB) groups, including, but not limited to, 2-aminoimidazole-5-methanol derivatives, heterocyclic PAB analogs, β-glucuronides, and ortho- or para-aminobenzyl acetals.

[0263] Preferably, the self-immolative linker moiety has any of the following structures: [ka]

[0264] In the formula, ( * Atoms labeled with X are points of attachment to additional spacers, releasable linker units, cytotoxic agents, and / or cell binding molecules (CBAs); 1 , Y 1 , Z 2 , and Z 3 is independently NH, O or S; Z 1 is independently H, NH, O, or S; v is 0 or 1; U1 are independently H, OH, and C 1 -C 6 Alkyl, (OCH 2 CH 2 ) n , F, Cl, Br, I, OR 5 , S.R. 5 , N.R. 5 R 5’ , N=NR 5 , N=R 5 , N.R. 5 R 5’ , NO 2 , SOR 5 R 5’ , S.O. 2 R 5 , S.O. 3 R 5 , O.S.O. 3 R 5 , P.R. 5 R 5’ , POR 5 R 5’ , P.O. 2 R 5 R 5’ , OPO(OR 5 )(OR 5’ ), or OCH 2 PO(OR 5 )(OR 5’ ) in which R 5 and R 5’ is as above; preferably R 5 and R 5’ are independent of each other, H, C 1 -C 8 Alkyl;C 2 -C 8 Alkenyl, alkynyl, heteroalkyl; C 3 -C 8 It is selected from aryl, heterocycle, carbocycle, cycloalkyl, heterocycloalkyl, heteroaralkyl, alkylcarbonyl; or a medicinal cationic salt.

[0265] The non-self-immolative linker moiety has any of the following structures: [ka] TIFF2024541059000076.tif129170

[0266] In the formula, ( * ) are atoms marked with an additional spacer R 1 , which is the point of attachment to the releasable linker unit, the cytotoxic agent, and / or the binding molecule; X 1 , Y 1 , U 1 , R 5 , R 5’ is as defined above; r is 0 to 100; m and n are independently 0 to 6;

[0267] More preferably, L 1 Or L 2 may be comprised of one or more linker moieties that are 6-maleimidocaproyl ("MC"), maleimidopropanoyl ("MP"), valine-citrulline ("val-cit" or "vc"), alanine-phenylalanine ("ala-phe" or "af"), p-aminobenzyloxycarbonyl ("PAB"), 4-thiopentanoic acid ("SPP"), 4-(N-maleimidomethyl)cyclohexane-1 carboxylate ("MCC"), (4-acetyl)aminobenzoic acid ("SIAB"), 4-thiobutyric acid (SPDB), 4-thio-2-hydroxysulfonylbutyric acid (2-sulfo-SPDB), or a natural or unnatural peptide having from 1 to 8 natural or unnatural amino acid units.

[0268] More preferably, L 1 Or L 2may be a releasable conjugate. The term releasable conjugate refers to a conjugate that includes at least one bond that can be broken down under physiological conditions, such as pH labile, acid labile, base labile, oxidatively labile, metabolically labile, biochemically labile, or enzymatically labile. Physiological conditions that cause the breakdown of such bonds do not necessarily include biological or metabolic processes, but may instead include standard chemical reactions, such as hydrolysis or substitution reactions, for example disulfide bond exchange reactions with intracellular thiols such as glutathione, which is abundant in the millimolar range in endosomes, which have a lower pH than the cytosolic pH, and / or in malignant cells.

[0269] The releasable conjugate (L, L 1 Or L 2 ) examples include, but are not limited to: -(CR 5 R 6 ) m (Aa) r (CR 7 R 8 ) n (OCH 2 CH 2 ) t -, -(CR 5 R 6 ) m (CR 7 R 8 ) n (Aa) r (OCH 2 CH 2 ) t -, -(Aa) r -(CR 5 R 6 ) m (CR 7 R 8 ) n (OCH 2 CH 2 ) t -, -(CR 5 R 6 ) m (CR 7 R 8 ) n (OCH 2 CH 2 ) r(Aa) t -、-(CR 5 R 6 ) m -(CR 7 =CR 8 )(CR 9 R 10 ) n (Aa) t (AND 2 CH 2 ) r -、-(CR 5 R 6 ) m (No. 11 CO)(Aa) t (CR 9 R 10 ) n (AND 2 CH 2 ) r -、-(CR 5 R 6 ) m (Aa) t (No. 11 CO)(CR 9 R 10 ) n (AND 2 CH 2 ) r -、-(CR 5 R 6 ) m (OCO)(Aa) t (CR 9 R 10 ) n (AND 2 CH 2 ) r -、-(CR 5 R 6 ) m (OCNR 7 )(Aa) t (CR 9 R 10 ) n (AND 2 CH 2 ) r -、-(CR 5 R 6 ) m (CO)(Aa) t -(CR 9 R 10 ) n (AND2 CH 2 ) r -、-(CR 5 R 6 ) m (No. 11 CO)(Aa) t (CR 9 R 10 ) n (AND 2 CH 2 ) r -、-(CR 5 R 6 ) m -(OCO)(Aa) t (CR 9 R 10 ) n -(AND 2 CH 2 ) r -、-(CR 5 R 6 ) m (OCNR 7 )(Aa) t (CR 9 R 10 ) n (AND 2 CH 2 ) r -、-(CR 5 R 6 ) m (CO)(Aa) t (CR 9 R 10 ) n (AND 2 CH 2 ) r -、-(CR 5 R 6 ) m -phenyl-(CO)(Aa) t (CR 7 R 8 ) n -、-(CR 5 R 6 ) m -フリル-(CO)(Aa) t (CR 7 R 8 ) n -、-(CR 5 R 6 ) m-Oxazolyl-(CO)(Aa) t (CR 7 R 8 ) n -, -(CR 5 R 6 ) m -Thiazolyl-(CO)(Aa) t (CR 7 R 8 ) n -, -(CR 5 R 6 ) t -Thienyl-(CO)(CR 7 R 8 ) n -, -(CR 5 R 6 ) t -Imidazolyl-(CO)(CR 7 R 8 ) n -, -(CR 5 R 6 ) t -Morpholino-(CO)(Aa) t (CR 7 R 8 ) n -, -(CR 5 R 6 ) t -Piperazino-(CO)(Aa) t (CR 7 R 8 ) n -, -(CR 5 R 6 ) t -N-Methylpiperazine-(CO)(Aa) t (CR 7 R 8 ) n -, -(CR 5 R 6 ) m -(Aa) t Phenyl-, -(CR 5 R 6 ) m -(Aa) t Frill-,-(CR 5 R 6 ) m -Oxazolyl (Aa) t -, -(CR 5 R 6 )m -Thiazolyl (Aa) t -, -(CR 5 R 6 ) m -Thienyl (Aa) t -, -(CR 5 R 6 ) m -Imidazolyl (Aa) t -, -(CR 5 R 6 ) m -Morpholino-(Aa) t -, -(CR 5 R 6 ) m -Piperazino-(Aa) t -, -(CR 5 R 6 ) m -N-Methylpiperazino-(Aa) t -, -K(CR 5 R 6 ) m (Aa) r (CR 7 R 8 ) n (OCH 2 CH 2 ) t -, -K(CR 5 R 6 ) m (CR 7 R 8 ) n (Aa) r (OCH 2 CH 2 ) t -, -K(Aa) r (CR 5 R 6 ) m (CR 7 R 8 ) n (OCH 2 CH 2 ) t -, -K(CR 5 R 6 ) m (CR 7 R 8 ) n (OCH 2 CH 2 ) r (Aa)t -、-K(CR 5 R 6 ) m (CR 7 =R 8 )(CR 9 R 10 ) n (Aa) t (AND 2 CH 2 ) r -、-K(CR 5 R 6 ) m (No. 11 CO)(Aa) t (CR 9 R 10 ) n (AND 2 CH 2 ) r -、-K(CR 5 R 6 ) m (Aa) t (No. 11 CO)(CR 9 R 10 ) n (AND 2 CH 2 ) r -、-K(CR 5 R 6 ) m (OCO)(Aa) t (CR 9 R 10 ) n (AND 2 CH 2 ) r -、-K(CR 5 R 6 ) m (OCNR 7 )(Aa) t (CR 9 R 10 ) n (AND 2 CH 2 ) r -、-K(CR 5 R 6 ) m (CO)(Aa) t (CR 9 R 10 ) n (AND2 CH 2 ) r -、-K(CR 5 R 6 ) m (No. 11 CO)(Aa) t (CR 9 R 10 ) n (AND 2 CH 2 ) r -、-K(CR 5 R 6 ) m (OCO)(Aa) t (CR 9 R 10 ) n (AND 2 CH 2 ) r -、-K(CR 5 R 6 ) m (OCNR 7 )(Aa) t (CR 9 R 10 ) n (AND 2 CH 2 ) r -、-K(CR 5 R 6 ) m (CO)(Aa) t (CR 9 R 10 ) n (AND 2 CH 2 ) r -、-K(CR 5 R 6 ) m -phenyl-(CO)(Aa) t (CR 7 R 8 ) n -、-K(CR 5 R 6 ) m -フリル-(CO)(Aa) t (CR 7 R 8 ) n -、-K(CR 5 R 6 ) m-Oxazolyl-(CO)(Aa) t (CR 7 R 8 ) n -, -K(CR 5 R 6 ) m -Thiazolyl-(CO)(Aa) t (CR 7 R 8 ) n -, -K(CR 5 R 6 ) t -Thienyl-(CO)(CR 7 R 8 ) n -, -K(CR 5 R 6 ) t -Imidazolyl-(CO)(CR 7 R 8 ) n -, -K(CR 5 R 6 ) t -Morpholino-(CO)(Aa) t (CR 7 R 8 ) n -, -K(CR 5 R 6 ) t -Piperazino-(CO)(Aa) t (CR 7 R 8 ) n -, -K(CR 5 R 6 ) t -N-Methylpiperazine-(CO)(Aa) t (CR 7 R 8 ) n -, -K(CR 5 R 6 ) m -(Aa) t Phenyl-, -K(CR 5 R 6 ) m -(Aa) t Frill-,-K(CR 5 R 6 ) m -Oxazolyl (Aa) t -, -K(CR 5 R6 ) m -Thiazolyl (Aa) t -, -K(CR 5 R 6 ) m -Thienyl (Aa) t -, -K(CR 5 R 6 ) m -Imidazolyl (Aa) t -, -K(CR 5 R 6 ) m -Morpholino-(Aa) t -, -K(CR 5 R 6 ) m -Piperazino-(Aa) t G-, -K(CR 5 R 6 ) m -N-Methylpiperazino-(Aa) t .

[0270] In the formula, m, Aa, m, n, R 3 , R 4 , and R 5 is defined as above; t and r are independently 0 to 100; R 6 , R 7 , and R 8 are independently: H; halide; C 1 ~C 8 Selected from alkyl, aryl, alkenyl, alkynyl, ether, ester, amine, or amide, and one or more of halide, CN, NR 1 R 2 , C.F. 3 , OR 1 , aryl, heterocycle, S(O)R 1 , S.O. 2 R 1 , -CO 2 H, -SO 3 H, -OR 1 , -CO 2 R 1 , -CONR 1 , -PO 2 R 1 R 12 , -PO 3 H or P(O)R 1R 2 R 3’ K is optionally substituted with NR 1 , -SS-, -C(=O)-, -C(=O)NH-, -C(=O)O-, -C=NH-O-, -C=N-NH-, -C(=O)NH-NH-, O, S, Se, B, or C 3 ~C 6 It is a heteroaromatic ring.

[0271] The connector L 1 and L 2 The structure may be the following structures or combinations thereof: [ka] TIFF2024541059000078.tif238170TIFF2024541059000079.tif241170TIFF2024541059000080.tif252170 TIFF2024541059000081.tif237170TIFF2024541059000082.tif236170TIFF2024541059000083.tif145170

[0272] In the formula, X 2 , X 3 , X 4 , X 5 , or X 6 are independently NH, NHNH;N(R 12 );N(R 12 )N(R 12’ );O;S;C 1 ~C 6 Alkyl;C 2 ~C 6 Heteroalkyl, alkylcycloalkyl, heterocycloalkyl;C 3 ~C 8 Aryl, Ar-alkyl, heterocycle, carbocycle, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl;CH 2 OR 12 , C.H. 2 S.R. 12 , C.H. 2 NHR12 or 1 to 8 amino acids; 12 and R 12’ are independent, H;C 1 ~C 8 Alkyl;C 2 ~C 8 Heteroalkyl, alkylcycloalkyl, heterocycloalkyl;C 3 ~C 8 Aryl, Ar-alkyl, heterocycle, carbocycle, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl; or an ester, ether, or amide having 1 to 8 carbon atoms; or a group of the formula (OCH 2 CH 2 ) p Or (OCH 2 CH(CH 3 )) p where p is an integer from 0 to about 100; or a combination thereof.

[0273] Preferably, L 1 and L 2 are independently selected from: [ka] TIFF2024541059000085.tif65170

[0274] During the ceremony, [ka] is a drug or linker L 1 Or L 2 "#" is the site of attachment to an antibody S (thiol), O (phenol), NH (amino), CHO (aldehyde), C(=O) (ketone), C(O)(NH) (amide), C(O)(OH) (carboxylate); Aa is an L- or D-natural or unnatural amino acid.

[0275] R 1is H, C 1 -C 8 Alkyl, OH, CH 2 OH, CH 2 CH 2 OH, NH 2 , S.H., S.C.H. 3 , C.H. 2 COOH, CH 2 CH 2 COOH, CH 2 CH 2 CH 2 CH 2 NH 2 , C 6 H 5 , C.H. 2 C 6 H 5 , C.H. 2 C 6 H 4 OH, CH(OH)CH 3 , C.H. 2 C(O)NH 2 , C.H. 2 CH 2 C(O)NH 2 , C.H. 2 CH 2 CH 2 NHC(=NH)NH 2 It is.

[0276] r is 0 to 12; if r is not 0, (Aa)r is an amino acid or peptide unit of the same or different sequence;

[0277] m 1 =1-18;m 2 =1-100;m 3 =1-8;m 4 =0-8;m 5 =1-8.

[0278] Y 7 NH, OCH 2 NH, NHC(=O), NHNH, C(=O)NH, N(R 1 ), SO 2 , P(O)(OH), NHS(O) 2 , NHS(O) 2 NH, NHS(O) 2NHC(O), NHS(O) 2 NHC(O)O, NHS(O) 2 NHC(O)NH, NHP(O)(OH), NHP(O)(OH)NH, OP(O)(OH)O, NHP(O)(OH)O, OP(O)(OH)NH, S, O, OP(O)(OH)OP(O)(OH)NH, NHP(O)(OH)OP(O)(OH)NH, NHP(O)(OH)OP(O)(OH)O, OCH 2 CH 2 O, OCH 2 CH 2 NH, N(CH 2 CH 2 ) 2 N, N.H.C. 6 H 4 N.H., C.H. 2 is;

[0279] Y 8 is NHC(=O), NHS(O 2 ), NH(SO), NHS(O 2 )NH, NHP(O)(OH)NH, or C(O)NH;

[0280] R 9 is H, (O=)CR 1 , (O=)CNHR 1 , R 1 COOH, R 1 (COCH 2 NH)m 2 H, R 1 (Aa) r or R 1 (COCH 2 NCH 3 ) m2 H, where R 1 is defined above.

[0281] Lv 1’ is selected from: [ka] TIFF2024541059000088.tif252170TIFF2024541059000089.tif221170TIFF20245410590 00090.tif247170TIFF2024541059000091.tif248170TIFF2024541059000092.tif206170 During the ceremony, [ka] is a drug or linker L 1 Or L 2 "#" is a site that binds to S (thiol), O (phenol), NH (amino), CHO (aldehyde), C(=O) (ketone), C(O)(NH) (amide), C(O)(OH) (carboxylate) of the antibody; 1 , X 1 ', and X 2 ' is as above; X is O, NH, S, CH 2 is the bond in the middle of two atoms [ka] means that it can bond to either of the two atoms;

[0282] More preferably, L 1 and L 2 are independently selected from: [ka]

[0283] During the ceremony, Aa is an L- or D-natural or unnatural amino acid; R 1 is H, C 1 -C 8 Alkyl, OH, CH 2 OH, CH 2 CH2 OH, NH 2 , S.H., S.C.H. 3 , C.H. 2 COOH, CH 2 CH 2 COOH, CH 2 CH 2 CH 2 CH 2 NH 2 , C 6 H 5 , C.H. 2 C 6 H 5 , C.H. 2 C 6 H 4 OH, CH(OH)CH 3 , C.H. 2 C(O)NH 2 , C.H. 2 CH 2 C(O)NH 2 , C.H. 2 CH 2 CH 2 NHC(=NH)NH 2 is; r is 0 to 12; if r is not 0, (Aa)r is an amino acid or peptide unit of the same or different sequence; m 1 =1-18;m 2 =1-100;m 3 =1-8;m 4 =0-8;m 5 = 1-8; Y 7 NH, OCH 2 NH, NHC(=O), NHNH, C(=O)NH, N(R 1 ), SO 2 , P(O)(OH), NHS(O) 2 , NHS(O) 2 NH, NHS(O) 2 NHC(O), NHS(O) 2 NHC(O)O, NHS(O) 2NHC(O)NH, NHP(O)(OH), NHP(O)(OH)NH, OP(O)(OH)O, NHP(O)(OH)O, OP(O)(OH)NH, S, O, OP(O)(OH)OP(O)(OH)NH, NHP(O)(OH)OP(O)(OH)NH, NHP(O)(OH)OP(O)(OH)O, OCH 2 CH 2 O, OCH 2 CH 2 NH, N(CH 2 CH 2 ) 2 N, N.H.C. 6 H 4 N.H., C.H. 2 is; Y 8 is NHC(=O), NHS(O 2 ), NH(SO), NHS(O 2 )NH, NHP(O)(OH)NH, or C(O)NH; R 9 is H, (O=)CR 1 , (O=)CNHR 1 , R 1 COOH, R 1 (COCH 2 NH)m 2 H, R 1 (Aa) r or R 1 (COCH 2 NCH 3 ) m2 H, where R 1 is defined above.

[0284] In certain embodiments, the conjugates of formula (I), (II), and (III) are prepared via a conjugation reaction of an antibody with a compound having the following formula (IV), (V), and (VI), respectively: [ka]

[0285] Lv 1 And Lv 2 is a reactive group, independently selected from: [ka] TIFF2024541059000098.tif238170TIFF2024541059000099.tif231170TIFF2024541059000100.tif252170TIFF2024541059000101.tif246170 TIFF2024541059000102.tif237170TIFF2024541059000103.tif247170TIFF2024541059000104.tif247170TIFF2024541059000105.tif156170

[0286] In the formula, X 1 ' and X2 ' are independently F, Cl, Br, I, OTf, OMs, OC 6 H 4 (NO 2 ), O.C. 6 H 3 (NO 2 ) 2 , O.C. 6 F 5 , O.C. 6 HF 4 , or Lv 3 X 2 is O, NH, N(R 1 ), or C.H. 2 R 3 and R 5 are independently H, R 1 , aromatic, heteroaromatic, or aromatic group, and one or several H atoms are independently selected from -R 1 , -Halogen, -OR 1 , -SR 1 , -NR 1 R 2 , -NO 2 , -S(O)R 1 , -S(O) 2 R 1 , or -COOR 1 Replaced by; Lv 3 and Lv 3' is independently a leaving group selected from F, Cl, Br, I, nitrophenoxyl, N-hydroxysuccinimide (NHS), phenoxyl, benzenethiol, dinitrophenoxyl, pentafluorophenoxyl, tetrafluorophenoxyl, difluorophenoxyl, monofluorophenoxyl, pentachlorophenoxyl, triflate, imidazole, dichlorophenoxyl, tetrachlorophenoxyl, 1-hydroxybenzotriazole, tosylate, mesylate, 2-ethyl-5-phenylisoxazolium-3'-sulfonate, anhydrides formed with themselves or other anhydrides, acetyl anhydride, formyl anhydride, or an intermediate molecule generated with a condensation reagent for a peptide coupling reaction or a Mitsunobu reaction.

[0287] In formula (V) and formula (VI), [ka] can be selected from the following: [ka] TIFF2024541059000108.tif225170TIFF2024541059000109.tif248170TIFF2024541059000110.tif236170TIFF2024541059 000111.tif237170TIFF2024541059000112.tif248170TIFF2024541059000113.tif247170TIFF2024541059000114.tif55170

[0288] In the formula, Lv 3 , Lv 3 ', X 1 ', and X 2 ' is as above. The bond in the middle of two atoms [ka] means that it can bond to either of the two atoms in question.

[0289] In some embodiments, under the method of the present patent invention, a conjugate having formula (VII), (VIII), or (IX) as shown below can be first reacted with a selectively reduced thiol in an antibody or antibody-like protein (usually the thiol between the heavy and light chains if the antibody or antibody-like protein is an IgG antibody) and then condensed with a cytotoxic agent or cytotoxic agent / conjugate to form a conjugate of formula (I), (II), or (III) as shown above: [ka]

[0290] In the formula, L 1 , L 2 , E 1 , Lv 1 , and Lv 2 has the same definition as above for formulas (I), (II), (III), (IV), (V), and (VI).

[0291] Lv 5 And Lv 6 is independently selected from:

[0292] [ka] TIFF2024541059000118.tif33170

[0293] In the formula, X1' is F, Cl, Br, I, OTs (tosylate), OTf (triflate), OMs (mesylate), OC 6 H 4 (NO 2 ), O.C. 6 H 3 (NO 2 ) 2 , O.C. 6 F 5 , O.C. 6 HF4 , or Lv 3 X 2’ is O, NH, N(R 1 ), or CH 2 R 3 and R 5 are independent, H, R 1 , aromatic, heteroaromatic, or one or several H atoms are independently -R 1 , -Halogen, -OR 1 , -SR 1 , -NR 1 R 2 , -NO 2 , -S(O)R 1 , -S(O) 2 R 1 , or -COOR 1 is an aromatic group substituted by; Lv 3 And Lv 3’ are independently a leaving group selected from F, Cl, Br, I, nitrophenoxyl; N-hydroxysuccinimide (NHS); phenoxyl; benzenethiol, dinitrophenoxyl; pentafluorophenoxyl; tetrafluorophenoxyl; difluorophenoxyl; monofluorophenoxyl; pentachlorophenoxyl; triflate; imidazole; dichlorophenoxyl; tetrachlorophenoxyl; 1-hydroxybenzotriazole; tosylate; mesylate; 2-ethyl-5-phenylisoxazolium-3'-sulfonate, anhydrides formed with themselves or with other anhydrides selected from acetyl anhydride and formyl anhydride, or intermediate molecules generated with condensation reagents for peptide coupling reactions or Mitsunobu reactions; wherein the reaction is 1 Or Lv 2 As long as the reaction between the functional group Lv and the antibody, especially the thiols in the antibody, is at least 1-fold faster or slower than the reaction between the functional group Lv 5 And / or Lv 6 can also react with thiols in cytotoxic agents.

[0294] In some embodiments, the method of the present patent involves introducing a specific thiol into the antibody-like protein, typically generating a thiol between the heavy and light chains when the antibody or antibody-like protein is an IgG antibody, and then reacting the thiol simultaneously or sequentially with a conjugate of the following formula (VII), (VIII), or (IX) to form an antibody / conjugate conjugate molecule of the following formula (X), (XI), or (XII), followed by the addition of a drug D. 1 Or D 2 to form a conjugate of formula (I), (II), or (III):

[0295] [ka]

[0296] In the formula, Lv 5 , Lv 6 , L 1 , L 2 , E 1 , Lv 1’ , Lv 2’ , mAb, n, and n' are as defined above.

[0297] In some embodiments, under the method of the present invention, the conjugate of formula (VII), (VIII), or (IX) above can be first reacted with a cytotoxic agent to form a cytotoxic agent / conjugate molecule of formula (IV), (V), or (VI), and then reacted independently with a reduced thiol in an antibody or antibody-like protein to form a conjugate of formula (I), (II), or (III). The first step condensation reaction with a cytotoxic agent of formula (VII), (VIII), or (IX) can be carried out in a separate pot, and the resulting cytotoxic agent / conjugate conjugate of formula (IV), (V), or (VI) can be optionally purified by chromatography, extraction, or precipitation before coupling with a reduced thiol in an antibody-like protein. The first step of specific reduction of disulfide bonds in the antibody-like protein and the coupling reaction with formula (IV), (V), or (VI) are preferably carried out in the same pot without separation of intermediates.

[0298] Lv against cytotoxic agents 5 And / or Lv 6 and the reaction between Lv and thiols in antibody-like proteins. 1 And / or Lv 2To distinguish between the reactions between the above, the reactions involving the linker of formula (VIII), (IX), or (X) can be carried out in the same or different reaction pods under different conditions. For example, a drug having an amino group can be condensed with a carboxyl group in the linker in the presence of a condensation reagent such as EDC, TBTU, BroP, etc. to obtain a modified drug / linker conjugate of formula (IV), (V), or (VI) having an amide bond. This condensation reaction can be carried out in a physiological buffer by activating the carboxyl group at one end of the linker of formula (VII), (VIII), or (IX) with n-hydroxyimide (NHS), pentafluorophenyl, dinitrophenyl ester, or carboxylic acid chloride, etc., which can react with a drug having an amino group to provide a drug / linker conjugate of formula (III), (IV), or (V), and then simultaneously or sequentially conjugating with a thiol of an antibody-like protein according to the method of the present invention to form a conjugate of formula (I), (II), or (III). In another embodiment, a thiol-reactive group (e.g., maleimide, vinylsulfonyl, haloacetyl, acryl, substituted propiolic) is provided at one end, and a group capable of reacting with a drug (e.g., hydroxysuccinimidyl (NHS), pentafluorophenyl, dinitrophenyl ester, amino, alkyloxyamino, or a group capable of click chemistry (e.g., azide, alkyne, dibenzocyclooctyne, BCN ((1R,8S,9s)-bicyclo[6.1.0]non-4-yn-9-ylmethanol)) is provided at the other end, comprising formula (VII), (VIII), or (VIII). The conjugate of formula (IX) can be conjugated to a thiol of an antibody-like protein (preferably an antibody) by applying the method of the present application in a buffer solution at pH 4.5-7.5 and 2° C.-40° C. (preferably 2° C.-8° C.) with or without the addition of 0-30% water-miscible organic solvent to obtain an antibody conjugate of formula (X), (XI), or (XII), respectively. Then, a drug capable of reacting with a reactive group in the antibody-conjugate conjugate of formula (X), (XI), or (XII) can be added sequentially or simultaneously to the reaction solution to obtain a conjugate of formula (I), (II), or (III).Optionally, the antibody-conjugate conjugate of formula (X), (XI), or (XII) can be purified prior to condensation with the drug, and the condensation conditions in the second step can be modified, for example, by adjusting the pH to 6.5 to 8.5 and / or the temperature to 20 to 45° C.

[0299] In some embodiments, during the conjugation method, prior to conjugation with the drug, the antibody-like protein is conjugated with an amine to sulfhydryl (succinimidyl (NHS) ester / maleimide, NHS ester / pyridyldithiol, NHS ester / haloacetyl) conjugation, a diazirine (SDA) to sulfhydryl conjugation, an azide to sulfhydryl conjugation, an alkyne to sulfhydryl conjugation, a sulfhydryl to carbohydrate (maleimide / hydrazide, pyridyl) conjugation, or a thiol conjugation. The hydroxyl to sulfhydryl (isocyanate / maleimide) linkages may be modified by conjugation with a heterobifunctional cross-linker of formula (X), (XI), or (XII), such as a hydroxyl to sulfhydryl (maleimide / psoralen, pyridyldithiol / psoralen, haloacetyl / psoralen) linkage, a sulfhydryl to carboxyl (carbodiimide) linkage.

[0300] The reactive group of the drug / cytotoxic agent that reacts with the modified antibody-conjugate complex of formula (X), (XI), or (XII) to give the final conjugate can be by different methods accordingly. For example, conjugates linked via disulfide bonds can be achieved by a first step of conjugating a conjugate of formula (VI), (VII), or (VIII) to an antibody at 2° C.-8° C. and pH 4.5-6.0 by reduction and conjugation of an antibody-like protein according to the invention, followed by disulfide exchange between a drug containing a free thiol group and a disulfide bond (e.g., a pyridyldithio moiety) in the conjugate attached to the modified antibody at pH 6.5-8.0 and 20° C.-40° C. Prior to the addition of the drug containing a free thiol for conjugation, it is preferred to remove excess reducing agent (such as TCEP or tri(3-hydroxylpropyl)phosphine) from the reaction vessel or quench by addition of an azide compound (e.g., 4-(azidomethyl)benzoic acid). The synthesis of thioether linked conjugates is accomplished by first reacting a linker containing both thiol reactive ends, maleimide, haloacetyl, ethylsulfonyl, or substituted propionyl groups, with a thiol in an antibody reduced by the methods of this patent application at 2° C.-8° C. and pH 4.5-6.0 to produce an antibody-conjugate conjugate of formula (X), (XI), or (XII), followed by reaction with a thiol containing drug at pH 6.5-8.0 and 20° C.-40° C. to yield a conjugate of formula (I), (II), or (III). If the same pH and / or temperature conditions are selected for the two-step reaction of the thioether linked conjugates, the linker containing the reactive dual end thiol is used for the conjugation at 4-fold or more equivalents. It should be noted that the preferred method of synthesis of disulfide or thiol-ether bond conjugates is by first chemical synthesis of a drug-conjugate conjugate having a disulfide or thiol-ether bond of formula (IV), (V), or (VI), followed by reaction with a thiol in a protein (antibody) according to the methods of the present invention.Synthesis of conjugates having acid-labile hydrazone bonds can be achieved by reacting a carbonyl group with a hydrazide moiety in the linker by methods known in the art (e.g., P. Hamann et al., Cancer Res. 53, 3336-34, 1993; B. Laguzza et al., J. Med. Chem., 32; 548-55, 1959; P. Trail et al., Cancer Res., 57; 100-5, 1997). The synthesis of conjugates with triazole bonds can be achieved by reacting the 1-yne group of the drug with the azide moiety in the linker via click chemistry (Huisgen cycloaddition) (Lutz, JF. et al, 2008, Adv. Drug Del Rev. 60, 958-70; Sletten, EM et al 2011, Acc Chem. Research 44, 666-76). The synthesis of conjugates linked via oximes is achieved by reaction of modified antibody-like proteins containing ketones or aldehydes with drugs containing oxyamine groups. Drugs bearing hydroxyl or thiol groups can be reacted in the presence of a weak base, e.g., at pH 8.0-9.5, with modified linkers of formula (X), (XI), or (XII) bearing halogens, particularly alpha halides of carboxylic acids, to give modified drug / linker conjugates bearing ether or thiol ether bonds of formula (IV), (V), or (VI). A drug containing a hydroxyl group can be condensed with a linker of formula (X), (XI), or (XII) bearing a carboxyl group in the presence of a dehydrating agent such as EDC or DCC to form an ester bond, and the drug / linker conjugate can then be conjugated to an antibody under the methods of the present invention.Drugs containing an amino group can be condensed with NHS, imidazole, nitrophenoxyl carboxyl ester; N-hydroxysuccinimide (NHS); methylsulfonylphenoxyl; dinitrophenoxyl; pentafluorophenoxyl; tetrafluorophenoxyl; difluorophenoxyl; monofluorophenoxyl; pentachlorophenoxyl; triflate; imidazole; dichlorophenoxyl; tetrachlorophenoxyl; 1-hydroxybenzotriazole; tosylate; mesylate; 2-ethyl-5-phenylisoxazolium-3'-sulfonate on the antibody-conjugate of formula (X), (XI), or (XII) to obtain an amide bond conjugate of formula (I), (II), or (III). Many routine chemical and biochemical processes for antibody-drug conjugation are known in the art (e.g., Mada, Y. and Mendelsohn, BA, Expert Opin Biol Ther. 2021, 21(7): 963-975; Puthenveetil, S., Methods Mol Biol. 2020, 2078: 99-112; van Delft, F., and Lambert, JM, eds., Chemical Linkers in Antibody-Drug Conjugates (ADCs), Royal Soc. Chem. Pub., 22. Dec. 2021, ISBN 978-1-83916-263-3, doi:10.1039 / 9781839165153; Tumey, LN, ed., Antibody-Drug Conjugates, Methods and Protocols, Springer Pub., 2020, ISBN: 978-1-4939 -9929-3; Khongorzul, P. et al, Mol Cancer Res. 2020,18(1):3-19; and many references incorporated within these books and articles).

[0301] In some embodiments, the BCMA antibody conjugates are preferably prepared via a homogeneous conjugation method that includes three key steps: (a) An effective amount of zinc cation-amino chelate / complex (Zn(NR 1 R 2 R 3 ) m1 2+ incubating the antibody in a buffer system (e.g., PBS, Mes, Bis-Tris, Bis-Tris propane, Pipes, Aces, Mopso, Bes, Mops, Hepes, Tes, Pipps, Dipso, Tapso, Heppso, Tris-up, Tris-HCl, Tricine, Hepps, Gly-Gly, Bicine, Taps, Hepee, Acetate, Histidine, Citrate, MES, or Borate) in the presence of PBS, Mes-Tris, Bis-Tris propane, Pipes, Aces, Mops, Bes, Mops, Hepes, Tes, Pipps, Dipso, Tapso, Heppso, Tris-up, Tris-HCl, Tricine, Hepps, Gly-Gly, Bicine, Taps, Hepee, Acetate, Histidine, Citrate, MES, or Borate); (b) adding an effective amount of a conjugate of formula (VII), (VIII) or (IX), or a payload / conjugate complex / assembly of formula (IV), (V) or (VI), having a thiol-reactive group (e.g., a drug containing a maleimide terminus) to react with the thiol group generated in step (a); and (c) adding an effective amount of an oxidizing agent (e.g., dehydroascorbic acid) to reoxidize unreacted thiol groups, and then purifying the resulting conjugate; (d) Step (c) can optionally be replaced by adding an effective amount of cystine to quench excess conjugates or conjugate / payload complexes containing thiol-reactive groups (e.g., maleimides) and simultaneously or sequentially adding an azide compound (e.g., 4-(azidomethyl)benzoic acid) or a disulfide compound (e.g., cystine) to quench unreacted reducing agent (e.g., TCEP or tris(hydroxypropyl)phosphine). The addition of cystine to quench unreacted reducing agent (e.g., TCEP) can form a cysteine ​​that can simultaneously quench excess conjugates or conjugate / payload complexes containing thiol-reactive groups (e.g., maleimides).

[0302] Formula Zn(NR 1 R 2 R3 ) m1 2+ R in 1 , R 2 and R 3 are each independently, C 1 -C 8 Alkyl, C 2 -C 8 Heteroalkyl, alkylcycloalkyl, heterocycloalkyl;C 3 -C 8 m1 is selected from aryl, Ar-alkyl, heterocycle, carbocycle, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, or heteroaryl. m1 is 1, 2, 3, 4, 5, 6, 7, or 8. Preferably, m1 is 1, 2, 3, or 4.

[0303] In addition, (NR 1 R 2 R 3 ) m1 can form dimers, trimers, tetramers, pentamers, or hexamers, and these polymers are N,R 1 , R 2 and / or R 3 They can be covalently linked to form a heterocycle, a carbocycle, a heterobicycle, or a carbobicycle.

[0304] The transition metal cation-amino complex M(NR 1 R 2 R 3 ) m1 m2+ The concentration of is 0.5 to 20 equivalents per mole of protein and is selected from ethanol, methanol, propanol, propanediol, DMA, DMSO, THF, or CH 3 The reaction solution may be added together with a water-miscible organic solvent selected from CN.

[0305] The reducing agent is an organic phosphine, preferably selected from tris(2-carboxyethyl)phosphine (TECP) or tris(hydroxypropyl)phosphine, and is used in a concentration of 0.02 mM to 1.0 mM in the reaction solution or in an amount of 1.0 to 20 equivalents per mole of protein. The oxidizing agent added in step (c) is DHAA, Fe 3+ , I 2 , Cu 2+ , Mn 3+ , MnO 2 , or Fe 3+ / I - The oxidizing agent is used in the reaction solution at a concentration of 0.02 mM to 1.0 mM or at 1 to 100 equivalents per mole of protein. The optimum pH for the conjugation reaction is usually about 5.0 to 8.0, preferably about 5.5 to 7.5. The optimum temperature for the binding reaction is usually about -5 to about 40°C, preferably about 0 to 37°C, more preferably about 2 to 8°C. The optimum time for the conjugation reaction is usually about 15 minutes to about 48 hours, preferably about 30 minutes to overnight (10 to 16 hours). The optimum reaction conditions (pH, temperature, buffer, reactant concentration, etc.) depend in particular on the antibody-like protein, payload / conjugate-linker complex, reducing agent and / or M(NR 1 R 2 R 3 ) m1 m2+ Depends on.

[0306] In a further embodiment, under homogeneous conjugation method, the obtained conjugate of formula (I), (II), or (III) is more than 75% conjugated to the cysteine ​​sites between the heavy and light chains of the antibody, and less than 15% is conjugated to the cysteine ​​sites (hinge region) between the heavy and heavy chains of the antibody. Typically, for formula (I), (II), or (III), when the drug / antibody ratio (DAR) is set to 4, the percentage distribution of the number of drugs in the antibody is as follows: D0<1%, D2<10%, D4>65%, D6<10%, D8<10%; for formula (VII).

[0307] The resulting conjugate can be purified by standard biochemical methods, such as gel filtration on Sephadex G25 or Sephacryl S300 columns, adsorption chromatography, ion (cation or anion) exchange, or dialysis (ultrafiltration or hyperfiltration (UF) and diafiltration (DF)). In some cases, when a small antibody-like protein (e.g., <100 KD) is conjugated with a small molecule drug as a cell-binding molecule, it can be purified by chromatography, such as HPLC, medium pressure column chromatography, or ion exchange chromatography.

[0308] In general, conjugates of formula (I), (II), or (III) are preferably generated from drug / conjugate conjugates of formula (IV), (V), or (VI) as a one-pot reaction. When a thiol reduced from an antibody-like protein reacts with a thiol-reactive group at the terminus of a drug / conjugate conjugate of formula (IV), (V), or (VI), Ellman's reagent can be optionally used to monitor efficient reduction of disulfide bonds and conjugation of thiols through measurement of the number of free thiols in the reaction. UV spectroscopy at wavelengths ranging from 190 to 390 nm, preferably 240 to 380 nm, more preferably 240 to 330 nm, is preferably used to assist the reaction (through monitoring of conjugation). Thus, the conjugation reaction can be measured or carried out in a quartz cell or Pyrex flask in a temperature-controlled environment. The drug / protein (antibody) ratio (DAR) of the conjugate can also be measured by UV at wavelengths ranging from 240-380 nm, by measuring the integrated area of ​​each drug / protein fragment, by hydrophobic interaction chromatography (HIC-HPLC), or by capillary electrophoresis (CE), and / or LC-MS, LC-MS / MS, or CE-MS (a combination of liquid chromatography (LC) or CE with mass spectrometry (MS) via measuring both the integrated area of ​​LC or CE and the peak intensity of MS for each drug / protein fragment). In the conjugation method of the present invention, if the drug or drug / conjugate complex is not well soluble in the aqueous buffer, up to 30% of a water-miscible (miscible) organic solvent, such as DMA, DMF, ethanol, methanol, acetone, acetonitrile, THF, isopropanol, dioxane, propylene glycol, or ethylenediol, can be added to the aqueous buffer as a co-solvent.

[0309] Aqueous solutions for modification of antibody-like proteins are buffered to a pH between 4-9, preferably 6.0-7.5, and may contain non-nucleophilic buffer salts useful for these pH ranges. Exemplary buffers include phosphate, acetate, triethanolamine HCl, HEPES, and MOPS buffers, and may further include additional components such as cyclodextrin, hydroxypropyl-β-cyclodextrin, polyethylene glycol, sucrose, and salts (e.g., NaCl, KCl). Other biological buffers used in conjugation methods are listed in the definitions section. The progress of the reaction can be monitored by measuring the decrease in absorbance at 252 nm or the increase in absorbance at 280 nm, or changes at other suitable wavelengths. After the reaction is complete, isolation of the modified cell-binding agent can be performed by conventional methods, such as gel filtration chromatography, ion exchange chromatography, adsorption chromatography, or column chromatography on silica gel or alumina, crystallization, preparative thin layer chromatography, ion exchange chromatography, or HPLC.

[0310] When disulfide exchange reactions are used to modify antibody-like proteins, the degree of modification can be assessed by measuring the absorbance of the nitropyridinethione, dinitropyridinedithione, pyridinethione, carboxyamidopyridinedithione, and dicarboxyamidopyridinedithione groups emitted through the UV spectrum. In conjugates without chromophore groups, the modification or conjugation reaction can be monitored by LC-MS, preferably UPLC-QTOF mass spectrometry, or capillary electrophoresis (CEMS). The side chain crosslinkers described herein have a variety of functional groups that can react with any drug, preferably a cytotoxic drug, with the appropriate substituent. For example, modified cell-binding molecules with amino or hydroxy substituents can react with drugs with N-hydroxysuccinimide (NHS) esters, and modified cell-binding molecules with thiol substituents can react with drugs with maleimide or haloacetyl groups. Additionally, modified cell-binding molecules with carbonyl substituents (ketones or aldehydes) can react with drugs with hydrazides or alkyloxyamines. One of skill in the art can readily determine which linker to use based on the known reactivity of the available functional groups on the linker.

[0311] Formulation and Application

[0312] The BCMA antibody conjugates of the present application are suitable to be formulated in a liquid or lyophilized and then reconstituted into a liquid formulation. The conjugate in a liquid formulation or in a formulated lyophilized powder may comprise 0.01% to 99% by weight as the major component in the formulation. In general, a pre-lyophilized liquid formulation containing the conjugate active ingredient at a concentration of 0.1 g / L to 300 g / L for delivery to a patient without high levels of antibody aggregation may contain one or more polyols (e.g., sugars), a buffer having a pH of 4.5 to 7.5, a surfactant (e.g., polysorbate 20 or 80), an antioxidant (e.g., ascorbic acid and / or methionine), an isotonicity agent (e.g., mannitol, sorbitol, or NaCl), a chelating agent such as EDTA, a metal complex (e.g., Zn-protein complex); a biodegradable polymer such as a polyester; a preservative (e.g., benzyl alcohol), and / or free amino acids.

[0313] Buffers suitable for use in the formulation include, but are not limited to, organic acid salts such as sodium, potassium, ammonium, or trihydroxyethylamino salts of citric acid, ascorbic acid, gluconic acid, carbonic acid, tartaric acid, succinic acid, acetic acid, or phthalic acid; Tris, tromethamine hydrochloride, sulfate, or phosphate buffers. Additionally, amino acid cationic components can be used as buffers. Such amino acid components include, but are not limited to, arginine, glycine, glycylglycine, and histidine. Arginine buffers include arginine acetate, arginine chloride, arginine phosphate, arginine sulfate, arginine succinate, and the like. In one embodiment, the arginine buffer is arginine acetate. Examples of histidine buffers include histidine chloride-arginine chloride, histidine acetate-arginine acetate, histidine phosphate-arginine phosphate, histidine sulfate-arginine sulfate, histidine succinate-arginine succinate, and the like. The pH of the buffer formulation is 4.5 to 7.5, preferably about 4.5 to about 6.5, more preferably about 5.0 to about 6.2. In some embodiments, the concentration of the organic acid salt in the buffer is about 10 mM to about 500 mM.

[0314] "Polyols," which are optionally included in the formulation, are substances that have multiple hydroxyl groups. Polyols can be used as stabilizing excipients and / or isotonicity agents in both liquid and lyophilized formulations. Polyols can protect biopharmaceuticals from physical and chemical degradation pathways. Preferentially excluded cosolvents increase the effective surface tension of the solvent at the protein interface, such that the most energetically favorable structural conformation is the one with the smallest surface area. Polyols include sugars (reducing and nonreducing sugars), sugar alcohols, and sugar acids. "Reducing sugars" contain hemiacetal groups that can reduce metal ions or covalently react with lysine and other amino groups of proteins, while "nonreducing sugars" do not possess these properties of reducing sugars. Examples of reducing sugars are fructose, mannose, maltose, lactose, arabinose, xylose, ribose, rhamnose, galactose, and glucose. Nonreducing sugars include sucrose, trehalose, sorbose, melezitose, and raffinose. The sugar alcohol is selected from mannitol, xylitol, erythritol, maltitol, lactitol, erythritol, threitol, sorbitol, and glycerol. The sugar acid includes L-gluconate and its metal salts. The polyol in the liquid formulation or the prepared lyophilized solid can be 0.0% to 20% by weight. Preferably, a non-reducing sugar, sucrose or trehalose, at a concentration of about 0.1% to 15%, is selected in the formulation, with trehalose being preferred over sucrose due to the solution stability of trehalose.

[0315] Surfactants that are optionally included in the formulation include polysorbates (such as polysorbate 20, polysorbate 40, polysorbate 65, polysorbate 80, polysorbate 81, polysorbate 85, etc.), poloxamers (such as poloxamer 188, poly(ethylene oxide)-poly(propylene oxide), poloxamer 407, or polyethylene-polypropylene glycol, etc.); Triton; sodium dodecyl sulfate (SDS); sodium lauryl sulfate; sodium octyl glycoside; lauryl-, myristyl-, linoleyl-, or stearyl-sulfobetaine; lauryl-, myristyl-, linoleyl-, or stearyl-sarcosine; linoleyl-, myristyl-, or cetyl-betaine; lauroamidopropyl-, cocamidopropyl-, oleamide ... The surfactant is selected from the group consisting of propyl-, linoleamidopropyl-, myristamidopropyl-, palmimidopropyl-, or isostearamidopropyl-betaine (e.g., lauroamidopropyl); myristamidopropyl-, palmidopropyl-, or isostearamidopropyl-dimethylamine; sodium methyl cocoyl or sodium methyl oleyl taurate; dodecyl betaine, dodecyl dimethylamine oxide, cocamidopropyl betaine, and cocoamphoglycinate, the "MONAQUAT" (trademark) series (e.g., isostearyl ethylimidonium ethosulfate); polyethyl glycol, polypropylene glycol, and copolymers of ethylene and propylene glycol (e.g., Pluronics, PF68, etc.). Suitable surfactants are polyoxyethylene sorbitan fatty acid esters, such as polysorbate 20, 40, 60, or 80 (Tween 20, 40, 60, or 80). The concentration of the surfactant in the formulation ranges from 0.0% to about 2.0% by weight. In certain embodiments, the concentration of the surfactant is from about 0.01% to about 0.2%. In one embodiment, the concentration of the surfactant is about 0.02%.

[0316] A "preservative" that is optionally included in the formulation is a compound that essentially reduces bacterial activity. Examples of potential preservatives include octadecyl dimethyl benzyl ammonium chloride, hexamethonium chloride, benzalkonium chloride (a mixture of alkyl benzyl dimethyl ammonium chlorides where the alkyl group is a long-chain compound), and benzethonium chloride. Other types of preservatives include aromatic alcohols such as phenol, butyl and benzyl alcohol, alkyl parabens such as methyl or propyl paraben, catechol, resorcinol, cyclohexanol, 3-pentanol, and m-cresol. The preservative in the liquid formulation or in the prepared lyophilized powder can be 0.0% to 5.0% by weight. In one embodiment, the preservative herein is benzyl alcohol.

[0317] Suitable free amino acids as tonicity or osmolality adjusters in bulk materials or formulations are selected from, but not limited to, arginine, cystine, glycine, lysine, histidine, ornithine, isoleucine, leucine, alanine, glycerol glutamic acid or aspartic acid. Basic amino acids, i.e., those containing arginine, lysine, and / or histidine, are preferred. When the composition contains histidine, it can function as both a buffer and a free amino acid, but when a histidine buffer is used, it typically contains a non-histidine free amino acid, e.g., a histidine buffer and lysine. The amino acids may be either D- and / or L-, but are usually in the L-form. The amino acids may be present in any stable salt, e.g., hydrochloride salts such as arginine-HCl. The amino acids in the liquid formulation or the prepared lyophilized powder can be 0.0% to 30% by weight.

[0318] The antioxidant in the formulation can optionally be selected from methionine, glutathione, cysteine, cystine, or ascorbic acid at a concentration of up to about 5 mg / ml in the liquid formulation or 0.0% to 5.0% by weight in the formulated lyophilized powder. The formulation can optionally include a metal chelator, e.g., EDTA, EGTA, etc., at a concentration of up to about 2 mM in the liquid formulation or 0.0% to 0.3% in the formulated lyophilized powder.

[0319] The final formulation should be prepared by adding buffer adjusters (e.g., HCl, H 2 SO 4 , Acetic acid, H 3 PO 4 , citric acid, or NaOH, KOH, NH 4 The preferred pH can be adjusted with a base such as ethanolamine, diethanolamine, triethanolamine, sodium phosphate, potassium phosphate, trisodium citrate, tromethamine, etc. The formulation must be controlled to be "isotonic". This means that the formulation has essentially the same osmolarity as human blood. Isotonic formulations generally have an osmolarity of about 250-350 mOsm. Isotonicity can be measured, for example, using a vapor pressure or ice type osmometer. The isotonic agent is selected from mannitol, sorbitol, sodium acetate, potassium chloride, sodium phosphate, potassium phosphate, trisodium citrate, or NaCl. In general, both the buffer salt and the isotonic agent can account for up to 30% by weight in the formulation.

[0320] Other excipients that may be useful in either the liquid or lyophilized formulations of the patent application include, for example, fucose, cellobiose, maltotriose, melibiose, octulose, ribose, xylitol, arginine, histidine, glycine, alanine, methionine, glutamic acid, lysine, imidazole, glycylglycine, mannosylglycerate, Triton X-100, Pluoronic F-127, cellulose, cyclodextrin, (2-hydroxypropyl)-β-cyclodextrin, dextran (10, 40 or 70 kD), polydextrose, maltodextrin, ficoll, gelatin, hydroxypropyl methacrylate, sodium phosphate, potassium phosphate, ZnCl 2 , zinc, zinc oxide, sodium citrate, trisodium citrate, tromethamine, copper, fibronectin, heparin, human serum albumin, protamine, glycerin, glycerol, EDTA, metacresol, benzyl alcohol, phenol, polyhydric alcohols or polyhydric alcohols, hydrogenated forms of carbohydrates having a carbonyl group reduced to a primary or secondary hydroxyl group.

[0321] Other contemplated excipients that may be utilized in the aqueous pharmaceutical compositions of the present patent application include, for example, flavoring agents, antimicrobial agents, sweeteners, antioxidants, antistatic agents, lipids such as phospholipids or fatty acids, steroids such as cholesterol, protein excipients such as serum albumin (human serum albumin), recombinant human albumin, gelatin, casein, salt forming counterions such as sodium, etc. These and additional known pharmaceutical excipients and / or additives suitable for use in the formulations of the present invention are described in, for example, "The Handbook of Pharmaceutical Excipients, 4 th edition, Rowe et al., Eds., American Pharmaceuticals Association (2003); and Remington: The Science and Practice of Pharmacy, 21 thedition, Gennaro, Ed., Lippincott Williams & Wilkins (2005).”

[0322] A pharmaceutical container or container is used to hold any pharmaceutical formulation of the conjugate of the patent application. The container is a vial, a bottle, a prefilled syringe, a prefilled, or an autoinjector syringe. Liquid formulations can be lyophilized or drum dried into cake or powder form in borosilicate or soda lime glass vials. Solid powders can also be prepared by efficient spray drying and then packaged in vials or pharmaceutical containers for storage and distribution.

[0323] In a further embodiment, the present invention provides a method for preparing a formulation, comprising the steps of: (a) lyophilizing a formulation comprising a conjugate, an excipient, and a buffer system; and (b) reconstituting the lyophilized mixture of step (a) in a reconstitution medium such that the reconstituted formulation is stable. The formulation of step (a) may further comprise a stabilizer and one or more excipients selected from the group comprising bulking agents, salts, surfactants, and preservatives described above. As the reconstitution medium, some diluted organic acid or water, i.e., sterile water or bacteriostatic water for injection (BWFI), can be used. The reconstitution medium can be selected from the group consisting of water, i.e., sterile water, bacteriostatic water for injection (BWFI), or acetic acid, propionic acid, succinic acid, sodium chloride, magnesium chloride, an acidic solution of sodium chloride, an acidic solution of magnesium chloride, and an acidic solution of arginine, in an amount of about 10 to about 250 mM.

[0324] Liquid pharmaceutical formulations of the present conjugates should exhibit various predefined properties. One of the main concerns for liquid pharmaceuticals is stability, since proteins / antibodies tend to form soluble and insoluble aggregates during preparation and storage. Furthermore, various chemical reactions (deamination, oxidation, clipping, isomerization, etc.) can occur in solution, leading to increased levels of degradation products and loss of biological activity. Preferably, the conjugates in liquid or lyophilized formulations should exhibit a shelf life of more than 6 months at 25°C. More preferably, the conjugates in either liquid or lyophilized formulations should exhibit a shelf life of more than 12 months at 25°C. Most preferred liquid formulations should exhibit a shelf life of about 24-36 months at 2-8°C, and lyophilized formulations should exhibit a shelf life of up to about 60 months at 2-8°C. Both liquid and lyophilized formulations should exhibit a shelf life of at least 2 years at -20°C or -70°C.

[0325] In certain embodiments, the formulation is stable after freezing (e.g., at -20°C or -70°C) and thawing, e.g., after one, two, or three freeze-thaw cycles. Stability can be qualitatively and / or quantitatively assessed by a variety of methods, such as assessment of drug / antibody (protein) ratio and aggregate formation (e.g., using ultraviolet, size exclusion chromatography, measuring turbidity, and visual inspection, etc.); assessment of charge heterogeneity using cation exchange chromatography, electric focusing or capillary zone electrophoresis such as image capillaries; amino-terminus or carboxyl-terminus sequence analysis; mass spectrometry, or matrix-assisted laser desorption / ionization / time-of-flight mass spectrometry (MALDI / TOF MS), or HPLC-MS / MS; SDS-PAGE analysis for comparison of antibody reduction or integrity; analysis of peptide maps (e.g., trypsin or lysine-carbon); evaluation of antibody biological activity or antigen-binding function; etc. Instability can include any one or more of the following: aggregation, deamidation (e.g., Asn deamidation), oxidation (e.g., Met oxidation), isomerization (e.g., Asp isomerization), shearing / hydrolysis / cleavage (e.g., hinge rupture), succinimide formation, unpaired cysteines, N-terminal extensions, C-terminal processing, and catabolic glycosylation.

[0326] A stable conjugate should also "retain its biological activity" in the pharmaceutical formulation, e.g., the biological activity of the conjugate at a particular time, e.g., 12 months, should be within about 20%, preferably within about 10% (within the error of the assays) of the biological activity at the time the pharmaceutical formulation is prepared, as determined by, e.g., antigen binding assays and / or in vitro cytotoxicity assays.

[0327] For in vivo clinical application, the conjugate of the present invention may be provided in the form of a solution or in the form of a lyophilized solid that can be redissolved in sterile water for injection. Examples of suitable conjugate administration methods are as follows: The conjugate is administered as an intravenous bolus once daily, weekly, every two weeks, every three weeks, every four weeks, or once a month for 8 to 108 weeks. The bolus dose is dissolved in 50 to 1000 mL of saline, to which human serum albumin can be optionally added (e.g., 0.5 to 1 ml of concentrated human serum albumin solution at 100 mg / ml). The drug dosage is about 50 μg to 20 mg / kg body weight per week, and is administered intravenously (each injection in the range of 10 μg to 200 mg / kg). After 4 to 108 weeks of treatment, the patient can receive a second course of treatment. The detailed treatment method, including the route of administration, excipients, diluents, dosage, and treatment period, can be determined by an experienced surgeon.

[0328] Examples of treating diseases by selectively killing cell populations by in vivo or ex vivo methods include any type of cancer, autoimmune diseases, transplant rejection, and infectious diseases (including viral, bacterial, or parasitic).

[0329] The amount of conjugate required to achieve a desired biological effect will vary depending on many factors, including the chemical properties, potency and bioavailability of the conjugate, the type of disease, the patient's race, the patient's pathological condition, the route of administration, all of which determine the required dosage, delivery and regimen.

[0330] In general terms, the conjugate via the linker of the present invention may be a formulation for parenteral administration dissolved in a physiological buffer so as to contain the conjugate at a concentration of 0.1 to 10% w / v. Typical dosage ranges are 1 μg / kg body weight to 0.1 g / kg body weight per day, week, 2 weeks, 3 weeks, or month, with preferred dosage ranges being 0.01 mg / kg body weight to 25 mg / kg body weight per week, 2 weeks, 3 weeks, or month. The preferred drug dosage may suitably depend on variables such as, for example, the type and extent of progression of the disease or disorder, the overall health of the individual patient, the relative biological activity of the selected drug, the dosage form of the compound, the mode of administration (intravenous, intramuscular, or other), the pharmacokinetic properties of the drug in the selected mode of administration, and the administration rate (single injection or continuous infusion) and administration schedule (frequency of administration within a given time period).

[0331] In some embodiments, when the reconstituted conjugate is injected subcutaneously, into muscle, or into other tissues of the body, hyaluronidase (HAase) is preferably administered with the conjugate, where hyaluronidase is used to help absorb the injected conjugate into the patient's body. Hyaluronidase is used synergistically at doses of 20-200 units, preferably 60-160 units.

[0332] The conjugates of the present invention may be administered in unit doses, where "unit dose" refers to a single dose administered to one patient, and may be used in simple and convenient packaging, while maintaining a physically and chemically stable unit dose, either as the active conjugate itself or as a pharma- ceutically acceptable composition, as described below. Thus, a typical daily / weekly / biweekly / monthly total dosage range is 0.01-100 mg / kg. As a general guide, unit doses for humans range from 1-3000 mg per day, week, biweekly, 3 weeks, or month. Preferably, the unit dosage range is 1 mg-500 mg administered 1-4 times per month, more preferably 1 mg-100 mg administered once per week, once every 2 weeks, or once every 3 weeks. The conjugates provided herein may be prepared by adding one or more pharma- ceutically acceptable excipients to a pharmaceutical composition. The unit dosage of the drug can be administered orally as a tablet, simple capsule or soft capsule; for intranasal administration as a powder, nasal drops or aerosol; or for dermal administration as an ointment, cream, lotion, gel or spray or skin patch. The composition can be conveniently administered in unit dosage form and can be prepared by any method known in the pharmaceutical art, for example, as described in "Remington: The Science and Practice of Pharmacy, 21st ed.; Lippincott Williams & Wilkins: Philadelphia, PA, 2005".

[0333] Preferred formulations include pharmaceutical compositions in which the compounds of the present invention are formulated for oral or parenteral administration. For example, oral dosage forms such as tablets, pills, powders, capsules, lozenges, etc. may contain one or more of the following ingredients or other compounds of similar nature: binders such as microcrystalline cellulose or gum tragacanth; diluents such as starch or lactose; dispersants such as starch and cellulose derivatives; lubricants such as magnesium stearate; lubricants such as colloidal silica; sweeteners such as sucrose or saccharin; flavorings such as peppermint or methyl salicylate. Capsules are available in hard or soft form, mixed with gelatin or optionally mixed with plasticizers, as are starch capsules. Furthermore, dosage units may contain various different materials, such as sugar coatings, shellacs, or enteric coatings, to modify their physical form. Other oral dosage forms, such as syrups or elixirs, may contain sweeteners, preservatives, pigments, colorants, and flavorings. Additionally, the active compound can be processed and formulated to produce fast dissolving, sustained release or extended release dosage forms, preferably such sustained release formulations being dual mode.A preferred tablet formulation contains a mixture of lactose, cornstarch, magnesium silicate, croscarmellose sodium, povidone, magnesium stearate, or talc, or combinations thereof.

[0334] Liquid preparations for parenteral administration include sterile aqueous or non-aqueous solutions, suspensions and emulsions. Liquid preparations may contain binders, buffers, preservatives, chelating agents, sweeteners, flavorings and coloring agents, and the like. Non-aqueous solvents include alcohol, propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and organic acid esters such as ethyl oleate. Aqueous carriers include mixtures of water and ethanol, buffers and saline. In particular, biocompatible, biodegradable lactide polymers, lactide / glycolide copolymers, or polyoxyethylene / polyoxypropylene copolymers can be used as materials to control the release of active compounds. Vehicles for intravenous administration include fluid and nutrient replenishers and electrolyte replenishers based on Ringer's dextrose, and the like. Other possible parenteral delivery systems for these active compounds include ethylene-vinyl acetate copolymer particles, osmotic pumps, implantable infusion systems, and liposomes.

[0335] Alternative methods of administration include inhalants, which include dry powders, aerosols and drops. Inhalants may be, for example, aqueous solutions containing polyoxyethylene-9-lauryl ether, glycocholate, deoxycholate, or oily solutions for administration in the form of nasal drops or as a gel for intranasal application. Formulations for intraoral administration, for example tablets or lozenges, may contain flavorings such as sucrose or gum arabic and other additives such as glycocholate. Formulations suitable for rectal administration are preferably given as unit-dose suppositories, which may contain, for example, a solid base such as cocoa butter and may also contain salicylic acid. Topical skin preparations are preferably in the form of ointments, creams, lotions, pastes, gels, sprays, aerosols or oils. Bases include petrolatum, lanolin, polyethylene glycols and alcohols, and mixtures thereof. Formulations suitable for transdermal administration may be given as separate patches and may be lipophilic emulsions or buffered aqueous solutions, dissolved or dispersed in a polymer or adhesive.

[0336] Further, in another embodiment, a pharmaceutical composition comprising a therapeutically effective amount of a conjugate of formula (I), (II), (III), or any conjugate described throughout this patent, can be coadministered with other therapeutic agents, such as chemotherapeutic agents, radiotherapeutic agents, immunotherapeutic agents, autoimmune disorder agents, anti-infective agents, or other conjugates, for synergistically effective treatment or prevention of cancer, autoimmune disease, or infectious disease. As used herein, the term "coadministration" refers to administration of one or more additional therapeutic agents and an antibody or ADC, or an antibody or ADC-containing composition described herein, close enough in time, or vice versa, that the antibody or ADC can have an enhanced effect. In this regard, the antibody or ADC, or a composition comprising same, can be administered first and one or more additional therapeutic agents administered second, or vice versa. For example, an antibody or ADC, or a composition comprising same, can be administered in combination with another agent (e.g., as an adjuvant) for the treatment or prevention of multiple myeloma. In this regard, the antibody or ADC, or antibody- or ADC-containing composition, can be used in combination with at least one other anti-cancer agent, including, for example, any suitable chemotherapeutic agent known in the art, ionizing radiation, small molecule anti-cancer agents, cancer vaccines, biological therapies (e.g., other monoclonal antibodies, cancer-killing viruses, gene therapy, and adoptive T cell transfer), and / or surgery. In one embodiment, the synergist or radiation therapy can be administered at least 1 hour, 12 hours, 1 day, 1 week, 2 weeks, 3 weeks, 1 month, or in further embodiments, several months prior to or after administration of the conjugate.

[0337] Said synergist is preferably selected from one or several of the following drugs: abatacept, abiraterone acetate, abraxane, acetaminophen / hydrocodone, acalabrutinib, aducanumab, adalimumab, ADXS31-142, ADXS-HER2, afatinib dimaleate, aldesleukin, alectinib, alemtuzumab, alitretinoin, adotrastuzumab emtansine, amphetamine / dextroamphetamine, anastrozole, aripiprazole, anthracycline, aripiprazole, atazanavir, atezolizumab, atorvastatin, avelumab, axicabtagene ciloleucel, axitinib, belinostat, live BCG, zumab, bexarotene, blinatumomab, bortezomib, bosutinib, brentuximab Vedotin, brigatinib, budesonide, budesonide / formoterol, buprenorphine, cabazitaxel, cabozantinib, capmatinib, capecitabine, carfilzomib, chimeric antigen receptor-modified T (CAR-T) cells, celecoxib, ceritinib, cetuximab, chidamide, cyclosporine, cinacalcet, crizotinib, cobimetinib, cosentyx, crizotinib, CTL01 9. Dabigatran, dabrafenib, dacarbazine, daclizumab, dacomitinib, daptomycin, daratumumab, darbepoetin alfa, darunavir, dasatinib, denileukin diftitox, denosumab, depakote, dexlansoprazole, methylphenidate, dexamethasone, dinutuximab, doxycycline, duloxetine, duvelisib, durval Umab, elotuzumab, emtricitabine / rilpivirine / tenofovir, disoproxil fumarate, emtricitabine / tenofovir / efavirenz, enoxaparin, ensartinib, enzalutamide, epoetin alfa, erlotinib, esomeprazole, eszopiclone, everolimus, exemestane, everolimus, exenatide ER, ezetimibe, ezetimibe / simvastatin, fenofibrate, filgrastim, fingolimod, fluticasone propionate, fluticasone / salmeterol, fulvestrant, gaziva, gefitinib, glatiramer, goserelin acetate, icotinib, imatinib, ibritumomabTiuxetan, ibrutinib, idelarib, ifosfamide, infliximab, imiquimod, Immucyst, immune BCG, iniparib, insulin aspart, insulin detemir, insulin glargine, insulin lispro, interferon alpha, interferon alpha 1b, interferon alpha 2a, interferon alpha alpha 2b, interferon beta, interferon beta 1a, interferon beta 1b, interferon gamma 1a, iapatinib, ipilimumab, ipratropium bromide / salbutamo oxal, ixazomib, Kanuma, lanreotide acetate, lenalidomide, lenalidomide, lenvatinib mesylate, letrozole, levothyroxine, lidocaine, linezolid, liraglutide, examfetamine, LN-144, lorlatinib, memantine, methylphenidate, metoprolol, mekinist, melicitabine / rilpivirine / tenofovir, modafinil, mometasone, misidac-C, necitumumab, neratinib, nilotinib, niraparib, nivolumab, ofatumumab, obinutuzumab, ora Paribu, olmesartan, olmesartan / hydrochlorothiazide, omalizumab, omega-3 fatty acid ethyl ester, oncoline, oseltamivir, osimertinib, oxycodone, palbociclib, palivizumab, panitumumab, panobinostat, pazopanib, pembrolizumab, PD-1 antibody, PD-L1 antibody, pemetrexed, pertuzumab, pneumococcal conjugate vaccine, pomalidomide, poziotinib, pregabalin, proskavax, propranolol, quetiapine, rabeprazole, radium chloride M223, raloxifene, raltegravir, ramucirumab, ranibizumab, regorafenib, rituximab, rivaroxaban, romidepsin, rosuvastatin, ruxolitinib phosphate, salbutamol, savolitinib, semaglutide, sildenafil, siltuximab, sipulcel-T, sitagliptin, sitagliptin / metformin, solifenacin, solanezumab, sonidegib, sorafenib, sunitinib, tacrolimus, taclimus, tadalafil, tamoxifen, tafinlar, talimogene laherparepvec, talazoparib, telaporebrib, IR / emtricitabine, tenofovirDisoproxil fumarate, testosterone gel, thalidomide, TICE BCG, tiotropium bromide, tisagencleucel, toremifene, trametinib, trastuzumab, trastuzumab deruxtecan, trabectedin (ecteinascidin 743), trametinib, tremelimumab, trifluridine / tipiracil, tretinoin, ustekinumab, valsartan, veliparib, vandetanib, vemurafenib, venetoclax, vorinostat, dibuaflibercept, zostavax, and their analogues, derivatives, pharma- ceutically acceptable salts, carriers, diluents or excipients, or combinations thereof.

[0338] In some embodiments, the present disclosure also provides a composition comprising the above-mentioned antibody or antibody-drug conjugate and a pharma- ceutically acceptable (e.g., physiologically acceptable) carrier. Any suitable carrier known in the art can be used within the scope of the present invention. The choice of carrier will be determined, in part, by the particular site to which the composition may be administered and the particular method used to administer the composition. The composition may optionally be sterile. The composition may be produced according to conventional techniques, for example, as described in Remington: The Science and Practice of Pharmacy, 21st Edition, Lippincott Williams & Wilkins, Philadelphia, PA (2001).

[0339] The compositions of the invention desirably comprise an amount of an antibody or ADC effective to treat or prevent multiple myeloma. Thus, the present disclosure provides a method of killing multiple myeloma cells, comprising contacting a multiple myeloma cell expressing BCMA with an antibody or ADC as described herein, or with a composition comprising an antibody or ADC as described herein, whereby the antibody or ADC binds to BCMA on the multiple myeloma cell and kills the multiple myeloma cell. The present disclosure also provides the use of an antibody or ADC as described herein, or a composition comprising an antibody or ADC, in the manufacture of a medicament for treating multiple myeloma. As discussed herein, multiple myeloma, also known as plasma cell myeloma or Köhler's disease, is a cancer of plasma cells, a type of white blood cell that is normally responsible for the production of antibodies (Raab et al., Lancet, 374: 324-329 (2009)). Multiple myeloma affects 1-4 per 100,000 people annually. For reasons that remain unclear, the disease is more prevalent in men and is generally twice as prevalent in African-Americans as in white Americans. Multiple myeloma is the least common hematologic malignancy (14%), accounting for 1% of all cancers. Treatment for multiple myeloma typically involves high-dose chemotherapy followed by hematopoietic stem cell transplantation (allogeneic or autologous). However, multiple myeloma patients who undergo such treatment generally experience a high relapse rate. As discussed above, BCMA is highly expressed by multiple myeloma cells.

[0340] As demonstrated herein, BCMA is also expressed on multiple myeloma stem cells. Thus, the present disclosure provides a method of killing multiple myeloma stem cells, comprising contacting multiple myeloma stem cells expressing BCMA with a composition comprising an antibody-drug conjugate as described herein, or an ADC as described herein, whereby the antibody-drug conjugate binds to BCMA on the multiple myeloma stem cells and kills the multiple myeloma stem cells. Multiple myeloma stem cells can be identified in the bone marrow of multiple myeloma patients by the surface expression of CD19 and the lack of surface expression of CD138 (see, e.g., Matsui et al., Blood, 103:2332-6 (2004)). These cells have unique clonogenicity and engraft in immunodeficient mice, whereas myeloma plasma cells, defined as CD138+CD19-, do not. Multiple myeloma stem cells are also resistant to current therapies (Matsui et al., Cancer Res., 68: 190-7 (2008)). Thus, the present invention provides a method of treating a patient having or at risk of having a BCMA-expressing cancer, comprising administering to the patient an effective regimen of a BCMA antibody or BCMA ADC as described above. Optionally, the cancer is a hematological cancer. Optionally, the hematological cancer is a myeloma, leukemia, or lymphoma. Optionally, the hematological cancer is multiple myeloma. Optionally, the hematological cancer is non-Hodgkin's lymphoma (NHL) or Hodgkin's lymphoma. Optionally, the hematological cancer is myelodysplastic syndrome (MDS), myeloproliferative syndrome (MPS), Waldenstrom's macroglobulinemia, or Burkett's lymphoma.

[0341] As used herein, the terms "treatment", "treating" and the like refer to obtaining a desired pharmacological and / or physiological effect. Preferably, the effect is therapeutic, i.e., the effect partially or completely cures the disease and / or adverse symptoms resulting from the disease. To this end, the method of the invention includes administering a "therapeutically effective amount" of an antibody or ADC, or a composition comprising an antibody or ADC and a pharma- ceutically acceptable carrier. A "therapeutically effective amount" refers to an amount effective, at dosages and for periods of time necessary, to achieve a desired therapeutic result. A therapeutically effective amount may vary depending on factors such as the individual's condition, age, sex, weight, ability of the antibody or ADC to elicit a desired response in the individual, and the like. For example, a therapeutically effective amount of an ADC of the invention is an amount that binds to and destroys BCMA on multiple myeloma cells.

[0342] The pharmacological and / or physiological effect of the treatment may be prophylactic, i.e., the effect may be to completely or partially prevent the disease or its symptoms. In this regard, the method of the invention comprises administering a "prophylactically effective amount" of an ADC or a composition comprising an ADC to a mammal predisposed to multiple myeloma. A "prophylactically effective amount" refers to an amount effective, at dosages and for periods of time necessary, to achieve the desired prophylactic result (e.g., prevention of disease onset). The therapeutic or prophylactic effect may be monitored by periodic evaluation of the treated patient. In one embodiment, the ADC described herein inhibits or suppresses the proliferation of BCMA-expressing myeloma cells by at least about 10% (e.g., at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, or at least about 100%). Cell proliferation can be measured using any suitable method known in the art, such as measuring the incorporation of labeled nucleosides (e.g., 3H-thymidine or bromodeoxyuridine Brd(U)) into genomic DNA (see, e.g., Madhavan, HN, J. Stem Cells Regen. Med. 3(1): 12-14 (2007)).

[0343] In some embodiments, the BCMA antibody and BCMA ADC invention further provides a method of treating a patient having or at risk of having an immune disorder mediated by immune cells expressing BCMA by administering to the patient any of the above antibodies or ADCs in an effective dosing regimen. Optionally, the disorder is a B cell mediated disorder. Optionally, the immune disorder is rheumatoid arthritis, systemic lupus E (SLE), type I diabetes, asthma, atopic dermatitis, allergic rhinitis, thrombocytopenic purpura, multiple sclerosis, psoriasis, Sjogren's syndrome, Hashimoto's thyroiditis, Graves' disease, primary biliary cirrhosis, Wegener's granulomatosis, tuberculosis, and graft-versus-host disease.

[0344] In some embodiments, the BCMA antibody and BCMA ADC invention further provides methods of treating a patient having or at risk of having cancer, an autoimmune disease, an infectious disease, a viral disease, or a pathogenic infection by administering to the patient any of the above antibodies or ADCs, or by administering to the patient any of the above antibodies or ADCs in combination with other therapeutic agents, such as chemotherapeutic agents, radiation therapy, immunotherapy agents, autoimmune disorder agents, anti-infective agents, or other conjugates, in an effective dosing regimen.

[0345] Target cancers include, but are not limited to, adrenal cortical carcinoma, anal cancer, bladder cancer, brain tumors (adult, brain stem glioma, childhood, cerebellar astrocytoma, cerebral astrocytoma, ependymoma, medulloblastoma, supratentorial primitive neuroectodermal and pineal tumors, visual pathway and hypothalamic gliomas), breast cancer, carcinoid tumors, gastrointestinal, carcinoma of unknown primary site, cervical carcinoma, colorectal carcinoma, endometrial cancer, esophageal cancer, extrahepatic bile duct cancer, Ewing family tumors Tumors (PNET), Extracranial Malignant Germ Cell Tumors, Eye Cancer, Intraocular Melanoma, Gallbladder Cancer, Gastric Cancer (Stomach), Germ Cell Tumors, Extragonadal, Gestational Trophoblastic Tumor, Head and Neck Cancer, Hypopharyngeal Cancer, Pancreatic Islet Cell Carcinoma, Kidney Cancer (Renal Cell Carcinoma), Laryngeal Carcinoma, Leukemia (Acute Lymphoblastic, Acute Myeloid, Chronic Lymphocytic, Chronic Myeloid, Hairy Cell), Lip and Oral Cavity Cancer, Liver Cancer, Lung Cancer (Non-Small Cell, Small Cell, Lymphoma (AIDS-Related, Central Nervous System, Cutaneous T Cell) , Hodgkin's disease, non-Hodgkin's disease, malignant mesothelioma, melanoma, Merkel cell carcinoma, metastatic squamous neck cancer of unknown primary site, multiple myeloma and other plasma cell neoplasms, mycosis fungoides, myelodysplastic syndromes, myeloproliferative disorders, nasopharyngeal cancer, neuroblastoma, oral cancer, pharyngeal cancer, osteosarcoma, ovarian cancer (epithelial, germ cell tumors, low malignant potential tumors), pancreatic cancer (exocrine gland, islet cell carcinoma), paranasal sinus and nasal cancer, parathyroid cancer, penile cancer, brown These include chromocytoma cancer, pituitary cancer, plasmacytoma, prostate cancer, rhabdomyosarcoma, rectal cancer, renal cell carcinoma (kidney cancer), renal pelvis and ureter (transitional cell), salivary gland cancer, Sezary syndrome, skin cancer, skin cancer (cutaneous T-cell lymphoma, Kaposi's sarcoma, melanoma), small intestine cancer, soft tissue sarcoma, stomach cancer, testicular cancer, thymoma (malignant), thyroid cancer, urethral cancer, uterine cancer (sarcoma), abnormal cancer of children, vaginal cancer, vulvar cancer, and Wilms' tumor.

[0346] The autoimmune diseases include, but are not limited to, autoimmune achlorhydria chronic active hepatitis, acute disseminated encephalomyelitis, acute hemorrhagic leukoencephalitis, Addison's disease, agammaglobulinemia, alopecia areata, amyotrophic lateral sclerosis, ankylosing spondylitis, anti-GMB / TBM nephritis, antiphospholipid syndrome, antisynthetase syndrome, arthritis, atopic allergy, atopic dermatitis, autoimmune aplastic anemia, autoimmune cardiomyopathy, autoimmune hemolytic anemia, autoimmune hepatitis, autoimmune inner ear disease, autoimmune lymphoproliferative syndrome, autoimmune peripheral nervous system disease, autoimmune pancreatitis, multiple myelopathy, ... Autoimmune endocrinopathies types I, II, III, autoimmune progesterone dermatitis, autoimmune thrombocytopenic purpura, autoimmune uveitis, Barlow's disease / Barlow's concentric sclerosis, Behçet's disease, Berger's disease, Bickerstaff encephalitis, Blau syndrome, bullous pemphigoid, Castleman's disease, Chagas disease, chronic fatigue immune dysfunction syndrome, chronic inflammatory demyelinating polyneuropathy, chronic relapsing multifocal osteomyelitis, chronic Lyme disease, chronic obstructive pulmonary disease, allergic granulomatous vasculitis, cicatricial pemphigoid, celiac disease, Cogan's syndrome, cold agglutinin disease, complement component C Type 2 deficiency, cranial arteritis, CREST syndrome, Crohn's disease (idiopathic inflammatory bowel disease), Cushing's syndrome, cutaneous leukocytoclastic vasculitis, malignant atrophic papulosis, painful adiposity, dermatitis herpetiformis, dermatomyositis, type 1 diabetes mellitus, diffuse cutaneous scleroderma, myocardial infarction, discoid lupus erythematosus, eczema, endometriosis, enthesitis-associated arthritis, eosinophilic fasciitis, epidermolysis bullosa acquisita, erythema nodosum, idiopathic mixed cryoglobulinemia, Evans syndrome, fibrodysplasia ossificans progressiva, fibromyalgia, fibromyositis, fibrosing alveolitis, gastritis, gastrointestinal pemphigoid, giant cell arteritis, glomerulonephritis, Goodpasture's disease syndrome, Graves' disease, Guillain-Barre syndrome, Hashimoto's encephalopathy, Hashimoto's thyroiditis, hemolytic anemia, allergic purpura, herpes gestationis, hidradenitis suppurativa, Hughes' syndrome (antiphospholipid syndrome), hypogammaglobulinemia, idiopathic inflammatory demyelinating disease, idiopathic pulmonary fibrosis, idiopathic thrombocytopenic purpura (autoimmune thrombocytopenic purpura), IgA nephropathy (Berger's disease), inclusion body myositis, inflammatory demyelinating polyneuropathy, interstitial cystitis, irritable bowel syndrome, juvenile idiopathic arthritis, juvenile rheumatoid arthritis, mucocutaneous lymph node syndrome, Lambert-Eaton myasthenic syndrome, leukocytoclastic vasculitis,Lichen planus, Lichen sclerosus, Linear IgA disease (LAD), Lou Gehrig's disease (amyotrophic lateral sclerosis), Lupus-like hepatitis, Lupus erythematosus, Blau syndrome, Meniere's disease, Microscopic polyangiitis, Miller Fisher syndrome, Mixed connective tissue disease, Scleroderma, Mucha-Jakob disease, Muckle-Wells syndrome, Multiple myeloma, Multiple sclerosis, Myasthenia gravis, Myositis, Narcolepsy, Neuromyelitis optica (Devic's disease), Neuromuscular, Ocular cicatricial Pemphigoid, opsoclonus-myoclonus syndrome, Ord's thyroiditis, relapsing rheumatism, Panda syndrome (childhood autoimmune neuropsychiatric disorder complicated by streptococcal infection), neoplastic cerebellar degeneration, paroxysmal nocturnal hemoglobinuria, Parry-Romberg syndrome, Parsonage-Georgia syndrome, pars plana inflammation, pemphigus, pemphigus vulgaris, pernicious anemia, perivenous encephalomyelitis, POEMS syndrome, polyarteritis nodosa, polymyalgia rheumatica, polymyositis, genotype Primary biliary cirrhosis, primary sclerosing cholangitis, progressive inflammatory neuropathy, psoriasis, psoriatic arthritis, pyoderma gangrenosum, pure red cell aplastic anemia, Rasmussen's encephalitis, Raynaud's disease, relapsing polychondritis, Reiter's syndrome, restless legs syndrome, retroperitoneal fibrosis, rheumatoid arthritis, rheumatic fever, sarcoidosis, schizophrenia, Schmidt's syndrome, Schnitzler's syndrome, scleritis, scleroderma, Sjogren's syndrome, spondyloarthropathy, viscous hematoma group, Still's disease, stiff-man syndrome, subacute bacterial endocarditis, Susac syndrome, acute febrile neutrophilic dermatosis, Sydenham's chorea, sympathetic ophthalmia, Takayasu's arteritis, temporal arteritis (giant cell arteritis), Tolosa-Hunt syndrome, transverse myelitis, ulcerative colitis (idiopathic inflammatory bowel disease), undifferentiated connective tissue disease, undifferentiated spondyloarthropathy, vasculitis, vitiligo, Wegener's granulomatosis, Wilson's syndrome, and Westcott-Aldrich syndrome.

[0347] Such infections include, but are not limited to, Acinetobacter infections, Actinomycosis, African sleeping sickness (African trypanosomiasis), AIDS (Acquired Immune Deficiency Syndrome), amebiasis, Anaplasmosis, Bacillus anthracis, bacterial tuberculosis infection, Argentine hemorrhagic fever, Ascariasis, Aspergillosis, Astrovirus infection, Babesiosis, Bacillus cereus infection, bacterial pneumonia, bacterial vaginosis, Bacteroides infection, Balantidiosis, Bailey's roundworm infection, BK virus infection, Black sand worm, Blastocystis hominis infection, Blastomyces, Bolivian hemorrhagic fever, Borrelia infection, Botulinum toxin infection, and the like. Infections (and infant botulism), Brazilian hemorrhagic fever, brucellosis, Burkholderia infection, Buruli ulcer, Calicivirus infection (Norovirus, Sapovirus), Campylobacter infection, Candida infection (candidiasis, thrush), cat scratch disease, cellulitis, Chagas disease (American trypanosomiasis), chancroid, chickenpox, Clostridium difficile infection, pneumonia, cholera, chromomycosis, Clonorchiasis, Clostridium difficile infection, coccidioidomycosis, Colorado tick fever, cold (acute viral nasopharyngitis, acute rhinitis), Creutzfeldt-Jakob disease, Limia-Congo hemorrhagic fever, Cryptococcus, Cryptosporidium, cutaneous larval migration, Cyclospora infection, Cysticercosis, Cytomegalovirus infection, Dengue fever, Dinula amebiasis, Diphtheria, Diphyllobothriasis, Guinea worm disease, Ebola hemorrhagic fever, Hydatid disease, Ehrlichiosis, Pinworms (pinworm infection), Enterococcus infection, Enterovirus infection, Typhus, Erythema infectiosum (fifth disease), acute exanthema in children, Fasciola fascioliasis, Hepatic malaria, Fatal familial insomnia, Filariasis, Food poisoning caused by Clostridium perfringens, Nonparasitic amoeba infection, Fusobacterium infection, Gas gangrene (Clostridium perfringens) rhizal myonecrosis), diotrichosis, Gerstmann-Sträussler-Scheinker syndrome, Giardiasis, glanders, gnathostomiasis, gonorrhea, inguinal granuloma (donovanosis), group A streptococcal infection, group B streptococcal infection, Haemophilus influenzae infection, hand, foot and mouth disease (HFMD), Hantavirus pulmonary syndrome, Helicobacter pylori infection, hemolytic uremic syndrome, hemorrhagic fever with renal syndrome, hepatitis A, hepatitis B, hepatitis C, hepatitis D, hepatitis E, herpes simplex, histoplasmosis, hookworm infection, human Balkan virus infection, human ehrlichiosis Evans, human granulocytic anaplasmosis,Human metapneumovirus infection, human monocytic amycosis, human papillomavirus infection, human parainfluenza virus infection, taeniasis, influenza, isosporiasis, Kawasaki disease, mononucleosis, Mycobacterium kurui, Lassa fever, Legionnaires' disease, Legionnaires' disease, leishmaniasis, leprosy, leptospirosis, listeriosis, Lyme disease, lymphatic filariasis, lymphocytic choriomeningitis, malaria, Marburg hemorrhagic fever, measles, melioidosis (Whitmore disease) ), meningitis, meningococcal disease, metagonism, microsporidiosis, molluscum contagiosum, mumps, typhus (endemic typhus), mycoplasma pneumonia, mycetoma, myiasis, neonatal conjunctivitis (ophthalmia neonatorum), Creutzfeldt-Jakob disease (vCJD, nvCJD), nocardiosis, onchocerciasis (blinding filariasis), paracoccidioidomycosis (South American blastomycosis), paragonimiasis, pasteurellosis, head lice (head lice), body lice disease (body lice), pubic lice disease (pubic lice, Crarbice), pelvic inflammatory disease, Whoopingcough, Plague, Pneumococcal infection, Pneumonia carinii, Pneumonia, Polio, Prevotella infection, PAME, Progressive multifocal leukoencephalopathy, Psittacosis, Q fever, Rabies, Rat bite fever, Respiratory syncytial virus infection, Rhinovirus infection, Rickettsia infection, Rickettsia, Rift Valley fever, Rocky Mountain spotted fever, Rotavirus infection, Rubella, Salmonellosis, SARS (Severe acute respiratory syndrome), Scabies, Schistosomiasis, Sepsis, Diarrhea (dysentery), Shingles (Herpeszoster), Smallpox, Sporotrichum, Staphylococcus aureus food poisoning , Staphylococcal infection, nematodes, syphilis, taeniasis, tetanus (opening disorder), Barber's sitch, tinea manus, pityriasis nigricans, tinea pedis, tinea unguium, tinea versicolor, toxocariasis (ocular larva migrans), toxocariasis (visceral larva migrans), toxoplasmosis, trichinella, trichomoniasis, cryptobiosis (whipworm infection), pulmonary tuberculosis, tularemia, urea-degrading mycoplasma infection, Venezuelan equine encephalitis, Venezuelan hemorrhagic fever, viral pneumonia, West Nile fever, rhizobial disease, pseudotuberculosis, yersiniosis, yellow fever, and zygomycosis.

[0348] The pathogenic strains include, but are not limited to, Acinetobacter baumannii, Actinomyces israelii, Actinomyces odontolyticus, Propionibacterium propionicus, Trypanosoma brucei, HIV (human immunodeficiency virus), Entamoeba histolytica, Anaplasma spp., Bacillus anthracis, Arcanobacterium haemolyticum, Junin virus, Roundworm, Aspergillus, Astroviridae, Babesia spp., Bacillus cereus spp., Mammalian rabies, Bacillus casei ... Rupture bacteria, Bacteroides spp., Colonic pouch ciliates, Baileyi roundworm spp., BK virus, Piedraia hortae, Blastocystis hominis, Dermatophytosis, Macupo virus, Borrelia spp., Clostridium botulinum, Sabia, Brucella spp., usually Burkholderia cepacia and other Burkholderia species, Mycobacterium ulcerans, Caliciviridae family, Campylobacter spp., usually Candida albicans and other Candida species, Bartonella hense lae), Group A Streptococcus and Staphylococcus, Trypanosoma cruzi, Haemophilus ducreyi, Varicella zoster virus (VZV), Chlamydia trachomatis, Chlamydia pneumoniae, Vibrio cholerae, Fonsecae pedrosoi, Clonorchiasis, Clostridium difficile, Coccidioides immitis, Coccidioides posadaci, Colorado tick fever virus, Rhinovirus, Coronavirus, Creutzfeldt-Jakob disease prion, Crimean-Congo hemorrhagic fever virus, Cryptococcus neoformans, Cryptosporidium spp., Cat hookworm, Co-parasites, Schizosaccharomyces cerevisiae ... Clospora, Taenia solium, Cytomegalovirus, Dengue virus (DEN-1, DEN-2, DEN-3 and DEN-4)-Flavivirus, Dictyonaeba, Corynebacterium diphtheriae, Diphyllobothrix spp., Dracunculus medinensis, Ebola virus, Echinococcus spp., Ehrlichia spp., Pinworms, Enterococcus spp., Enterovirus spp., Typhi rickettsia, Parvovirus B19, Human herpesvirus type 6, Human herpesvirus type 7, Fasciola fasciata, Fasciola gigantica and Fasciola gigantica, FFI prion, Filaria head superfamily,Clostridium perfringens, Fusobacterium, Clostridium perfringens, other Clostridium spp., Geotrichum candidum, GSS prion, Giardia lamblia, Burkholderia mallei, Gnathostoma stenophylla, Gnathostoma niger, Neisseria gonorrhoeae, Borrelia granulomatosis, Streptococcus pyogenes, Streptococcus agalactiae, Haemophilus influenzae, enteric viruses, most Coxsackie A viruses, enteric virus 71, Sin Nombre virus, Helicobacter pylori, Escherichia coli O158:H7, Bunyaviridae, Hepatitis A virus, Hepatitis B virus, Hepatitis C virus Hepatitis virus, Hepatitis D virus, Hepatitis E virus, Herpes simplex virus type 1, Herpes simplex virus type 2, Histoplasma capsulatum, Ancylostoma niger, American hookworm, Haemophilus influenzae, Boca human virus, Ehrlichia ewingii, Anaplasma phagocytophilum, Human metapneumovirus, Ehrlichia chaffeensis, Human papillomavirus, Human parainfluenza virus, Tapeworm dwarf, Tapeworm diminutum, Epstein-Barr virus, Orthomyxoviridae, Isospora nigra Isosporabelli, Kingella kingae, Klebsiella pneumoniae, Klebsiella ozena, Klebsiella rhinoscleromotis, Coulour purion, Lassa fever virus, Legionella pneumophila, Leishmania, Mycobacterium lepromatosis and Mycobacterium lepromatosis, Leptospira spp., Listeria monocytogenes, Borreliosis and other Borrelia species, Wuchereria bancrofti and Brugia malayi, Lymphocytic choriomeningitis virus (LCMV), Plasmodium genus, Marburg virus, measles virus, Burkholderia pseudomallei, Neisseria meningitidis, Fasciola yokogawai, Microsporidia, Molluscum contagiosum virus (MCV), Mumps virus, Rickettsia typhi, Mycoplasma pneumoniae, various bacteria (actinomycetoma) and fungi (mycosis fungi), Diptera parasitic fly larvae, Chlamydia trachomatis and Neisseria gonorrhoeae, vCJD prion,Nocardia asteroides and other Nocardia species, Onchocerca volvulus, Blastomyces braziliensis, Paragonimus and other Paragonimus species, Pasteurella spp., Head lice, Body lice, Phthiruspubis, Bordetella pertussis, Yersinia pestis, Streptococcus pneumoniae, Pneumocystis cysticercosis, Poliovirus, Prevotella spp., Naegleria ameba, JC virus, Chlamydia psittacosis, Coxiella burnetii, Rabies virus, Bead-chain E. coli and Rat-bite fever spirochetes, Respiratory syncytial virus, Rhinosporidium sayberi, Rhinovirus , Rickettsia spp., Rickettsia mites, Rift Valley fever virus, Rocky Mountain spotted fever rickettsia, Rotavirus, Rubella virus, Salmonella spp., Atypical pneumonia coronavirus, Scabies mites, Schistosoma spp., Shigella, Varicella zoster virus, Variola major or minor, Sporothrix schenckii, Staphylococcus spp., Staphylococcus aureus, Streptococcus pyogenes, Strongyloides stercoralis, Syphilis spirochete, Cestodes spp., Clostridium tetani, Tinea, Trichophyton tonsurans, Tinea, Epidermophyton floccosum, Tinea rubra and Tinea mentagrophytes, Tinea rubra, Holtea wernecki, Tinea, Malassezia spp. , Roundworms of dogs and cats, Toxoplasma gondii, Trichinella spiralis, Trichomonas vaginalis, Trichuris trichiura, Mycobacterium tuberculosis, Tula hot francis bacteria, Ureaplasma urealyticum, Venezuelan equine encephalitis virus, Vibrio cholerae, Guanarito virus, West Nile virus, Mycobacterium tuberculosis, Mycobacterium pseudotuberculosis, Yersinia enterocolitica, Yellow fever virus, Mucorales (mucormycosis) and Insecta mesh mold (entomophthorosis), Pseudomonas aeruginosa, Campylobacter fetus (Vibrio), Aeromonas bacteria, Edwardsiella spp., Tarda, Yersinia pestis, Shigella dysenteriae, Shigella sonnei, Salmonella typhimurium, Treponema pertussis ne, Treponema carateneum, phenenburgdorferi, Borrelia burgdorferi, Leptospira hemorrhagic jaundice, Pneumocystis carinii, Brucella abortus, Brucella abortus, Malta fever, Mycoplasma spp., Rickettsia typhi, Rickettsia tsutsugamushi, Chlamydia spp., Pathogenic fungi (Aspergillus fumigatus, Candida albicans, Histoplasma capsulatum); Protozoa (Amoeba histolytica, Trichomonas vaginalis, Trichomonas venoides, Trypanosoma gambiens, Trypanosoma rhodesiense, Leishmania donovani, Leishmania tropicalis,Leishmania braziliensis, Pneumocystis carinii pneumoniae, Plasmodium vivax, Plasmodium falciparum, and Plasmodium malignum; or helminths (Schistosoma japonicum, Schistosoma mansoni, Schistosoma haematobium, and Ancylostoma fasciata).

[0349] Pathogenic viruses include, but are not limited to, Poxyiridae, Herpesviridae, Adenoviridae, Papovaviridae, Enteroviridae, Picornaviridae, Parvoviridae, Reoviridae, Retroviridae, influenza virus, parainfluenza virus, mumps, measles, respiratory syncytial virus, rubella, arbovirus, rhabdovirus, Arenaviridae, non-A / non-B hepatitis virus, rhinovirus, coronavirus. , rotavirus, tumor viruses [e.g., HBV (hepatocellular carcinoma), HPV (cervical cancer, anal cancer), Kaposi's sarcoma-associated herpesvirus (Kaposi's sarcoma), EB virus (nasopharyngeal carcinoma, Burkitt's lymphoma, primary central nervous system lymphoma), MCPyV (Merkel cell carcinoma), SV40 (simian virus 40), HCV (hepatocellular carcinoma), HTLV-I (adult T-cell leukemia / lymphoma)]; immune diseases caused by viruses: [e.g., human immunodeficiency virus (AIDS)], CNS viruses: [e.g., JCV (progressive multifocal leukemia, encephalopathy), MeV (subacute sclerosing panencephalitis), LCV (lymphocytic choriomeningitis), arboviral encephalitis, Orthomyxoviridae (presumed) (sluggish encephalitis), RV (rabies), vesicular stomatitis, herpesvirus meningitis, Ramsay Hunt syndrome type II; poliovirus (acute poliomyelitis, post-polio syndrome), HTLV-I (tropical spastic paralysis)]; cytomegalovirus (CMV retinitis, HSV (herpetic keratitis)); cardiovascular disease viruses [e.g. CBV (pericarditis, myocarditis)]; respiratory system / acute nasopharyngitis / viral lung inflammation: [EBV (EBV infection / infectious mononucleosis), cytomegalovirus, SARS coronavirus (severe acute respiratory syndrome), Orthomyxoviridae: influenza virus A / B / C (influenza / avian influenza), Paramyxovirus: human parainfluenza virus (parainfluenza), RSV (human respiratory syncytial virus), hMPV]; digestive viruses [MuV (mumps), cytomegalovirus (CMV esophagitis); Adenovirus (adenovirus infection);Such pathogenic viral diseases include rotavirus, norovirus, astrovirus, coronavirus, HBV (hepatitis B virus), CBV, HAV (hepatitis A virus), HCV (hepatitis C virus), HDV (hepatitis D virus), HEV (hepatitis E virus), HGV (hepatitis G virus)]; urogenital viruses [e.g., BK virus, MuV (mumps)];

[0350] According to a further object, the present invention also relates to a pharmaceutical composition for treating cancer, infectious diseases, or autoimmune diseases, comprising the conjugate of the present invention together with a pharma- ceutically acceptable carrier, diluent, or excipient. The method for treating cancer, infectious diseases, and autoimmune diseases can be performed in vitro, in vivo, or ex vivo. Examples of in vitro therapies include cell culture treatments to kill all cells except for desired variants that do not express the target antigen, or to kill variants that express undesired antigens. Examples of ex vivo therapies include treatment of hematopoietic stem cells (HSCs) prior to performing a transplant (HSCT) and returning them to the same patient to kill diseased or malignant cells. For example, clinical ex vivo treatments to remove cancer cells or lymphocyte cells from bone marrow prior to autologous transplantation in the treatment of cancer and autoimmune diseases, or to remove T cells and other lymphocyte cells from allogeneic bone marrow or tissue prior to transplantation to prevent graft-versus-host disease, can be performed by: After obtaining bone marrow cells from a patient or other individual, they are cultured at 37°C for 30 minutes to about 48 hours in serum-containing medium to which the conjugate of the present invention has been added, in a concentration range of about 1 pM to 0.1 mM. The exact concentration conditions and culture time (= dose) can be easily determined by an experienced clinician. After the culture is completed, the bone marrow cells are washed with serum-containing medium and returned to the body by known methods such as intravenous injection. If the patient has received other treatments (e.g., abolition chemotherapy or total body irradiation) between obtaining and reinjecting the bone marrow cells, the processed bone marrow cells are cryopreserved in liquid nitrogen using standard medical equipment.

[0351] All references cited in this specification, including publications, patent applications, and patents, are herein incorporated by reference to the same extent as if each reference was individually and specifically indicated to be incorporated by reference and was set forth in its entirety herein.

[0352] The use of the terms "a" and "an", "the" and "at least one" and similar referents in the context of describing the present invention (particularly in the context of the claims) should be construed to include both the singular and the plural, unless otherwise indicated herein or clearly contradicted by the context. The use of the term "at least one" following a list of one or more items (e.g., "at least one of A and B") should be construed to mean one item selected from the listed items (A or B), or any combination of two or more of the listed items (A and B), unless otherwise indicated herein or clearly contradicted by the context. The terms "comprise", "have", "include", and "includes" should be construed as open-ended terms (i.e., meaning "including but not limited to"), unless otherwise indicated. The recitation of ranges of values ​​herein is merely intended to serve as a shorthand method of individually referring to each individual value falling within the range, unless otherwise indicated herein, and each individual value is incorporated into the specification as if it were individually set forth herein. All methods described herein can be performed in any suitable order unless otherwise indicated herein or clearly contradicted by context. Any examples provided herein, or the use of exemplary language (e.g., "etc.") are merely intended to facilitate a better understanding of the invention and do not impose limitations on the scope of the invention unless otherwise claimed. No language in the specification should be construed as indicating any non-claimed element as essential to the practice of the invention.

[0353] Preferred embodiments of the invention are described herein, including the best mode known to the inventors for carrying out the invention. Variations of these preferred embodiments will become apparent to those skilled in the art upon reading the foregoing description. The inventors expect that such variations will be adopted by those skilled in the art as appropriate, and it is intended that the invention be practiced otherwise than as specifically described herein. Accordingly, this invention includes all modifications and equivalents of the subject matter recited in the claims appended hereto as permitted by applicable law. Moreover, this invention encompasses any combination of the above-described elements in all possible variations thereof unless otherwise indicated herein or clearly contradicted by context.

[0354] The following examples further illustrate the invention but, of course, should not be construed as in any way limiting its scope. EXAMPLES

[0355] The present invention is further illustrated by the following examples, which are not intended to limit the scope of the present application. The cell lines described in the following examples were maintained in culture media according to conditions specified by the American Type Culture Collection (ATCC), the German Collection of Microbial Cell Cultures (DMSZ), Braunschweig, Germany, or the Shanghai Institute of Cell Culture, Chinese Academy of Sciences, unless otherwise stated. All cell culture reagents were provided by Invitrogen, unless otherwise stated. All anhydrous solvents were commercially available and stored in nitrogen-sealed bottles. PEG compounds were purchased from Biomatrik Inc, Jiaxing, China. Some compounds were provided by Chinese contract research organizations (CROs) (e.g., Wuxi Apptec, Haoyuan Chemexpress, Raybow Pharma) when not referred to synthesis. Experimental animals were purchased from the National Resource Center of Model Mice via GemPharmatech Co., Ltd (Najin, China) and Shanghai SLAC Laboratory Animal Co., Ltd. (Shanghai, China). T-DM1 was purchased from Roche via a pharmacy in Hong Kong, China. All other reagents and solvents were purchased according to the highest standards and used without further purification. Preparative HPLC separation was performed on a Varain PreStar HPLC. HPLC analysis was performed on an Agilent 1260. Mass spectral data were acquired on a Waters Xwvo QTOF equipped with a Waters Acquity UPLC separation module and an Acquity TUV detector. NMR spectra were recorded on a Zhongke-niujin WNMR-I 400 MHz instrument at the Department of Chemistry, Zhejiang University of Technology. Chemical shifts (δ) were in parts per million (ppm) and were referenced to tetramethylsilane at 0.00, and coupling constants (J) were in Hz. Elemental analysis of C, H, and / or N was provided by the Department of Chemistry, Zhejiang University of Technology and performed on an Elementar UNICUBE. Quantitative analysis of metal atoms was performed on an Agilent ICPOES 730 ICP-MS.

[0356] Example 1. Generation of monoclonal antibodies against B cell maturation antigen (BCMA)

[0357] Six-week-old BalB / c mice were given four subcutaneous injections of purified recombinant human (rHu)TrxA-BCMA according to the RIMMS immunization regimen described by Kilpatrick et al., Hybridoma, 16(4): 381-389 (1997). Mice were immunized over a 13-day period with 2- to 3-day intervals. For each round of immunization, mice were first anesthetized with isoflurane. Immunogens were emulsified in complete or incomplete Freund's adjuvant. Gold adjuvant (Sigma-Aldrich) was injected bilaterally at multiple sites. Test bleeds were taken on day 13 and analyzed by antigen ELISA. Mice with good serum titers were given an intraperitoneal prefusion boost and sacrificed on day 17. Spleen cells were harvested and fused with the myeloma cell line P3-X63-Ag8.653 according to the polyethylene glycol fusion method (Roche Diagnostics) to generate stable hybridomas.

[0358] Anti-BCMA specific hybridomas were identified by screening hybridoma supernatants in a direct binding ELISA followed by FACS on BCMA expressing RPMI-8226-BCMA cells. Positive hybridomas were further tested for their ability to bind and internalize into RPMI-8226-BCMA cells in vitro by FACS binding to BCMA expressed on the cell line.

[0359] After sorting and subcloning, BCMA-A2-6H4-5D2 was selected and antibody binding affinity was determined by Elisa assay with anti-BCMA antibody J6M0 (described in U.S. Pat. No. 9,273,141 and referred to herein as belantamab or DXA009B), the results of which are shown in FIG. 1. Deposit at the China Center for Typical Culture Collection (CCTCC) was made under the Budapest Treaty on June 23, 2022. The CCTCC is located at Wuhan University, Wuhan, Hubei Province, China, zip code 430000. The CCTCC deposit was assigned the accession number CCTCC C2022188.

[0360] The amino acid sequences of the heavy and light chain variable regions of monoclonal antibody BCMA-A2-6H4-5D2 are shown in Table 1.

[0361] The results of this example demonstrate the production of monoclonal antibodies against BCMA. [Table 1]

[0362] Example 2. Generation of humanized monoclonal antibodies of BCMA-A2-6H4-5D2

[0363] Chimeric antibody c5D2 HC (SEQ ID NO: 13) constructed by fusing the VH of BCMA-A2-6H4-5D2 (SEQ ID NO: 10) with the human IgG1 HC constant domain (SEQ ID NO: 12, encoded by SEQ ID NO: 23), and chimeric antibody c5D2 LC (SEQ ID NO: 15) constructed by fusing BCMA-A2-6H4-5D2 (SEQ ID NO: 11) with the human kappa LC constant domain (SEQ ID NO: 14, encoded by SEQ ID NO: 26). Humanized antibody hu5D2 HC (SEQ ID NO: 8) generated by substitution of the corresponding amino acid in 5D2 with human germline gene amino acid, humanized antibody hu5D2 LC (SEQ ID NO: 9) generated by substitution of the corresponding amino acid in 5D2 with human germline gene amino acid. The affinity of hu5D2 is shown in FIG. 2.

[0364] The amino acid sequences of the heavy and light chain variable regions of monoclonal antibodies c5D2 and hu5D2 are shown in Table 2.

[0365] [Table 2]

[0366] Example 3. A tridational method for producing an antibody-drug conjugate (ADC) comprising a BCMA monoclonal antibody conjugated to a cytotoxic agent with a maleimide group terminus.

[0367] The hu5D2 monoclonal antibody was conjugated with maleimide-terminated cytotoxic agents. Specifically, purified antibody was incubated with 3.2-4.2 molar excess of reducing agent TCEP (tris(2-carboxyethyl)phosphine) in PBS pH 7.2, 1 mM EDTA (ethylenediaminetetraacetic acid) for 1 hour at 37°C. Approximately 6.5-9.0 equivalents of maleimide-terminated cytotoxin payload from a stock solution in 10% (v / v) DMA or DMSO was then added, followed by incubation at room temperature for 1-3 hours with gentle agitation. The conjugation reaction was quenched, if necessary, by adding 4 molar equivalents (over payload) of N-acetylcysteine. After incubation, reducing agent and excess payload / conjugate complex were removed by dialysis 2-10 times in PBS pH 5.0-7.2 at 4°C using a 10,000 MWCO dialysis cassette.

[0368] The conjugation method may result in the formation of 0.1-10% aggregates. Polymeric aggregates, conjugation reagents containing cysteine-quenched payloads, can be removed using ceramic hydroxyapatite type II chromatography (CHT), for example, as described in Thompson et al., J. Control Release, 236: 100-116 (2016). The ADC was formulated in 25 mM histidine-HCl, 7% sucrose, 0.02% polysorbate-20 or 80, pH 6, as appropriate.

[0369] To determine monomer content, aggregates, and fragments, analytical size-exclusion chromatography (SEC-HPLC) was performed using 100 μL (100 μL volume) of antibody or ADC loaded onto a TSKgel.RTM.G3000WXL column (Tosoh Biosciences, Tokyo, Japan). The mobile phase consisted of 0.1 M sodium sulfate, 0.1 M sodium phosphate, and 10% isopropanol, pH 6.8, with a flow rate of 1 mL / min, and each analysis was performed for 10–45 min at room temperature. Hydrophobic interaction chromatography (HIC-HPLC) was used to evaluate conjugation and drug load distribution and was performed using a butyl nonporous resin (NPR) column (4.6 mm ID × 3.5 cm, 2.5 μm, Tosoh Biosciences). Mobile phase A consisted of 25 mM Tris-HCl, 1.5 M (NH 4 ) 2 SO 4Mobile phase B consisted of 25 mM Tris-HCl and 5% isopropanol, pH 8.0. 100 μL of antibody or ADC at a concentration of 1 mg / mL was loaded and eluted at a flow rate of 1 mL / min with a gradient from 5% B to 100% B over 10-30 min. To confirm chain-specific conjugation, reducing reversed-phase chromatography (rRP-HPLC) was used. Antibodies and ADCs were reduced with 42 mM dithiothreitol (DTT) in PBS (pH 7.2) for 20 min at 37 °C. Ten micrograms of reduced antibody or ADC was loaded onto a polymeric reversed-phase media (PLRP-S) 1000A column (2.1 × 50 mm) (Agilent Technologies, Santa Clara, CA) and eluted with a gradient of 5% B to 100% B (mobile phase A: 0.1% trifluoroacetic acid in water; mobile phase B: 0.1% trifluoroacetic acid in acetonitrile) over 20–35 min at 80 °C and a flow rate of 1 mL / min.

[0370] Conjugation and drug / antibody ratios (DAR) on the heavy and light chains were determined by reduced liquid chromatography mass spectrometry (rLCMS) performed on an Agilent 1290 series uHPLC connected to an Agilent 6230 TOF (Agilent Technologies, Santa Clara, CA). 2 μg of reduced antibody or ADC was loaded onto a ZORBAX.RTM. and eluted at a flow rate of 0.5 mL / min using a step gradient of 80% B (mobile phase A: 0.1% formic acid in water and mobile phase B: 0.1% formic acid in acetonitrile) after 2.1 min using a ZORBAX.RTM.-Rapid Resolution High Resolution (RRHD) 300-Diphenyl column (2.1×50 mm, 1.8 μm) (Agilent Technologies, Santa Clara, CA). Positive time-of-flight MS scans were acquired and data collection and processing were performed using MassHunter software (Agilent Technologies, Santa Clara, CA).

[0371] Example 4. Method for generating BCMA-expressing cell lines

[0372] Stable cell lines were developed by transfecting RPMI-8226 cells with either full-length BCMA clones or an empty vector co-expressing GFP protein. Flow cytometry confirmed positive expression of BCMA on the surface of transfected BCMA (RPMI-8226-BCMA). These cell lines were then used as a tool to confirm the specificity of the cloned BCMA antibodies.

[0373] Example 5. Binding affinity of monoclonal BCMA antibodies and ADCs described herein to soluble and membrane-bound BCMA

[0374] Binding of BCMA-A2-6H4-5D2, hu5D2, and c5D2 to soluble BCMA was determined using an Elisa assay. Elisa assays were performed by coating with 1 μg / ml soluble BCMA (50 μL / well) for 1 hour at 37°C, followed by blocking with PBS+2% BSA. A range of antibody concentrations were diluted and added to the pre-coated Elisa plate and incubated for 1 hour at 37°C, followed by three PBST washes (BioTek 405), after which 50 μL / well of goat anti-human IgG (Fab specific)-peroxidase (Sigma-Aldrich) (1:20000 dilution) was added for detection. Prior to detection, plates were washed three times with PBST, followed by the addition of TMB and 2M H 2 SO 4 The results of this experiment are shown in Figure 2.

[0375] Binding of hu5D2, c5D2, and hu5D2-tub196 to membrane-bound human BCMA was assessed using flow cytometry in a multiple myeloma cell line (NCI-H929) that endogenously expresses BCMA. Binding assays were performed by incubating anti-BCMA antibodies with 200,000 cells for 30 minutes at 4°C, followed by washing twice with PBS+2% FBS (FACS buffer). A range of antibody concentrations was assessed using an 11-point 4-fold dilution series. Cells were then incubated with 5 μg / mL goat anti-human IgG Fc secondary antibody, PE (Thermo Fisher Scientific) at 4°C, followed by washing twice with PBS+2% FBS. Cells were resuspended in 200 μL of PBS+2% FBS.

[0376] Fluorescence of viable single cells was measured using a Guava easyCyte HT cytometer. Mean fluorescence intensity values ​​were used to determine percent binding and EC50s were determined using Prism software. The results of this experiment are shown in Figure 3.

[0377] Example 6. Methods for killing multiple myeloma cells in vitro using antibody-drug conjugates

[0378] Killing of multiple myeloma and plasma cell leukemia cell lines by antibody-drug conjugates containing hu5D2 or its affinity-optimized clones conjugated with tubulysin analogs such as compound 322 or 390 was evaluated in vitro using the protocol recommended in the CCK8 kit (Dojindo Laboratories, Japan). Briefly, 5000 cells in 180 μL RPMI+10% FBS were added to the inner wells of a 96-well plate. The following BCMA-expressing cell lines were tested: RPMI-8226-BCMA, NCI-H929, MM.1S, and Jurkat were also tested. Antibody-drug conjugates were diluted to 10x stocks (100 μg / mL) in RPMI+10% FBS. Therapeutic agents were then serially diluted 1:10 in RPMI+10% FBS. 20 μL of this series was added in triplicate to the cells, resulting in an eight-point dose curve of antibody-drug conjugate ranging from a high of 10 μg / mL to a low of 0 μg / mL. The plate was incubated at 37°C and 5% CO 2 The wells were incubated at 450 nm for 96 h. At the end of the incubation period, 10 μL of substrate solution was added to each well. The absorbance at 450 nm was measured using a SpectraMax i3x plate reader (Molecular Device, USA). Data were analyzed and graphed using GraphPad Prismor Excel software, and the 50% inhibitory concentration (IC 50 The results of this experiment are shown in Figures 4A, 4B, 4C, and 4D.

[0379] Example 7. Glycosylation site analysis of BCMA antibodies by LC-MS

[0380] Sample information: Recombinant humanized anti-BCMA monoclonal antibody (DXA009 DS), batch number is 009A2201B (manufactured by the applicant of this application, Hangzhou DAC Biotechnology Co., Ltd.). Sample preparation: Glycopeptide EEQYNSTYR (SEQ ID NO: 34) with a glycan attached at the asparagine position. Recombinant humanized anti-BCMA monoclonal antibody (DXA009 DS, batch number 009A2201B) was denatured and reduced with 6 M urea, 10 mM dithiothreitol at 56 °C for approximately 40 minutes, alkylated (approximately 30 mM iodoacetamide, in the dark, at room temperature for 40 minutes), and purified with 50 mM NH4HCO 3 and digested with trypsin (1 / 50, enzyme / substrate weight ratio, 4 hours, 37°C).

[0381] The glycopeptide masses and responses are shown in Table 3. The percentages of glycopeptide species (including Man5, G0F-GlcNAc, G0, G0F, G1, G1F, and G2F) are shown in Table 4. The MS / MS daughter ion or product ion spectra of the glycopeptides are shown in Figure 5 ((a)-(h)). [Table 3]

[0382] The position of the N-glycoside is therefore confirmed at Asn(N)-300.

[0383] [Table 4]

[0384] UPLC conditions: LC system: Waters ACQUITY UPLC H-Class system; detector: ACQUITY UPLC TUV; absorption wavelength: 214 nm; trap column: ACQUITY UPLC BEH C18 1.7 μm, 2.1 × 100 mm column. Mobile phase A: 0.1% formic acid (FA) in water, mobile phase B: 0.1% formic acid (FA) in ACN; chromatographic separation was performed at a flow rate of 0.25 ml / min using a linear gradient of mobile phase B (0.1% FA in ACN) from 1% to 40% in 95 min, followed by 40% to 80% in 10 min, then 80% to 80% in 5 min. MS conditions: MS system: Waters Xevo-G2XS Q-TOF; Ionization mode: ESI positive; Sensitivity mode; Data acquisition: MS E Mass range: m / z 100-2500 Da. Informatics: Data analysis using UNIFI V1.8.2.169 software (Waters).

[0385] Example 8. Molecular Weight Reduction and DAR Analysis of Deglycosylated BCMA-Tub ADCs by LC-MS

[0386] Sample preparation: ADC (e.g. (DXC009 DP)) was reduced using 5 mM dithiothreitol for approximately 2 hours at 37° C., followed by a deglycosylation step using PNGase F overnight at 37° C., resulting in six fragments as shown in Figures 6 and 7. HC and LC were present in naked or conjugated form carrying up to three payloads. The mass of each ADC fragment and the average DAR of the ADC are shown in Table 5. The following formula was used to calculate the average DAR of conventionally conjugated ADCs:

[0387] [Table 5]

[0388] Average DAR=L1 / (L0+L1)×2+H1 / (H0+H1+H2+H3)×2+H2 / (H0+H1+H2+H3)×2+ H3 / (H0+H1+H2+H3)×2

[0389] Method conditions: UPLC system: Waters ACQUITY UPLC H-Class system; Detector: ACQUITY UPLC TUV; Absorption wavelength: 280 nm; Trap column: ACQUITY UPLC C4 1.7 μm, 2.1×50 mm column. Chromatographic separation was performed at a flow rate of 0.4 ml / min using a linear gradient of mobile phase B from 5% to 25% in 2 min, followed by 25% to 45% in 8 min, then 45% to 85% in 2 min. MS conditions: MS system: Waters Xevo-G2XS Q-TOF; ionization mode: ESI positive; mass range: m / z 500-4000 Da. Informatics: Data analysis using UNIFI V1.8.2.169 software (Waters).

[0390] Example 9. Drug binding site analysis of BCMA-ADC by LC-MS

[0391] Sample preparation: Recombinant humanized anti-BCMA monoclonal antibody-tubulysin B conjugate (ex. DXC009DP), batch number 22030251, pack size 100mg / bottle, manufactured by Hangzhou DAC Biotechnology Co., Ltd. The ADC sample was denatured and reduced (6M urea, 10mM dithiothreitol, 56℃, about 40min), alkylated (about 30mM iodoacetamide, dark, room temperature, 40min), diluted with 50mM HEPES, and digested with trypsin (1 / 50, enzyme / substrate weight ratio, 4h, 37℃).

[0392] The drug-loaded peptides of the ADCs are shown in Table 6. The mass and average DAR of each ADC fragment is shown in Figure 7. The MS / MS daughter or product ion spectra of the drug-loaded peptides of the ADCs are shown in Figure 2.

[0393] [Table 6]

[0394] Method conditions: LC system: Waters ACQUITY UPLC H-Class system; Detector: ACQUITY UPLC TUV; Absorption wavelength: 214 nm; Trap column: ACQUITY UPLC C4 1.7 μm, 2.1×100 mm column. Chromatographic separation was performed at a flow rate of 0.25 ml / min using a linear gradient of mobile phase B (0.1% FA in ACN) from 1% to 40% in 95 min, followed by 40% to 80% in 10 min, then 80% to 80% in 5 min, of mobile phase B (0.1% FA in ACN). MS conditions: MS system: Waters Xevo-G2XS Q-TOF; Ionization mode: ESI positive, sensitivity mode; Data acquisition: MS E Mass range: m / z 100-2500 Da. Informatics: Data analysis using UNIFI V1.8.2.169 software (Waters).

[0395] Example 10. Synthesis of meso-2,3-bis((2,4-dimethoxybenzyl)amino)succinic acid (2) [ka]

[0396] To a solution of meso-2,3-dibromosuccinic acid (500 g, 1.80 mol) in ethanol (3.6 L) was added trimethylamine (729 g, 7.20 mol) followed by 2,4-dimethoxybenzylamine (903 g, 5.4 mol). After the addition was complete, the mixture was heated to 90° C. and stirred under reflux overnight. The mixture was cooled to room temperature and the solid formed was filtered, rinsed with ethanol and dried to give meso-2,3-bis(2,4-dimethoxybenzylamino)succinic acid (600 g, 72% yield).

[0397] Example 11. Synthesis of meso-2,3-diaminosuccinic acid (3) [ka]

[0398] A solution of meso-2,3-bis(2,4-dimethoxybenzylamino)succinic acid (800 g, 1.78 mol) in dichloromethane (100 mL) was treated with trifluoroacetic acid (2035 g, 17.8 mol) at room temperature overnight. The mixture was concentrated, then 1N NaOH (6 L) was added slowly and stirred for 30 min. The precipitate was filtered off and the solution was adjusted to pH 5-6 with aqueous hydrochloric acid. The resulting white precipitate was collected by filtration, rinsed with water, and dried to give meso-2,3-diaminosuccinic acid (217 g, 78% yield).

[0399] Example 12. Synthesis of meso-2,3-bis(((benzyloxy)carbonyl)amino)succinic acid (4) [ka]

[0400] Meso-2,3-diaminosuccinic acid (386 g, 2.6 mol) was dissolved in 2N NaOH (6.5 L) and mixed with 1,4-dioxane (2.1 L). The solution was cooled to 0° C. and benzyl chloroformate (1333 g, 7.8 mol) was added at such a rate as to maintain the internal temperature below 5° C. After the addition was complete, the mixture was stirred for 3 h and warmed to 0° C. Stirred at room temperature overnight. The reaction was diluted with water (2 L) and washed with ethyl acetate (2×1 L). The aqueous layer was acidified with concentrated hydrochloric acid. HCl was added until a pH of 3 was reached, then extracted with ethyl acetate (3×2 L). The organic phases were combined, washed with water (1 L), dried over anhydrous Na2SO4, filtered, and concentrated. The residue was triturated with dichloromethane / petroleum ether (1:1) and filtered to give meso-2,3-bis(((benzyloxy)carbonyl)amino)succinic acid (900 g, 83%).

[0401] Example 13: Synthesis of di-tert-butyl 4,4'-((2,3-bis(((benzyloxy)carbonyl)amino)succinyl)bis(azanediyl))dibutyrate (6) [ka]

[0402] To a solution of compound 4 (10 g, 28.7 mmol) and tert-butyl aminobutyrate hydrochloride (11.2 g, 57.4 mol) in tetrahydrofuran (200 mL) was added HATU (32.8 g, 86.3 mmol) and diisopropylethylamine (19 mL, 115 mmol), the reaction was stirred at room temperature overnight, diluted with water (400 mL), stirred for 10 min, filtered and oven dried to give a white solid (17.7 g, >100% yield). MS-ESI (m / z): [M + H] + C36H51N4O10 calcd, 699.35; found, 699.35.

[0403] Example 14. Synthesis of di-tert-butyl 4,4'-((2,3-diaminosuccinyl)bis(azanediyl))dibutyrate (7) [ka]

[0404] Compound 6 (16 g, 22.9 mmol) was dissolved in methanol (200 mL) and then Pd / C (2.0 g) was added. The reaction flask was evacuated, refilled with hydrogen, heated to 60° C., and stirred until the reaction was complete. Filtration and concentration gave product 7 (9.8 g, 100% yield). MS-ESI (m / z): [M + H] + C20H39N4O6 calculated, 431.28; found, 431.28.

[0405] Example 15. Synthesis of Compound 9 [ka]

[0406] Compound 7 (9.8 g, 22.8 mmol) was dissolved in dichloromethane (200 mL) and exo-3,6-epoxy-1,2,3,6-tetrahydrophthalic anhydride (7.5 g, 45.5...

Claims

1. 1. An isolated monoclonal antibody or antigen-binding fragment that binds to B cell maturation antigen (BCMA or CD269), comprising: The antibody or antigen-binding fragment thereof (i) a VH domain comprising a CDR H1 comprising SEQ ID NO:1 (TSFLIHW), a CDR H2 comprising SEQ ID NO:2 (FIIPGNGGTKYNQKFQ), and a CDR H3 comprising SEQ ID NO:3 (YDGSFEGYFDV); and (ii) a VL domain comprising CDR L1 comprising SEQ ID NO:4 (SSQSLVVHSDGNTYLH), CDR L2 comprising SEQ ID NO:5 (KVSNRDS), and CDR L3 comprising SEQ ID NO:6 (SQSTHWPWT); Including, The isolated monoclonal antibody or antigen-binding fragment, wherein the medical disorder associated with the presence of pathogenic B cells is a cancer of plasma cells or a cancer of B-lymphatic cells.

2. 2. The isolated monoclonal antibody or antigen-binding fragment of claim 1, wherein the cancer of plasma cells is multiple myeloma, plasmacytoma, Waldenstrom's macroglobulinemia, or plasma cell leukemia, or the cancer of B lymphocytes is Hodgkin's disease.

3. 2. The isolated monoclonal antibody or antigen-binding fragment of claim 1, wherein the VL and VH domains are fused to constant domains of human Ig kappa (encoded by SEQ ID NO:14 or SEQ ID NO:26) and IgG1 (encoded by SEQ ID NO:12 or SEQ ID NO:24), respectively.

4. comprising a VH domain comprising SEQ ID NO:10 or SEQ ID NO:16 and a VL domain comprising SEQ ID NO:11 or SEQ ID NO:17; or a heavy chain comprising SEQ ID NO:8 or SEQ ID NO:13; or 2. The isolated monoclonal antibody or antigen-binding fragment of claim 1, comprising a light chain comprising SEQ ID NO:9 or SEQ ID NO:

15.

5. 2. The isolated monoclonal antibody or antigen-binding fragment thereof of claim 1, which is a chimeric, humanized, partially humanized, bispecific, or single chain antibody, or a combination thereof.

6. (i) a VH domain comprising a sequence encoded by SEQ ID NO:20 or SEQ ID NO:22; and (ii) a VL domain comprising a sequence encoded by SEQ ID NO:21 or SEQ ID NO:23, or comprising a heavy chain encoded by SEQ ID NO:18 or SEQ ID NO:25; 2. The isolated monoclonal antibody or antigen-binding fragment of claim 1, comprising a light chain encoded by SEQ ID NO:19 or SEQ ID NO:

27.

7. 2. The isolated monoclonal antibody or antigen-binding fragment of claim 1, which is glycosylated.

8. The isolated monoclonal antibody or antigen-binding fragment thereof of claim 7, comprising an N-linked oligosaccharide chain at Asn300 of the heavy chain.

9. 1. An antibody-drug conjugate (ADC) against B-cell maturation antigen (BCMA) comprising a monoclonal antibody or antigen-binding fragment thereof conjugated to a cytotoxin, The monoclonal antibody is (a) a heavy chain variable region comprising a complementarity determining region 1 (CDR H1) having the amino acid sequence of SEQ ID NO:1, a CDR H2 having the amino acid sequence of SEQ ID NO:2, and a CDR H3 having the amino acid sequence of SEQ ID NO:3; and (b) a light chain variable region comprising a complementarity determining region 1 (CDR L1) having the amino acid sequence of SEQ ID NO:4, a CDR L2 having the amino acid sequence of SEQ ID NO:5, and a CDR L3 having the amino acid sequence of SEQ ID NO:6; An antibody-drug conjugate (ADC) comprising:

10. 10. The antibody-drug conjugate of claim 9 having the formula: 【Chemistry 1】 In the formula, D, D １ , and D ２ is a small molecule cytotoxin or functional small molecule, commonly referred to as a payload; L, L １ , and L ２ is a conjugate; mAb is a monoclonal antibody or antigen-binding fragment thereof according to claim 1.

11. The cytotoxin or functional small molecule D, D １ , and D ２ The antibody-drug conjugate of claim 10, wherein is independently selected from the group consisting of: 1) A chemotherapeutic agent selected from the group consisting of: a) alkylating agents selected from the group consisting of: nitrogen mustards: chlorambucil, chlornaphazine, cyclophosphamide, dacarbazine, estramustine, ifosfamide, mechlorethamine, mechlorethamine oxide hydrochloride, mannomustine, mitobronitol, melphalan, mitolactol, pipobroman, nobembitine, phenesterine, prednimustine, thiotepa, trofosfamide, uracil mustard; CC-1065 and synthetic analogs of adozelesin, carzelesin and bizeresin; duocarmycin and its synthetic analogs KW-2189 and CBI-TMI; benzodiazepine dimers or pyrrolobenzodiazepine (PBD) dimers, tomaymycin dimers, indolinobenzodiazepine dimers. benzodiazepines, imidazobenzothiadiazepine dimers, or oxazolidinobenzodiazepine dimers; nitrosourea compounds including carmustine, lomustine, chlorozotocin, fotemustine, nimustine, ranimustine; alkylsulfonates including busulfan, treosulfan, improsulfan, and piposulfan; triazenes or dacarbazines; platinum-containing compounds including carboplatin, cisplatin, and oxaliplatin; aziridines, benzodopa, carboquone, meturedopa, and uredopa; ethylenimines and methylamelamines including altretamine, triethylenemelamine, triethylenephosphoramide, triethylenethiophosphoramine, trimethylolomelamine; b) plant alkaloids selected from the group consisting of: vinca alkaloids including vincristine, vinblastine, vindesine, vinorelbine, navelbine; taxoids including paclitaxel and docetaxel and analogs thereof; maytansinoids including DM1, DM2, DM3, DM4, DM5, DM6, DM7, maytansine, ansamitocins and analogs thereof; cryptophycins including the cryptophycin 1 and cryptophycin 8 groups; epothilones, eleutherobin, discodermolide, bryostatins, dolostatins, auristatins, tubulysins, cephalostatins; pancratistatin; sarcodictiin; spongistatin; c) DNA topoisomerase inhibitors selected from: epipodophyllins, including 9-aminocamptothecin, camptothecin, crisnatol, daunomycin, etoposide, etoposide phosphate, irinotecan, mitoxantrone, novantrone, retinoic acid (retinols), teniposide, topotecan, 9-nitrocamptothecin or RFS 2000; and mitomycins and their analogs; d) Antimetabolites selected from: {[Antifolates: (DHFR inhibitors: including methotrexate, trimetrexate, denopterin, pteropterin, aminopterin (4-aminopteroic acid), or other folate analogs); IMP dehydrogenase inhibitors (including mycophenolic acid, tiazofurin, ribavirin, EICAR); Ribonucleotide reductase inhibitors (including hydroxyurea, deferoxamine)]; [Pyrimidine analogs uracyl analogues (including ancitabine, azacitidine, 6-azauridine, capecitabine, carmofur, cytarabine, dideoxyuridine, doxifluridine, enocitabine, 5-fluorouracil, floxuridine, raltitrexed); cytosine analogues (including cytarabine, cytosine arabinoside, fludarabine); purine analogues (including azathioprine, fludarabine, mercaptopurine, thiamiprine, thioguanine); folic acid supplements such as folinic acid}, and nicotinamide phosphate ribonuclease (NAMPT) inhibitors; e) Hormonal therapy agents selected from: Receptor antagonists: [Antiestrogen: (including megestrol, raloxifene, tamoxifen); LHRH agonists: (including goserelin, leuprolide acetate); Antiandrogens: (including bicalutamide, flutamide, calsterone, dromostanolone propionate, epithiostanol, goserelin, leuprolide, mepitiostane, nilutamide, testolactone, trilostane, and other similar antineoplastic agents] ). retinoids / deltoid: [vitamin D3 analogs: (including CB1093, EB1089, KH1060, cholecalciferol, ergocalciferol); photodynamic therapy agents: (including verteporfin, phthalocyanines, photosensitizer Pc4, demethoxyhypocrelin A); cytokines: (including interferon alpha, interferon gamma, tumor necrosis factor (TNF), TNF domain-containing human proteins)]}; f) Kinase inhibitors selected from: BIBW2992 (anti-EGFR / Erb2), imatinib, gefitinib, pegaptanib, sorafenib, dasatinib, sunitinib, erlotinib, nilotinib, lapatinib, axitinib, pazopanib, vandetanib, E7080 (anti-VEGFR2), mubritinib, ponatinib, bafetinib, bosutinib, cabozantinib, vismodegib, iniparib, ruxolitinib, CYT387, axitinib, tivozanib, sorafenib, bevacizumab, cetuximab, trastuzumab, ranibizumab, panitumumab, ispinesib; g) a poly(ADP-ribose)polymerase (PARP) inhibitor selected from the group consisting of olaparib, niraparib, iniparib, talazoparib, veliparib, CEP9722 (Cephalon), E7016 (Eisai), BGB-290 (Beigene), or 3-aminobenzamide; h) enediyne antibiotics (selected from the calicheamicins, calicheamicin gamma 1, delta 1, alpha 1, or beta 1; dynemicins including dynemicin A and deoxydynemicin; esperamicin, kedarcidin, C-1027, maduropeptin, or neocarzinostatin chromophore and related chromoprotein enediyne antibiotic chromophores), aclacinomycins, actinomycin, anthramycin, azaserine, bleomycins, cactinomycin, carabicin, carminomycin, carzinophilin; chromomycins, dactinomycin, daunomycin, Antibiotics selected from rubicin, detorubicin, 6-diazo-5-oxo-L-norleucine, doxorubicin, morpholino-doxorubicin, cyanomorpholino-doxorubicin, 2-pyrrolino-doxorubicin and deoxydoxorubicin, epirubicin, eribulin, esorubicin, idarubicin, marcelomycin, nitomycins, mycophenolic acid, nogalamycin, olivomycins, peplomycin, potofilomycin, puromycin, queramycin, rodorubicin, streptonigrin, streptozocin, tubercidin, ubenimex, zinostatin, zorubicin; i) Polyketides (acetogenins), bullatacin and bullatacinone; gemcitabine, epoxomicins and carfilzomib, bortezomib, thalidomide, lenalidomide, pomalidomide, tosedostat, zybrestat, PLX4032, STA-9090, Stimuvax, allovectin-7, zygeva, probenge, elvoy, isoprenylation inhibitors and lovastatin, dopaminergic neurotoxins and 1-methyl-4-phenyl Pyridine ion, cell cycle inhibitors (including staurosporine), actinomycins (including actinomycin D and dactinomycin), amanitins, bleomycins (including bleomycin A2, bleomycin B2, and peplomycin), anthracyclines (including daunorubicin, doxorubicin (adriamycin), idarubicin, epirubicin, pirarubicin, and zorubicin), mitoxantrone, MDR inhibitors or verapamil, Ca ２＋ ATP inhibitors or thapsigargin, histone deacetylase inhibitors (including vorinostat, romidepsin, panobinostat, valproic acid, mocetinostat (MGCD0103), belinostat, PCI-24781, entinostat, SB939, resminostat, gibinostat, AR-42, CUDC-101, sulforaphane, trichostatin A); thapsigargin, celecoxib, glitazones, epigallocatechin gallate, disulfiram, salinosporamide A, anti-adrenal drugs (selected from the group consisting of aminoglutethimide, mitotane, trilostanca); aceglatone; aldophosphamide glycoside; aminolevulinic acid; amsacrine; arabinoside, bestravsil; bisanto len; edatrexate; defofamine; demecolcine; diaziquone; elfornitine (DFMO), elfomitine; elliptinium acetate; etocurside, gallium nitrate, gacytosine, hydroxyurea; ibandronate, lentinan; lonidamine; mitoguazone; mopidamol; nitracrine; pentostatin; phenamet; pirarubicin; podophyllic acid; 2-ethylhydrazide; procarbazine; PSK (registered trademark); razoxane; rhizoxin; sizofiran; spirogermanium; tenuazonic acid; triaziquone; 2,2',2''-trichlorotriethylamine; trichothecenes (including T2 toxin, verrucarin A, roridin A, and anguidine); urethane, siRNA, antisense medicines, nucleases; 2) Anti-autoimmune disease drugs: cyclosporine, cyclosporine A, aminocaproic acid, azathioprine, bromocriptine, chlorambucil, chloroquine, cyclophosphamide, corticoids (including amcinonide, betamethasone, budesonide, hydrocortisone, flunisolide, fluticasone propionate, fluocortolone danazol, dexamethasone, triamcinolone acetonide, and beclomethasone dipropionate), DHEA, etanercept, hydroxychloroquine, infliximab, meloxicam, methotrexate, mofetil, mycophenolic acid, prednisone, sirolimus, and tacrolimus; 3) Anti-infectives, including but not limited to: a) Aminoglycosides: amikacin, astromycin, gentamicin (netilmicin, sisomicin, isepamicin), hygromycin B, kanamycin (amikacin, arbekacin, bekanamycin, dibekacin, tobramycin), neomycin (framycin, paromomycin, ribostamycin), netilmicin, spectinomycin, streptomycin, tobramycin, verdamycin; b) Amphenicols: azidamphenicol, chloramphenicol, florfenicol, thiamphenicol; c) Ansamycins: geldanamycin, herbimycin; d) Carbapenems: biapenem, doripenem, ertapenem, imipenem, cilastatin, meropenem, panipenem; e) Cephalosporins: carbacephems (loracarbef), cephacetrile, cefaclor, cephradine, cefadroxil, cephalonium, cephaloridine, cephalothin or cephalothin, cephalexin, cephaloglycin, cefamandole, cephapirin, cefatrizine, cefazaflur, cefazedone, cefazolin, cefbuperazone, cefcapene, cefdaloxime, cefepime, cefminox, cefoxitin, cefprozil, cefroxadine, ceftezole, cefuroxime, cefixime, cefdinir, cefditoren, cefepime, Fetameth, cefmenoxime, cefodizime, cefonicid, cefoperazone, ceforanide, cefotaxime, cefotiam, cefozopran, cephalexin, cefpimizole, cefpiramide, cefpirome, cefpodoxime, cefprozil, cefquinome, cefsulodin, ceftazidime, cefteram, ceftibuten, ceftiolene, ceftizoxime, ceftobiprol, ceftriaxone, cefuroxime, cefuzonam, cephamycins (including cefoxitin, cefotetan, cefmetazole), oxacephems (flomoxef, latamoxef); f) Glycopeptides: bleomycin, vancomycin (including oritavancin, telavancin), teicoplanin (dalbavancin), ramoplanin; g) Glycylcyclines: Tigecycline; h) β-lactamase inhibitors: penams (sulbactam, tazobactam), clavams (clavulanic acid); i) Lincosamides: clindamycin, lincomycin; j) Lipopeptides: daptomycin, A54145, calcium-dependent antibiotic (CDA); k) Macrolides: azithromycin, cethromycin, clarithromycin, dirithromycin, erythromycin, flurithromycin, josamycin, ketolides (telithromycin, cethromycin), midecamycin, miokamicin, oleandomycin, rifamycin (rifampicin, rifampin, rifabutin, rifapentine), rokitamycin, roxithromycin, spectinomycin, spiramycin, tacrolimus (FK506), troleandomycin, telithromycin; l) Monobactams: aztreonam, tigemonam; m) Oxazolidinones: Linezolid; n) Penicillins: amoxicillin, ampicillin, pivampicillin, hetacillin, bacampicillin, methampicillin, talampicillin, azidocillin, azlocillin, benzylpenicillin, benzathine benzylpenicillin, benzathine phenoxymethylpenicillin, clometocillin, procaine benzylpenicillin, carbenicillin (carindacillin), cloxacillin, dicloxacillin, epicillin, flucloxacillin, mecillinam (pivmecillinam), mezlocillin, methicillin, nafcillin, oxacillin, penamecillin, penicillin, phenethicillin, phenoxymethylpenicillin, piperacillin, propicillin, sulbenicillin, temocillin, ticarcillin; o) Polypeptides: bacitracin, colistin, polymyxin B; p) quinolones: alatrofloxacin, balofloxacin, ciprofloxacin, clinafloxacin, danofloxacin, difloxacin, enoxacin, enrofloxacin, floxin, garenoxacin, gatifloxacin, gemifloxacin, grepafloxacin, canotrovafloxacin, levofloxacin, lomefloxacin, marbofloxacin, moxifloxacin, nadifloxacin, norfloxacin, orbifloxacin, ofloxacin, pefloxacin, trovafloxacin, grepafloxacin, sitafloxacin, sparfloxacin, temafloxacin, tosufloxacin, trovafloxacin; q) Streptogramins: pristinamycin, quinupristin / dalfopristin; r) Sulfonamides: mafenide, prontosil, sulfacetamide, sulfamethizole, sulfanilamide, sulfasalazine, sulfisoxazole, trimethoprim, trimethoprim-sulfamethoxazole (co-trimoxazole); s) Steroidal antibacterial agents: fusidic acid, etc.; t) Tetracyclines: doxycycline, chlortetracycline, clomocycline, demeclocycline, lymecycline, meclocycline, methacycline, minocycline, oxytetracycline, penimepicycline, rolitetracycline, tetracycline, glycylcycline (such as tigecycline); u) Other antibiotics: annonacin, arsphenamine, bactoprenol inhibitors (bacitracin), DADAL / AR inhibitors (cycloserine), dictyostatin, discodermolide, eleutherobin, epothilone, ethambutol, etoposide, faropenem, fusidic acid, furazolidone, isoniazid, laulimalide, metronidazole, mupirocin, mycolactone, NAM synthesis inhibitors (fosfomycin), nitrofurantoin, paclitaxel, platensimycin, pyrazinamide, quinupristin / dalfopristin, rifampicin (rifampin), tazobactamcinidazole, uvaricin; 4) Antiviral drugs: a) Entry / fusion inhibitors: aplaviroc, maraviroc, vicriviroc, gp41 (enfuvirtide), PRO140, CD4 (ibalizumab); b) Integrase inhibitors: raltegravir, elvitegravir, globoidnan A; c) Maturation inhibitors: Bevirimat, Vivecon; d) Neuraminidase inhibitors: oseltamivir, zanamivir, peramivir; e) Nucleosides and Nucleotides: Abacavir, Acyclovir, Adefovir, Amdoxovir, Apricitabine, Brivudine, Cidofovir, Clevudine, Dexerbucitabine, Didanosine (DDI), Elvucitabine, Emtricitabine (FTC), Entecavir, Famciclovir, Fluorouracil (5-FU), 3'-Fluoro substituted 2',3'-deoxynucleoside analogues (e.g., 3'-Fluoro-2',3'-dideoxythymidine (FLT) and 3'-Fluoro-2',3'-deoxythymidine (FLT), dideoxyguanosine (FLG), fomivirsen, ganciclovir, idoxuridine, lamivudine (3TC), L-nucleosides (e.g., including the group consisting of β-L-thymidine and β-L-2'-deoxycytidine), penciclovir, rasvir, ribavirin, stampidine, stavudinecet (d4T), taribavirin (viramidine), telbivudine, tenofovir, trifluridine, valacyclovir, valganciclovir, zalcitabine (ddC), zidovudine (AZT); f) Non-nucleosides: amantadine, ateviridine, capravirine, diarylpyrimidines (etravirine, rilpivirine), delavirdine, docosanol, emivirine, efavirenz, foscarnet (phosphorylformate), imiquimod, interferon alpha, loviride, rhodenosine, methisazone, nevirapine, NOV-205, peginterferon alpha, podophyllotoxin, rifampicin, rimantadine, resiquimod (R-848), tromantadine; g) Protease inhibitors: amprenavir, atazanavir, boceprevir, darunavir, fosamprenavir, indinavir, lopinavir, nelfinavir, pleconaril, ritonavir, saquinavir, telaprevir (VX-950), tipranavir; h) Other antivirals: abzyme, arbidol, calanolide A, ceragenin, cyanovirin-N, diarylpyrimidine, epigallocatechin gallate (EGCG), foscarnet, griffithsin, taribavirin (viramidine), hydroxyurea, KP-1461, miltefosine, pleconaril, portmanteau inhibitors, ribavirin, seliciclib; (5) The drugs used in the conjugates prepared according to the present invention also include radioisotopes. Examples of radioisotopes (radionuclides) are: ３ H. １１ C. １４ C. １８ F. ３２ P. ３５ S. ６４ Cu, ６８ G.A. ８６ Y. ９９ Tc, １１１ In, １２３ I, １２４ I, １２５ I, １３１ I, １３３ Xe, １７７ Lu, ２１１ At, or ２１３ Bi; 6) Chromophore molecules capable of absorbing UV light, fluorescent light, IR light, near-IR light, and visible light; xanthophores, erythrophores, iridophores, leucophores, melanophores, cereus; a class or subclass of fluorescent molecules, which are fluorescent chemicals that re-emit light with light; a class or subclass of visual light-sensitive molecules; a class or subclass of light-emitting molecules; a class or subclass of luminescent molecules; and a class or subclass of luciferin compounds; Non-proteinaceous organic fluorophores selected from: xanthene derivatives (including fluorescein, rhodamine, Oregon Green, eosin, and Texas Red); cyanine derivatives (including cyanine, indocarbocyanine, oxacarbocyanine, thiacarbocyanine, and merocyanine); squalene derivatives and ring-substituted squaraines (including Seta, SeTau, and Square dyes); naphthalene derivatives (including dansyl and prodan derivatives); coumarin derivatives; oxadiazole derivatives (including pyridyloxazole, nitrobenzoxadiazole, and benzoxadiazole); anthracene derivatives (including anthraquinones, including DRAQ5, DRAQ7, and CyTRAK Orange); pyrene derivatives (cascade blue); oxazine derivatives (including Nile Red, Nile Blue, cresyl violet, oxazine 170); acridine derivatives (including proflavine, acridine orange, acridine yellow). Arylmethine derivatives (including auramine, crystal violet, and malachite green). Tetrapyrrole derivatives (including porphines, phthalocyanines, bilirubins); Fluorophore compound analogs and derivatives including: CF dyes, DRAQ and CyTRAK probes, BODIPY, Alexa Fluor, DyLight Fluor, Atto and Trancey, FluoProbes, Abberior dyes, DY and MegaStokes dyes, SulfoCy dyes, HiLyte Fluor, Seta, SeTau and Square dyes, Quasar and Cal Flour dyes, SureLight dyes (APC, RPEPerCP, phycobilisome), APC, APCXL, RPE, BPE, allophycocyanin (APC), aminocoumarin, APC-Cy7 conjugates, BODIPY-FL, Cascade Blue, Cy2, Cy3, Cy3.5, Cy3B, Cy5, Cy5.5, Cy7, fluorescein, FluorX, hydroxycoumarin, Lissamine rhodamine B, Lucifer Yellow, methoxycoumarin, NBD, Pacific Blue, Pacific Orange, PE-Cy5 conjugate, PE-Cy7 conjugate, PerCP, R-Phycoerythrin (PE), Red613, Seta-555-azide, Seta-555-DBCO, Seta-555-NHS, Seta-580-NHS, Seta-680-NHS, Seta-780-NHS, Seta-APC-780, Seta-PerCP-680, Seta-R-PE-670, SeTau380-NHS, SeTau405-maleimide, SeTau405-NHS, SeTau425-NHS, SeTau647-NHS, Texas Red, TRITC, T ruRed, X-rhodamine, 7-AAD (7-aminoactinomycin D, CG selective), acridine orange, chromomycin A3, CyTRAK orange (red excitation dark), DAPI, DRAQ5, DRAQ7, ethidium bromide, Hoechst 33258, Hoechst 33342, LDS751, mithramycin, propidium iodide (PI), SYTOX blue, SYTOX green, SYTOX orange, thiazole orange, TO-PRO: cyanine monomer, TOTO-1, TO-PRO-1, TOTO-3, TO-PRO-3, YOYO-1, YO-1;Fluorescent dyes including the following compounds or their derivatives: DCFH (2′7′-dichlorodihydro-fluorescein, oxidized form), DHR (dihydrorhodamine 123, oxidized form, photocatalytic oxidation), Fluo-3 (AM ester, pH>6), Fluo-4 (AM ester, pH 7.2), Indo-1 (AM ester, low / high calcium (Ca; ２＋ )), SNARF (pH 6 / 9), Allophycocyanin (APC), AmCyan1 (tetramer, Clontech), AsRed2 (tetramer, Clontech), Azami Green (monomer), Azurite, B-Phycoerythrin (BPE), Cerulean, CyPet, DsRed monomer (Clontech), DsRed2 ("RFP"), EBFP, EBFP2, ECFP, EGFP (weak dimer), Emerald (weak dimer), EYFP (weak dimer), GFP (S65A mutant), GFP (S65C mutant), GFP (S65L mutant), GFP (S65T mutant), GFP (Y66F mutant), GFP (Y66H mutant), GFP (Y66W mutant), GFP ｕｖ , HcRed1, J-Red, Katyusha, Kusabira Orange (monomer, MBL), mCFP, mCherry, mCitrine, Midriishi Cyan (dimer, MBL), mKate (TagFP635, monomer), mKeima-Red (monomer), mKO, mOrange, mPlum, mRaspberry, mRFP1 (monomer), mStrawberry, mTFP1, mTurquoise2, P3 (phycobilisome complex), Peridinin Chlorophyll (PerCP), R-phycoerythrin (RPE), T-Sapphire, TagCFP (dimer), TagGFP (dimer), TagRFP (dimer), TagYFP (dimer), tdTomato (tandem dimer), Topaz, TurboFP602 (dimer), TurboFP635 (dimer), TurboGFP (dimer), TurboRFP (dimer), TurboYFP635 (dimer), Venus, native GFP, YPet, Zs Green 1 (tetramer), Zs Yellow 1 (tetramer), and derivatives thereof; 7) Cell receptor agonists or cell stimulating molecules selected from the following: folic acid derivatives; glutamic acid urea derivatives; somatostatin and its analogs (selected from the group consisting of octreotide (sandostatin) and lanreotide (somatilin); aromatic sulfonamides; pituitary adenylate cyclase activating peptide (PACAP) (PAC1); vasoactive intestinal peptide (VIP / PACAP) (VPAC1, VPAC2); melanocyte stimulating hormone (α-MSH); cholecystokinin (CCK) / gastrin receptor agonists, bombesin (Pyr-Gln-Arg-Leu-Gly-Asn-Gln-Trp-Ala-Val-Gly-His-Leu-Met-NH ２ Neurotensin receptor ligands (NTR1, NTR2, NTR3), substance P (NK1 receptor) ligands; neuropeptide Y (Y1-Y6); homing peptides including RGD (Arg-Gly-Asp), NGR (Asn-Gly-Arg), dimeric and multimeric cyclic RGD peptides (cRGDfV), TAASGVRSMH, and LTLRWVGLMS (chondroitin sulfate proteoglycan NG2 receptor ligand), and F3 peptide; cell membrane permeable peptides Peptides (CPPs); peptide hormones, luteinizing hormone releasing hormone (LHRH) agonists and antagonists, and gonadotropin releasing hormone (GnRH) agonists that act by targeting follicle stimulating hormone (FSH) and luteinizing hormone (LH) as well as testosterone production, such as buserelin (Pyr-His-Trp-Ser-Tyr-D-Ser(OtBu)-Leu-Arg-Pro-NHEt), gonadorelin (Pyr-His-Trp-Ser-Tyr-Gly-Leu-Arg-Pro-Gly-NH ２ ), goserelin (Pyr-His-Trp-Ser-)Tyr-D-Ser(OtBu)-Leu-Arg-Pro-AzGly-NH ２ ), histrelin (Pyr-His-Trp-Ser-Tyr-D-His(N-benzyl)-Leu-Arg-Pro-NHEt), leuprolide (Pyr-His-Trp-Ser-Tyr-D-Leu-Leu-Arg-Pro-NHEt), nafarelin (Pyr-His-Trp-Ser-Tyr-2Nal-Leu-Arg-Pro-Gly-NH ２ ), Triptorelin (Pyr-His-Trp-Ser-Tyr-D-Trp-Leu-Arg-Pro-Gly-NH ２ ), nafarelin, deslorelin, abarelix (Ac-D-2Nal-D-4-chloroPhe-D-3-(3-pyridyl)Ala-Ser-(N-Me)Tyr-D-Asn-Leu-isopropylLys-Pro-DAla-NH ２ ), Cetrorelix (Ac-D-2Nal-D-4-chloroPhe-D-3-(3-pyridyl)Ala-Ser-Tyr-D-Cit-Leu-Arg-Pro-D-Ala-NH ２ ), degarelix (Ac-D-2Nal-D-4-chloroPhe-D-3-(3-pyridyl)Ala-Ser-4-aminoPhe(L-hydroorotyl)-D-4-aminoPhe(carbamoyl)-Leu-isopropylLys-Pro-D-Ala-NH ２ ), and ganirelix (Ac-D-2Nal-D-4-chloroPhe-D-3-(3-pyridyl)Ala-Ser-Tyr-D-(N9,N10-diethyl)-homoArg-Leu-(N9,N10-diethyl)-homoArg-Pro-D-Ala-NH ２ ); pattern recognition receptors (PRRs) selected from Toll-like receptor (TLR) ligands, C-type lectins, and Nodlike receptor (NLR) ligands; calcitonin receptor agonists; integrin receptors and their receptor subtypes (α V β 1 , α V β 3 , α V β 5 , α V β 6 , α 6 β 4 , α 7 β 1 , α L β 2 , α IIb β 3 agonists (selected from the group consisting of GRGDSPK, cyclo(RGDfV)(L1) and its derivatives [cyclo(-N(Me)R-GDfV), cyclo(R-Sar-DfV), cyclo(RG-N(Me)D-fV), cyclo(RGD-N(Me)f-V), cyclo(RGDf-N(Me)V-)(cilengitide)]; anticalins (derivatives of lipocalin); adnectins (tenth FN3 (fibronectin)); designed ankyrin repeat proteins (DARPins); avimers; EGF receptor or VEGF receptor agonists; (8) A pharma- ceutically acceptable salt, acid, derivative, hydrate or hydrated salt; or a crystal structure; or an optical isomer, racemate, diastereomer, or enantiomer, of any of the above drugs.

12. The cytotoxins D, D １ and D. ２ are independently an anti-microtubule agent, a DNA minor groove binder, an RNA polymerase II inhibitor, a DNA topoisomerase I or III inhibitor, or a DNA alkylating agent, and are selected from the group consisting of tubulysin, amanitin, auristatin (including analogs of AFP, MMAF, MMAE, AEB, AEBA, E), calicheamicin, camptothecin (including SN-38, topotecan, or exatecan), etoposide, teniposide, daunomycin, doxorubicin (including morpholino-doxorubicin, cyanomorpholino-doxorubicin), duocarmycin (including CC1605 analogs, DC1, DC4, CBI-dimer), dolastatin, enediyne, eribulin, lexitropsin, taxane, puromycin, 11. The antibody-drug conjugate of claim 10, wherein the conjugate is selected from the group consisting of sirolimusin, maytansinoids, vinca alkaloids, paclitaxel, docetaxel, pyrrolobenzodiazepines (including PBD, IGN), rhizoxin, dolastatin, echinomycin, combrestatin, vinca alkaloids, calicheamacycin, maytansine (DM1, DM4, DM21), geldanamycin, vinblastine, methotrexate, hemiasterin, netropsin, nicotinamide phosphoribosyltransferase inhibitors (NAMPT), spliceostatin, pladienolide, protein kinase inhibitors, MEK inhibitors, proteinase inhibitors, immunotoxins, cell receptor agonists, and derivatives or analogs thereof. (a) Tubulysin and its analogs, or derivatives having one or more isotopes, or pharma- ceutically acceptable salts, hydrates, or hydrated salts, or polymorphic crystal structures, or optical isomers, racemates, diastereomers, or enantiomers thereof, having the following formula (IV): 【Chemistry 2】 During the ceremony, 【Chemistry 3】 One or two of them are independently L １ And / or L ２ is a binding site capable of binding to; Two 【Chemistry 4】 L １ and L ２ When R １ and R ２ , or Z ２ and Z ３ is preferably the double bond site. R １ , R １’ , R ２ , R ３ , and R ４ are independently H, C １ ~C ８ Alkyl; C ２ ~C ８ Heteroalkyl or heterocycle; C ３ ~C ８ Ar is aryl, Ar-alkyl, cycloalkyl, alkylcycloalkyl, heterocycloalkyl, heteroalkylcycloalkyl, carbocycle, or alkylcarbonyl; or R １ R ２ , R １ R ３ , R ２ R ３ , R ３ R ４ , R ５ R ６ , R １１ R １２ , or R １３ R １４ form a 3- to 7-membered carbocyclic, cycloalkyl, heterocyclic, heterocycloalkyl, aromatic, or heteroaromatic ring system; L １ Or L ２ When independently or simultaneously bound to R １ and R ２ may be independently absent; R ５ , R ６ , R ８ , R １０ , and R １１ are independently H or C １ ~C ４ is alkyl or heteroalkyl; R ７ are independently H, R １４ , -R １４ C(=O)X １ R １５ or -R １４ X １ R １５ is; １ is O, S, SS, NH, CH ２ , or NR １４ is; R ９ is H, OH, =O, -OR １４ , -OC(=O)R １４ , -OC(=O)NHR １４ -, -OC(=O)R １４ R １５ -, OP (=O) (OR １４ ) ２ -, -OC(=O)NR １４ R １５ OR １４ OP(=O)(OR １５ ) ２ R is selected from ９ L １ Or L ２ When bonded to R ９ is -O-, -OC(=O)NH-, or -OC(=O)N(R １４ )-is; R １１ are independently H, R １４ , -R １４ C(=O)R １５ , -R １４ C(=O)X ２ R １５ wherein X ２ is -O-, -S-, -NH-, or -N(R １４ )-is; R １２ -COOH, -COSH, -CONH ２ , C.O.N.H. ２ , C.O.N.H.R. １５ , -CONH(R １５ ), -COOR １５ , -R １５ C.O.R. １６ , -R １５ COOR １６ , -R １５ C(O)NH ２ , -R １５ C(O)NHR １６ , -COSR １５ , R １５ S (=O) ２ R １６ , -R １５ P(=O)(OR １７ ) ２ , -R １５ OP(=O)(OR １７ ) ２ , -COOCH ２ OP(=O)(OR １７ ) ２ , -COX ２ SO ２ R １７ , -COOR １５ X ２ R １６ , tetrazole, imidazole, or triazole; X ２ is -O-, -S-, -NH-, -N(R １５ ) -, -OR １５ -, -SR １５ -, CH ２ , or -NHR １５ -is; R １２ L １ Or L ２ When bonded to R １２ is -C(O)O-, -C(O)NH-, -C(=O)NHS(O) ２ R １５ -, or -C(=O)N(R １５ )-is; R １３ and R １４ is independent, C １ ~C ８ Alkyl, heteroalkyl: C ２ ~C ８ Alkenyl, alkynyl, heteroalkyl, heterocycloalkyl; ３ ~C ８ Aryl, Ar-alkyl; Z ２ and Z ３ are independently H, O, S, NH, N(R １５ ), NHNH, -OH, -SH, -NH ２ , NH, NHNH2, -NH(R １５ ), -OR １５ , CO, -COX2, -COX2R １６ , R １７ ,F,Cl,Br,I,SR １６ , N.R. １６ R １７ , N=NR １６ , N=R １６ , NO2, SOR １６ R １７ , SO2R １６ , SO3R １６ , OSO3R １６ , P.R. １６ R １７ , P.O.R. １６ R １７ , PO2R １６ R １７ , OP(O)(OR １７ )2, OCH2OP(O)(OR １７ ) 2, O.C.(O)R １７ ,OC(O)OP(O)(OR １７ ) 2, PO (OR １６ ) (OR １７ ), OP(O)(OR １７ ) OP (O) (OR １７ )2, OC(O)NHR １７ -O-(C ４ ~C １２ Glycoside), -N-(C ４ ~C １２ Glycoside); C １ ~C ８ Alkyl, heteroalkyl; ２ ~C ８ Alkenyl, alkynyl, heteroalkyl, heterocycloalkyl; ３ ~C ８ Aryl, Ar-alkyl, carbocyclic, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl, or ester, ether, or amide of 2 to 8 carbon atoms; or a peptide containing 1 to 8 amino acids (NH(Aa) １～８ Or CO(Aa) １～８ (N-terminus or C-terminus (Aa) ｒ , r=1 to 8 identical or different amino acid sequences), or ２ CH ２ ) ｐ Or (OCH ２ CH (CH ３ )) ｐ where p is an integer from 0 to about 1000, or a combination of the above groups; X ２ is O, S, SS, NH, CH ２ , O.H., S.H., and N.H. ２ , CHR １５ , or NR １５ is; R １５ , R １６ , and R １７ are independently H, C １ ~C ８ Alkyl, heteroalkyl; ２ ~C ８ Alkenyl, alkynyl, heteroalkyl, heterocycloalkyl; ３ ~C ８ Aryl, Ar-alkyl, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl, alkylcarbonyl, or Na ＋ , K ＋ , Cs ＋ , Li ＋ , Ca ２＋ , Mg ＋ , Zn ２＋ , N ＋ (R １ ) (R ２ ) (R ３ ) (R ４ ), or HN ＋ (C ２ H ５ O.H. ３ It is salt; Y １ and Y ２ is independently N or CH; q is 0 or 1; when q=0, Y ３ does not exist, and Y ４ , Y ５ , Y ６ , and Y ７ are independently CH, N, NH, O, S, or N(R1), so that Y ２ , Y ４ , Y ５ , Y ６ , and Y ７ form a heteroaromatic ring of furan, pyrrole, thiophene, thiazole, oxazole, imidazole, pyrazole, triazole, tetrazole, thiadiazole; when q=1, Y ３ , Y ４ , Y ５ , Y ６ , and Y ７ are independently CH or N, so that Y ２ , Y ３ , Y ４ , Y ５ , Y ６ , and Y ７ forms an aromatic ring in benzene, pyridine, pyridazine, pyrimidine, pyrazine, triazine, tetrazine, or pentazine; In particular, tubulysin analogs have the structure shown below: 【Chemistry 5】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 During the ceremony, R ２０ is H;C １ ~C ８ A linear or branched alkyl or heteroalkyl group of ２ ~C ８ Straight-chain or branched alkenyl, alkynyl, alkylcycloalkyl, heterocycloalkyl; ３ ~C ８ Ar-alkyl, heterocycle, carbocycle, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl; carbonate (-C(O)OR １７ ), carbamates (-C(O)NR １７ R １８ or a carboxylate, ester, ether, or amide having 1 to 8 carbon atoms; or an amino acid having 1 to 8 carbon atoms; or a group represented by the formula (OCH ２ CH ２ ) ｐ Or (OCH ２ CH (CH ３ )) ｐ where p is an integer from 0 to about 1000; or ２０ is absent and the oxygen atom forms a ketone, or a combination of the above; Z ２ and Z ３ are independently H, OH, NH ２ , O, NH, COOH, COO, C(O), C(O), C(O)NH, C(O)NH ２ , R １８ , O.C.H. ２ OP(O)(OR １８ ) ２ ,OC(O)OP(O)(OR １８ ) ２ , OPO (OR １８ ) ２ , NHPO (OR １８ ) ２ , OP(O)(OR １８ ) OP (O) (OR １８ ) ２ , O.C.(O)R １８ , O.C.(O)N.R. １８ , OSO ２ (OR １８ ), O-(C ４ ~C １２ -glycoside), C １ ~C ８ Linear or branched alkyl or heteroalkyl of C ２ ~C ８ Straight-chain or branched alkenyl, alkynyl, alkylcycloalkyl, heterocycloalkyl; ３ ~C ８ Ar-alkyl, heterocycle, carbocycle, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl; carbonate (-C(O)OR １７ ), carbamates (-C(O)NR １７ R １８ ) is; R １７ and R １８ are independently H, linear or branched alkyl or heteroalkyl, C ２ ~C ８ Straight-chain or branched alkenyl, alkynyl, alkylcycloalkyl, heterocycloalkyl; ３ ~C ８ Ar-alkyl, heterocycle, carbocycle, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl; carbonate (-C(O)OR １７ ), carbamates (-C(O)NR １７ R １８ ) is; R １９ is H, OH, NH ２ , OSO ２ (OR １８ ), X.H.H. ２ OP(O)(OR １８ ) ２ , XPO(OR １８ ) ２ ,XC(O)OP(O)(OR １８ ) ２ , XC(O)R １８ , XC(O)NHR １８ , C １ ~C ８ Alkyl or carboxylate; C ２ ~C ８ Alkenyl, alkynyl, alkylcycloalkyl, heterocycloalkyl; C ３ ~C ８ aryl or alkylcarbonyl; or a pharmaceutical salt; X is O, S, NH, NHNH, or CH ２ is; R ７ is the same as defined above; Bonding sites in formulae (IV-01) to (IV-94) 【Chemistry 6】 is omitted here but follows in the same position as shown in formula (IV). (b) calicheamicin and its related enediyne antibiotics, or isotopes of chemical elements, or pharma-ceutically acceptable salts, hydrates, or hydrated salts, or polymorphic crystal structures, or optical isomers, racemates, diastereomers, or enantiomers thereof, having the following structural formula: 【Chemistry 7】 (c) geldanamycin, having the following structure: 【Chemistry 8】 In the formula, X is O, NH, CH ２ and 【Chemistry 9】 L １ Or L ２ This is the binding site for (d) a maytansinoid, which is maytansine or a derivative thereof having the formula: 【Chemistry 10】 (e) Camptothecin (CPT) or a derivative thereof having the following formula: 【Chemistry 11】 During the ceremony, R １ , R ２ , and R ４ are independently H, F, Cl, Br, CN, NO ２ , C １ ~C ８ Alkyl; O-C １ ~C ８ Alkyl; NH-C １ ~C ８ Alkyl; C ２ ~C ８ Heteroalkyl, alkylcycloalkyl, heterocycloalkyl; C ３ ~C ８ Aryl, Ar-alkyl, heterocycle, carbocycle, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl; or an ester, ether, amide, carbonate, urea, or carbamate having 2 to 8 carbon atoms; R ３ is H, OH, NH ２ , C １ ~C ８ Alkyl; O-C １ ~C ８ Alkyl; NH-C １ ~C ８ Alkyl; C ２ ~C ８ heteroalkyl, alkylcycloalkyl, heterocycloalkyl; or an ester, ether, amide, carbonate, urea, or carbamate having from 2 to 8 carbon atoms; Or, R １ R ２ , R ２ R ３ , and R ３ R ４ independently form a 5-7 membered carbocyclic, heterocyclic, heterocycloalkyl, aromatic, or heteroaromatic ring system; 【Chemistry 12】 is an intramolecular site capable of binding to L1 or L2; Said camptothecin (CPT) or its derivatives (Cpt-I), or isotopes of one or more chemical elements, or pharma- ceutically acceptable salts, hydrates or hydrated salts, or polymorphic crystal structures of these compounds, or optical isomers, racemates, diastereomers or enantiomers, in particular have the following formula: 【Chemistry 13】 【change】 【change】 【change】 【change】 During the ceremony, P １ is H, OH, NH ２ , COOH, C(O)NH ２ , O.C.H. ２ OP(O)(OR １８ ) ２ ,OC(O)OP(O)(OR １８ ) ２ , OPO (OR １８ ) ２ , NHPO (OR １８ ) ２ , O.C.(O)R １８ , OP(O)(OR １８ ) OP (O) (OR １８ ) ２ , O.C.(O)N.R. １８ , O C (O) N (C ２ H ４ ) ２ NCH ３ , OSO ２ (OR １８ ), O-(C ４ -C １２ -glycoside), OC(O)N(C ２ H ４ ) ２ CH ２ N (C ２ H ４ ) ２ CH ３ , O-(C １ ~C ８ (straight-chain or branched alkyl) １ ~C ８ linear or branched alkyl or heteroalkyl of ２ ~C ８ Straight-chain or branched alkenyl, alkynyl, alkylcycloalkyl, heterocycloalkyl; ３ ~C ８ Ar-alkyl, heterocyclic, carbocyclic, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl; carbonate (-C(O)OR １７ ), carbamates (-C(O)NR １７ R １８ ); R １７ and R １８ are independently H, linear or branched alkyl or heteroalkyl; C ２ ~C ８ Straight-chain or branched alkenyl, alkynyl, alkylcycloalkyl, heterocycloalkyl; ３ ~C ８ Ar-alkyl, heterocyclic, carbocyclic, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl; １７ ), carbamates (-C(O)NR １７ R １８ ) is; R １ and R ２ are independently H; O-C １ ~C ８ Alkyl; C ２ ~C ８ Heteroalkyl, alkylcycloalkyl, heterocycloalkyl; C ３ ~C ８ Ar-aryl, Ar-aralkyl; Connection parts in the above CPT structures (Icp-01 to Icp-24) 【Chemistry 14】 is omitted here, but has the same definition as the position indicated by (Cpt-I). (f) Combretastatin having the formula: or one or more elemental isotopic substitutions, or a pharma- ceutically acceptable salt, hydrate, or hydrated salt, or a polycrystalline structure of these compounds, or an optical isomer, racemate, non-enantiomer, or enantiomer: 【Chemistry 15】 (g) Taxanes and derivatives thereof having the following formula: or one or more elemental isotopic substitutions, or pharma- ceutically acceptable salts, hydrates, or hydrated salts, or polycrystalline structures of these compounds, or chiral isomers, racemates, enantiomers, or enantiomers: 【Chemistry 16】 During the ceremony, 【Chemistry 17】 L １ Or L ２ Ar and Ar' are independently aryl or heteroaryl. (h) Anthracyclines and derivatives thereof having the following formula: or one or more elemental isotopic substitutions, or pharma- ceutically acceptable salts, hydrates, or hydrated salts, or polycrystalline structures of these compounds, or optical rotators, racemates, enantiomers, or enantiomers: 【Chemistry 18】 【change】 (i) Vinca alkaloids and derivatives thereof having the following formula: or one or more elemental isotopic substitutions, or pharma- ceutically acceptable salts, hydrates, or hydrated salts, or polycrystalline structures of these compounds, or optical rotators, racemates, enantiomers, or enantiomers: 【Chemistry 19】 【change】 (j) dolastatins, auristatins, and derivatives thereof having the following formulae (Ih-01), (Ih-02), (Ih-03), (Ih-04), (Ih-05), (Ih-06), (Ih-07), (Ih-08), (Ih-09), (Ih-10), and (Ih-11), or one or more elemental isotope substitutes, or pharma- ceutically acceptable salts, hydrates, or hydrated salts, or polycrystalline structures of these compounds, or optical rotators, racemates, non-enantiomers, or enantiomers: 【Chemistry 20】 【change】 During the ceremony, R １ , R ２ , R ３ , R ４ , and R ５ are independently H; C １ ~C ８ a linear or branched alkyl, aryl, heteroaryl, heteroalkyl, alkylcycloalkyl, ester, ether, amide, amine, heterocycloalkyl, or acyloxylamine of the formula (OCH ２ CH ２ ) ｐ Or (OCH ２ CH (CH ３ )) ｐ where p is an integer from 1 to about 5000; １ R ２ , R ２ R ３ , R １ R ３ , or R ３ R ４ may be taken together to form a 3-8 membered ring of an alkyl, aryl, heteroaryl, heteroalkyl, or alkylcycloalkyl group; Y １ and Y ２ is a connection site (which binds independently to L1 and / or L2) 【Chemistry 21】 and independently O, NH, NHNH, NR ５ , S, C(O)O, C(O)NH, OC(O)NH, OC(O)O, NHC(O)NH, NHC(O)S, OC(O)N(R １ ), N(R １ ) C(O)N(R ２ ), C(O)NHNHC(O), and C(O)NHR １ or a connection site 【Chemical 22】 If not bonded to OH, NH ２ , N.H.N.H. ２ , N.H.R. ５ , SH, C(O)OH, C(O)NH ２ , OC(O)NH ２ , OC(O)OH, NHC(O)NH ２ , NHC(O)SH, OC(O)NH(R １ ), N(R １ )C(O)NH(R ２ ), C(O)NHNHC(O)OH, and C(O)NHR １ is; R １２ は、OH、NH ２ 、NHR １ NHNH ２ 、NHNHCOOH、O-R １ -COOH、NH-R １ -COOH、NH-(Aa) ｎ COOH, O(CH ２ CH ２ O) ｐ CH ２ CH ２ Hmm, Hmm ２ CH ２ O) ｐ CH ２ CH ２ NH ２ NH(CH ２ CH ２ O) ｐ CH ２ CH ２ NH ２ NR １ R １’ 、NHOH、NHOR １ O(CH ２ CH ２ O) ｐ CH ２ CH ２ COOH, NH(CH ２ CH ２ O) ｐ CH ２ CH ２ COOH、NH-Ar-COOH、NH-Ar-NH ２ O(CH ２ CH ２ O) ｐ CH ２ CH ２ P.S. ３ H,NH(CH ２ CH ２ O) ｐ CH ２ CH ２ P.S. ３ H １ -NHSO ３ H １ -NHSO ３ H、O(CH ２ CH ２ O) ｐ CH ２ CH ２ NHPO ３ H ２ 、NH(CH ２ CH ２ O) ｐ CH ２ CH ２ NHPO ３ H ２ 、OR １ 、R １ -NHPO ３ H ２ 、R １ -OPO ３ H ２ 、O(CH ２ CH ２ O) ｐ CH ２ CH ２ OPO ３ H ２ 、OR １ -NHPO ３ H ２ 、NH-R １ -NHPO ３ H ２ 、NH(CH ２ CH ２ NH) ｐ CH ２ CH ２ NH ２ 、NH(CH ２ CH ２ S) ｐ CH ２ CH ２ NH ２ 、NH(CH ２ CH ２ NH) ｐ CH ２ CH ２ OH、NH(CH ２ CH ２ S) ｐ CH ２ CH ２ OH、NH-R １ -NH ２ 、或NH(CH ２ CH ２ O) ｐ CH ２ CH ２ NHPO ３ H ２ where Aa is 1 to 8 identical or different amino acids; p is 1 to 5000; R １ , R ２ , R ３ , R ４ , R ５ , R ５’ , Z １ , Z ２ , and n are as defined above. (k) Hemiasterlin and its derivatives, or one or more elemental isotope substitutions, or pharma- ceutically acceptable salts, hydrates, or hydrated salts, or polycrystalline structures of these compounds, or optical isomers, racemates, non-enantiomers, or enantiomers, having the following formula: 【Chemistry 23】 During the ceremony, 【Chemistry 24】 L １ Or L ２ R １ , R ２ , R ３ , R ４ , and R ５ are independently H; C １ ~C ８ linear or branched alkyl, aryl, heteroaryl, heteroalkyl, alkylcycloalkyl, ester, ether, amide, amine, heterocycloalkyl, or acyloxylamine; or a peptide containing 1 to 8 amino acids, or a peptide of the formula (OCH ２ CH ２ ) ｐ Or (OCH ２ CH (CH ３ )) ｐ where p is an integer from 1 to about 5000; ２ R ３ can form a 3-8 membered ring of an alkyl, aryl, heteroaryl, heteroalkyl, or alkylcycloalkyl group. (l) Eribulin and derivatives thereof having the following formula: or one or more elemental isotope substitutions, or a pharma- ceutically acceptable salt, hydrate, or hydrated salt, or a polycrystalline structure of these compounds, or an optical rotator, racemate, non-enantiomer, or enantiomer: 【Chemistry 25】 (m) Nicotinamide phosphoribosyltransferase (NAMPT) inhibitors having the following formulas NP01, NP02, NP03, NP04, NP05, NP06, NP07, NP08, and NP09, or one or more isotopes of chemical elements, or pharma- ceutically acceptable salts, hydrates, or hydrated salts, or polymorphic crystal structures of these compounds, or optical isomers, racemates, diastereomers, or enantiomers: 【Chemistry 26】 During the ceremony, 【Chemistry 27】 is the same as above; X ５ is F, Cl, Br, I, OH, OR １ , R １ , O.P.O. ３ H ２ , OSO ３ H, N.H.R. １ , O.C.O.R. １ , NHCOR １ is; (n) Benzodiazepine dimers and analogs thereof having the formula PB01, PB02, PB03, PB04, PB05, PB06, PB07, PB08, PB09, PB10, PB11, PB12, PB13, PB14, PB15, PB16, PB17, PB18, PB19, PB20, or one or more isotopes of chemical elements, or pharma- ceutically acceptable salts, hydrates, or hydrated salts, or polymorphic crystal structures of these compounds, or optical isomers, racemates, diastereomers, or enantiomers: 【Chemistry 28】 【change】 【change】 【change】 During the ceremony, X １ , X ２ , Y １ , and Y ２ , Z １ , Z ２ and n is as defined above; preferably, X １ , X ２ , Y １ , and Y ２ are independently O, N, NH, NHNH, NR ５ , S, C(O)O, C(O)NH, OC(O)NH, OC(O)O, NHC(O)NH, NHC(O)S, OC(O)N(R １ ), N(R １ ) C(O)N(R １ ), CH, C(O)NHNHC(O), and C(O)NR １ is; R １ , R ２ , R ３ , R １’ , R ２’ , and R ３’ are independently H; F; Cl; =O; =S; OH; SH; C １ -C ８ Linear or branched alkyl, aryl, alkenyl, heteroaryl, heteroalkyl, alkylcycloalkyl, ester (COOR ５ or -OC(O)R ５ ), ester (OR ５ ), amide (CONR ５ ), carbamates (OCONR ５ ), amine (NHR ５ , N.R. ５ R ５’ ), heterocycloalkyl, or acyloxylamine (-C(O)NHOH, -ONHC(O)R ５ ); or a peptide comprising 1 to 20 natural or unnatural amino acids, or a peptide of the formula (OCH ２ CH ２ ) ｐ Or (OCH ２ CH (CH ３ )) ｐ where p is an integer from 1 to about 1000; １ R ２ , R ２ R ３ , R １ R ３ , R １’ R ２’ , R ２’ R ３’ , or R １’ R ３’ can independently form a 3-8 membered ring of an alkyl, aryl, heteroaryl, heteroalkyl, or alkylcycloalkyl group; X ３ and Y ３ are independently N, NH, CH, or CR １ or X ３ and Y ３ Any one of may be absent; R １ and R ２ is C １ ~C ８ Linear or branched alkyl, heteroalkyl; ３ ~C ８ aryl, heteroaryl, alkylcycloalkyl, acyloxyl, alkylaryl, alkylaryloxyl, alkylarylamino, alkylarylthiol; or 1 to 6 identical or different amino acid / peptide sequences (Ar)r (r=1-6); R ４ , R ５ , R ５’ , R ６ , R １２ , and R １２’ are independently H, OH, NH ２ , NH(CH ３ ), N.H.N.H. ２ , COOH, SH, OZ ３ , S.Z. ３ , F, Cl, or C １ -C ８ Linear or branched alkyl, aryl, heteroaryl, heteroalkyl, alkylcycloalkyl, acyloxylamine; Z ３ is H, OP(O)(OM １ ) (O.M. ２ ), O.C.H. ２ OP(O)(OM １ ) (O.M. ２ ), OSO ３ M １ , or O-glycosides (glucosides, galactosides, mannosides, glucuronosides / glucuronides, allosides, fructosides, etc.), NH-glycosides, S-glycosides, or CH ２ - is a glycoside; M １ and M. ２ are independently H, Na, K, Ca, Mg, NH ４ , N.R. １ R ２ R ３ is; X ６ are CH, N, P(O)NH, P(O)NR １ , CHC(O)NH,C ３ -C ８ aryl, heteroaryl, alkylcycloalkyl, acyloxyl, alkylaryl, alkylaryloxyl, alkylarylamino, or Aa (amino acid, preferably selected from Lys, Phe, Asp, Glu, Ser, Thr, His, Cys, Tyr, Trp, Gln, Asn, Arg); 【Chemical 29】 is the same as defined above; (o) CC-1065 analogs and doucarmycin analogs and derivatives thereof having the structures of formulas CC01, CC02, CC03, CC04, CC05, CC06, and CC07: 【Chemistry 30】 【change】 During the ceremony, X １ , X ２ , Y １ , and Y ２ is the connection site 【Chemistry 31】 and independently O, NH, NHNH, NR ５ , S, C(O)O, C(O)NH, OC(O)NH, OC(O)O, NHC(O)NH, NHC(O)S, OC(O)N(R １ ), N(R １ ) C(O)N(R ２ ), C(O)NHNHC(O), and C(O)NR １ or a connection site 【Chemistry 32】 If not bonded to OH, NH ２ , N.H.N.H. ２ , N.H.R. １ , SH, C(O)OH, C(O)NH ２ , OC(O)NH ２ , OC(O)OH, NHC(O)NH ２ , NHC(O)SH, OC(O)NH(R １ ), N(R １ )C(O)NH(R ２ ), C(O)NHNHC(O)OH, and C(O)NHR １ is; Z ３ , H, PO (OM １ ) (O.M. ２ ), S.O. ３ M １ , C.H. ２ PO (OM １ ) (O.M. ２ ), C.H. ３ N (CH ２ CH ２ ) ２ NC(O)-, O(CH ２ CH ２ ) ２ N-C(O)-, R １ or a glycoside; R １ , R ２ , R ３ , M. １ , M. ２ , and n are as defined above. (p) Amatoxin and its analogs, or isotopic substitutions of one or more chemical elements, or pharma- ceutically acceptable salts, hydrates, or hydrated salts, or polycrystalline structures of these compounds, or optical isomers, racemates, diastereomers, or enantiomers, having the following formulae Am01, Am02, and Am03: 【Chemical Formula 33】 During the ceremony, X １ and Y １ are independently O, NH, NHNH, NR ５ , S, C(O)O, C(O)NH, OC(O)NH, OC(O)O, NHC(O)NH, NHC(O)S, OC(O)N(R １ ), N(R １ ) C(O)N(R １ ), C.H. ２ , CHNH, CH ２ O, C(O)NHNHC(O), and C(O)NR １ is; R ７ , R ８ , and R ９ are independently H, OH, OR １ , N.H. ２ , N.H.R. １ , C １ ~C ６ Alkyl, or lacking; Y ２ , O, O ２ , N.R. １ , NH, or absent; R １０ is CH ２ , O., N.H., N.R. １ , NHC(O), NHC(O)NH, NHC(O)O, OC(O)O, C(O), OC(O), OC(O)(NR １ ), (NR １ ) C(O)(NR １ ), C(O)R １ , or lacking; R １１ は、OH、NH ２ 、NHR １ NHNH ２ 、NHNHCOOH、O-R １ -COOH、NH-R １ -COOH、NH-(Aa) ｒ COOH, O(CH ２ CH ２ O) ｐ CH ２ CH ２ Hmm, Hmm ２ CH ２ O) ｐ CH ２ CH ２ NH ２ NH(CH ２ CH ２ O) ｐ CH ２ CH ２ NH ２ NR １ R ２ O(CH ２ CH ２ O) ｐ CH ２ CH ２ -COOH, A. ２ CH ２ O) ｐ CH ２ CH ２ COOH、NH-Ar-COOH、NH-Ar-NH ２ O(CH ２ CH ２ O) ｐ CH ２ CH ２ -NHSO ３ H,NH(CH ２ CH ２ O) ｐ CH ２ CH ２ P.S. ３ H １ -NHSO ３ H １ -NHSO ３ H、O(CH ２ CH ２ O) ｐ -CH ２ CH ２ P. ３ H ２ , NH(CH ２ CH ２ O) ｐ CH ２ CH ２ N.H.P.O. ３ H ２ , OR １ , R １ -NHPO ３ H ２ , R １ -OPO ３ H ２ , O(CH ２ CH ２ O) ｐ CH ２ CH ２ O.P.O. ３ H ２ , OR １ -NHPO ３ H ２ , N.H.-R. １ -NHPO ３ H ２ , or NH(CH ２ CH ２ O) ｐ CH ２ -CH ２ N.H.P.O. ３ H ２ where: (Aa) ｒ is 1 to 8 amino acids; n and m １ is independently 1 to 20; p is 1 to 5000; R １ , R ２ , Ar, and 【Chemical 34】 is the same as the definition above. (q) spliceostatins and pladienolides and derivatives thereof, or isotopic substitutions of one or more chemical elements, or pharma- ceutically acceptable salts, hydrates, or hydrated salts, or polycrystalline structures of these compounds, or optical isomers, racemates, diastereomers, or enantiomers, having the following formula: 【Chemistry 35】 (r) Protein kinase inhibitors and derivatives thereof having the following formulas PK01-PK40, or isotopic substitutions of one or more chemical elements, or pharma- ceutically acceptable salts, hydrates, or hydrated salts, or polycrystalline structures of these compounds, or optical isomers, racemates, diastereomers, or enantiomers: 【Chemical 36】 【change】 【change】 【change】 【change】 【change】 During the ceremony, Z ５ and Z ５’ are independently O, NH, NHNH, NR ５ , S, C(O)O, C(O)NH, OC(O)NH, OC(O)O, NHC(O)O, NHC(O)NH, NHC(O)S, OC(O)N(R １ ), N(R １ ) C(O)N(R ２ ), C(O)NHNHC(O), and C(O)NR １ Selected from: 【Chemical 37】 L １ Or L ２ is the binding site for (s) MEK inhibitors and derivatives thereof, or isotopic substitutions of one or more chemical elements, or pharma- ceutically acceptable salts, hydrates, or hydrated salts, or polycrystalline structures of these compounds, or optical isomers, racemates, diastereomers, or enantiomers, having the following formula: 【Chemical 38】 During the ceremony, Z ５ is O, NH, NHNH, NR ５ , S, C(O)O, C(O)NH, OC(O)NH, OC(O)O, NHC(O)O, NHC(O)NH, NHC(O)S, OC(O)N(R １ ), N(R １ ) C(O)N(R ２） , C(O)NHNHC(O), and C(O)NR １ Selected from: 【Chemical 39】 L １ Or L ２ is the binding site for (t) Protease inhibitors and derivatives thereof, or isotopic substitutions of one or more chemical elements, or pharma- ceutically acceptable salts, hydrates, or hydrated salts, or polycrystalline structures of these compounds, or optical isomers, racemates, diastereomers, or enantiomers, having the following formula: 【Chemistry 40】 (u) Immunotoxins useful as payloads of the conjugates of the invention are selected from diphtheria toxin (DT), cholera toxin (CT), trichosanthin (TCS), dianthin, Pseudomonas exotoxin A (ETA), erythrogenic toxin, diphtheria toxin, AB toxin, type III exotoxin, proaerolysin, and topsalysin; the immunotoxins are assembled with the antibodies of the invention via bioengineering of protein fusions or via free amino, thiol, or carboxyl groups of amino acids of the immunotoxin; (v) Cell-binding molecules / ligands or cell receptor agonists and derivatives thereof, or isotopic substitutions of one or more chemical elements, or pharma- ceutically acceptable salts, hydrates, or hydrated salts, or polycrystalline structures of these compounds, or optical isomers, racemates, diastereomers, or enantiomers, having the following formula: 【Chemistry 41】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 During the ceremony, 【Chemistry 42】 L １ Or L ２ is a binding site for ４ and Y １ are independently O, NH, NHNH, NR １ , S, C(O)O, C(O)NH, OC(O)NH, OC(O)O, NHC(O)NH, NHC(O)S, OC(O)N(R １ ), N(R １ ) C(O)N(R １ ), C.H. ２ , C(O)NHNHC(O), and C(O)NR １ It is. (w) one or more DNA, RNA, mRNA, small interfering RNA (siRNA), microRNA (miRNA), and PIWI-interacting RNA (piRNA) conjugated to the BCMA antibody of the invention as a chemotherapeutic agent / functional compound, having the following structure: 【Chemistry 43】 During the ceremony, 【Chemistry 44】 is the attachment site for the side chain linker of this patent; 【Chemistry 45】 is a single or double strand of DNA, RNA, mRNA, siRNA, miRNA, or piRNA; X １ and Y is independently O, NH, NHNH, NR １ , S, C(O)O, C(O)NH, OC(O)NH, OC(O)O, NHC(O)NH, NHC(O)S, OC(O)N(R １ ), N(R １ ) C(O)N(R １ ), C.H. ２ , C(O)NHNHC(O), and C(O)NR １ It is.

13. Connector L １ and L ２ The antibody-drug conjugate of claim 10, wherein is independently selected from the group consisting of: O, NH, S, NHNH, N(R ３ ), N(R ３ ) N (R ３ '), or the formula (OCH ２ CH ２ ) ｐ OR ３ , (OCH ２ CH (CH ３ )) ｐ OR ３ , NH(CH ２ CH ２ O) ｐ OR ３ , NH(CH ２ CH (CH ３ ) O) pR3 ｐ OR ３ , N[(CH ２ CH ２ O) ｐ R ３ ][(CH ２ CH ２ O) ｐ’ R ３’ , (OCH ２ CH ２ ) ｐ COOR ３ , or C.H. ２ CH ２ (O.C.H. ２ CH ２ ) ｐ COOR ３ where p and p′ are independently an integer selected from 0 to about 1000, or a combination thereof; １ -C ８ Alkyl; C ２ -C ８ Heteroalkyl, alkylcycloalkyl, heterocycloalkyl; C ３ -C ８ aryl, Ar-alkyl, heterocycle, carbocycle, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl; ３ and R ３’ are independently H, C １ -C ８ Alkyl; C ２ -C ８ heteroalkyl, alkylcycloalkyl, aryl of heterocycloalkyl, Ar-alkyl, heterocycle, carbocycle, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl; or an ester, ether, or amide having 1 to 8 carbon atoms; or 1 to 8 natural or unnatural amino acids as described in the definitions; or a group of the formula (OCH ２ CH ２ ) ｐ Or (OCH ２ CH (CH ３ )) ｐ where p is an integer from 0 to about 1000, or a combination thereof; L １ Or L ２ may contain self-immolative or non-self-immolative moieties, peptidyl units, hydrazone bonds, disulfide, ester, oxime, amide, or thioether bonds, said self-immolative units including para-aminobenzylcarbamoyl (PAB) groups, including 2-aminoimidazole-5-methanol derivatives, heterocyclic PAB analogs, β-glucuronides, and ortho- or para-aminobenzyl acetals; The self-immolative linker moiety has any of the following structures: 【Chemistry 46】 In the formula, ( ＊ Atoms labeled with X are points of attachment to additional spacers, releasable linker units, cytotoxic agents, and / or cell binding molecules (CBAs); １ , Y １ , Z ２ , and Z ３ is independently NH, O or S; Z １ is independently H, NH, O, or S; v is 0 or 1; U １ are independently H, OH, C １ -C ６ Alkyl, (OCH ２ CH ２ ) ｎ ,F,Cl,Br,I,OR ５ , S.R. ５ , N.R. ５ R ５’ , N=NR ５ , N=R ５ , N.R. ５ R ５’ , NO ２ , SOR ５ R ５’ , S.O. ２ R ５ , S.O. ３ R ５ , OSO ３ R ５ , P.R. ５ R ５’ , P.O.R. ５ R ５’ , P.O. ２ R ５ R ５’ , OPO (OR ５ ) (OR ５’ ), or OCH ２ PO (OR ５ ) (OR ５’ ) in which R ５ and R ５’ is as defined above; preferably R ５ and R ５’ are independently H, C １ -C ８ Alkyl; C ２ -C ８ Alkenyl, alkynyl, heteroalkyl; ３ -C ８ aryl, heterocyclic, carbocyclic, cycloalkyl, heterocycloalkyl, heteroaralkyl, alkylcarbonyl; or a medicinal cationic salt; The non-self-immolative linker moiety has any of the following structures: 【Chemistry 47】 【change】 In the formula, ( ＊ ) are atoms marked with an additional spacer R １ , a releasable linker unit, a cytotoxic agent, and / or a point of attachment to a binding molecule; X １ , Y １ , U １ , R ５ , R ５’ is as defined above; r is 0 to 100; m and n are independently 0 to 6; L １ Or L ２ may be independently composed of one or more linker components which are 6-maleimidocaproyl ("MC"), maleimidopropanoyl ("MP"), valine-citrulline ("val-cit" or "vc"), alanine-phenylalanine ("ala-phe" or "af"), p-aminobenzyloxycarbonyl ("PAB"), 4-thiopentanoic acid ("SPP"), 4-(N-maleimidomethyl)cyclohexane-1 carboxylate ("MCC"), (4-acetyl)aminobenzoic acid ("SIAB"), 4-thiobutyric acid (SPDB), 4-thio-2-hydroxysulfonylbutyric acid (2-sulfo-SPDB), or a natural or unnatural peptide having from 1 to 8 natural or unnatural amino acid units. L １ Or L ２ The releasable linker (L) may be a linker that includes at least one bond that can be broken down under physiological conditions, such as being pH labile, acid labile, base labile, oxidatively labile, metabolically labile, biochemically labile, or enzymatically labile. １ Or L ２ Examples of ) include: -(CR ５ R ６ ) ｍ (Aa) ｒ (CR ７ R ８ ) ｎ (O.C.H. ２ CH ２ ) ｔ -, - (CR ５ R ６ ) ｍ (CR ７ R ８ ) ｎ (Aa) ｒ (O.C.H. ２ CH ２ ) ｔ -, -(Aa) ｒ - (CR ５ R ６ ) ｍ (CR ７ R ８ ) ｎ (O.C.H. ２ CH ２ ) ｔ -, - (CR ５ R ６ ) ｍ (CR ７ R ８ ) ｎ (O.C.H. ２ CH ２ ) ｒ (Aa) ｔ -, - (CR ５ R ６ ) ｍ - (CR ７ =CR ８ ) (CR ９ R １０ ) ｎ (Aa) ｔ (O.C.H. ２ CH ２ ) ｒ -, - (CR ５ R ６ ) ｍ (NR) １１ CON ｔ (CR) ９ R １０ ) ｎ (OCH) ２ CH ２ ) ｒ -、-(CR ５ R ６ ) ｍ (A) ｔ (NR) １１ CR) ９ R １０ ) ｎ (OCH) ２ CH ２ ) ｒ -、-(CR ５ R ６ ) ｍ (O) (A) ｔ (CR) ９ R １０ ) ｎ (OCH) ２ CH ２ ) ｒ -、-(CR ５ R ６ ) ｍ (OCNR) ７ )(Aa) ｔ (CR) ９ R １０ ) ｎ (OCH) ２ CH ２ ) ｒ -、-(CR ５ R ６ ) ｍ (C) (A) ｔ -(CR) ９ R １０ ) ｎ (OCH) ２ CH ２ ) ｒ -、-(CR ５ R ６ ) ｍ (NR) １１ CON ｔ (CR) ９ R １０ ) ｎ (OCH) ２ CH ２ ) ｒ -、-(CR ５ R ６ ) ｍ -(OCO)(Aa) ｔ (CR ９ R １０ ) ｎ - (OCH ２ CH ２ ) ｒ -, - (CR ５ R ６ ) ｍ (OCNR ７ ) (Aa) ｔ (CR ９ R １０ ) ｎ (O.C.H. ２ CH ２ ) ｒ -, - (CR ５ R ６ ) ｍ (CO) (Aa) ｔ (CR ９ R １０ ) ｎ (O.C.H. ２ CH ２ ) ｒ -, - (CR ５ R ６ ) ｍ -phenyl-(CO)(Aa) ｔ (CR ７ R ８ ) ｎ -, - (CR ５ R ６ ) ｍ -furyl-(CO)(Aa) ｔ (CR ７ R ８ ) ｎ -, - (CR ５ R ６ ) ｍ -oxazolyl-(CO)(Aa) ｔ (CR ７ R ８ ) ｎ -, - (CR ５ R ６ ) ｍ -Thiazolyl-(CO)(Aa) ｔ (CR ７ R ８ ) ｎ -, - (CR ５ R ６ ) ｔ -Thienyl-(CO)(CR ７ R ８ ) ｎ -, - (CR ５ R ６ ) ｔ -imidazolyl-(CO)(CR ７ R ８ ) ｎ -, - (CR ５ R ６ ) ｔ -Morpholino-(CO)(Aa) ｔ (CR ７ R ８ ) ｎ -, - (CR ５ R ６ ) ｔ -piperazino-(CO)(Aa) ｔ (CR ７ R ８ ) ｎ -, - (CR ５ R ６ ) ｔ -N-methylpiperazine-(CO)(Aa) ｔ (CR ７ R ８ ) ｎ -, - (CR ５ R ６ ) ｍ -(Aa) ｔ Phenyl-, -(CR ５ R ６ ) ｍ -(Aa) ｔ Frill-,-(CR ５ R ６ ) ｍ -Oxazolyl (Aa) ｔ -, - (CR ５ R ６ ) ｍ -Thiazolyl (Aa) ｔ -, - (CR ５ R ６ ) ｍ -Thienyl (Aa) ｔ -, - (CR ５ R ６ ) ｍ -Imidazolyl (Aa) ｔ -, - (CR ５ R ６ ) ｍ -Morpholino-(Aa) ｔ -, - (CR ５ R ６ ) ｍ -Piperazino- (Aa) ｔ -, - (CR ５ R ６ ) ｍ -N-methylpiperazino- (Aa) ｔ -, -K(CR ５ R ６ ) ｍ (Aa) ｒ (CR ７ R ８ ) ｎ (O.C.H. ２ CH ２ ) ｔ -, -K(CR ５ R ６ ) ｍ (CR ７ R ８ ) ｎ (Aa) ｒ (O.C.H. ２ CH ２ ) ｔ -, -K(Aa) ｒ (CR ５ R ６ ) ｍ (CR ７ R ８ ) ｎ (O.C.H. ２ CH ２ ) ｔ -, -K(CR ５ R ６ ) ｍ (CR ７ R ８ ) ｎ (O.C.H. ２ CH ２ ) ｒ (Aa) ｔ -, -K(CR ５ R ６ ) ｍ (CR ７ = R ８ ) (CR ９ R １０ ) ｎ (Aa) ｔ (O.C.H. ２ CH ２ ) ｒ -, -K(CR ５ R ６ ) ｍ (NR) １１ CON ｔ (CR) ９ R １０ ) ｎ (OCH) ２ CH ２ ) ｒ -、-K(CR ５ R ６ ) ｍ (A) ｔ (NR) １１ CR) ９ R １０ ) ｎ (OCH) ２ CH ２ ) ｒ -、-K(CR ５ R ６ ) ｍ (O) (A) ｔ (CR) ９ R １０ ) ｎ (OCH) ２ CH ２ ) ｒ -、-K(CR ５ R ６ ) ｍ (OCNR) ７ )(Aa) ｔ (CR) ９ R １０ ) ｎ (OCH) ２ CH ２ ) ｒ -、-K(CR ５ R ６ ) ｍ (C) (A) ｔ (CR) ９ R １０ ) ｎ (OCH) ２ CH ２ ) ｒ -、-K(CR ５ R ６ ) ｍ (NR) １１ CON ｔ (CR) ９ R １０ ) ｎ (OCH) ２ CH ２ ) ｒ -、-K(CR ５ R ６ ) ｍ (OCO) (Aa) ｔ (CR ９ R １０ ) ｎ (O.C.H. ２ CH ２ ) ｒ -, -K(CR ５ R ６ ) ｍ (OCNR ７ ) (Aa) ｔ (CR ９ R １０ ) ｎ (O.C.H. ２ CH ２ ) ｒ -, -K(CR ５ R ６ ) ｍ (CO) (Aa) ｔ (CR ９ R １０ ) ｎ (O.C.H. ２ CH ２ ) ｒ -, -K(CR ５ R ６ ) ｍ -phenyl-(CO)(Aa) ｔ (CR ７ R ８ ) ｎ -, -K(CR ５ R ６ ) ｍ -furyl-(CO)(Aa) ｔ (CR ７ R ８ ) ｎ -, -K(CR ５ R ６ ) ｍ -oxazolyl-(CO)(Aa) ｔ (CR ７ R ８ ) ｎ -, -K(CR ５ R ６ ) ｍ -Thiazolyl-(CO)(Aa) ｔ (CR ７ R ８ ) ｎ -, -K(CR ５ R ６ ) ｔ -Thienyl-(CO)(CR ７ R ８ ) ｎ -, -K(CR ５ R ６ ) ｔ -imidazolyl-(CO)(CR ７ R ８ ) ｎ -, -K(CR ５ R ６ ) ｔ -Morpholino-(CO)(Aa) ｔ (CR ７ R ８ ) ｎ -, -K(CR ５ R ６ ) ｔ -piperazino-(CO)(Aa) ｔ (CR ７ R ８ ) ｎ -, -K(CR ５ R ６ ) ｔ -N-methylpiperazine-(CO)(Aa) ｔ (CR ７ R ８ ) ｎ -, -K(CR ５ R ６ ) ｍ -(Aa) ｔ Phenyl-, -K(CR ５ R ６ ) ｍ -(Aa) ｔ Frill-, -K (CR ５ R ６ ) ｍ -Oxazolyl (Aa) ｔ -, -K(CR ５ R ６ ) ｍ -Thiazolyl (Aa) ｔ -, -K(CR ５ R ６ ) ｍ -Thienyl (Aa) ｔ -, -K(CR ５ R ６ ) ｍ -Imidazolyl (Aa) ｔ -, -K(CR ５ R ６ ) ｍ -Morpholino-(Aa) ｔ -, -K(CR ５ R ６ ) ｍ -Piperazino- (Aa) ｔ G-, -K (CR ５ R ６ ) ｍ -N-methylpiperazino- (Aa) ｔ ; In the formula, m, Aa, m, n, R ３ , R ４ , and R ５ are defined above; t and r are independently 0 to 100; R ６ , R ７ , and R ８ are independently H; halide; C １ ~C ８ selected from alkyl, aryl, alkenyl, alkynyl, ether, ester, amine, or amide; １ R ２ , C.F. ３ , OR １ , aryl, heterocycle, S(O)R １ , S.O. ２ R １ , -CO ２ H, -SO ３ H, -OR １ , -CO ２ R １ , -CONR １ , -PO ２ R １ R １２ , -PO ３ H or P(O)R １ R ２ R ３’ K is optionally substituted with NR １ , -SS-, -C(=O)-, -C(=O)NH-, -C(=O)O-, -C=NH-O-, -C=N-NH-, -C(=O)NH-NH-, O, S, Se, B, or C ３ ~C ６ It is a heteroaromatic ring. The connector L １ And / or L ２ The structure may be the following structures or combinations thereof: 【Chemistry 48】 【change】 【change】 【change】 【change】 【change】 【change】 X ２ , X ３ , X ４ , X ５ , or X ６ are independently NH, NHNH; N(R １２ );N(R １２ ) N (R １２’ );O;S;C １ ~C ６ Alkyl; C ２ ~C ６ Heteroalkyl, alkylcycloalkyl, heterocycloalkyl; C ３ ~C ８ Aryl, Ar-alkyl, heterocycle, carbocycle, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl; CH ２ OR １２ , C.H. ２ S.R. １２ , C.H. ２ NHR １２ or 1 to 8 amino acids; １２ and R １２’ are independently H; C １ ~C ８ Alkyl; C ２ ~C ８ Heteroalkyl, alkylcycloalkyl, heterocycloalkyl; C ３ ~C ８ Aryl, Ar-alkyl, heterocycle, carbocycle, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl; or an ester, ether, or amide having 1 to 8 carbon atoms; or a group of the formula (OCH ２ CH ２ ) ｐ Or (OCH ２ CH (CH ３ )) ｐ where p is an integer from 0 to about 100; or a combination thereof;

14. Connector L １ and L ２ The antibody-drug conjugate of claim 10, wherein is independently selected from the group consisting of: 【Chemistry 49】 【change】 During the ceremony, 【Chemistry 50】 is a drug or a linker L １ Or L ２ "#" is the site of attachment to an antibody S (thiol), O (phenol), NH (amino), CHO (aldehyde), C(=O) (ketone), C(O)(NH) (amide), C(O)(OH) (carboxylate); Aa is an L- or D-natural or unnatural amino acid; R １ is H, C １ -C ８ Alkyl, OH, CH ２ O.H., C.H. ２ CH ２ O.H., N.H. ２ , S.H., S.C.H. ３ , C.H. ２ COOH, CH ２ CH ２ COOH, CH ２ CH ２ CH ２ CH ２ N.H. ２ , C ６ H ５ , C.H. ２ C ６ H ５ , C.H. ２ C ６ H ４ OH, CH(OH)CH ３ , C.H. ２ C(O)NH ２ , C.H. ２ CH ２ C(O)NH ２ , C.H. ２ CH ２ CH ２ NHC (=NH)NH ２ is; r is 0 to 12; if r is not 0, (Aa)r is an amino acid or peptide unit of the same or different sequence; m １ = 1 - 18; m ２ = 1 - 100; m ３ = 1 - 8; m ４ = 0 - 8; m ５ = 1-8; Y ７ yes ２ I'm sorry, I'm sorry, I'm sorry １ )、SO ２ 、P(O)(OH)、NHS(O) ２ 、NHS(O) ２ H, H ２ I'm sorry, I'm sorry ２ NHC(O)O、NHS(O) ２ NHC(O)NH、NHP(O)(OH)、NHP(O)(OH)NH、O P(O)(OH)O、NHP(O)(OH)O、OP(O)(OH)NH、S 、O、OP(O)(OH)OP(O)(OH)NH、NHP(O)(OH)O P(O)(OH)NH、NHP(O)(OH)OP(O)(OH)O、OCH ２ CH ２ O、OCH ２ CH ２ H ２ CH ２ ) ２ N、NHC ６ H ４ H ２ である; Y ８ are NHC(=O), NHS(O ２ ), NH(SO), NHS(O ２ )NH, NHP(O)(OH)NH, or C(O)NH; R ９ is H, (O=)CR １ , (O=)CNHR １ , R １ COOH, R １ (COCH ２ N H) m ２ H, R １ (Aa) r or R １ (COCH ２ NCH ３ ) ｍ２ H, where R １ is defined above. Lv １’ and Lv2' is selected from: 【Chemistry 51】 【change】 【change】 【change】 【change】 【change】 During the ceremony, 【Chemistry 52】 is a drug or a linker L １ Or L ２ "#" is a site of binding to an antibody with S (thiol), O (phenol), NH (amino), CHO (aldehyde), C(=O) (ketone), C(O)(NH) (amide), or C(O)(OH) (carboxylate); １ , X １ ', and X ２ ' is as above; X is O, NH, S, CH ２ is the bond in the middle of two atoms 【Chemistry 53】 means that it can bond to either of the two atoms;

15. Connector L １ and L ２ The antibody-drug conjugate of claim 10, wherein 【Chemical 54】 During the ceremony, Aa is an L- or D-natural or unnatural amino acid; R １ is H, C １ -C ８ Alkyl, OH, CH ２ O.H., C.H. ２ CH ２ O.H., N.H. ２ , S.H., S.C.H. ３ , C.H. ２ COOH, CH ２ CH ２ COOH, CH ２ CH ２ CH ２ CH ２ N.H. ２ , C ６ H ５ , C.H. ２ C ６ H ５ , C.H. ２ C ６ H ４ OH, CH(OH)CH ３ , C.H. ２ C(O)NH ２ , C.H. ２ CH ２ C(O)NH ２ , C.H. ２ CH ２ CH ２ NHC (=NH)NH ２ is; r is 0 to 12; if r is not 0, (Aa)r is an amino acid or peptide unit of the same or different sequence; m １ = 1 - 18; m ２ = 1 - 100; m ３ = 1 - 8; m ４ = 0 - 8; m ５ = 1-8; Y ７ yes ２ I'm sorry, I'm sorry, I'm sorry １ )、SO ２ 、P(O)(OH)、NHS(O) ２ 、NHS(O) ２ H, H ２ I'm sorry, I'm sorry ２ NHC(O)O、NHS(O) ２ NHC(O)NH、NHP(O)(OH)、NHP(O)(OH)NH、O P(O)(OH)O、NHP(O)(OH)O、OP(O)(OH)NH、S 、O、OP(O)(OH)OP(O)(OH)NH、NHP(O)(OH)O P(O)(OH)NH、NHP(O)(OH)OP(O)(OH)O、OCH ２ CH ２ O、OCH ２ CH ２ H ２ CH ２ ) ２ N、NHC ６ H ４ H ２ である; Y ８ are NHC(=O), NHS(O ２ ), NH(SO), NHS(O ２ )NH, NHP(O)(OH)NH, or C(O)NH; R ９ is H, (O=)CR １ , (O=)CNHR １ , R １ COOH, R １ (COCH ２ N H) m ２ H, R １ (Aa) r or R １ (COCH ２ NCH ３ ) ｍ２ H, where R １ is defined above.

16. The antibody-drug conjugate of claim 10, prepared by reacting a drug / conjugate complex having the following formula (IV), (V), or (VI), respectively, with an amino acid in an antibody of the invention to form a conjugate of formula (I), (II), or (III): 【Chemistry 55】 During the ceremony, Lv １ and Lv ２ is a reactive group and is independently selected from: 【Chemistry 56】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 During the ceremony, X １ ' and Ｘ２ ' are independently F, Cl, Br, I, OTf, OMs, OC ６ H ４ (NO ２ ), O.C. ６ H ３ (NO ２ ) ２ , O.C. ６ F ５ , O.C. ６ H.F. ４ , or Lv ３ is; ２ is O, NH, N(R １ ), or C.H. ２ is; R ３ and R ５ are independently H, R １ , aromatic, heteroaromatic, or aromatic group, and one or several H atoms are independently selected from -R １ , -halogen, -OR １ , -SR １ , -NR １ R ２ , -NO ２ , -S(O)R １ , -S(O) ２ R １ , or -COOR １ is replaced by; ３ and Lv ３ ' is independently a leaving group selected from F, Cl, Br, I, nitrophenoxyl, N-hydroxysuccinimide (NHS), phenoxyl, benzenethiol, dinitrophenoxyl, pentafluorophenoxyl, tetrafluorophenoxyl, difluorophenoxyl, monofluorophenoxyl, pentachlorophenoxyl, triflate, imidazole, dichlorophenoxyl, tetrachlorophenoxyl, 1-hydroxybenzotriazole, tosylate, mesylate, 2-ethyl-5-phenylisoxazolium-3'-sulfonate, anhydrides formed with themselves or with other anhydrides, acetyl anhydride, formyl anhydride, or an intermediate molecule generated with a condensation reagent for a peptide coupling reaction or a Mitsunobu reaction; L １’ and L ２’ are the same or different, and each independently represents O, NH, S, S-S, NHNH, N(R ３ ), N(R ３ ) N (R ３ '), C １ -C ８ Alkyl; C ２ -C ８ Heteroalkyl, alkylcycloalkyl, heterocycloalkyl; C ３ -C ８ Aryl, Ar-alkyl, heterocycle, carbocycle, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl; ２ -C ８ (2 to 8 carbon atoms) esters, ethers, or amides; 1 to 8 natural or unnatural amino acids as defined above; ２ CH ２ ) ｐ , (OCH ２ CH (CH ３ )) ｐ , (OCH ２ CH ２ ) ｐ OR ３ , (OCH ２ CH (CH ３ )) ｐ OR ３ , NH(CH ２ CH ２ O) ｐ OR ３ , NH(CH ２ CH (CH ３ ) O) pR3 ｐ OR ３ , N[(CH ２ CH ２ O) ｐ R ３ ][(CH ２ CH ２ O) ｐ R ３’ ], (OCH ２ CH ２ ) ｐ COOR ３ , or C.H. ２ CH ２ (O.C.H. ２ CH ２ ) ｐ COOR ３ where p and p′ are independently an integer selected from 0 to about 1000, or a combination thereof, ３ and R ３’ are independently H, C １ -C ８ Alkyl; C ２ -C ８ heteroalkyl, alkylcycloalkyl, aryl of heterocycloalkyl, Ar-alkyl, heterocycle, carbocycle, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, heteroaryl; or an ester, ether, or amide having 1 to 8 carbon atoms; or 1 to 8 natural or unnatural amino acids as described in the definitions; or a group of the formula (OCH ２ CH ２ ) ｐ Or (OCH ２ CH (CH ３ )) ｐ where p is an integer from 0 to about 1000, or a combination thereof; L １’ And / or L ２’ may comprise a self-immolative or non-self-immolative moiety, a peptidyl unit, a hydrazone bond, a disulfide, an ester, an oxime, an amide, or a thioether bond, wherein said self-immolative unit and non-self-immolative moiety are all as defined in claim 13. In formula (V) and formula (VI), 【Chemistry 57】 can be selected from: 【Chemistry 58】 【change】 【change】 【change】 【change】 【change】 【change】 In the formula, Lv ３ , Lv ３ ', X １ ', and X ２ ' is as above; the bond in the middle of two atoms 【Chemistry 59】 means that it can bond to either of the two atoms;

17. The antibody-drug conjugate of claim 10, which is prepared by first reacting a conjugate having formula (VII), (VIII), or (IX) as shown below with a cytotoxic agent to form a compound of formula (IV), (V), or (VI), which is then reacted with an amino acid in the antibody to form a conjugate of formula (I), (II), or (III): 【Chemistry 60】 In the ceremony, L １’ , L ２’ , E １ , Lv １ , and Lv ２ is defined as above for formulas (I), (II), (III), (IV), (V), and (VI); Lv ５ and Lv ６ is independently selected from: 【Chemistry 61】 【change】 During the ceremony, X1' is F, Cl, Br, I, OTs (tosylate), OTf (triflate), OMs (mesylate), OC ６ H ４ (NO ２ ), O.C. ６ H ３ (NO ２ ) ２ , O.C. ６ F ５ , O.C. ６ H.F. ４ , or Lv ３ is; ２’ is O, NH, N(R １ ), or C.H. ２ is; R ３ and R ５ are independently H, R １ , aromatic, heteroaromatic, or one or several H atoms are independently -R １ , -halogen, -OR １ , -SR １ , -NR １ R ２ , -NO ２ , -S(O)R １ , -S(O) ２ R １ , or -COOR １ is an aromatic group substituted by Lv ３ and Lv ３’ are independently a leaving group selected from F, Cl, Br, I, nitrophenoxyl; N-hydroxysuccinimide (NHS); phenoxyl; benzenethiol, dinitrophenoxyl; pentafluorophenoxyl; tetrafluorophenoxyl; difluorophenoxyl; monofluorophenoxyl; pentachlorophenoxyl; triflate; imidazole; dichlorophenoxyl; tetrachlorophenoxyl; 1-hydroxybenzotriazole; tosylate; mesylate; 2-ethyl-5-phenylisoxazolium-3′-sulfonate, anhydrides formed with themselves or with other anhydrides selected from acetyl anhydride and formyl anhydride, or intermediate molecules generated with condensation reagents for peptide coupling reactions or Mitsunobu reactions; １ Or Lv ２ As long as the reaction between the functional group Lv and the thiols in the antibody is at least 1-fold faster or slower than the reaction between the functional group Lv and the thiols in the antibody. ５ And / or Lv ６ can also react with thiols in cytotoxic agents.

18. A thiol group is generated in the antibody by reduction of a disulfide bond, and then the thiol is reacted simultaneously or sequentially with a conjugate of formula (VII), (VIII), or (IX) to form an antibody / conjugate conjugate molecule of formula (X), (XI), or (XII) below, followed by addition of a cytotoxic drug D. １ Or D ２ to form a conjugate of formula (I), (II), or (III): 【Chemistry 62】 In the formula, Lv ５ , Lv ６ , L1, L １ , L ２ , E １ , Lv １’ , and Lv ２’ , mAb, n, and n′ have the same definitions as in claim 17.

19. The antibody-drug conjugate of claim 10, produced by a homogeneous conjugation method comprising the steps of: (a) providing an effective amount of a zinc cation-amino chelate / complex (Zn(NR １ R ２ R ３ ) ｍ１ ｍ２＋ 2. Incubating the antibody of the present invention in a buffered system in the presence of 1,2-dichlorophenyl ether (pH 7.0) and a reducing agent; (b) adding an effective amount of a payload / conjugate conjugate of formula (IV), (V), or (VI) as set forth in claim 16, or a conjugate of formula (VII), (VIII), or (IX) as set forth in claim 17, to react with the thiol group generated in step (a); and (c) optionally adding an effective amount of an oxidizing agent (e.g., dehydroascorbic acid) to reoxidize any unreacted sulfhydryl groups; (d) and then purifying the resulting conjugate; (e) optional step (c) above can be replaced by the addition of an effective amount of cystine or a related disulfide compound to quench any unreacted reducing agent, while an effective amount of cysteine ​​or a related sulfhydryl compound can be added to quench excess conjugation of a linker or linker / payload conjugate containing a thiol reactive group to generate cysteine, or an effective amount of cysteine ​​or a related sulfhydryl compound can be added to quench excess linker or linker / payload conjugate molecules; The zinc cation-amino chelate / complex Zn(NR １ R ２ R ３ ) ｍ１ ｍ２＋ Inside R １ , R ２ and R ３ are each independently １ -C ８ Alkyl, C ２ -C ８ Heteroalkyl, alkylcycloalkyl, heterocycloalkyl; C ３ -C ８ selected from aryl, Ar-alkyl, heterocycle, carbocycle, cycloalkyl, heteroalkylcycloalkyl, alkylcarbonyl, or heteroaryl; m1 is 1, 2, 3, or 4; The Zn(NR １ R ２ R ３ ) ｍ１ ｍ２＋ is selected from the following; Zn(NH ２ CH ３ ) ２ ２＋ 、Zn(NH ２ CH ２ CH ３ ) ２ ２＋ 、Zn(NH ２ CH ２ CH ２ CH ３ ) ２ ２＋ 、Zn(NH ２ CH(CH ３ ) ２ ) ２ ２＋ 、Zn(NH ２ C(CH ３ ) ３ ) ２ ２＋ 、Zn(NH ２ CH ２ C(CH ３ ) ３ ) ２ ２＋ 、Zn(NH(CH ３ ) ２ ) ２ ２＋ 、Zn(NH(CH ２ CH ３ ) ２ ) ２ ２＋ 、Zn(NH(CH(CH ３ ) ２ ) ２ ) ２ ２＋ 、Zn(NH(C(CH ３ ) ３ ) ２ ) ２ ２＋ 、Zn(NH(CH(CH ２ CH3) ２ ) ２ ) ２ ２＋ 、Zn(NH(CH ２ C(CH ３ ) ３ ) ２ ) ２ ２＋ Zn(NH(CH ２ C(CH ２ CH ３ ) ３ ) ２ ) ２ ２＋ Zn(NH(CH ２ CH ２ C(CH ３ ) ３ ) ２ ) ２ ２＋ Zn(NH ２ CH ２ CH ２ OH) ２ ２＋ Zn(NH(CH ２ CH ２ OH) ２ ) ２ ２＋ Zn(N(CH ２ CH ２ OH) ３ ) ２ ２＋ Zn(NH ２ CH ２ COOH) ２ ２＋ Zn(NH ２ CH ２ P. ２ ) ２ ２＋ Zn(NH ２ CH ２ COOCH ３ ) ２ ２＋ Zn(NH ２ CH ２ COOCH ２ CH ３ ) ２ ２＋ Zn(NH ２ CH ２ P.S. ３ ) ３ ) ２ ２＋ Zn(NH ２ CH ２ P.S. ３ ) ２ ) ２ ２＋ Zn(NH ２ CH ２ CH ２ COOH) ２ ２＋ Zn(NH(CH ２ COOH) ２ ) ２ ２＋ Zn(N(CH ２ CH ２ COOH) ３ ) ２ ２＋ Zn(NH ２ CH ３ ) ４ ２＋ Zn(NH ２ CH ２ CH ３ ) ４ ２＋ Zn(NH ２ CH ２ CH ２ CH ３ ) ４ ２＋ Zn(NH ２ CH(CH ３ ) ２ ) ４ ２＋ Zn(NH ２ C(CH ３ ) ３ ) ４ ２＋ Zn(NH ２ CH ２ C(CH ３ ) ３ ) ４ ２＋ Zn(NH(CH ３ ) ２ ) ４ ２＋ Zn(NH(CH ２ CH ３ ) ２ ) ４ ２＋ Zn(NH(CH(CH ３ ) ２ ) ２ ) ４ ２＋ Zn(NH(C(CH ３ ) ３ ) ２ ) ４ ２＋ Zn(NH(CH(CH ２ CH ３ ) ２ ) ２ ) ４ ２＋ Zn(NH(CH ２ C(CH ３ ) ３ ) ２ ) ４ ２＋ Zn(NH(CH ２ C(CH ２ CH ３ ) ３ ) ２ ) ４ ２＋ Zn(NH(CH ２ CH ２ C(CH ３ ) ３ ) ２ ) ４ ２＋ Zn(NH ２ CH ２ CH ２ OH) ４ ２＋ Zn(NH(CH ２ CH ２ OH) ２ ) ４ ２＋ Zn(N(CH ２ CH ２ OH) ３ ) ４ ２＋ Zn(NH ２ CH ２ COOH) ４ ２＋ Zn(NH ２ CH ２ P. ２ ) ４ ２＋ Zn(NH ２ CH ２ COOCH ３ ) ４ ２＋ Zn(NH ２ CH ２ COOCH ２ CH ３ ) ４ ２＋ Zn(NH ２ CH ２ P.S. ３ ) ３ ) ４ ２＋ Zn(NH ２ CH ２ P.S. ３ ) ２ ) ４ ２＋ Zn(NH ２ CH ２ CH ２ COOH) ４ ２＋ Zn(NH(CH ２ COOH) ２ ) ４ ２＋ Zn(N(CH ２ CH ２ COOH) ３ ) ４ ２＋ 、 【Chemistry 63】 【change】 All of the complex cations are Cl － , Br － , I － , SO ４ ２－ , HSO ４ － , NO ３ － , PO ４ ３－ , HPO ４ ２－ , H ２ PO ４ － , CO ３ ２－ , HCO ３ － , HCOO － , CH ３ COO － , F ３ CCOO － , Cl ３ CCOO － , FCH ２ COO － , ClCH ２ COO － , F ２ CHCOO － , Cl ２ CHCOO － , BF ４ － , SO ３ ２－ , HSO ３ － , CH ３ SO ３ － , C ６ H ５ CH ２ SO ３ － , C ６ H ５ SO ３ － , C ６ H ５ COO － , C ６ H ５ CH ２ COO － , C ６ F ５ O － , C ６ H ４ (OH)COO － , C ６ H ２ F ３ O － , C ６ H ４ (NO ２ ) O － , and C ６ H ２ (NO ２ ) ３ O － may be formed as a salt with an anion selected from The concentration of the transition metal cation-amino complex in the reaction solution is 0.01 mM to 1.0 mM, or 0.5 to 20 equivalents per mole of antibody, and the reaction solution is preferably in ethanol, methanol, propanol, propanediol, DMA, DMSO, THF, or CH ３ CN may be added to the reaction solution together with a water-miscible organic solvent selected from the group consisting of toluene, ... The reducing agent is tris(2-carboxyethyl)phosphine (TCEP) (P(CH ２ CH ２ COOH) ３ ) or tris(hydroxypropyl)phosphine (P(CH ２ CH ２ CH ２ O.H. ３ ), which are used at 1.0 to 20 equivalents per mole of antibody; The oxidizing agent is DHAA, Fe ３＋ , I ２ , Cu ２＋ , Mn ３＋ , MnO ２ , or Fe ３＋ / I － the concentration of said oxidizing agent used in the reaction solution is 0.02 mM to 1.0 mM, or 1 to 100 equivalents per mole of antibody; The pH of the conjugation reaction is usually about 5.0 to 8.0, preferably about 5.5 to 7.5; as a co-solvent, up to 30% of a water-miscible organic solvent selected from DMA, DMF, ethanol, methanol, acetone, acetonitrile, THF, isopropyl alcohol, dioxane, propylene glycol, or ethylene glycol can be added to the aqueous buffer solution; The optimum temperature for the conjugation reaction is usually about -5°C to 40°C, preferably about 0°C to 37°C; more preferably about 2°C to 8°C; the optimum reaction time for the conjugation reaction is about 15 minutes to about 48 hours, preferably 30 minutes to 16 hours; The conjugate can be purified by Sephadex G25 or Sephacryl S300 column gel filtration, adsorption chromatography, ion (cation or anion) exchange chromatography, or dialysis (ultrafiltration or ultrafiltration (UF) and / or diafiltration (DF).

20. The resulting conjugate of formula (I), (II), (III), (X), (XI) or (XII) produced by the homogeneous conjugation method is predominantly (more than 60%) bound to a cysteine ​​site between the heavy and light chains of an antibody, and the drug percentage distribution in the antibody is D0<5%, D2<10%, D4>60%, D6<10%, D8<10%, when the drug / antibody ratio (DAR) is 4; 20. The antibody-drug conjugate of claim 19, wherein the drug / antibody ratio (DAR) of the conjugate is measured by UV (detection wavelength 240-380 nm), hydrophobic interaction chromatography (HIC-HPLC), reversed phase chromatography (RP-HPLC), capillary electrophoresis (CE), LC-MS, CE-MS, or LC-MS / MS.

21. The antibody-drug conjugate of claim 10 having the following structure: 【Chemistry 64】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 where mAb is an antibody of the invention and n=1-20.

22. The antibody-drug conjugate of claim 16, wherein the drug / conjugate complex of formulas (IV), (V), and (VI) has the following structure: 【Chemistry 65】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】 【change】

23. A pharmaceutical composition for treating multiple myeloma cells, a B cell-mediated or plasma cell-mediated disease, or an antibody-mediated disease or disorder, comprising a therapeutically effective amount of the BCMA antibody or antigen-binding fragment of claim 1 and a pharma- ceutical acceptable salt, carrier, diluent, or excipient, or a combination of said conjugates.

24. A pharmaceutical composition for treating multiple myeloma cells, a B cell-mediated or plasma cell-mediated disease, or an antibody-mediated disease or disorder comprising the antibody-drug conjugate of claim 9 and a pharma-ceutically acceptable salt, carrier, diluent, or excipient, or a combination of said conjugates.

25. 25. The pharmaceutical composition of claim 23 or 24, which is a liquid formulation or a lyophilized solid / powder formulation, comprising, by weight 0.01% to 99% of the BCMA antibody of claim 1 or the antibody-drug conjugate of claim 9; Polyol 0.0% to 20.0%; 0.0% to 2.0% of one or more surfactants; 0.0% to 5.0% of one or more preservatives; 0.0% to 30% of one or more amino acids; 0.0% to 5.0% of one or more antioxidants; 0.0% to 0.3% of one or more metal chelating agents; 0.0% to 30.0% of one or more buffer salts to adjust the pH of the formulation to 4.5 to 7.5; and one or more tonicity agents, 0.0% to 30.0%, to adjust the osmolality to between about 250 and 350 mOsm after reconstitution for administration to a patient; The polyol is selected from fructose, mannose, maltose, lactose, arabinose, xylose, ribose, rhamnose, galactose, glucose, sucrose, trehalose, sorbose, melezitose, raffinose, mannitol, xylitol, erythritol, maltitol, lactitol, erythritol, sorbitol, glycerol, or L-gluconate and its metal salts; The surfactant may be polysorbate 20, polysorbate 40, polysorbate 65, polysorbate 80, polysorbate 81, or polysorbate 85, poloxamer, poly(ethylene oxide)-poly(propylene oxide), polyethylene-polypropylene, Triton; sodium dodecyl sulfate (SDS), sodium lauryl sulfate; sodium octyl glucoside; lauryl-, myristyl-, linoleyl-, or stearyl-sulfobetaine; lauryl-, myristyl-, linoleyl-, or stearyl-sarcosine; linoleyl-, myristyl-, or cetyl-betaine; lauroamidopropyl-, cocamide propyl- myristamidopropyl-, palmidopropyl-, or isostearamidopropyl-dimethylamine; sodium methyl cocoyl-, or disodium methyl oleyl-taurate; dodecyl betaine, dodecyl dimethylamine oxide, cocamidopropyl betaine, and cocoamphoglycinate; or isostearyl ethylimidonium ethosulfate; polyethyl glycol, polypropyl glycol, and copolymers of ethylene and propylene glycol; The preservative is selected from benzyl alcohol, octadecyl dimethyl benzyl ammonium chloride, hexatonium chloride, benzalkonium chloride, benzethonium chloride, phenol, butyl alcohol, alkyl parabens such as methyl or propyl paraben, catechol, resorcinol, cyclohexanol, 3-pentanol, or m-cresol; The amino acids are selected from arginine, cystine, glycine, lysine, histidine, ornithine, isoleucine, leucine, alanine, glycine, glutamic acid, or aspartic acid; The antioxidant is selected from ascorbic acid, glutathione, cystine, or methionine; The chelating agent is selected from EDTA or EGTA; The buffer salt is selected from sodium, potassium, ammonium, or trihydroxyethylamino salts of citric acid, ascorbic acid, gluconic acid, carbonic acid, tartaric acid, succinic acid, acetate, or phthalic acid; tris or tromethamine hydrochloride, phosphate, or sulfate; anionic acetate, chloride, phosphate, sulfate, or succinate of arginine, glycine, glycylglycine, or histidine; The pharmaceutical composition, wherein the isotonicity agent is selected from mannitol, sorbitol, sodium acetate, potassium chloride, sodium phosphate, potassium phosphate, trisodium citrate, or sodium chloride.

26. 25. The pharmaceutical composition of claim 23 or 24, which is filled in a vial, a bottle, a prefilled syringe, or a prefilled autoinjector syringe in liquid or lyophilized solid form.

27. 25. The BCMA antibody or antigen-binding fragment of claim 1, the conjugate of claim 9, or the pharmaceutical composition of claim 23 or 24, which has cell killing activity in vitro, in vivo, or ex vivo.

28. A pharmaceutical composition of the BCMA antibody of claim 1, the conjugate of claim 9, or the pharmaceutical composition of claim 23 or 24, administered simultaneously with a chemotherapeutic agent, a radiotherapy, an immunotherapeutic agent, an autoimmune disease agent, an anti-infective agent, a synergistic agent, or another conjugate for synergistically treating multiple myeloma cells, B cell-mediated or plasma cell-mediated diseases.

29. The pharmaceutical composition, conjugate, or pharmaceutical composition of claim 28, wherein the synergist is selected from one or more of the following drugs: abatacept, abiraterone acetate, abraxane, acetaminophen / hydrocodone, acalabrutinib, aducanumab, adalimumab, ADXS31-142, ADXS-HER2, afatinib dimaleate, aldesleukin, alectinib, alemtuzumab, alitretinoin, adotrastuzumab emtansine, amphetamine / dextroamphetamine, anastrozole, aripiprazole, anthracycline, aripiprazole, atazanavir, atezolizumab, atorvastatin, avelumab, axicabtagene. Ciloreucel, axitinib, belinostat, live BCG, zumab, bexarotene, blinatumomab, bortezomib, bosutinib, brentuximab vedotin, brigatinib, budesonide, budesonide / formoterol, buprenorphine, cabazitaxel, cabozantinib, capmatinib, capecitabine, carfilzomib, chimeric antigen receptor modified T (CAR-T) cells, celecoxib, ceritinib, cetuximab, chidamide, cyclosporine, cinacalcet, crizotinib, cobimetinib, cosentyx, crizotinib, CTL01 9. Dabigatran, dabrafenib, dacarbazine, daclizumab, dacomitinib, daptomycin, daratumumab, darbepoetin alfa, darunavir, dasatinib, denileukin diftitox, denosumab, depakote, dexlansoprazole, methylphenidate, dexamethasone, dinutuximab, doxycycline, duloxetine, duvelisib, durvalUmab, elotuzumab, emtricitabine / rilpivirine / tenofovir, disoproxil fumarate, emtricitabine / tenofovir / efavirenz, enoxaparin, ensartinib, enzalutamide, epoetin alfa, erlotinib, esomeprazole, eszopiclone, everolimus, exemestane, everolimus, exenatide ER, ezetimibe, ezetimibe / simvastatin, fenofibrate, filgrastim, fingolimod, fluticasone propionate, fluticasone / salmeterol, fulvestrant, gaziva, gefitinib, glatiramer, goserelin acetate, icotinib, imatinib, ibritumomabTiuxetan, ibrutinib, idelarib, ifosfamide, infliximab, imiquimod, immucyst, immune BCG, iniparib, insulin aspart, insulin detemir, insulin glargine, insulin lispro, interferon alpha, interferon alpha 1b, interferon alpha 2a, interferon alpha 2b, interferon beta, interferon beta 1a, interferon beta 1b, interferon gamma 1a, iapatinib, ipilimumab, ipratropium bromide / salbutamo oxal, ixazomib, Kanuma, lanreotide acetate, lenalidomide, lenalidomide, lenvatinib mesylate, letrozole, levothyroxine, lidocaine, linezolid, liraglutide, examfetamine, LN-144, lorlatinib, memantine, methylphenidate, metoprolol, mekinist, melicitabine / rilpivirine / tenofovir, modafinil, mometasone, misidac-C, necitumumab, neratinib, nilotinib, niraparib, nivolumab, ofatumumab, obinutuzumab, ora Paribu, olmesartan, olmesartan / hydrochlorothiazide, omalizumab, omega-3 fatty acid ethyl ester, oncoline, oseltamivir, osimertinib, oxycodone, palbociclib, palivizumab, panitumumab, panobinostat, pazopanib, pembrolizumab, PD-1 antibody, PD-L1 antibody, pemetrexed, pertuzumab, pneumococcal conjugate vaccine, pomalidomide, poziotinib, pregabalin, proskavax, propranolol, quetiapine, rabeprazole, radium chloride M223, raloxifene, raltegravir, ramucirumab, ranibizumab, regorafenib, rituximab, rivaroxaban, romidepsin, rosuvastatin, ruxolitinib phosphate, salbutamol, savolitinib, semaglutide, sildenafil, siltuximab, sipulcel-T, sitagliptin, sitagliptin / metformin, solifenacin, solanezumab, sonidegib, sorafenib, sunitinib, tacrolimus, taclimus, tadalafil, tamoxifen, tafinlar, talimogene laherparepvec, talazoparib, telaporebrib, IR / emtricitabine, tenofovirDisoproxil fumarate, testosterone gel, thalidomide, TICE BCG, tiotropium bromide, tisagencleucel, toremifene, trametinib, trastuzumab, trastuzumab deruxtecan, trabectedin (ecteinascidin 743), trametinib, tremelimumab, trifluridine / tipiracil, tretinoin, ustekinumab, valsartan, veliparib, vandetanib, vemurafenib, venetoclax, vorinostat, dibuaflibercept, zostavax, and analogs, derivatives, pharma- ceutically acceptable salts, carriers, diluents or excipients thereof, or combinations thereof.

30. 1. A method for the treatment of a medical disorder associated with the presence of pathogenic B cells expressing B cell maturation antigen (BCMA) in a human subject, comprising: The method comprises administering to the human subject a BCMA antibody or antigen-binding fragment of claim 1, an antibody-drug conjugate of claim 9, or a pharmaceutical composition of claim 23 or 24; The medical disorder associated with the presence of pathogenic B cells is a cancer of plasma cells or a cancer of B-lymphatic cells.

31. 25. Use of the BCMA antibody or antigen-binding fragment of claim 1, the antibody-drug conjugate of claim 9, or the pharmaceutical composition of claim 23 or 24 in the manufacture of a medicament for treating a medical disorder associated with the presence of pathogenic B cells expressing B-cell maturation antigen (BCMA), comprising: The medical disorder associated with the presence of pathogenic B cells is a cancer of plasma cells or a cancer of B lymphatic cells.

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