WO2022078279A1 - Conjugué anticorps-médicament et son utilisation - Google Patents

Conjugué anticorps-médicament et son utilisation Download PDF

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WO2022078279A1
WO2022078279A1 PCT/CN2021/123020 CN2021123020W WO2022078279A1 WO 2022078279 A1 WO2022078279 A1 WO 2022078279A1 CN 2021123020 W CN2021123020 W CN 2021123020W WO 2022078279 A1 WO2022078279 A1 WO 2022078279A1
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antibody
pharmaceutically acceptable
acceptable salt
formula
stereoisomer
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PCT/CN2021/123020
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English (en)
Chinese (zh)
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丁会
柯天一
于海勇
劳芳
徐云雷
张西东
刘岩
荣鹏飞
王闯
范墨林
李凡
欧阳芳幸
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昆山新蕴达生物科技有限公司
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Priority to CN202180069652.9A priority Critical patent/CN116472064A/zh
Publication of WO2022078279A1 publication Critical patent/WO2022078279A1/fr

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/68—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an antibody, an immunoglobulin or a fragment thereof, e.g. an Fc-fragment
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents

Definitions

  • the present invention relates to the field of biomedicine, in particular, the present invention relates to a new type of linker structure, a drug-linker compound including the linker structure, and an antibody-drug conjugate including the drug-linker compound, the above-mentioned drugs - Preparation methods and applications of linker compounds and antibody-drug conjugates.
  • ADCs Antibody-drug conjugates
  • conjugates have shown unique advantages over pure antibody drugs, by combining them with tumor cell surface antigen binding specificity.
  • the monoclonal antibody is linked to a biologically active cytotoxin, thereby combining the tumor recognition targeting of the antibody with the high-efficiency killing effect of the cytotoxin. big flaw.
  • ADC can accurately target tumor cells while reducing the impact on normal cells, greatly improving the effectiveness and safety of anti-tumor drugs.
  • ADC generally consists of three parts: antibody, linker and toxin.
  • Antibodies are targeted functional macromolecules of ADCs, which play the role of enriching toxins near the tumor tissue site to improve the killing efficiency of toxins.
  • major popular targets such as HER-2, Trop-2, PDL-1, CD30, etc.
  • ADC linkers are divided into two types: cleavable and non-cleavable.
  • the ideal linker should meet the requirements of "good stability and high release efficiency", that is, the ADC remains stable in the blood circulation and can be quickly released after reaching the tumor cells. toxins, killing tumor cells.
  • the linker is crucial for the ADC to function. An unstable linker will lead to off-target ADC and increase the safety risk, while an overly stable linker will affect the release rate of the toxin, thereby affecting the efficacy of the drug.
  • the toxin part of ADC is a small drug molecule that plays a killing role, and generally kills tumor cells by inhibiting DNA or protein synthesis, inhibiting cell mitosis, and the like.
  • the toxins currently used for ADC development mainly include microtubule inhibitors maytansinoids (see EP0425235, US5208020, US5416064, US7276497) and auristatin (MMAE/MMAF, see US2016304621A).
  • the representative drugs currently on the market are T-DM1 developed by Genetech.
  • T-DM1 is a compound formed by a stable thioether linker MCC (4-[N-maleimidomethyl]cyclohexane-1 - carboxylate) conjugated to an ADC consisting of trastuzumab conjugated to the maytansinoid toxoid DM1 (US8337856).
  • MCC stable thioether linker
  • ADC maytansinoid toxoid DM1
  • Other classes of cytotoxins include Calicheamicin (see US5606040), benzodipyrrole derivatives (duocarmycin, see US7129261), pyrrolobenzodiazepines (PBDs, see WO2005/040170) and Derivatives of tree alkaloids.
  • camptothecin derivatives include SN-38, CPT-11, ixatecan, 9-nitrocamptothecin, 10-hydroxycamptothecin and the like.
  • ADC using camptothecin toxoid is DS-8201 developed by Daiichi Sankyo Co., Ltd. It uses ixatecan, which is highly cytotoxic, as the toxin part.
  • T-DM1 and DS-8201 still have the following shortcomings:
  • T-DM1 As far as T-DM1 is concerned, first of all, the efficacy of T-DM1 is insufficient, one is because its DAR is low, only 3-4, and the other is because it uses the linker of SMCC to connect with DM-1, and SMCC is non-degradable. linker, which reduces the efficacy of T-DM1; secondly, T-DM1 uses DM-1 as a toxin, which is a microtubule inhibitor, and the permeability of cell membranes is weak; thirdly, the presence of T-DM1 reduces white blood cells Serious side effects.
  • ixatecan is 10 times more toxic than SN-38, it cannot be used as a single drug due to its strong cell-killing activity. There is also only one enzymatic cleavage site, which also prolongs the onset time of ADC in cells to some extent. In addition, ixatecan has a short half-life in the blood, which reduces toxic side effects, but faces the risk of a short half-life of the drug.
  • camptothecin-based ADCs there is still a need to develop more effective and safe camptothecin-based ADCs in this field.
  • the preparation has a faster onset time, longer drug half-life, and at the same time, it has the advantages of stability, hydrophilicity and hydrophobicity, and anti-aggregation. Camptothecin ADCs with superior safety indicators are imminent.
  • the inventors designed a linker structure suitable for camptothecin derivatives, and used it as a linking structure between camptothecin derivatives and antibodies, so as to form a linker structure with faster onset time, longer drug half-life, and better drug resistance.
  • this ADC has excellent anti-tumor effect.
  • a first aspect of the present invention provides a compound represented by formula (I),
  • R 11 is a carboxy-substituted C 1 -C 6 alkyl group
  • R 12 is a cyano-substituted C 2 -C 6 alkynyl group, and 1-2 C atoms in X, Y, X' and Y' are replaced by N Atom substitution; preferably, R 11 is a carboxy-substituted C 1 -C 3 alkyl group, and R 12 is a cyano-substituted C 2 -C 3 alkynyl group.
  • X, Y, X' and Y&apos have and only 1 C atom substituted with a N atom.
  • 2 C atoms in X, Y, X' and Y' are substituted with N atoms, and only 1 C atom in X, Y is substituted with N atoms, and X', Y' There is one and only one C atom replaced by N atom.
  • the compound structure is shown below,
  • the compound is linked to the antibody as the linking unit in the antibody-drug conjugate through the formation of a thioether bond between the alkynyl carbon of R12 and the disulfide moiety present in the hinge portion of the antibody.
  • a second aspect of the present invention provides a method for preparing a compound represented by formula (I), comprising the steps of:
  • step (1) The reaction product of step (1) reacts with Pd(PPh 3 ) 2 Cl 2 , triethylamine, and propyn-3-ol in tetrahydrofuran;
  • step (2) reacts with TEMPO, PhI(OAC) 2 , and NH 4 OAC in a solution of CH 3 CN/H 2 O in a ratio of 9:1;
  • step (3) The product of step (3) is produced under the action of TFA/DCM.
  • the third aspect of the present invention provides an antibody-drug conjugate represented by formula (IV), a stereoisomer thereof or a pharmaceutically acceptable salt thereof, or the antibody-drug conjugate, a stereoisomer thereof
  • the solvate of the isomer or a pharmaceutically acceptable salt thereof characterized in that AB represents an antibody, T represents a compound represented by formula (II), and the antibody-drug conjugate is a compound (T) and an antibody ( AB) connected via a linker represented by the following formula (III):
  • R 2 is selected from hydrogen, halogen, hydroxy, nitro, amino, saturated or unsaturated C 1 -C 6 alkyl, C 1 -C 6 alkoxy, C 1 -C 6 alkane substituted by -NR 7 R 8 group or C 1 -C 6 alkyl substituted by C 2 -C 6 alkenyl;
  • R 4 is selected from hydrogen, halogen, hydroxyl, nitro, amino, C 1 -C 6 alkyl, or C 1 -C 6 alkoxy;
  • R1 and R2 can be linked together with the parent moiety to form a 5-6 membered ring optionally substituted with R9 ;
  • R and R can be linked together with the parent moiety to form a 5-6 membered oxygen-containing heterocycle optionally substituted with R ;
  • R 7 and R 8 are independently selected from hydrogen, C 1 -C 6 alkyl, C 1 -C 6 acyl substituted with hydroxy or amino; or R 7 and R 8 may be combined with the N atom to which they are attached taken together to form a 5-6 membered nitrogen-containing heterocycle optionally substituted by R;
  • each occurrence of R7 and R8 is independently selected from hydrogen, methyl, isopropyl, Alternatively R7 and R8 may together with the attached N atom form a 5-6 membered nitrogen-containing heterocycle optionally substituted by R9 ;
  • R 9 is independently selected from halogen, hydroxy, nitro, -NR 7 R 8 , C 1 -C 6 alkyl, C 1 -C 6 alkoxy, optionally C 1 -C 6 alkane substituted piperidinyl;
  • each occurrence of R 9 is independently selected from methyl, -NR 7 R 8 , piperidinyl;
  • R 13 represents a carboxy-substituted C 1 -C 6 alkyl group
  • L 2 represents valine residue, guanidine residue, phenylalanine residue, lysine residue, D-valine residue, glycine residue, alanine residue, aspartic acid Residues;
  • R 10 is independently selected from hydrogen, C 1 -C 6 alkyl optionally substituted with 1 or 2 hydroxy;
  • n 3 represents an integer from 1 to 4
  • n 4 represents an integer from 1 to 4
  • Aryl represents a C 6 -C 10 aryl optionally substituted by R 9 ;
  • W is a single key or Wherein, position 1 means connecting with -NR 10 -, and position 2 means connecting with (CH 2 CH 2 -O-)n 1 -;
  • Q 1 is the compound described in the first aspect of the present invention, which is connected by the carboxyl group of R 11 and the left-terminal amino group -NR 10 - in the formula of L 1 to form an amide bond, and the alkynyl carbon of R 12 and the disulfide of the hinge part of the antibody are connected. bond to form a thioether bond,
  • R 2 represents hydrogen, C 3 -C 4 alkenyl, nitro, amino, C 1 -C 4 alkyl substituted with -N(C 1 -C 4 alkyl) 2 or C 2 -C 4 alkenyl substituted C 1 -C 4 alkyl.
  • R 2 represents hydrogen, nitro, amino
  • R 3 represents hydrogen, halogen, hydroxy, C 1 -C 4 alkyl
  • R 3 represents hydrogen, F, hydroxyl, methyl
  • R 4 represents hydrogen or halogen.
  • R 4 represents hydrogen or F.
  • R 1 and R 2 are joined together to form the group shown below in A moiety represents a bond to the parent group.
  • R 1 and R 2 are joined together to form the group shown below in A moiety represents a bond to the parent group.
  • R and R are joined together to form the group shown below in A moiety represents a bond to the parent group.
  • the compound represented by formula (IV) is gimatecan or gimitecan, more preferably gimatecan:
  • L 2 represents a lysine residue
  • n 4 represents an integer from 1 to 2
  • R 10 represents hydrogen or C 1 -C 4 Alkyl (preferably methyl)
  • Aryl represents a benzene ring group, preferably, the -NR 10 - group and the -(CH 2 )n 4 - group are located in the para position of the benzene ring.
  • La represents
  • the linker represented by formula (III) is a group selected from the group consisting of:
  • the average number of linker-drug linkages is 2-8, preferably 4-8, more preferably 6-8, such as 3.3, 3.5, 5.5, 6.2, 6.5, 6.6, 6.8, 7.0, 7.1, 7.2, 7.4, 7.5 or 7.8.
  • the antibody (AB) is a full-length antibody or antigen-binding fragment thereof, or a bispecific antibody or antigen-binding fragment thereof.
  • the antibody is selected from the group consisting of anti-Her-2 antibody, Trop-2 antibody, EGFR antibody, B7-H3 antibody, PD-1 antibody, PD-L1 antibody, HER-3, HER-4 antibody, CD20 Antibodies, CD30 antibodies, CD19 antibodies, CD33 antibodies; preferably, the antibodies are murine antibodies, chimeric antibodies, and humanized antibodies; preferably, the humanized antibodies are fully human antibodies.
  • the antigen-binding fragment is selected from the group consisting of Fab, Fab', F(ab') 2 , single chain Fv (scFv), Fv and dsFv.
  • the antibody is an anti-TROP-2 antibody, wherein the complementarity determining region (CDR) of the light chain variable region of the anti-Trop-2 antibody comprises CDR1 consisting of the amino acid sequence of KASQDVSIVA, consisting of the amino acid sequence of SASYRYT CDR2 composed of sequence, and CDR3 composed of QQHYITPLT amino acid sequence; CDRs of heavy chain variable region include CDR1 composed of NYGMN amino acid sequence, CDR2 composed of WINTYTGEPTYTDDFKG amino acid sequence, and CDR3 composed of GGFGSSYWYFDV amino acid sequence; preferably,
  • the amino acid sequences of the light chain and heavy chain of the anti-Trop-2 antibody are shown in SEQ ID NO: 1 and SEQ ID NO: 2 respectively; preferably, the coding of the light chain and the heavy chain of the anti-Trop-2 antibody
  • the nucleotide sequences are shown in SEQ ID NO: 3 and SEQ ID NO: 4, respectively.
  • the antibody is an anti-Her-2 antibody, wherein the complementarity determining region (CDR) of the light chain variable region of the anti-Her-2 antibody comprises CDR1 consisting of the amino acid sequence of RASQDVNTAVA, consisting of the amino acid sequence of SASFLYS CDR2 composed of sequence, and CDR3 composed of QQHYTTPPT amino acid sequence; CDRs of heavy chain variable region include CDR1 composed of DTYIH amino acid sequence, CDR2 composed of RIYPTNGYTRY amino acid sequence, and CDR3 composed of WGGDGFYAMDY amino acid sequence; preferably, The amino acid sequences of the light chain and heavy chain of the anti-Her-2 antibody are shown in SEQ ID NO: 5 and SEQ ID NO: 6, respectively.
  • the fourth aspect of the present invention provides a linker-drug intermediate compound represented by formula (VI), characterized in that T represents a compound represented by formula (II), and the intermediate compound is compound (T) with the following The joints represented by formula (V) are connected:
  • R 1 , R 2 , R 3 and R 4 are as described in the specification of the present invention.
  • L 1 , L 2 and La are as described in the specification of the present invention.
  • Q 1 is the compound described in the first aspect of the present invention, which is connected by the carboxyl group of R 11 and the left-terminal amino group -NR 10 - in the formula L 1 to form an amide bond, and the compound represented by the formula (II) is represented by the 19-position hydroxyl group.
  • the compound of formula (II) is as previously described.
  • the linker-drug intermediate compound is a compound selected from the group consisting of,
  • the fifth aspect of the present invention provides the linker structure shown in general formula (III):
  • the sixth aspect of the present invention provides a method for preparing the antibody-drug conjugate of the third aspect of the present invention, the method comprising:
  • linker-drug intermediate compound represented by the formula (VI) is reacted with AB-SH to connect the linker-drug intermediate represented by the formula (VI) through a thioether bond formed by the disulfide bond moiety of the hinge portion of the antibody
  • the compound is linked to the antibody;
  • R 1 , R 2 , R 3 and R 4 are as described in the specification of the present invention.
  • T represents the compound represented by the formula (II), and the compound represented by the formula (II) uses the oxygen in the hydroxyl group at the 19th position as the linking site, or when R 3 or R 4 is a hydroxyl group, the oxygen in the hydroxyl group of R 3 or R 4 is used as a linking site.
  • AB-SH represents an antibody carrying a sulfhydryl group
  • AB represents an antibody
  • the seventh aspect of the present invention provides a method for preparing the linker-drug intermediate compound of the fourth aspect of the present invention, the method comprising:
  • N-[(9H-fluoren-9-ylmethoxy)carbonyl]-N'-[(4-methoxyphenyl)diphenylmethyl]-L-lysine (CN- CMTC-1) and PABOH were dissolved in dichloromethane: methanol solution, reacted under the action of EEDQ, recrystallized and purified to obtain the product;
  • step (2) treating the product of step (1) with a solution of piperidine in acetonitrile, and then purifying the product;
  • step (3) reacts with the product of step (2) to generate compound
  • Gematecan-Boc or Gematecan-Boc is treated with triphosgene, DMAP, and dichloromethane to form a formyl chloride compound, and then the reaction compound of step (4) is added, and then deprotected with TFA/DCM. ;
  • step (5) The product of step (5) is subjected to Click reaction with the compound described in the first aspect of the present invention, and the final product is obtained after being treated with TFA/DCM;
  • step (6) can also be replaced by the following steps: the product of step (5) and SM-1 are added to a solution of DMSO/H 2 O, then CuBr is added to catalyze the reaction, the reaction is complete, and after purification, TFA/DCM is added for deprotection , the final product is obtained;
  • the SM-1 is N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N
  • the eighth aspect of the present invention provides an antibody-drug conjugate represented by formula (VIII), a stereoisomer thereof or a pharmaceutically acceptable salt thereof, or the antibody-drug conjugate, a stereoisomer thereof
  • the solvate of the isomer or a pharmaceutically acceptable salt thereof characterized in that AB represents an antibody, T represents a compound represented by formula (II), and the antibody-drug conjugate is a compound (T) and an antibody ( AB) connected via a linker represented by the following formula (VII):
  • R 1 , R 2 , R 3 and R 4 are as described in the description of the present invention.
  • n 5' represents an integer of 1 to 3
  • each n 5 independently represents an integer of 1 to 8
  • W and W' represent an integer Or a single bond
  • the position 1 of W indicates that it is connected to Z
  • the position 2 indicates that it is connected to (CH 2 CH 2 -O-)n 5 -
  • the position 1 of W' indicates that it is connected to (CH 2 CH 2 -O-)n 5 - is connected
  • position 2 indicates that it is connected to -CH 2- of L 4
  • W and W' are not at the same time
  • Cyclo represents a cyclohexane group
  • L 3 represents or a single bond, each n independently represents an integer from 1 to 6 (eg 2);
  • LP represents a peptide residue consisting of 1 to 7 amino acids
  • R 10 is independently selected from hydrogen, C 1 -C 6 alkyl optionally substituted with 1 or 2 hydroxy (preferably methyl),
  • Q 2 represents -(succinimide-3-yl-N)-, or -Q 1 -NR 10 -, Q 1 is defined as the compound described in the first aspect of the present invention, and Q 1 passes through the carboxyl group of R 11 and - NR 10 - forms an amide bond and connects with L 3 ,
  • -Q 1 -NR 10 - represented by Q 2 is connected by forming a thioether bond between the alkynyl carbon of R 12 and the disulfide bond of the antibody hinge portion,
  • the compound represented by formula (II) is as previously described.
  • the peptide residue of LP is selected from the group consisting of alanine, phenylalanine, glycine, valine, lysine, citrulline, serine, glutamic acid, and aspartic acid peptide residues formed from amino acids.
  • the peptide residues of LP are formed from amino acids selected from the group consisting of phenylalanine, glycine, valine, lysine, citrulline, serine, glutamic acid, and aspartic acid peptide residues.
  • LP is a peptide residue consisting of 1-5 amino acids.
  • LP is a peptide residue selected from the group consisting of:
  • LP is a peptide residue selected from the group consisting of:
  • L 4 represents
  • L b represents
  • the -NR 10 - group and the -(CH 2 )n 8 - group are located in the para position of the benzene ring.
  • the linker represented by formula (VII) is a group selected from the group consisting of the following, wherein each n independently represents an integer of 1-8:
  • the average number of linker-drug linkages is 2-8, preferably 4-8, more preferably 6-8, such as 3.3, 3.5, 5.5, 6.2, 6.5, 6.6, 6.8, 7.0, 7.1, 7.2, 7.4, 7.5 or 7.8.
  • the antibody (AB) is a full-length antibody or antigen-binding fragment thereof, or a bispecific antibody or antigen-binding fragment thereof.
  • the antibody is selected from the group consisting of anti-Her-2 antibody, Trop-2 antibody, EGFR antibody, B7-H3 antibody, PD-1 antibody, PD-L1 antibody, HER-3, HER-4 antibody, CD20 Antibodies, CD30 antibodies, CD19 antibodies, CD33 antibodies; preferably, the antibodies are murine antibodies, chimeric antibodies, and humanized antibodies; preferably, the humanized antibodies are fully human antibodies.
  • the antigen-binding fragment is selected from the group consisting of Fab, Fab', F(ab') 2 , single chain Fv (scFv), Fv and dsFv.
  • the antibody is an anti-TROP-2 antibody, wherein the complementarity determining region (CDR) of the light chain variable region of the anti-Trop-2 antibody comprises CDR1 consisting of the amino acid sequence of KASQDVSIVA, consisting of the amino acid sequence of SASYRYT CDR2 composed of sequence, and CDR3 composed of QQHYITPLT amino acid sequence; CDRs of heavy chain variable region include CDR1 composed of NYGMN amino acid sequence, CDR2 composed of WINTYTGEPTYTDDFKG amino acid sequence, and CDR3 composed of GGFGSSYWYFDV amino acid sequence; preferably,
  • the amino acid sequences of the light chain and heavy chain of the anti-Trop-2 antibody are shown in SEQ ID NO: 1 and SEQ ID NO: 2 respectively; preferably, the coding of the light chain and the heavy chain of the anti-Trop-2 antibody
  • the nucleotide sequences are shown in SEQ ID NO:3 and SEQ ID NO:4, respectively.
  • the antibody is an anti-Her-2 antibody, wherein the complementarity determining region (CDR) of the light chain variable region of the anti-Her-2 antibody comprises CDR1 consisting of the amino acid sequence of RASQDVNTAVA, consisting of the amino acid sequence of SASFLYS CDR2 composed of sequence, and CDR3 composed of QQHYTTPPT amino acid sequence; CDRs of heavy chain variable region include CDR1 composed of DTYIH amino acid sequence, CDR2 composed of RIYPTNGYTRY amino acid sequence, and CDR3 composed of WGGDGFYAMDY amino acid sequence; preferably, The amino acid sequences of the light chain and heavy chain of the anti-Her-2 antibody are shown in SEQ ID NO: 5 and SEQ ID NO: 6, respectively.
  • the ninth aspect of the present invention provides a linker-drug intermediate compound represented by formula (X), characterized in that T represents a compound represented by formula (II), and the intermediate compound is compound (T) with the following The joints represented by formula (IX) are connected:
  • R 1 , R 2 , R 3 and R 4 are as described in the description of the present invention.
  • Q' 2 represents (maleimide-N)- or Q 1 -NR 10 -, and Q 1 is defined as the compound described in the first aspect of the present invention,;
  • L 3 , L 4 , L P and L b are as described in the specification of the present invention.
  • R 10 is independently selected from hydrogen, C 1 -C 6 alkyl optionally substituted with 1 or 2 hydroxy;
  • the nitrogen atom at the 1-position is connected to the methylene group in the linker containing this structure, or Q 1 -NR 10 - represented by Q' 2 , Q 1 forms an amide with -NR 10 - through the carboxyl group of R 11 key and connect with L3 ;
  • the compound of formula (II) is as previously described.
  • the linker-drug intermediate compound is a compound selected from the group consisting of, wherein each n independently represents an integer from 1 to 8:
  • a tenth aspect of the present invention provides a linker structure shown in the general formula (VII):
  • the eleventh aspect of the present invention provides a method for preparing the antibody-drug conjugate of the eighth aspect of the present invention, the method comprising:
  • linker-drug intermediate compound represented by the formula (X) is reacted with AB-SH to connect the linker-drug intermediate represented by the formula (X) through a thioether bond formed by the disulfide bond moiety of the hinge portion of the antibody
  • the compound is linked to the antibody;
  • R 1 , R 2 , R 3 and R 4 are as described in the description of the present invention.
  • T represents the compound represented by the formula (II), and the compound represented by the formula (II) uses the oxygen in the hydroxyl group at the 19th position as the linking site, or when R3 or R4 is a hydroxyl group, the compound represented by the formula (II) uses the oxygen in the hydroxyl group of R3 or R4 as a hydroxyl group .
  • AB-SH represents an antibody carrying a sulfhydryl group
  • AB represents an antibody
  • the twelfth aspect of the present invention provides a method for preparing the linker-drug intermediate compound of the ninth aspect of the present invention.
  • Boc-GGFG and PABOH are generated under the action of EEDQ, using dichloromethane and methanol as solvents, and stirring overnight at room temperature to generate Boc-GGFG-PABOH;
  • N 3 -PEGn-GGFG was condensed with N-Boc-N-methylethylenediamine, and then Boc was removed with TFA/DCM to obtain compound N 3 -PEGn-GGFG-NH-C 2 H 4 -NH -CH 3 ;
  • the thirteenth aspect of the present invention provides intermediate compounds of formula (XI), (XII):
  • the fourteenth aspect of the present invention provides a pharmaceutical composition
  • a pharmaceutical composition comprising the antibody-drug conjugate of the third aspect and the eighth aspect of the present invention, its stereoisomer or a pharmaceutically acceptable salt thereof, or the The antibody-drug conjugate, a solvate of a stereoisomer thereof, or a pharmaceutically acceptable salt thereof, and an optional pharmaceutically acceptable carrier.
  • the fifteenth aspect of the present invention provides a pharmaceutical preparation comprising the antibody-drug conjugate of the third aspect and the eighth aspect of the present invention, a stereoisomer thereof or a pharmaceutically acceptable salt thereof, Or a solvate of the antibody-drug conjugate, a stereoisomer thereof, or a pharmaceutically acceptable salt thereof.
  • the sixteenth aspect of the present invention provides the antibody-drug conjugates, stereoisomers or pharmaceutically acceptable salts thereof, or the antibody-drug conjugates of the third and eighth aspects of the present invention.
  • the antibody-drug conjugates, stereoisomers or pharmaceutically acceptable salts thereof, or the antibody-drug conjugates, stereoisomers thereof according to the third aspect and the eighth aspect of the present invention
  • the tumor or cancer is selected from breast cancer, colorectal cancer, lung cancer, pancreatic cancer, ovarian cancer, prostate cancer, cervical cancer, kidney cancer, urethral cancer, glioblastoma, melanoma, liver cancer , bladder cancer, gastric cancer, esophageal cancer; preferably, the cancer is carcinoma in situ or metastatic cancer; preferably, the breast cancer is Sanyinjiao breast cancer, lung cancer, pancreatic cancer, and colorectal cancer.
  • the seventeenth aspect of the present invention provides a method for preventing or treating tumor or cancer, which comprises administering an effective amount of the antibody-drug couple of the third aspect and the eighth aspect of the present invention to a subject in need thereof.
  • the conjugate, a stereoisomer thereof or a pharmaceutically acceptable salt thereof, or a solvate of the antibody-drug conjugate, a stereoisomer thereof or a pharmaceutically acceptable salt thereof, as described in the fourteenth aspect The pharmaceutical composition or the pharmaceutical preparation of the fifteenth aspect.
  • the tumor or cancer is selected from breast cancer, colorectal cancer, lung cancer, pancreatic cancer, ovarian cancer, prostate cancer, cervical cancer, kidney cancer, urethral cancer, glioblastoma, melanoma, liver cancer , bladder cancer, gastric cancer, esophageal cancer; preferably, the cancer is carcinoma in situ or metastatic cancer; preferably, the breast cancer is Sanyinjiao breast cancer, lung cancer, pancreatic cancer, and colorectal cancer.
  • the eighteenth aspect of the present invention provides the antibody-drug conjugate of the third aspect and the eighth aspect, its stereoisomer or a pharmaceutically acceptable salt thereof, or the antibody-drug conjugate, Use of a solvate of its stereoisomer or a pharmaceutically acceptable salt thereof, the pharmaceutical composition of the fourteenth aspect or the pharmaceutical preparation of the fifteenth aspect for the preparation of a reagent for use in Inhibits cancer cell growth, proliferation or migration.
  • the nineteenth aspect of the present invention provides the antibody-drug conjugate of the third aspect and the eighth aspect, its stereoisomer or a pharmaceutically acceptable salt thereof, or the antibody-drug conjugate, A solvate of its stereoisomer or a pharmaceutically acceptable salt thereof, the pharmaceutical composition of the fourteenth aspect or the pharmaceutical preparation of the fifteenth aspect, which are used for inhibiting the growth, proliferation or migrate.
  • the twentieth aspect of the present invention provides a method for inhibiting the growth, proliferation or migration of cancer cells, comprising administering to the cancer cells an effective amount of the antibody-drug conjugates of the third aspect, the eighth aspect of the present invention, Its stereoisomer or a pharmaceutically acceptable salt thereof, or the antibody-drug conjugate, a solvate of its stereoisomer or a pharmaceutically acceptable salt thereof, the drug described in the fourteenth aspect
  • the composition or the pharmaceutical preparation of the fifteenth aspect comprising administering to the cancer cells an effective amount of the antibody-drug conjugates of the third aspect, the eighth aspect of the present invention, Its stereoisomer or a pharmaceutically acceptable salt thereof, or the antibody-drug conjugate, a solvate of its stereoisomer or a pharmaceutically acceptable salt thereof, the drug described in the fourteenth aspect.
  • the twenty-first aspect of the present invention provides a kit for inhibiting the growth, proliferation or migration of cancer cells, which comprises the antibody-drug conjugates and stereoisomers thereof according to the third and eighth aspects of the present invention or a pharmaceutically acceptable salt thereof, or the antibody-drug conjugate, a solvate of a stereoisomer or a pharmaceutically acceptable salt thereof, the pharmaceutical composition of the fourteenth aspect or the tenth
  • a kit for inhibiting the growth, proliferation or migration of cancer cells which comprises the antibody-drug conjugates and stereoisomers thereof according to the third and eighth aspects of the present invention or a pharmaceutically acceptable salt thereof, or the antibody-drug conjugate, a solvate of a stereoisomer or a pharmaceutically acceptable salt thereof, the pharmaceutical composition of the fourteenth aspect or the tenth
  • a kit for inhibiting the growth, proliferation or migration of cancer cells which comprises the antibody-drug conjugates and stereoisomers thereof according to the third and eighth aspects of the present invention or a pharmaceutically acceptable salt thereof, or the antibody
  • a twenty-second aspect of the present invention provides an antibody-drug conjugate represented by formula (IV), a stereoisomer thereof or a pharmaceutically acceptable salt thereof, or the antibody-drug conjugate, a stereoisomer thereof
  • R 2 is selected from hydrogen, halogen, hydroxy, nitro, amino, saturated or unsaturated C 1 -C 6 alkyl, C 1 -C 6 alkoxy, or C 1 -C 6 substituted by -NR 7 R 8 alkyl;
  • R 4 is selected from hydrogen, halogen, hydroxyl, nitro, amino, C 1 -C 6 alkyl, or C 1 -C 6 alkoxy;
  • R1 and R2 can be linked together with the parent moiety to form a 5-6 membered ring optionally substituted with R9 ;
  • R and R can be linked together with the parent moiety to form a 5-6 membered oxygen-containing heterocycle optionally substituted with R ;
  • R 7 and R 8 are independently selected from hydrogen, C 1 -C 6 alkyl, C 1 -C 6 acyl substituted with hydroxy or amino; or R 7 and R 8 may be combined with the N atom to which they are attached taken together to form a 5-6 membered nitrogen-containing heterocycle optionally substituted by R;
  • R 9 is independently selected from halogen, hydroxy, nitro, -NR 7 R 8 , C 1 -C 6 alkyl, C 1 -C 6 alkoxy, optionally C 1 -C 6 alkane substituted piperidinyl;
  • R 13 represents a carboxy-substituted C 1 -C 6 alkyl group
  • L 2 represents valine residue, guanidine residue, phenylalanine residue, lysine residue, D-valine residue, glycine residue, alanine residue, aspartic acid Residues;
  • R 10 is independently selected from hydrogen, C 1 -C 6 alkyl optionally substituted with 1 or 2 hydroxyl groups, n 3 represents an integer of 1 to 4, and n 4 represents an integer of 1 to 4;
  • Aryl represents a C 6 -C 10 aryl optionally substituted by R 9 ;
  • W is a single key or Wherein, position 1 means connecting with -NR 10 -, and position 2 means connecting with (CH 2 CH 2 -O-)n 1 -;
  • Q 1 is the compound described in the first aspect of the present invention, which is connected by the carboxyl group of R 11 and the left-terminal amino group -NR 10 - in the formula of L 1 to form an amide bond, and the alkynyl carbon of R 12 and the disulfide of the hinge part of the antibody are connected. bond to form a thioether bond,
  • R 2 represents hydrogen, C 3 -C 4 alkenyl, nitro, amino, or C 1 -C 4 alkyl substituted with -N(C 1 -C 4 alkyl) 2 .
  • R 3 represents hydrogen, halogen, hydroxyl, or
  • R 4 represents hydrogen or halogen.
  • R 1 and R 2 are joined together to form the group shown below in A moiety represents a bond to the parent group.
  • R and R are joined together to form the group shown below in A moiety represents a bond to the parent group.
  • the compound represented by formula (II) is gimatecan or gimitecan, more preferably gimatecan:
  • L 2 represents a lysine residue
  • n 4 represents an integer from 1 to 2
  • R 10 represents hydrogen or C 1 -C 4
  • Aryl represents a benzene ring group, preferably, the -NR 10 - group and the -(CH 2 )n 4 - group are located in the para position of the benzene ring.
  • the linker represented by formula (III) is a group selected from the group consisting of:
  • the average number of linker-drug linkages is 2-8, preferably 4-8, more preferably 6-8.
  • the antibody (AB) is a full-length antibody or antigen-binding fragment thereof, or a bispecific antibody or antigen-binding fragment thereof.
  • the antibody is selected from the group consisting of anti-He-r2 antibody, Trop-2 antibody, EGFR antibody, B7-H3 antibody, PD-1 antibody, PD-L1 antibody, HER-3, HER4 antibody, CD20 antibody, CD30 antibody, CD19 antibody, CD33 antibody.
  • the antibody is a murine antibody, a chimeric antibody, a humanized antibody; preferably, the humanized antibody is a fully human antibody.
  • the antigen-binding fragment is selected from the group consisting of Fab, Fab', F(ab') 2 , single chain Fv (scFv), Fv and dsFv.
  • the antibody is an anti-TROP-2 antibody, wherein the complementarity determining region (CDR) of the light chain variable region of the anti-Trop-2 antibody comprises CDR1 consisting of the amino acid sequence of KASQDVSIVA, consisting of the amino acid sequence of SASYRYT CDR2 composed of sequence, and CDR3 composed of QQHYITPLT amino acid sequence; CDRs of heavy chain variable region include CDR1 composed of NYGMN amino acid sequence, CDR2 composed of WINTYTGEPTYTDDFKG amino acid sequence, and CDR3 composed of GGFGSSYWYFDV amino acid sequence; preferably,
  • the amino acid sequences of the light chain and heavy chain of the anti-Trop-2 antibody are shown in SEQ ID NO: 1 and SEQ ID NO: 2 respectively; preferably, the coding of the light chain and the heavy chain of the anti-Trop-2 antibody
  • the nucleotide sequences are shown in SEQ ID NO: 3 and SEQ ID NO: 4, respectively.
  • the antibody is an anti-Her-2 antibody, wherein the complementarity determining region (CDR) of the light chain variable region of the anti-Her2 antibody comprises CDR1 consisting of the amino acid sequence of RASQDVNTAVA, consisting of the amino acid sequence of SASFLYS CDR2, and CDR3 composed of QQHYTTPPT amino acid sequence; CDRs of heavy chain variable region include CDR1 composed of DTYIH amino acid sequence, CDR2 composed of RIYPTNGYTRY amino acid sequence, and CDR3 composed of WGGDGFYAMDY amino acid sequence; preferably, the The amino acid sequences of the light chain and heavy chain of the anti-Her2 antibody are shown in SEQ ID NO: 5 and SEQ ID NO: 6, respectively.
  • the twenty-third aspect of the present invention provides a linker-drug intermediate compound represented by formula (VI), wherein T represents a compound represented by formula (II), and the intermediate compound is compound (T) with the following formula ( The joints indicated by V) are connected:
  • R 1 , R 2 , R 3 and R 4 are as described in the twenty-second aspect of the present invention.
  • L 1 , L 2 and La are as described in the twenty-second aspect of the present invention.
  • Q 1 is the compound described in the first aspect of the present invention, which is connected by forming an amide bond between the carboxyl group of R 11 and the left-terminal amino group -NR 10 - in the formula of L 1 ,
  • the compound of formula (II) is described in the twenty-second aspect of the present invention.
  • the linker-drug intermediate compound is a compound selected from the group consisting of,
  • a twenty-fourth aspect of the present invention provides a linker, wherein it is represented by the following formula (III),
  • the linker is a linker selected from the twenty-second aspect of the present invention.
  • a twenty-fifth aspect of the present invention provides an antibody-drug conjugate represented by formula (VIII), a stereoisomer thereof or a pharmaceutically acceptable salt thereof, or the antibody-drug conjugate, a stereoisomer thereof
  • R 1 , R 2 , R 3 and R 4 are as described in the twenty-second aspect of the present invention.
  • L 3 represents -ZW-(CH 2 CH 2 -O)n 5 -W'- or a single bond
  • n 5 represents an integer from 1 to 8
  • W and W' represent Or a single bond
  • the position 1 of W indicates that it is connected to Z
  • the position 2 indicates that it is connected to (CH 2 CH 2 -O-)n 5 -
  • the position 1 of W' indicates that it is connected to (CH 2 CH 2 -O-)n 5 - is connected
  • position 2 indicates that it is connected to -CH 2- of L 4
  • W and W' are not at the same time
  • Cyclo represents a cyclohexane group
  • LP represents a peptide residue consisting of 2-7 amino acids
  • Aryl represents a C 6 -C 10 aryl group optionally substituted by R 9 , n 7 represents an integer of 1-4, and n 8 represents an integer of 1-4;
  • R 10 is independently selected from hydrogen, C 1 -C 6 alkyl optionally substituted with 1 or 2 hydroxy;
  • Q 2 represents -(succinimide-3-yl-N)-, or -Q 1 -NR 10 -, Q 1 is the compound described in the first aspect of the present invention, and Q 1 passes through the carboxyl group of R 11 and -NR 10 - form an amide bond to connect with L 3 ;
  • Q 2 is -(succinimide-3-yl-N)-, with the following formula:
  • the compound represented by formula (II) is as described in the twenty-second aspect of the present invention.
  • the peptide residues of LP are formed from amino acids selected from the group consisting of phenylalanine, glycine, valine, lysine, citrulline, serine, glutamic acid, and aspartic acid peptide residues.
  • LP is a peptide residue consisting of 2-5 amino acids.
  • LP is a peptide residue selected from the group consisting of:
  • the -NR 10 - group and the -(CH 2 )n 8 - group are located in the para position of the benzene ring.
  • the linker represented by formula (VII) is a group selected from the group consisting of:
  • the average number of linker-drug linkages is 2-8, preferably 4-8, more preferably 6-8.
  • the antibody (AB) is a full-length antibody or antigen-binding fragment thereof, or a bispecific antibody or antigen-binding fragment thereof.
  • the antibody is selected from the group consisting of anti-Her-2 antibody, Trop-2 antibody, EGFR antibody, B7-H3 antibody, PD-1 antibody, PD-L1 antibody, HER3, HER4 antibody, CD20 antibody, CD30 antibody , CD19 antibody, CD33 antibody.
  • the antibody is a murine antibody, a chimeric antibody, a humanized antibody; preferably, the humanized antibody is a fully human antibody.
  • the antigen-binding fragment is selected from the group consisting of Fab, Fab', F(ab') 2 , single chain Fv (scFv), Fv and dsFv.
  • the antibody is an anti-TROP-2 antibody, wherein the complementarity determining region (CDR) of the light chain variable region of the anti-Trop-2 antibody comprises CDR1 consisting of the amino acid sequence of KASQDVSIVA, consisting of the amino acid sequence of SASYRYT CDR2 composed of sequence, and CDR3 composed of QQHYITPLT amino acid sequence; CDRs of heavy chain variable region include CDR1 composed of NYGMN amino acid sequence, CDR2 composed of WINTYTGEPTYTDDFKG amino acid sequence, and CDR3 composed of GGFGSSYWYFDV amino acid sequence; preferably,
  • the amino acid sequences of the light chain and heavy chain of the anti-Trop-2 antibody are shown in SEQ ID NO: 1 and SEQ ID NO: 2 respectively; preferably, the coding of the light chain and the heavy chain of the anti-Trop-2 antibody
  • the nucleotide sequences are shown in SEQ ID NO:3 and SEQ ID NO:4, respectively.
  • the antibody is an anti-Her-2 antibody, wherein the complementarity determining region (CDR) of the light chain variable region of the anti-Her2 antibody comprises CDR1 consisting of the amino acid sequence of RASQDVNTAVA, consisting of the amino acid sequence of SASFLYS CDR2, and CDR3 composed of QQHYTTPPT amino acid sequence; CDRs of heavy chain variable region include CDR1 composed of DTYIH amino acid sequence, CDR2 composed of RIYPTNGYTRY amino acid sequence, and CDR3 composed of WGGDGFYAMDY amino acid sequence; preferably, the The amino acid sequences of the light chain and heavy chain of the anti-Her2 antibody are shown in SEQ ID NO: 5 and SEQ ID NO: 6, respectively.
  • the twenty-sixth aspect of the present invention provides a linker-drug intermediate compound represented by formula (X), wherein T represents a compound represented by formula (II), and the intermediate compound is compound (T) with the following formula ( IX) is formed by connecting the joints indicated:
  • R 1 , R 2 , R 3 and R 4 are as described in the twenty-second aspect of the present invention.
  • Q' 2 represents (maleimide-N)- or Q 1 -NR 10 -, and Q 1 is the compound described in the first aspect of the present invention,;
  • L 3 , L 4 , L P and L b are as described in the twenty-fifth aspect of the present invention.
  • R 10 is independently selected from hydrogen, C 1 -C 6 alkyl optionally substituted with 1 or 2 hydroxy;
  • Q' 2 represents (maleimide-N)-, the following formula structure:
  • the nitrogen atom at the 1-position is connected with the methylene group in the linker containing this structure, or Q' 2 represents Q 1 -NR 10 -, and Q 1 forms an amide bond with -NR 10 - through the carboxyl group of R 11 And connect with L3 ;
  • the compound of formula (II) is described in the twenty-second aspect of the present invention.
  • the linker-drug intermediate compound is a compound selected from the group consisting of:
  • a twenty-seventh aspect of the present invention provides a linker, wherein it is represented by the following formula (VII)
  • the linker is a structure selected from the twenty-fifth aspect of the present invention.
  • a twenty-eighth aspect of the present invention provides a pharmaceutical composition comprising the antibody-drug conjugate, a stereoisomer thereof, or a pharmaceutically acceptable compound thereof according to the twenty-second or twenty-fifth aspect of the present invention.
  • a twenty-ninth aspect of the present invention provides a pharmaceutical preparation comprising the antibody-drug conjugate of the twenty-second or twenty-fifth aspect of the present invention, a stereoisomer thereof, or a pharmaceutically acceptable compound thereof.
  • the thirtieth aspect of the present invention provides the antibody-drug conjugate of the twenty-second aspect or the twenty-fifth aspect of the present invention, a stereoisomer thereof or a pharmaceutically acceptable salt thereof, or the antibody - a drug conjugate, a solvate of a stereoisomer thereof or a pharmaceutically acceptable salt thereof, the pharmaceutical composition of the twenty-eighth aspect of the present invention and/or the pharmaceutical preparation of the twenty-ninth aspect of the present invention for the prevention and/or treatment of tumors or cancers.
  • the tumor or cancer is selected from breast cancer, colorectal cancer, lung cancer, pancreatic cancer, ovarian cancer, prostate cancer, cervical cancer, kidney cancer, urethral cancer, glioblastoma, melanoma, liver cancer , bladder cancer, gastric cancer, esophageal cancer; preferably, the cancer is carcinoma in situ or metastatic cancer; preferably, the breast cancer is Sanyinjiao breast cancer.
  • the thirty-first aspect of the present invention provides a method of preventing or treating cancer, comprising administering to a subject in need thereof a prophylactically or therapeutically effective amount of the method of the twenty-second or twenty-fifth aspect of the present invention
  • the antibody-drug conjugate, its stereoisomer or a pharmaceutically acceptable salt thereof, or a solvate of the antibody-drug conjugate, its stereoisomer or a pharmaceutically acceptable salt thereof the pharmaceutical composition of the twenty-eighth aspect of the present invention and/or the pharmaceutical preparation of the twenty-ninth aspect of the present invention.
  • the thirty-second aspect of the present invention provides the antibody-drug conjugate of the twenty-second or twenty-fifth aspect of the present invention, a stereoisomer thereof or a pharmaceutically acceptable salt thereof, or the Antibody-drug conjugate, solvate of its stereoisomer or pharmaceutically acceptable salt thereof, the pharmaceutical composition of the twenty-eighth aspect of the present invention and/or the drug of the twenty-ninth aspect of the present invention Use of a formulation for inhibiting cancer cell growth, proliferation or migration.
  • the thirty-third aspect of the present invention provides the antibody-drug conjugate of the twenty-second or twenty-fifth aspect of the present invention, a stereoisomer thereof or a pharmaceutically acceptable salt thereof, or the Antibody-drug conjugate, solvate of its stereoisomer or pharmaceutically acceptable salt thereof, the pharmaceutical composition of the twenty-eighth aspect of the present invention and/or the drug of the twenty-ninth aspect of the present invention Agents for inhibiting the growth, proliferation or migration of cancer cells.
  • the thirty-fourth aspect of the present invention provides a method for inhibiting the growth, proliferation or migration of cancer cells, comprising administering to the cancer cells an effective amount of the antibody-drug according to the twenty-second or twenty-fifth aspect of the present invention
  • the conjugate, its stereoisomer or its pharmaceutically acceptable salt, or the solvate of said antibody-drug conjugate, its stereoisomer or its pharmaceutically acceptable salt, the second of the present invention The pharmaceutical composition of the eighteenth aspect and/or the pharmaceutical preparation of the twenty-ninth aspect of the present invention.
  • the thirty-fifth aspect of the present invention provides a kit for inhibiting the growth, proliferation or migration of cancer cells, comprising the antibody-drug conjugate of the twenty-second or twenty-fifth aspect of the present invention, its A stereoisomer or a pharmaceutically acceptable salt thereof, or the antibody-drug conjugate, a solvate of a stereoisomer or a pharmaceutically acceptable salt thereof, the drug of the twenty-eighth aspect of the present invention.
  • the thirty-sixth aspect of the present invention provides the antibody-drug conjugate of the twenty-second aspect of the present invention, a stereoisomer thereof or a pharmaceutically acceptable salt thereof, or the antibody-drug conjugate , the preparation method of the solvate of its stereoisomer or its pharmaceutically acceptable salt, described method comprises:
  • linker-drug intermediate compound represented by the formula (VI) is reacted with AB-SH to connect the linker-drug intermediate represented by the formula (VI) through a thioether bond formed by the disulfide bond moiety of the hinge portion of the antibody
  • the compound is linked to the antibody;
  • R 1 , R 2 , R 3 and R 4 are as described in the twenty-second aspect of the present invention.
  • T represents the compound represented by the formula (II), and the compound represented by the formula (II) uses the oxygen in the hydroxyl group at the 19th position as the linking site, or when R 3 or R 4 is a hydroxyl group, the oxygen in the hydroxyl group of R 3 or R 4 is used as a linking site.
  • AB-SH represents an antibody carrying a sulfhydryl group
  • AB represents an antibody
  • the thirty-seventh aspect of the present invention provides a method for preparing a linker-drug intermediate compound of the twenty-third aspect of the present invention, the method comprising:
  • N-[(9H-fluoren-9-ylmethoxy)carbonyl]-N'-[(4-methoxyphenyl)diphenylmethyl]-L-lysine (CN- CMTC-1) and PABOH were dissolved in dichloromethane: methanol solution, reacted under the action of EEDQ, recrystallized and purified to obtain the product;
  • step (2) treating the product of step (1) with a solution of piperidine in acetonitrile, and then purifying the product;
  • step (3) reacts with the product of step (2) to generate compound
  • Gematecan-Boc or Gematecan-Boc is treated with triphosgene, DMAP, and dichloromethane to form a formyl chloride compound, and then the reaction compound of step (4) is added, and then deprotected with TFA/DCM. ;
  • step (5) The product of step (5) is subjected to Click reaction with the compound described in the first aspect of the present invention, and the final product is obtained after being treated with TFA/DCM;
  • step (6) can also be replaced by the following steps: the product of step (5) and SM-1 are added to a solution of DMSO/H 2 O, and then CuBr is added to catalyze the reaction to complete the reaction. After purification, TFA/DCM is added for deprotection , the final product is obtained;
  • the SM-1 is N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N
  • the thirty-eighth aspect of the present invention provides the antibody-drug conjugate of the twenty-fifth aspect of the present invention, a stereoisomer thereof or a pharmaceutically acceptable salt thereof, or the antibody-drug conjugate , the preparation method of the solvate of its stereoisomer or its pharmaceutically acceptable salt, described method comprises:
  • linker-drug intermediate compound represented by the formula (X) is reacted with AB-SH to connect the linker-drug intermediate represented by the formula (X) through a thioether bond formed by the disulfide bond moiety of the hinge portion of the antibody
  • the compound is linked to the antibody;
  • R 1 , R 2 , R 3 and R 4 are as described in the twenty-second aspect of the present invention.
  • Q 2 , Q' 2 , L 3 , L 4 , L P and La are as described in the twenty-fifth or twenty-sixth aspect of the present invention.
  • T represents the compound represented by the formula (II), and the compound represented by the formula (II) uses the oxygen in the hydroxyl group at the 19th position as the linking site, or when R 3 or R 4 is a hydroxyl group, the oxygen in the hydroxyl group of R 3 or R 4 is used as a linking site.
  • AB-SH represents an antibody carrying a sulfhydryl group
  • AB represents an antibody
  • the thirty-ninth aspect of the present invention provides a method for preparing the linker-drug intermediate compound of the twenty-sixth aspect of the present invention, the method comprising:
  • Boc-GGFG and PABOH are generated under the action of EEDQ, using dichloromethane and methanol as solvents, and stirring overnight at room temperature to generate Boc-GGFG-PABOH;
  • N 3 -PEGn-GGFG was condensed with N-Boc-N-methylethylenediamine, and then Boc was removed with TFA/DCM to obtain compound N 3 -PEGn-GGFG-NH-C 2 H 4 -NH -CH 3 ;
  • N 3 -PEGn-GGFG-NH-C 2 H 4 -NH-CH 3 and gimatecan-PNP (or gimatecan-PNP, SN-38-PNP) in the condition of TEA, DMF The following reaction is carried out to obtain the compound N 3 -PEGn-GGFG-NH-C 2 H 4 -N(CH 3 )-C(O)-gemitecan (or SN-38, or gematecan),
  • FIG. 1 shows the SEC-HPLC results of ADC-1.
  • Figure 2 is the SEC-HPLC profile of ADC-5.
  • Figure 3 is the SEC-HPLC profile of ADC-6.
  • Figure 4 is the SEC-HPLC profile of ADC-8.
  • Figure 5 is the SEC-HPLC profile of ADC-10.
  • Figure 6 is the SEC-HPLC profile of ADC-11.
  • Figure 7 is the SEC-HPLC profile of ADC-12.
  • Figure 8 is the SEC-HPLC profile of ADC-13.
  • Figure 9 is the SEC-HPLC profile of ADC-14.
  • Figure 10 is the SEC-HPLC profile of ADC-15.
  • Figure 11 is the SEC-HPLC profile of ADC-16.
  • Figure 12 is the SEC-HPLC profile of ADC-17.
  • Figure 13 is a release-peak localization map of ADC-5 small molecules.
  • Figure 14 is a release-line graph of ADC-5 small molecule.
  • Figure 15 is the percent change in the area of the main peak in the CL2A-CM sample.
  • Figure 16 is the CM peak area percent change in CL2A-CM samples.
  • Figure 17 is the percent change in the area of the main peak in ADC-5 samples.
  • Figure 18 is the CM peak area percent change in ADC-5 samples.
  • Figure 19 shows the results of inhibition of BXPC-3 cell activity by four ADCs.
  • Figure 20 shows the IC50 of the test drug on BxPC-3.
  • Figure 21 shows the IC50 of the test drug on COLO 205.
  • Figure 22 shows the IC50 of the tested drugs on Calu-3.
  • Figure 23 shows the IC50 of the tested drugs on Calu-6.
  • Figure 24 shows the IC50 of the tested drugs on NCI-N87.
  • Figure 25 is the antitumor activity of ADC-5 in the BxPC-3 tumor model.
  • Figure 26 shows the effect of ADC-5 on the body weight of the BxPC-3 model.
  • Figure 27 shows the anti-tumor activity of ADC-137 and ADC-5 in COLO 205 tumor model.
  • Figure 28 shows the effects of ADC-137 and ADC-5 on the body weight of the COLO 205 model.
  • Figure 29 shows the antitumor activity of ADC-137 and ADC-5 in BxPC-3 tumor model.
  • Figure 30 shows the effects of ADC-137 and ADC-5 on the body weight of the BxPC-3 model.
  • Figure 31 shows the antitumor activity of ADC-137 and ADC-5 in Calu-3 tumor model.
  • Figure 32 shows the effect of ADC-137 and ADC-5 on the body weight of Calu-3 model.
  • Figure 33 shows the antitumor activity of ADC-137 and ADC-5 in Capan-1 tumor model.
  • Figure 34 shows the effects of ADC-137 and ADC-5 on the body weight of the Capan-1 model.
  • Figure 35 shows the effect of ADC-16, ADC-5 and ADC-17 on body weight of the COLO 205 model.
  • Figure 36 is the anti-tumor activity of ADC-16, ADC-5 and ADC-17 in the COLO 205 tumor model.
  • Figure 37 is the antitumor activity of ADC-8, ADC-11, ADC-5 and ADC-12 in the BxPC-3 tumor model.
  • Figure 38 is the effect of ADC-8, ADC-11, ADC-5 and ADC-12 on body weight of the BxPC-3 model.
  • antibody refers to immunoglobulin, which is a tetrapeptide chain structure composed of two identical heavy chains and two identical light chains connected by interchain disulfide bonds.
  • the amino acid composition and sequence of the immunoglobulin heavy chain constant region are different, so their antigenicity is also different. Accordingly, immunoglobulins can be divided into five classes, or isotypes of immunoglobulins, namely IgM, IgD, IgG, IgA, and IgE, whose corresponding heavy chains are ⁇ , ⁇ , and ⁇ chains, respectively. , alpha chains, and epsilon chains.
  • IgG can be divided into different subclasses according to the difference in the amino acid composition of the hinge region and the number and position of disulfide bonds in the heavy chain.
  • IgG can be divided into IgG1, IgG2, IgG3, and IgG4.
  • Light chains are classified into kappa chains or lambda chains by the difference in the constant region.
  • Each of the five classes of Ig can have a kappa chain or a lambda chain.
  • the antibody light chain of the present invention may further comprise a light chain constant region comprising human or murine ⁇ , ⁇ chains or variants thereof.
  • the antibody heavy chain of the present invention may further comprise a heavy chain constant region comprising human or murine IgG1, IgG2, IgG3, IgG4 or variants thereof.
  • variable region The sequence of about 110 amino acids near the N-terminus of the antibody heavy and light chains varies greatly, and is the variable region (Fv region); the remaining amino acid sequences near the C-terminus are relatively stable and are the constant region.
  • the variable region includes three hypervariable regions (HVR) and four relatively conserved framework regions (FR). Three hypervariable regions determine the specificity of antibodies, also known as complementarity determining regions (CDRs).
  • CDRs complementarity determining regions
  • Each light chain variable region (LCVR) and heavy chain variable region (HCVR) consists of 3 CDR regions and 4 FR regions. The order from the amino terminus to the carboxy terminus is: FR1, CDR1, FR2, CDR2 , FR3, CDR3, FR4.
  • the three CDR regions of the light chain are referred to as LCDR1, LCDR2, and LCDR3; the three CDR regions of the heavy chain are referred to as HCDR1, HCDR2, and HCDR3.
  • the number and position of CDR amino acid residues in the LCVR and HCVR regions of the antibodies or antigen-binding fragments of the present invention conform to the known Kabat numbering rules (LCDR1-3, HCDR1-3).
  • Antibodies of the present invention include murine antibodies, chimeric antibodies, humanized antibodies, preferably humanized antibodies.
  • an "antibody fragment” or “antigen-binding fragment” of an antibody refers to any portion of a full-length antibody that is less than full-length, but which comprises at least a portion of the variable region (eg, one or more of the variable region of said antibody that binds an antigen) CDRs and/or one or more antibody binding sites), and thus retain binding specificity and at least part of the specific binding capacity of the full-length antibody.
  • an antigen-binding fragment refers to an antibody fragment comprising an antigen-binding portion that binds to the same antigen as the antibody from which the antibody fragment is derived.
  • Antibody fragments include antibody derivatives produced by enzymatic treatment of full-length antibodies, as well as synthetically produced derivatives, eg, recombinantly produced derivatives.
  • Antibodies include antibody fragments. Examples of antibody fragments include, but are not limited to, Fab, Fab', F(ab') 2 , single-chain Fv (scFv), Fv, dsFv, diabodies, Fd and Fd' fragments, and other fragments, including modified fragments (see, For example, Methods in Molecular Biology, Vol 207: Recombinant Antibodies for Cancer Therapy Methods and Protocols (2003); Chapter 1; p 3-25, Kipriyanov).
  • the fragments may comprise multiple chains linked together, eg, by disulfide bonds and/or by peptide linkers.
  • Antibody fragments generally comprise at least or about 50 amino acids, and typically at least or about 200 amino acids.
  • Antigen-binding fragments include any antibody fragment that, when inserted into the antibody framework (eg, by substituting the corresponding region), results in an antibody that immunospecifically binds (ie, exhibits a Ka of at least or at least about 107-108 M -1 ) to an antigen .
  • a "functional fragment” or “analog of an anti-Her-2 antibody” is a fragment or analog that prevents or substantially reduces the ability of the receptor to bind a ligand or initiate signal transduction.
  • a functional fragment generally has the same meaning as an "antibody fragment” and, with respect to an antibody, may refer to a fragment that prevents or substantially reduces the ability of the receptor to bind a ligand or initiate signal transduction, eg, Fv, Fab , F(ab') 2 , and so on.
  • the "Fv" fragment consists of a dimer ( VH - VL dimer) formed by non-covalent association of the variable domains of a heavy chain and the variable domains of a light chain.
  • the three CDRs of each variable domain interact to define the target binding site on the surface of the VH - VL dimer, as is the case with intact antibodies.
  • the six CDRs collectively confer the target-binding specificity of the intact antibody.
  • a single variable domain or half of an Fv that includes only 3 target-specific CDRs
  • BsAb Bispecific antibody
  • a bispecific antibody and/or an antigen-binding molecule contains Two antigen binding sites, each of which is specific for a different antigenic determinant.
  • the bispecific antibody and/or antigen binding molecule is capable of binding two antigenic determinants simultaneously, particularly two antigenic determinants expressed on two different cells.
  • monoclonal antibody or “monoclonal antibody” refers to a population of the same antibody, meaning that each individual antibody molecule in the monoclonal antibody population is identical to other antibody molecules. This property is in contrast to that of polyclonal populations of antibodies, which comprise antibodies with a variety of different sequences.
  • Monoclonal antibodies can be prepared by a number of well-known methods. For example, monoclonal antibodies can be prepared by immortalizing B cells, eg, by fusion with myeloma cells to generate hybridoma cell lines or by infecting B cells with a virus such as EBV. Recombinant techniques can also be used to prepare antibodies from clonal populations of host cells in vitro by transforming the host cells with a plasmid carrying an artificial sequence of nucleotides encoding the antibody.
  • a full-length antibody has two full-length heavy chains (eg VH-CH 1 -CH 2 -CH 3 or VH-CH 1 -CH 2 -CH 3 -CH 4 ) and two full-length light chains ( VL-CL) and hinge region antibodies, such as antibodies naturally produced by antibody-secreting B cells as well as synthetically produced antibodies having the same domains.
  • two full-length heavy chains eg VH-CH 1 -CH 2 -CH 3 or VH-CH 1 -CH 2 -CH 3 -CH 4
  • VL-CL full-length light chains
  • chimeric antibody refers to an antibody in which the variable region sequences are derived from one species and the constant region sequences are derived from another species, such as in which the variable region sequences are derived from a mouse antibody and the constant region sequences are derived from a human antibody of antibodies.
  • Humanized antibodies refer to non-human (eg, mouse) forms of antibodies that are chimeric immunoglobulins, immunoglobulin chains, or fragments thereof (eg, Fv, Fab, Fab', F(ab') 2 or other antigen-binding subsequences of antibodies) containing minimal sequence derived from non-human immunoglobulins.
  • the humanized antibody is a human immunoglobulin (recipient antibody) in which the complementarity determining region (CDR) residues of the recipient antibody are derived from a non-human species with the desired specificity, affinity and capacity ( donor antibody) such as mouse, rat or rabbit CDR residue substitutions.
  • CDR complementarity determining region
  • telomeres can be mutated amino acid residues within the CDR1, CDR2 and/or CDR3 regions of VH and/or VL, thereby improving one or more binding properties (eg, affinity) of the antibody .
  • PCR-mediated mutagenesis can be performed to introduce mutations whose effect on antibody binding or other functional properties can be assessed using the in vitro or in vivo assays described herein. Typically, conservative mutations are introduced. Such mutations can be amino acid substitutions, additions or deletions.
  • an antibody that immunospecifically binds (or specifically binds) an antigen has an affinity constant Ka of about or 1x107 M -1 or 1x108 M -1 or greater (or 1x10-7 M or 1x A dissociation constant (Kd) of 10 ⁇ 8 M or lower binds the antigen.
  • Affinity constants can be determined by standard kinetic methods of antibody response, eg, immunoassays, surface plasmon resonance (SPR), isothermal titration calorimetry (ITC), or other kinetic interaction assays known in the art. Instruments and methods for detecting and monitoring binding rates in real time are known and commercially available.
  • nucleic acid molecules refer to oligomers or polymers comprising at least two linked nucleotides or nucleotide derivatives, including usually linked together by phosphodiester bonds Deoxyribonucleic acid (DNA) and ribonucleic acid (RNA).
  • DNA deoxyribonucleic acid
  • RNA ribonucleic acid
  • nucleic acid molecule is intended to include DNA molecules and RNA molecules. Nucleic acid molecules can be single-stranded or double-stranded, and can be cDNA.
  • an isolated nucleic acid molecule is one that is separated from other nucleic acid molecules present in the natural source of the nucleic acid molecule.
  • An "isolated" nucleic acid molecule such as a cDNA molecule, can be substantially free of other cellular material or culture medium when prepared by recombinant techniques, or substantially free of chemical precursors or other chemical components when chemically synthesized.
  • Exemplary isolated nucleic acid molecules provided herein include isolated nucleic acid molecules encoding the provided antibodies or antigen-binding fragments.
  • operably linked with respect to nucleic acid sequences, regions, elements or domains means that the nucleic acid regions are functionally related to each other.
  • a promoter can be operably linked to a nucleic acid encoding a polypeptide such that the promoter regulates or mediates transcription of the nucleic acid.
  • expression refers to the process of producing a polypeptide by transcription and translation of a polynucleotide.
  • Expression levels of a polypeptide can be assessed using any method known in the art, including, for example, methods that determine the amount of polypeptide produced from a host cell. Such methods may include, but are not limited to, quantification of polypeptides in cell lysates by ELISA, Coomassie blue staining followed by gel electrophoresis, Lowry protein assay, and Bradford protein assay.
  • a "host cell” is a cell for receiving, maintaining, replicating and amplifying a vector.
  • Host cells can also be used to express the polypeptide encoded by the vector. When the host cell divides, the nucleic acid contained in the vector replicates, thereby amplifying the nucleic acid.
  • Host cells can be eukaryotic cells or prokaryotic cells. Suitable host cells include, but are not limited to, CHO cells, various COS cells, HeLa cells, HEK cells such as HEK 293 cells.
  • a "vector” is a replicable nucleic acid from which one or more heterologous proteins can be expressed when transformed into an appropriate host cell.
  • References to vectors include those into which nucleic acids encoding polypeptides or fragments thereof can be introduced, typically by restriction digestion and ligation. References to vectors also include those that contain nucleic acid encoding a polypeptide. Vectors are used to introduce nucleic acid encoding a polypeptide into a host cell, to amplify the nucleic acid, or to express/display the polypeptide encoded by the nucleic acid. Vectors generally remain episomal, but can be designed to integrate the gene or portion thereof into the chromosome of the genome. Also contemplated are artificial chromosome vectors, such as yeast artificial vectors and mammalian artificial chromosomes. The selection and use of such vehicles is well known to those skilled in the art.
  • the vector also includes "viral vector” or "viral vector”.
  • a viral vector is an engineered virus that is operably linked to a foreign gene to transfer (either as a vehicle or shuttle) the foreign gene into a cell.
  • an "expression vector” includes a vector capable of expressing DNA operably linked to regulatory sequences, such as promoter regions, capable of affecting the expression of such DNA fragments. Such additional fragments may include promoter and terminator sequences, and optionally, one or more origins of replication, one or more selectable markers, enhancers, polyadenylation signals, and the like. Expression vectors are typically derived from plasmid or viral DNA, or may contain elements of both. Thus, an expression vector refers to a recombinant DNA or RNA construct, such as a plasmid, phage, recombinant virus, or other vector, which, when introduced into an appropriate host cell, results in the expression of cloned DNA. Appropriate expression vectors are well known to those skilled in the art and include those that are replicable in eukaryotic and/or prokaryotic cells as well as those that remain episomal or that integrate into the host cell genome.
  • drug (drug compound) in the present invention, that is, “toxin” refers to a cytotoxic drug, that is, a compound represented by formula (I) (anti-tumor compound), which can strongly disrupt the normal growth of tumor cells. chemical molecules.
  • cytotoxic drugs can kill tumor cells at a high enough concentration, but due to the lack of specificity, they can also lead to normal cell apoptosis while killing tumor cells.
  • the term includes toxins, such as small molecule toxins or enzymatically active toxins of fungal, bacterial, plant or animal origin, radioisotopes (eg I 131 , Y 90 , Re 186 , I 125 ), toxic drugs, chemotherapeutic drugs, antibiotics and nucleolytic agents
  • the enzyme is preferably a toxic drug, more preferably a camptothecin derivative, more preferably gimitecan and gimatecan.
  • C a -C b (a and b represent an integer of 1 or more, a ⁇ b) includes any specific case of a to b carbons, for example, C 1 -C 6 includes C 1 and C 2 , C 3 , C 4 , C 5 , C 6 , also including any one range of a to b, for example, C 1 -C 6 includes C 1 -C 3 , C 1 -C 4 , C 1 -C 5 , C 2 -C 5 , C 2 -C 4 , C 3 -C 6 , etc.; similarly, "ab-membered ring” (a and b represent an integer of 1 or more, a ⁇ b) represents that the number of ring atoms is a to b
  • the ring structure for example, 3-6 membered ring includes 3-membered ring, 4-membered ring, 5-membered ring, 6-membered ring, and also includes any range from a to b, for
  • halogen refers to fluorine, chlorine, bromine and iodine.
  • C 1 -C 6 hydrocarbon group refers to a straight-chain or branched alkyl group derived from an alkane moiety containing 1-6 carbon atoms by removing one hydrogen atom
  • C 1 -C 6 hydrocarbyl may be saturated, ie "C 1 -C 6 alkyl”
  • C 1-6 alkyl includes but is not limited to methyl, ethyl, n-propyl, isopropyl, n-butyl , isobutyl, sec-butyl, tert-butyl, n-pentyl, isopentyl, 2-methylbutyl, neopentyl, 1-ethylpropyl, n-hexyl, isohexyl, 4-methylpentyl , 3-methylpentyl, 2-methylpentyl, 1-methylpentyl, 3,3-dimethylbutyl, 2,2-dimethylbutyl,
  • the "C 1 -C 6 hydrocarbyl” in the present invention may be unsaturated, such as “C 2 -C 6 alkenyl”, “C 3 -C 4 alkenyl”, and “C 3 -C 4 alkenyl” including but not limited to Not limited to propenyl, 1-butenyl, 2-butenyl, and the like.
  • C 1-6 alkoxy refers to a group in which "C 1-6 alkyl” as defined above is connected to the rest of the molecule via an oxygen atom, namely "C 1-6 alkyl-O"-” groups, specifically, include, but are not limited to, for example, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, sec-butoxy, tert-butoxy group, neopentyloxy, n-hexyloxy, etc.; the "C 1-4 alkoxy” refers to a group in which the above-defined "C 1-4 alkyl” is connected to the rest of the molecule through an oxygen atom , namely "C 1-4 alkyl-O-” group, specifically, including but not limited to, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy group, sec-butoxy, tert-butoxy.
  • 5-6 membered ring refers to a non-aromatic cyclic structure with 5-6 ring atoms, and the ring atoms can be all carbon atoms, thereby forming a carbocyclic ring; it can also contain 1- 3 ring heteroatoms each independently selected from N, O, or S, thereby forming a heterocycle (eg, oxygen-containing heterocycle, nitrogen-containing heterocycle, sulfur-containing heterocycle); the 5-6 membered ring may be saturated
  • the structure can also be an unsaturated structure containing 1 or 2 carbon-carbon double bonds or carbon-carbon triple bonds.
  • 5-6 membered nitrogen-containing heterocycle includes but is not limited to piperidine and piperazine, preferably piperidine.
  • C 6 -C 10 aryl group refers to an aromatic cyclic hydrocarbon group having 6-10 ring-forming carbon atoms, which can be a monovalent group or a divalent or higher group as required, including Monocyclic aryl group and fused-ring aryl group, "fused-ring aryl group” refers to an aryl group containing multiple rings (eg, containing 2) in which each ring in the group shares an adjacent pair of ring carbon atoms with other rings. .
  • the "C 6 -C 10 -membered aryl group” specifically includes a phenyl group and a naphthyl group.
  • linker refers to a chemical structural fragment or bond that is connected to an antibody at one end and a drug (drug compound) at the other end. Other linkers can also be connected. It is then linked to the drug compound.
  • linker structure of the present invention can be synthesized by methods known in the art, or can be synthesized using the methods described in the present invention.
  • the "antibody-drug conjugate" of the present invention refers to a ligand linked to a biologically active drug through a stable linking unit. In the present invention, it refers to linking the monoclonal antibody or fragment with the biologically active toxic drug through the linker structure.
  • salts refer to relatively nontoxic acid addition salts or base addition salts of the conjugates of the present invention.
  • the acid addition salts are salts formed by the conjugates of the present invention with suitable inorganic or organic acids, and these salts can be prepared by subjecting the conjugates of the present invention with suitable organic or inorganic acids in a suitable solvent reaction to prepare.
  • Representative acid addition salts include hydrobromide, hydrochloride, sulfate, bisulfate, sulfite, acetate, oxalate, valerate, oleate, palmitate, stearic acid Salt, laurosilicate, borate, benzoate, lactate, nitrate, phosphate, hydrogen phosphate, carbonate, bicarbonate, toluate, citrate, maleic acid Salt, fumarate, succinate, malate, ascorbate, tannate, pamoate, alginate, naphthalene sulfonate, tartrate, benzoate, mesylate, p-toluene Sulfonate, gluconate, lactobionate and lauryl sulfonate, etc.
  • the base addition salts are the salts formed by the conjugates of the present invention and suitable inorganic or organic bases, and these salts can be carried out by making the conjugates of the present invention and suitable inorganic or organic bases in a suitable solvent. reaction to prepare.
  • Representative base addition salts include, for example, salts formed with alkali metal, alkaline earth metal, quaternary ammonium cations, such as sodium, lithium, potassium, calcium, magnesium, tetramethylquaternary ammonium, tetraethylquaternary ammonium salts, etc.; amine salts, including salts formed with ammonia (NH 3 ), primary, secondary or tertiary amines, such as methylamine salts, dimethylamine salts, trimethylamine salts, triethylamine salts, ethylamine salts, and the like.
  • quaternary ammonium cations such as sodium, lithium, potassium, calcium, magnesium, tetramethylquaternary ammonium,
  • the conjugates of the present invention may exist in specific geometric or stereoisomeric forms.
  • the chiral center may exist in the antitumor compound (the compound represented by formula (I)), or may exist in the
  • the linker structure (the linker represented by formula (II)) may also exist in antibodies and derivatives thereof.
  • Optically active (R)- and (S)-isomers can be prepared by chiral synthesis or chiral reagents or other conventional techniques. If one enantiomer of the conjugates of the present invention is desired, it can be prepared by asymmetric synthesis or derivatization with a chiral auxiliary, wherein the resulting mixture of diastereomers is separated and the auxiliary group is cleaved to provide Pure desired enantiomer.
  • a diastereomeric salt is formed with an appropriate optically active acid or base, followed by conventional methods known in the art
  • the diastereoisomers were resolved and the pure enantiomers recovered.
  • separation of enantiomers and diastereomers is usually accomplished by the use of chromatography employing a chiral stationary phase, optionally in combination with chemical derivatization (eg, from amines to amino groups) formate).
  • solvates eg, hydrates
  • suitable solvates include solvates of the conjugate of the present invention with acetone, 2-butanol, 2-propanol, ethanol, ethyl acetate, tetrahydrofuran, diethyl ether, and the like. Hydrates or ethanolates can also be cited.
  • treating an individual suffering from a disease or condition means that the individual's symptoms are partially or completely alleviated, or remain unchanged after treatment.
  • treatment includes prevention, treatment and/or cure.
  • Prevention refers to preventing an underlying disease and/or preventing the worsening of symptoms or the development of a disease.
  • Treatment also includes any pharmaceutical use of the provided ADCs as well as the pharmaceutical compositions, pharmaceutical formulations provided herein.
  • therapeutic effect means the effect resulting from the treatment of an individual, which alters, generally ameliorates or ameliorates the symptoms of a disease or disease condition, or cures a disease or disease condition.
  • a “therapeutically effective amount” or “therapeutically effective dose” refers to an amount of a substance, compound, material or composition comprising a compound that is at least sufficient to produce a therapeutic effect after administration to a subject. Thus, it is an amount necessary to prevent, cure, ameliorate, retard or partially retard the symptoms of a disease or disorder.
  • a prophylactically effective amount or “prophylactically effective dose” refers to an amount of a substance, compound, material or composition comprising a compound that, when administered to a subject, will have a desired prophylactic effect, eg, prevent or delay a disease or symptom occurrence or recurrence, and reduce the likelihood of occurrence or recurrence of disease or symptoms.
  • a fully prophylactically effective dose need not occur by administering one dose, and may occur only after administering a series of doses.
  • a prophylactically effective amount can be administered in one or more administrations.
  • the antitumor compound is not particularly limited as long as it is a compound having an antitumor effect or a compound having a substituent capable of being linked to a linker structure.
  • a part or the whole of the linker is cleaved in tumor cells to free the antitumor compound part, thereby exhibiting an antitumor effect.
  • the linker is cleaved with the linking part of the drug, the antitumor compound is released in its original structure, and its original antitumor effect is exerted.
  • the antitumor compound in the present invention is a compound represented by the following formula (II).
  • R 2 is selected from hydrogen, halogen, hydroxy, nitro, amino, saturated or unsaturated C 1 -C 6 alkyl, C 1 -C 6 alkoxy, or C 1 -C 6 substituted by -NR 7 R 8 alkyl;
  • R 4 is selected from hydrogen, halogen, hydroxyl, nitro, amino, C 1 -C 6 alkyl, or C 1 -C 6 alkoxy;
  • R1 and R2 can be linked together with the parent moiety to form a 5-6 membered ring optionally substituted with R9 ;
  • R and R can be linked together with the parent moiety to form a 5-6 membered oxygen-containing heterocycle optionally substituted with R ;
  • R 7 and R 8 are independently selected from hydrogen, C 1 -C 6 alkyl, C 1 -C 6 acyl substituted with hydroxy or amino; or R 7 and R 8 may be combined with the N atom to which they are attached taken together to form a 5-6 membered nitrogen-containing heterocycle optionally substituted by R;
  • R 9 is independently selected from halogen, hydroxy, nitro, -NR 7 R 8 , C 1 -C 6 alkyl, C 1 -C 6 alkoxy, optionally C 1 -C 6 alkane substituted piperidinyl;
  • R 13 represents a carboxy-substituted C 1 -C 6 alkyl group.
  • R 2 represents hydrogen, C 3 -C 4 alkenyl, nitro, amino, or C 1 -C 4 alkyl substituted with -N(C 1 -C 4 alkyl) 2 .
  • R 3 represents hydrogen, halogen, hydroxyl, or
  • R4 represents hydrogen or halogen.
  • R1 and R2 are joined together to form the group shown below in A moiety represents a bond to the parent group.
  • R3 and R4 are joined together to form the group shown below in A moiety represents a bond to the parent group.
  • the compound represented by formula (I) is a compound selected from the group consisting of:
  • the compound represented by formula (I) is gimatecan or gimatecan:
  • the number of linker-drug linkages (drug loading (DAR, drug load ratio)) connected to one molecule of the antibody affects the effectiveness and safety of the conjugate.
  • the production of antibody-drug conjugates can be carried out by specifying reaction conditions such as the amount of raw materials and reagents used for the reaction in order to make the number of linker-drug linkages constant, but it is different from the chemical reaction of low-molecular-weight compounds. , usually obtained as a mixture of linked different numbers of drugs. Therefore, in the present invention, the number of linker-drug linkages linked to the antibody per molecule is represented by the average value, that is, the average number of drug linkages.
  • the The number of connections refers to the average.
  • the number of antitumor compounds linked to the antibody molecule can be controlled, and as the average number of drug linkages per antibody, about 1 to 10 antitumor compounds can be linked, preferably 2 to 8, more preferably 4 to 8, More preferably, it is 6-8. It should be noted that those skilled in the art can design the reaction of linking the necessary number of drugs on the antibody according to the description of the examples of the present application, and can obtain the antibody with the number of links of the anti-tumor compound controlled.
  • the number of free sulfhydryl groups attached to each antibody molecule is not actually measured.
  • the average number m of sulfhydryl groups attached to each antibody molecule is 6- 8.
  • the present invention provides a compound represented by formula (I),
  • R 11 is a C 1 -C 6 carboxyalkyl group
  • R 12 is a C 2 -C 6 cyanoalkynyl group
  • 1-2 C atoms in X, Y, X' and Y' are substituted by N atoms; preferably Typically, R 11 is C 1 -C 3 carboxyalkyl, and R 12 is C 2 -C 3 cyanoalkynyl.
  • X, Y, X' and Y' have one and only one C atom replaced by a N atom;
  • X, Y, X' and Y' have 2 C atoms substituted with N atoms, and X, Y have only 1 C atom substituted with N atoms, and X' , Y' has and only one C atom is replaced by N atom;
  • the structures of the compounds are shown below, designated CN-A, CN-B, CN-C and CN-D, respectively
  • the compound of formula (I) is used as the linking unit in the antibody-drug conjugate through a thioether formed by the alkynyl carbon of R12 and the disulfide moiety present in the hinge portion of the antibody bond to the antibody, that is, the alkynyl group of R 12 reacts with the disulfide bond of the hinge part of the antibody so that the alkynyl carbon of R 12 is connected to the reduced sulfhydryl group (-SH-) of the hinge part of the antibody, through the carboxyl group of R 11 It forms an amide bond with the amino group present at the end of the linker to connect with other linking units in the linker.
  • a preparation method of the compound of formula (I) (1) Effect of SM-A (5-bromopyridine-2-carboxylic acid) on Boc 2 O, DMAP, t-BuOH (2) A-1 reacted with Pd(PPh 3 ) 2 Cl 2 , triethylamine and propyn-3-ol in tetrahydrofuran at 70 °C for 12 hours to obtain the compound A-2(3) A-2 reacted with TEMPO, PhI(OAC) 2 , NH 4 OAC in a solution of CH 3 CN/H 2 O 9:1 at room temperature for 12 hours to obtain compound A-3; (4) A-3 generates compound CN-A under the action of TFA/DCM.
  • CN-B and CN-C can be prepared by replacing SM-A with SM-B (6-bromonicotinic acid) or SM-C (5-bromopyrimidine-2-carboxylic acid).
  • the linker of the present invention has the structure shown in the following formula (III):
  • L 2 represents valine residue, guanidine residue, phenylalanine residue, lysine residue, D-valine residue, glycine residue, alanine residue, aspartic acid Residues;
  • R 10 is independently selected from hydrogen, C 1 -C 6 alkyl optionally substituted with 1 or 2 hydroxyl groups, n 3 represents an integer of 1 to 4, and n 4 represents an integer of 1 to 4;
  • Aryl represents a C 6 -C 10 aryl optionally substituted by R 9 ;
  • W is a single key or Wherein, position 1 means connecting with -NR 10 -, and position 2 means connecting with (CH 2 CH 2 -O-)n 1 -;
  • Q 1 is linked through the formation of an amide bond between the carboxyl group of R 11 and the left-terminal amino group -NR 10 - in the formula of L 1 .
  • n 1 represents 6, 7, 8, 9, 10, 11 or 12 and n 2 represents 1 or 2.
  • R 10 represents hydrogen or C 1 -C 4 alkyl.
  • L 2 represents a single amino acid residue, such as valine residue, guanidine residue, phenylalanine residue, lysine residue, D-valine residue, glycine residue, alanine residue base, aspartic acid residue.
  • L2 represents a lysine residue
  • the compound represented by formula (II) uses the oxygen in the hydroxyl group at position 19 as the linking site, or when R3 or R4 is a hydroxyl group, the oxygen in the hydroxyl group of R3 or R4 is used as the linking site .
  • R 3 or R 4 when R 3 or R 4 is a hydroxyl group, the compound represented by formula (II) is connected to the above-mentioned La represented by the oxygen in the hydroxyl group of R 3 or R 4 as a linking site.
  • La represents a structure derived from 4-aminobenzyl alcohol.
  • the C-terminus of the amino acid represented by L 2 is attached to the terminal amino group of the group represented by L a .
  • the linker represented by formula (III) is a group selected from the group consisting of:
  • the linker of the present invention has the structure shown in the following formula (VII):
  • L 3 represents -ZW-(CH 2 CH 2 -O)n 5 -W'- or a single bond
  • n 5 represents an integer of 1 to 8
  • W and W' represent Or a single bond
  • the position 1 of W indicates that it is connected to Z
  • the position 2 indicates that it is connected to (CH 2 CH 2 -O-)n 5 -
  • the position 1 of W' indicates that it is connected to (CH 2 CH 2 -O-)n 5 - is connected
  • position 2 indicates that it is connected to -CH 2- of L 4
  • W and W' are not at the same time
  • Cyclo represents a cyclohexane group
  • LP represents a peptide residue consisting of 2 to 7 amino acids
  • Aryl represents a C 6 -C 10 aryl group optionally substituted by R 9 ,
  • n 7 represents an integer of 1-4, n 8 represents an integer of 1-4,
  • R 10 is independently selected from hydrogen, C 1 -C 6 alkyl optionally substituted with 1 or 2 hydroxy groups,
  • Q 2 represents -(succinimide-3-yl-N)-, with the following formula:
  • the alkynyl carbon of R 12 and the disulfide bond of the antibody hinge part form a thioether bond to connect, that is, the alkynyl group of R 12 reacts with the disulfide bond of the antibody hinge part to connect the alkynyl carbon of R 12 It is linked to L 3 by forming an amide bond between the carboxyl group of R 11 and -NR 10 - on the reduced sulfhydryl group (-SH-) of the antibody hinge.
  • LP represents a peptide residue consisting of 2 to 7 amino acids. That is, it consists of oligopeptide residues in which 2-7 amino acids are linked by peptide bonds.
  • the amino acid constituting LP is not particularly limited, and is, for example, an L- or D-amino acid, preferably an L -amino acid.
  • amino acids with structures such as ⁇ -alanine, ⁇ -aminocaproic acid, and ⁇ -aminobutyric acid may be used, and, for example, non-natural amino acids such as N-methylated amino acids may be used. type of amino acid.
  • the amino acid sequence of the LP moiety is not particularly limited, and the constituent amino acids include phenylalanine (Phe; F), tyrosine (Tyr; Y), leucine (Leu; L ), and glycine (Gly). ; G), alanine (Ala; A), valine (Val; V), lysine (Lys; K), citrulline (Cit; C), serine (Ser; S), glutamic acid (Glu; E), aspartic acid (Asp; D) and the like.
  • phenylalanine, glycine, valine, lysine, citrulline, serine, glutamic acid, and aspartic acid are preferably used.
  • the pattern of drug release can be controlled according to the type of amino acid.
  • the number of amino acids can be 2-7.
  • the peptide residue represented by LP is linked to the L2 moiety at the N-terminus and to the La moiety at the C-terminus.
  • LP is a peptide residue consisting of 2-5 amino acids.
  • LP is a peptide residue selected from the group consisting of:
  • the compound represented by formula (II) uses the oxygen in the hydroxyl group at position 19 as the linking site, or when R3 or R4 is a hydroxyl group, the oxygen in the hydroxyl group of R3 or R4 is used as the linking site .
  • R 3 or R 4 when R 3 or R 4 is a hydroxyl group, the compound represented by formula (II) is connected to the above-mentioned L represented by the oxygen in the hydroxyl group of R 3 or R 4 as a linking site.
  • L represented by the oxygen in the hydroxyl group of R 3 or R 4 as a linking site.
  • L b represents a structure derived from 4-aminobenzyl alcohol.
  • the disabled C-terminus of the peptide represented by LP is attached to the group represented by Lb , more specifically, the C-terminus is attached to the terminal amino group in the group represented by Lb .
  • the linker represented by formula (VII) is a group selected from the group consisting of:
  • linker intermediate compound of the present invention is shown in the following formulas (XI) and (XII):
  • R 1 , R 2 , R 3 , R 4 , Q 1 , Q′ 2 L 1 , L 2 , L 3 , L 4 , L P , La and L b are as described in the specification of the present invention.
  • the linker-drug intermediate compound of the present invention is a compound represented by the formula (II) represented by T, and the intermediate compound is formed by connecting the compound (T) and the linker represented by the following formula (V):
  • R 1 , R 2 , R 3 and R 4 are as described in the description of the present invention.
  • the linker-drug intermediate compound is a compound selected from the group consisting of,
  • the linker-drug intermediate compound of the present invention is that T represents the compound represented by the formula (II), and the intermediate compound is formed by connecting the compound (T) and the linker represented by the following formula (IX):
  • R 1 , R 2 , R 3 and R 4 are as described in the description of the present invention.
  • Q' 2 represents (maleimide-N)- or Q 1 -NR 10 -,
  • Q 1 -NR 10 - represented by Q 2 , Q 1 is connected to L 3 through the carboxyl group of R 11 and -NR 10 - forming an amide bond;
  • Q 1 , R 10 , L 3 , L 4 , L P and L b are as described in the specification of the present invention.
  • the linker-drug intermediate compound is a compound selected from the group consisting of,
  • gemnotecan when the compound represented by the formula (II) is gemnotecan, when L a and L b represent -NR 10 -(CH 2 )n 3 -, gemnotecan as an antitumor compound and the formula (III) ) and (VII) shown in the linker structure are connected by the structure -N-CH 2 -O-, and gemitecan has two connection sites at the 19th position and the 10th position, which provides more choices for ADC.
  • the antibody-drug conjugate (IV) of the present invention is formed by connecting the compound (T) and the antibody (AB) via a linker represented by the following formula (III):
  • R 1 , R 2 , R 3 and R 4 are as described in the specification of the present invention; the definitions of Q 1 , L 1 , L 2 and La are as described in the specification of the present invention; AB represents an antibody.
  • the antibody-drug conjugate (VIII) of the present invention is formed by connecting the compound (T) and the antibody (AB) via a linker represented by the following formula (VII):
  • R 1 , R 2 , R 3 and R 4 are as described in the description of the present invention; the definitions of Q 2 , L 3 , L 4 , LP and L b are as described in the description of the present invention; AB represents an antibody.
  • an antibody-drug conjugate in which an antibody and a linker structure are linked via a thioether can be produced, for example, by the following method.
  • AB-SH represents an antibody carrying a sulfhydryl group
  • AB represents an antibody
  • the compounds represented by formulas (VI) and (X) are the above-mentioned linker-drug intermediate compounds of the present invention.
  • the compound shown in VIII) is the antibody-drug conjugate of the present invention.
  • the compounds represented by formulae (IV) and (VIII) are described in the form of a structure in which one structural moiety from the drug to the end of the linker is linked to one antibody, but in fact, relative to In many cases, a plurality of these structural moieties are linked to one antibody molecule.
  • 2 to 8, preferably 4 to 8, more preferably 6 to 8 linker-drug intermediate compounds are linked to one antibody molecule.
  • the average number of linker-drugs linked to each molecule of antibody is expressed as the average number of drug linkages.
  • the antibody-drug conjugate represented by formula (IV) can be produced by reacting the above-mentioned linker-drug intermediate compound of the present invention with the antibody AB-SH having a thiol group.
  • Antibodies having thiol groups can be obtained by methods known to those skilled in the art (Hermanson, G.T, Bioconjugate Techniques, pp.56-136, pp.456-493, Academic Press (1996)). For example, the following methods are exemplified: allowing Traut to react with the amino group of the reagent and the antibody; and allowing N-succinimidyl S-acetylthioalkanoate to react with the amino group of the antibody After reacting with hydroxylamine; after reacting N-succinimidyl 3-(pyridyldithio)propionate, reacting reducing agent; reacting dithiothreitol, 2-mercaptoethanol, tri( A reducing agent such as 2-carboxyethyl) phosphine hydrochloride (TCEP) acts on the antibody to reduce the disulfide bond in the hinge portion of the antibody to generate a sulfhydryl group; etc., but not limited to these methods.
  • TCEP partial or A sulfhydryl group-carrying antibody
  • a chelating agent ethylenediaminetetraacetic acid (EDTA), diethylenetriaminepentaacetic acid (DTPA), etc. are mentioned, for example. They can be used in concentrations of 1 mM to 20 mM.
  • EDTA ethylenediaminetetraacetic acid
  • DTPA diethylenetriaminepentaacetic acid
  • the buffer solution sodium phosphate, sodium borate, sodium acetate solution or the like can be used.
  • an antibody AB-SH partially or completely reduced to have a thiol group can be obtained.
  • 2-20 molar equivalents of the compounds represented by the formulae (VI) and (X) can be used for each antibody AB-SH having a thiol group to produce an antibody-drug in which 2 to 8 drugs are linked to one antibody.
  • Conjugates (IV), (VIII). Specifically, to the buffer solution containing the antibody AB-SH having a thiol group, a solution in which the compounds represented by the formulae (VI) and (X) are dissolved is added and allowed to react.
  • the buffer solution sodium acetate solution, sodium phosphate, sodium borate, or the like can be used as the buffer solution.
  • the pH during the reaction is 5 to 9, and the reaction is more preferably near pH 7.
  • the compound (2) that is, the compound represented by the formula (II)
  • the organic solvent solution in which the compounds represented by formula (VI) and (X) are dissolved can be added to the buffer solution containing the thiol group-containing antibody AB-SH at 1 to 20% v/v, and the reaction can be carried out.
  • the reaction temperature is 0 to 37°C, more preferably 10 to 25°C, and the reaction time is 0.5 to 2 hours.
  • the reaction can be terminated by inactivating the reactivity of the unreacted compounds represented by the formulae (VI) and (X) with a thiol-containing reagent.
  • Thiol-containing reagents are, for example, cysteine or N-acetyl-L-cysteine (NAC). More specifically, 1 to 2 molar equivalents of NAC are added to the compounds represented by the formulae (VI) and (X) to be used, and the reaction is completed by incubating at room temperature for 10 to 30 minutes.
  • the container put the antibody or antibody-drug conjugate solution, use a centrifuge for centrifugation (for example, centrifuge at 2000G-3800G for 5-20 minutes), and concentrate the antibody or antibody-drug conjugate solution.
  • a centrifuge for centrifugation for example, centrifuge at 2000G-3800G for 5-20 minutes
  • Antibody concentration was measured using a UV analyzer according to the manufacturer's instructions.
  • Phosphate buffer eg, 10 mM, pH 6.0
  • sodium chloride eg, 137 mM
  • EDTA ethylenediaminetetraacetic acid
  • PBS6.0/EDTA ethylenediaminetetraacetic acid
  • Phosphate buffer eg, 50 mM, pH 6.5, also referred to herein as PBS6.5/EDTA
  • sodium chloride eg, 50 mM
  • EDTA eg, 2 mM
  • PBS6.5/EDTA Phosphate buffer
  • the NAP-25 column using Sephadex G-25 vector was equilibrated.
  • One of the NAP-25 columns was packed with 2.5 mL of aqueous antibody solution, and then a fraction (3.5 mL) eluted with PBS6.5/EDTA 3.5 mL was obtained by separation. This fraction was concentrated by common procedure A, and the antibody concentration was measured by common procedure B, and then the antibody concentration was adjusted to 5 mg/mL using PBS6.5/EDTA.
  • phosphate buffer eg, PBS7.4
  • sodium phosphate buffer eg, 10 mM, pH 6.0; also referred to in this specification as PBS6.0
  • sodium chloride eg, 137 mM
  • the NAP-25 column is equilibrated with any of the acetic acid buffers (eg, 10 mM, pH 5.5; also referred to herein as ABS) containing sorbitol (eg, 5%).
  • An antibody-drug conjugate reaction aqueous solution for example, about 1.5 mL
  • the antibody fraction is obtained by separation by eluting with an amount of buffer specified by the manufacturer.
  • phosphate buffer eg, PBS7.4
  • sodium phosphate buffer eg, 10 mM, pH 6.0; also referred to in this specification as PBS6.0
  • sodium chloride eg, 137 mM
  • An AKTA column filler: sephadex G 25
  • any of the acetate buffers eg, 10 mM, pH 5.5; also referred to as ABS in this specification
  • sorbitol eg, 5%
  • the injector is loaded with an antibody-drug conjugate reaction aqueous solution (eg, about 2 mL), eluted with an amount of buffer specified by the manufacturer, and the antibody fractions are obtained by separation.
  • the unconnected drug linker low molecular compound (tris(2-carboxyethyl)phosphine hydrochloride (TCEP), N-acetyl-L-cysteine (NAC), dimethyl sulfoxide) antibody-drug conjugates.
  • TCEP tris(2-carboxyethyl)phosphine hydrochloride
  • NAC N-acetyl-L-cysteine
  • dimethyl sulfoxide dimethyl sulfoxide
  • the preparation method of the linker-drug intermediate compound comprises the following steps:
  • CN-CMTC-1 N-[(9H-fluoren-9-ylmethoxy)carbonyl]-N'-[(4-methoxyphenyl)diphenylmethyl]-L-lysine
  • EEDQ 2-ethoxy-1-ethoxycarbonyl-1,2-dihydroquinoline
  • the target product with a purity of more than 95% is obtained after preparation and purification.
  • Gemitecan-Boc (or SN-38-Boc, or Gematecan) is reacted with triphosgene, DMAP, and dichloromethane to form the formyl chloride compound of Gemitecan, and then CN-CMTC-5 is added to react After about 5 mins, after quenching with methanol, the column was shorted to obtain the crude product of CN-CMTC-6 compound. After Pre-HPLC purification, pure CN-CMTC-6 was obtained, which was then treated with TFA/DCM for deprotection to obtain CN-CMTC-7, which was then subjected to Click reaction with CN-A, B, C, and D to obtain the compound CN-CMTC-8. , Compound 8 was treated with TFA/DCM to obtain the final product CN-CMTC (see the figure below).
  • CMTC represents gemnotecan.
  • gemnotecan can be replaced by SN-38 Or camptothecin derivatives such as gimatecan (GMTC).
  • the preparation method of the linker-drug intermediate compound comprises the following steps:
  • CN-CMTC-7 and SM-1 were added to DMSO/H2O solution, and then CuBr was added, and then stirred at room temperature for about 30 mins. HPLC detected that the reaction was complete, resulting in SMCC-PEG8-Lys(MMt)-PABC-CMTC, Pre-HPLC After purification, TFA/DCM was added for deprotection to obtain the target product SMCC-PEG8-Lys-PABC-CMTC.
  • the preparation method of the linker-drug intermediate compound comprises the following steps:
  • the preparation method of the linker-drug intermediate compound comprises the following steps:
  • Boc-GGFG and PABOH, dichloromethane and methanol were used as solvents, and stirred overnight at room temperature to generate Boc-GGFG-PABOH;
  • the preparation method of the linker-drug intermediate compound comprises the following steps:
  • DIEA is a base to obtain the compound N 3 -PEGn-GGFG, which is purified by Pre-HPLC and then condensed with N-Boc-N-methylethylenediamine.
  • the condensing agent is HATU, pyridine is used as a base, and DMF is used as a solvent to obtain the target product;
  • CMTC germane-maleimide
  • GMTC gematecan
  • the antibody (AB) is a full-length antibody or an antigen-binding fragment thereof, or a bispecific antibody or an antigen-binding fragment thereof.
  • the antibody is selected from the group consisting of Her-2 antibody, anti-Trop-2 antibody, EGFR antibody, B7-H3 antibody, PD-1 antibody, PD-L1 antibody, HER-3, HER- 4 antibody, CD20, CD30 antibody, CD19 antibody, CD33 antibody.
  • TROP-2 antibody belongs to the TACSTD family and is a cell surface glycoprotein encoded and expressed by the TACSTD2 gene, also known as tumor-associated calcium signal transducer 2 (TACSTD2), epidermal glycoprotein 1 (EGP-1), and gastrointestinal tumor-associated antigen. (GA733-1), Surface Marker 1 (M1S1). TROP-2 is overexpressed in various malignant tumors and is an oncogene related to the occurrence, invasion and metastasis of malignant tumors.
  • TACSTD2 tumor-associated calcium signal transducer 2
  • EGP-1 epidermal glycoprotein 1
  • M1S1 Surface Marker 1
  • TROP-2 of the natural sequence in the present invention can be isolated from nature, and can also be prepared by recombinant DNA technology, chemical synthesis method or a combination thereof.
  • the antibody used in the present invention is preferably an anti-human TROP-2 antibody.
  • the CDR1, CDR2 and/or CDR3 of the heavy and light chains in the anti-human TROP-2 antibody are CDR1, CDR2 and/or CDR3 of the heavy and light chains of the RS7 mAb, respectively.
  • the anti-human TROP-2 antibody may be a humanized antibody or a fully human antibody.
  • the complementarity determining regions (CDRs) of the light chain variable region of the anti-Trop-2 antibody include CDR1 consisting of the amino acid sequence of KASQDVSIAVA, CDR2 consisting of the amino acid sequence of SASYRYT, and consisting of the amino acid sequence of QQHYITPLT CDR3 composed of;
  • the CDRs of the heavy chain variable region include CDR1 composed of the NYGMN amino acid sequence, CDR2 composed of the WINTYTGEPTYTDDFKG amino acid sequence, and CDR3 composed of the GGFGSSYWYFDV amino acid sequence;
  • the light chain of the anti-Trop-2 antibody and the amino acid sequences of the heavy chain are shown in SEQ ID NO: 1 and SEQ ID NO: 2 respectively;
  • the coding nucleotide sequences of the light chain and heavy chain of the anti-Trop-2 antibody are respectively shown in SEQ ID NO: 3 and SEQ ID NO: shown in 4;
  • Her-2 antibodies or antigen-binding fragments thereof are Her-2 antibodies or antigen-binding fragments thereof, including bispecific antibodies and antibody functional derivatives.
  • Her-2 is also known as human epidermal growth factor receptor-2 (human epidermal growth factor receptor 2), or receptor tyrosine protein kinase erbB-2, also known as CD340 (cluster of differentiation 340), proto-oncogene Neu, Erbb2 (rodent) or ERBB2 (human), is a protein encoded by the ERBB2 gene in humans.
  • Her-2 overexpression has been shown to play an important role in the development and progression of certain aggressive types of breast cancer.
  • Her-2 overexpression occurs in approximately 15-30% of breast cancers. In recent years, this protein has become an important biomarker and therapeutic target in about 30% of breast cancer patients.
  • Her-2 overexpression also occurs in ovarian cancer, intestinal gastric cancer, and invasive forms of uterine cancer such as serous endometrial cancer.
  • the natural sequence Her-2 in the present invention can be isolated from nature, and can also be prepared by recombinant DNA technology, chemical synthesis method or a combination thereof.
  • the antibody used in the present invention is preferably an anti-human Her-2 antibody.
  • the CDR1, CDR2 and/or CDR3 of the heavy and light chains in the anti-human Her-2 antibody are CDR1, CDR2 and/or CDR3 of the heavy and light chains of the RS7 mAb, respectively.
  • the anti-human Her-2 antibody may be a humanized antibody or a fully human antibody.
  • the Her-2 antibody is the trastuzumab antibody described in US5821337, and the complementarity determining region (CDR) of its light chain variable region includes CDR1 consisting of the amino acid sequence of RASQDVNTAVA; consisting of the amino acid sequence of SASFLYS and CDR3 composed of the QQHYTTPPT amino acid sequence, and the CDRs of its heavy chain variable region include CDR1 composed of the DTYIH amino acid sequence; CDR2 composed of the RIYPTNGYTRY amino acid sequence; and CDR3 composed of the WGGDGFYAMDY amino acid sequence.
  • the light chain sequence and heavy chain sequence of the trastuzumab antibody are shown in SEQ ID NO: 5 and SEQ ID NO: 6, respectively. Also included are those antibodies that retain Her-2 binding activity after conservative amino acid substitutions to the above-mentioned antibodies.
  • the antibody-drug conjugates of the present invention can be preferably administered to mammals, more preferably humans.
  • the substance to be used in the pharmaceutical composition containing the antibody-drug conjugate of the present invention can be appropriately selected from formulation additives or others commonly used in the art in consideration of the dose and concentration to be administered.
  • the antibody-drug conjugates of the present invention may be administered in the form of a pharmaceutical composition or pharmaceutical formulation containing one or more pharmaceutically suitable ingredients.
  • the above-mentioned pharmaceutical compositions or pharmaceutical preparations may typically contain one or more pharmaceutically acceptable carriers (such as sterile liquids (such as water and oils (including petroleum, animal, vegetable, or synthetic origin) Oil (such as peanut oil, soybean oil, mineral oil, sesame oil, etc.))).
  • sterile liquids such as water and oils (including petroleum, animal, vegetable, or synthetic origin) Oil (such as peanut oil, soybean oil, mineral oil, sesame oil, etc.)
  • Oil such as peanut oil, soybean oil, mineral oil, sesame oil, etc.
  • water is a more representative carrier.
  • saline solution as well as aqueous dextrose and glycerol solutions are also Can be used as liquid carrier, especially can be used for injection solution.Appropriate pharmaceutical excipients are known in this field.As required, the above-mentioned composition can also contain a trace amount of wetting agent or emulsifying agent, or pH buffering agent Examples of suitable pharmaceutical carriers are described in "Examples of W. Martin Carriers Parmaceutical Sciences" by E.W. Martin. The prescription corresponds to the mode of administration.
  • the introduction method includes, but is not limited to, intradermal, intramuscular, intraperitoneal, intravenous, and subcutaneous routes. Administration, for example, can be by infusion or bolus injection. In certain preferred embodiments, the administration of the antibody-drug conjugates described above is performed by infusion. Parenteral administration is the preferred route of administration.
  • the above-mentioned pharmaceutical composition is formulated into a pharmaceutical composition for intravenous administration to humans, and is formulated according to conventional procedures.
  • compositions for intravenous administration are solutions in sterile isotonic aqueous buffer.
  • the above-mentioned pharmaceutical composition may further contain a solubilizer and a local anesthetic (eg, lidocaine) for relieving pain at the injection site, as required.
  • a solubilizer eg, lidocaine
  • the above ingredients may be supplied either as a dry freeze-dried powder or anhydrous concentrate in a sealed container (eg, an ampule or sachet, etc., which indicates the amount of active agent), respectively, Or mixed together in unit dosage form.
  • the above-mentioned medicine When the above-mentioned medicine is intended to be administered by infusion, for example, the above-mentioned medicine may be put into an infusion bottle containing sterilized pharmaceutical-grade water or saline.
  • the above-mentioned drugs When the above-mentioned drugs are administered by injection, ampoules of sterile water for injection or saline may be provided so that, for example, the above-mentioned components are mixed before administration.
  • the pharmaceutical composition or pharmaceutical preparation of the present invention may be a pharmaceutical composition or pharmaceutical preparation containing only the antibody-drug conjugate of the present invention, or may be a pharmaceutical composition or pharmaceutical preparation containing the antibody-drug conjugate and at least one other cancer therapeutic agent pharmaceutical composition.
  • the antibody-drug conjugate of the present invention can also be administered together with other cancer therapeutic agents, whereby the anticancer effect can be enhanced.
  • Other anticancer agents used for this purpose may be administered to the individual simultaneously, separately or sequentially with the antibody-drug conjugate, or may be administered at varying intervals.
  • cancer therapeutic agents include albumin-bound paclitaxel, carboplatin, cisplatin, gemcitabine, irinotecan (CPT-11), paclitaxel, pemetrexed, sorafenib, vinblastine, or international publications.
  • Such pharmaceutical compositions can be formulated in the form of freeze-dried preparations or liquid preparations as preparations having a selected composition and necessary purity.
  • a preparation When a preparation is formed as a freeze-dried preparation, it may be a preparation containing appropriate preparation additives available in the art.
  • a preparation can be formed as a liquid preparation containing various preparation additives usable in the art.
  • composition and concentration of the pharmaceutical composition vary depending on the administration method, but the affinity of the antibody-drug conjugate contained in the pharmaceutical composition of the present invention for the antigen of the antibody-drug conjugate, that is, the affinity for the antigen, varies. Considering the dissociation constant (Kd value), when the affinity is higher (the Kd value is lower), the drug effect can be exhibited even with a small dose. Therefore, when determining the administration amount of the antibody-drug conjugate, the administration amount can also be set based on the state of the affinity of the antibody-drug conjugate with the antigen.
  • the antibody-drug conjugate of the present invention When the antibody-drug conjugate of the present invention is administered to a human, for example, it may be administered once at about 0.001 to 100 mg/kg, or may be administered multiple times at intervals of 1 to 180 days.
  • the antibody-drug conjugates, pharmaceutical compositions, and pharmaceutical preparations of the present invention can be used to prevent and/or treat tumors or cancers.
  • the tumor or cancer targeted for prevention and/or treatment may be any cancer cell that expresses a protein recognized by the antibody in the antibody-drug conjugate.
  • the tumor or cancer is selected from breast cancer, colorectal cancer, lung cancer, pancreatic cancer, ovarian cancer, prostate cancer, cervical cancer, kidney cancer, urethral cancer, glioblastoma, melanoma tumor, liver cancer, bladder cancer, gastric cancer, esophageal cancer; preferably, the cancer is carcinoma in situ or metastatic cancer; preferably, the breast cancer is Sanyinjiao breast cancer.
  • a prophylactically or therapeutically effective amount of an antibody-drug conjugate, pharmaceutical composition or pharmaceutical formulation of the invention is administered to a subject in need thereof for inhibiting the growth, proliferation or migrate.
  • kits for inhibiting the growth, proliferation or migration of cancer cells comprising the antibody-drug conjugate, pharmaceutical composition or pharmaceutical formulation of the present invention.
  • the antibody-drug conjugate of the present invention has fast and efficient tumor cell killing activity, and at the same time has good biocompatibility, low immunogenicity, biological safety and stability.
  • the linker structure of the present invention such as formula (II) has the following advantages: (1) the linker structure of the present invention has suitable molecular weight and hydrophobicity, and has a higher drug loading (DAR, drug to antibody ratio)>7; (2) (3) the present invention The release of the linker structure is especially suitable for the preferred toxins gimatecan, jimitecan, and more preferably jimitecan, which are matched with their cytotoxicity, pharmacokinetics, and tumor inhibitory properties, and the linker is self-cleaving.
  • the speed is faster, which is conducive to the rapid release of toxin molecules, and greatly enhances the drug efficacy; (4)
  • the size, physicochemical properties and coupling sites of the joint designed by the present invention will not affect the physiological activity of the antibody; (5)
  • the present invention The synthesis method of the linker compound is simple and suitable for industrial production.
  • the anti-tumor compound of the present invention selects gimatecan and gimatecan.
  • the toxicity of gimatecan is about 10 times that of SN-38, which is comparable to that of ixatecan, but its safety is much better than that of ixatecan.
  • Satecan which is available alone as an oral preparation.
  • gemnotecan belongs to the camptothecin toxoid, which is substituted at the 10-position hydroxyl group and the 9-position allyl group, and has good antitumor activity.
  • the present invention connects gimatecan and gimatecan with a certain hydrophilic linker by optimizing the design of the linker, so as to increase the targeted drug delivery and solve the problem of water solubility of the toxin molecule. problem.
  • the linker molecule of formula (I) designed by the present invention has a more stable connection with the antibody, reduces the possibility of falling off at the non-target position, improves the safety, and improves the safety. Can achieve high drug loading.
  • the linker gimatecan has a strong ability to penetrate cell membranes, allowing them to kill nearby cancer cells after killing the cancer cells that engulfed the ADC.
  • trasstuzumab antibody (Herceptin antibody) was purchased from Genentech Inc.
  • ADC-137 was only used as a control, and was purchased from Changzhou Chenhong Biotechnology Co., Ltd., CAS#: 1279680-68-0.
  • hRS7 antibody was produced in CHO cells.
  • the expression vectors containing the hRS7 antibody gene were constructed by conventional molecular biology methods, and the nucleotide sequences of the light chain and heavy chain of the hRS7 antibody were shown in SEQ ID NO: 3 and SEQ ID NO: 4, respectively. Insert the above two sequences into the same expression vector, extract and prepare transfection plasmids in large quantities, and transfect them into CHO-K1 cells (ATCC CCL-61).
  • the specific transfection and antibody preparation processes are as follows:
  • Cell culture CHO-K1 cells were grown in suspension in ActiPro (GE HyClone) medium at 37°C, 7% CO 2 , 140 rpm, and 90% relative humidity;
  • the highly expressed cell fluid cultured in shake flasks was collected and purified by protein A affinity (GE, Mab Select SuRe) and ion exchange (GE, Capto S).
  • the molecular weight and purity of the purified antibodies were analyzed by SDS-PAGE and SEC-HPLC.
  • SDS-PAGE showed that the molecular weight of the prepared hRS7 was in line with expectations, and the purity of the antibody was 99.1% measured by SEC-HPLC.
  • tert-butyl 5-(cyanoethynyl)picolinate A-3 (1.2 g, 5.3 mmol) in DCM (80 mL) was added TFA (80 mL). The mixture was stirred at room temperature for 3 hours. The reaction was monitored by LCMS. The solution was concentrated under reduced pressure. The residue was triturated with ether (150 ml). The mixture was filtered and the filter cake was washed with ether (50 ml). The filter cake was dried in vacuo to give tert-butyl 5-(cyanoethynyl)picolinate target A (600 mg, 66% yield) as a white solid.
  • tert-Butyl 6-bromonicotinate B-1 (5 g, 19.46 mmol) and 2-yn-1-propanol (3.26 g, 25.3 mmol) were added to the flask followed by Pd( PPh3 ) 2Cl2 ( 0.98 g, 1.4 mmol) and CuI (0.45 g, 2.3 mmol). After purging the flask with nitrogen 3 times to remove oxygen. DMF (64 mL) and TEA (10 mL) were added via syringe. The reaction was stirred at 80°C overnight and monitored by LCMS. After the reaction was consumed, the reaction was quenched with saturated ammonium chloride (300 mL).
  • the hRS7 antibody of Example 1 was first reduced in 5 mg/mL pH 7.2 PBS/EDTA solution with 6 times the amount of TCEP at room temperature for 2 hours. Next, a 16-fold amount of the compound CN-C-CMTC dissolved in DMSO (final DMSO concentration 10%) was added to the antibody solution. The reaction was stirred at room temperature and protected from light for 3 hours to obtain the conjugated crude product ADC-1.
  • the coupling reaction crude product is detected by SEC, and the SEC chromatographic conditions are as follows:
  • the purified ADC-1 is shown in Figure 1.
  • the samples were concentrated to 5 mg/mL by ultrafiltration and lyophilized for storage.
  • the higher peak represents the antibody part
  • the lower peak represents the camptothecin compound part.
  • the retention time positions of the two peaks are the same, and it can be seen that the two have formed ADC conjugates.
  • the absorbance values of conjugate and naked antibody at 280nm and 363nm were measured by UV spectrophotometer.
  • the concentration of gemnotecan in the conjugate was calculated from the absorbance at 363 nm according to the standard curve.
  • the concentration of antibody in the conjugate was calculated by subtracting the absorbance of gemitecan at 280 from the absorbance at 280 nm.
  • the DAR value was calculated from the ratio of these two concentrations to be 6.5. That is, n is 6.5.
  • CN-C in this embodiment can be replaced by CN-A, CN-B, CN-D; gemitecan (CMTC) can be replaced by SN-38, gematecan (GMTC).
  • CMTC gemitecan
  • GMTC gematecan
  • the hRS7 antibody of Example 1 was first reduced in 5 mg/mL pH 7.2 PBS/EDTA solution with 6 times the amount of TCEP at room temperature for 2 hours. Next, a 16-fold amount of the compound SMCC-PEG2-GGFG-PABC-CMTC dissolved in DMSO (final DMSO concentration 10%) was added to the antibody solution. The reaction was stirred at room temperature and protected from light for 1 hour to obtain the conjugated crude product ADC-2.
  • the detection method is as described in step IV-3 of Example 4.
  • the purified ADC-2 was similar to ADC-1 of Figure 1 .
  • the samples were concentrated to 5 mg/mL by ultrafiltration and lyophilized for storage.
  • DAR was determined as described in Example 4, Step IV-5.
  • the DAR value of ADC-2 is 7.0. That is, n is 7.0.
  • CMTC Gematecan
  • GMTC gematecan
  • the hRS7 antibody of Example 1 was first reduced with 6 times the amount of TCEP in a 5 mg/mL solution of pH 7.2 PBS/EDTA for 2 hours at room temperature. Next, a 16-fold amount of the compound CN-C-PEG 2 -GGFG-PABC-CMTC dissolved in DMSO was added to the antibody solution (final DMSO concentration 10%). The reaction was stirred at room temperature and protected from light for 3 hours to obtain the conjugated crude product ADC-3.
  • the detection method is as described in step IV-3 of Example 4.
  • the purified ADC-3 is similar to ADC-1 of Figure 1 .
  • the samples were concentrated to 5 mg/mL by ultrafiltration and lyophilized for storage.
  • DAR was determined as described in Example 4, Step IV-5.
  • the DAR value of ADC-3 is 6.8. That is, n is 6.8.
  • CN-C in this embodiment can be replaced by CN-A, CN-B, CN-D; gemitecan (CMTC) can be replaced by SN-38, gematecan (GMTC).
  • CMTC gemitecan
  • GMTC gematecan
  • the hRS7 antibody of Example 1 was first reduced in 5 mg/mL pH 7.2 PBS/EDTA solution with 6 times the amount of TCEP at room temperature for 2 hours. Next, a 16-fold amount of the compound MC-GGFG-CMTC dissolved in DMSO (final DMSO concentration 10%) was added to the antibody solution. The reaction was stirred at room temperature and protected from light for 1 hour to obtain the conjugated crude product ADC-4.
  • the detection method is as described in step IV-3 of Example 4.
  • the purified ADC-4 is similar to ADC-1 of Figure 1 .
  • the samples were concentrated to 5 mg/mL by ultrafiltration and lyophilized for storage.
  • DAR was determined as described in Example 4, Step IV-5.
  • the DAR value of ADC-4 is 7.4. That is, n is 7.4.
  • the hRS7 antibody prepared in Example 1 was first reduced in 5 mg/mL pH 6.5 10 mM phosphate solution with 5 times the amount of TCEP at room temperature for 2 hours. Next, an 8-fold amount of Compound A dissolved in DMF (final DMF concentration 15%) was added to the antibody solution. The reaction was stirred at room temperature and protected from light for 1 hour to obtain the coupled crude product ADC-X-a.
  • the Herceptin antibody was first reduced in 6 mg/mL pH 7.2 PBS solution with 9.5 times the amount of TCEP for 30 minutes at room temperature. Next, a 12-fold substance amount of Compound A dissolved in DMF (final DMF concentration 10%) was added to the antibody solution. The reaction was stirred and protected from light at 4°C for 3 hours to obtain the coupled crude product ADC-X-b.
  • the coupling reaction crude product is detected by SEC, and the SEC chromatographic conditions are as follows:
  • the absorbance values of conjugate and naked antibody at 280nm and 363nm were measured by UV spectrophotometer.
  • the concentration of Gimatecan in the conjugate was calculated from the absorbance at 363 nm according to the standard curve.
  • the concentration of antibody in the conjugate was calculated by subtracting the absorbance of gimatecan at 280 from the absorbance at 280 nm. The result is as shown.
  • the DAR value of ADC-X-a was calculated from the ratio of these two concentrations to be 2.7, ie, n was 2.7.
  • Herceptin antibody When the antibody is Herceptin antibody:
  • the DAR value of ADC-X-b was calculated from the ratio of these two concentrations to be 7.3, ie, n was 7.3.
  • the hRS7 antibody prepared in Example 1 was first reduced in 5 mg/mL pH 6.5 10 mM phosphate solution with 7 times the amount of TCEP at 37°C for 2 hours. Next, a 16-fold amount of Compound G dissolved in DMSO (final DMSO concentration 10%) was added to the antibody solution. The reaction was stirred at room temperature and protected from light for 20 minutes to obtain the conjugated crude product ADC-5.
  • the detection method is as described in step I-3 of Example 9.
  • DAR was determined as described in Example 9, steps 1-5.
  • the SEC-HPLC profile of ADC-5 is shown in FIG. 2 .
  • the DAR value of ADC-5 is 7.8. That is, n is 7.8.
  • the DAR value of ADC-6 is 5.5. That is, n is 5.5.
  • ADC-8 has a DAR of 3.3. That is, n is 3.3.
  • the DAR value of ADC-10 is 3.5. That is, n is 3.5.
  • the DAR value of ADC-11 is 7.1. That is, n is 7.1.
  • the DAR value of ADC-12 is 6.2. That is, n is 6.2.

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  • Epidemiology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Organic Chemistry (AREA)
  • Peptides Or Proteins (AREA)
  • Immunology (AREA)
  • Medicinal Preparation (AREA)

Abstract

L'invention concerne un conjugué anticorps-médicament, son procédé de préparation et son utilisation. Le conjugué anticorps-médicament est préparé à l'aide d'un composé de liaison (T) avec un anticorps (AB) par l'intermédiaire d'un lieur représenté par la formule suivante (III) ou (VII), dans laquelle T représente un composé représenté par la formule (II). Le conjugué anticorps-médicament a un temps d'apparition plus rapide, une demi-vie de médicament plus longue, une stabilité plus modérée, une bonne biocompatibilité, une faible immunogénicité et est sans danger, et peut empêcher l'agrégation et présente d'excellents effets antitumoraux. AB-S-Q1-L1-L2-La-T (IV) ; AB-S-Q2-L3-L4-LP-Lb-T(VIII) ; -Q1-L1-L2-La- (III) ; -Q2-L3-L4-LP-Lb- (VII)
PCT/CN2021/123020 2020-10-12 2021-10-11 Conjugué anticorps-médicament et son utilisation WO2022078279A1 (fr)

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US11607459B1 (en) 2020-09-30 2023-03-21 Duality Biologics (Suzhou) Co., Ltd. Anti-tumor compound and preparation method and use thereof
US11814394B2 (en) 2021-11-16 2023-11-14 Genequantum Healthcare (Suzhou) Co., Ltd. Exatecan derivatives, linker-payloads, and conjugates and thereof
US11999748B2 (en) 2021-11-16 2024-06-04 Genequantum Healthcare (Suzhou) Co., Ltd. Exatecan derivatives, linker-payloads, and conjugates and thereof

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WO2016030791A1 (fr) * 2014-08-28 2016-03-03 Pfizer Inc. Lieurs modulant la stabilité destinés à être utilisés avec des conjugués anticorps-médicament
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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US11607459B1 (en) 2020-09-30 2023-03-21 Duality Biologics (Suzhou) Co., Ltd. Anti-tumor compound and preparation method and use thereof
US11685742B2 (en) 2020-09-30 2023-06-27 Duality Biologics (Suzhou) Co., Ltd. Anti-tumor compound and preparation method and use thereof
US11952384B2 (en) 2020-09-30 2024-04-09 Duality Biologics (Suzhou) Co., Ltd. Anti-tumor compound and preparation method and use thereof
US11814394B2 (en) 2021-11-16 2023-11-14 Genequantum Healthcare (Suzhou) Co., Ltd. Exatecan derivatives, linker-payloads, and conjugates and thereof
US11999748B2 (en) 2021-11-16 2024-06-04 Genequantum Healthcare (Suzhou) Co., Ltd. Exatecan derivatives, linker-payloads, and conjugates and thereof

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