Selective release of drugs from internalized conjugates of biologically active compounds

Abstract

The present invention relates to conjugates of biologically active compounds, wherein such conjugates comprise an amino acid sequence comprising a tripeptide that confers selective cleavage by tumor tissue homogenates to release free drug and / or enhanced biodistribution to tumor tissue compared to normal tissue homogenates from the same species, wherein the normal tissue is the site of adverse events associated with administration of a therapeutically effective amount of a comparative conjugate to a human subject in need thereof, wherein the amino acid sequence of the comparative conjugate is a dipeptide known to be selectively cleavable by cathepsin B.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims priority to U.S. Provisional Application No. 62 / 902,888, filed September 19, 2019, the disclosure of which is incorporated herein by reference in its entirety. Background Art

[0003] This invention relates to ligand drug conjugates (LDCs) and compositions thereof, said ligand drug conjugates including antibody drug conjugates (ADCs) and having enhanced selectivity for targeted cells compared to non-targeted cells.

[0004] Traditional ligand-drug conjugates exhibit biological activity in target cells that display a target moiety recognized by the ligand unit of the conjugate. This biological activity is achieved by the conjugate binding to the target moiety and then entering the cell through internalization of the bound conjugate. The superior selectivity for target cells compared to non-target cells is primarily achieved through traditional ligand-drug conjugates because the target moiety is present in greater abundance on target cells compared to non-target normal cells (cells not intended to be acted upon by the conjugate). When the conditional release of cytotoxic conjugates in their free form is influenced by intracellular proteases, the internalization of the bound conjugate is followed by enzymatic processing of the peptide-based linker unit of the conjugate.

[0005] Premature release of cytotoxic compounds from conventional dipeptide-based ligand drug conjugates is reduced by optimizing selectivity for specific lysosomal proteases (which are thought to be upregulated in cancer cells), which could otherwise lead to undesirable side effects. Since the proteases responsible for intracellular processing of conventional ligand drug conjugates are universal across all cells, selectivity for target cells is primarily due to the greater abundance of the target moiety on the cells intended to be acted upon by the conjugate, despite differences in intracellular activity levels of the processing proteases in targeted cancer cells and non-targeted normal cells. However, this method does not account for the potential differences in exposure of released cytotoxic compounds between tumor and normal tissues, differences that the ligand drug conjugates of this invention now utilize.

[0006] Therefore, the dipeptide sequences of conventional ligand-drug conjugates are designed to be selectively acted upon by intracellular proteases (which are upregulated in cancer cells of tumor tissue) but can still be restricted to protease action within normal tissues. This action can occur in the microenvironment of normal tissues or after immune-specific or non-specific uptake by normal tissue cells (leading to on-target or off-target toxicity, respectively). These toxicities are a more pressing issue to address for the targeted delivery of highly cytotoxic compounds. Therefore, it is believed that ligand-drug conjugates with improved peptide sequences offer lower exposure to normal tissues compared to conventional dipeptide-based ligand-drug conjugates, thus reducing exposure to cytotoxic compounds released from the ligand-drug conjugates, while maintaining the efficacy provided by these conventional conjugates and improving treatment tolerability.

[0007] Furthermore, it is believed that ligand drug conjugates with improved peptide sequences are more readily proteased by tumor tissues than by normal tissues, and will also reduce exposure to released cytotoxic compounds, which may contribute to improved treatment tolerance. Identifying these proteasing differences using tissue homogenates should capture those differences driven by the microenvironment of these tissues and / or post-internalization.

[0008] To provide a solution to this problem in the art, this paper discloses ligand-drug conjugates having peptide-based linker units whose sequences result in more selective exposure of tumor-targeted cells to cytotoxic compounds released from the conjugates compared to the exposure of cells in normal tissues to free cytotoxins, thereby improving tolerance to the conjugates while retaining the efficacy of conventional dipeptide-based conjugates in treating cancer in mammalian subjects. This difference in exposure may be due to greater selectivity in the proteolysis of the ligand-drug conjugates compared to the proteolysis of conventional dipeptide-based conjugates, which conjugates have peptide sequence selectivity conferred within tumor tissues. Because altering the peptide sequence can also affect the biochemical properties of the conjugate compound, greater exposure can occur, respectively, from increased biodistribution to tumor tissues rather than normal tissues and / or increased distribution once distributed to normal tissues, which preferentially retains the conjugate compound in tumor tissues and / or preferentially eliminates the conjugate compound from normal tissues. These biodistribution effects may even become determinants of preferential proteolysis, which may be difficult to observe in vivo.

[0009] Therefore, conjugates having peptide sequences that provide enhanced exposure to tumor tissues compared to normal tissues for the release of free cytotoxic compounds should exhibit reduced undesirable toxicity, since the peptide sequences are generally less sensitive to proteolysis in normal tissues or their cells compared to proteolysis in tumors, and / or due to improved pharmacokinetic properties of conjugates containing peptide sequences that are more favorable to tumor tissues than normal tissues.

[0010] Therefore, the ligand-drug conjugates of the present invention exhibit two levels of selectivity for targeted cells relative to non-targeted normal cells: (1) selective entry into target cells, and (2) reduced exposure of normal tissues to the conjugate compound compared to tumor tissues. From the second level of selectivity, the reduced toxicity to normal tissues is expected to decrease adverse events associated with conventional targeted therapy. Summary of the Invention

[0011] One main embodiment of the present invention provides a ligand-drug conjugate composition represented by Formula 1:

[0012] L-[LU-D'] p (1)

[0013] Or its salts, especially pharmaceutically acceptable salts, in which

[0014] L is the ligand unit;

[0015] LU is a connector unit; and

[0016] D' represents one or more drug units (D) in each drug junction portion of the formula -LU-D'; and

[0017] The subscript p is a number from 1 to 12, from 1 to 10, or from 1 to 8, or approximately 4 or approximately 8.

[0018] The ligand unit is an antibody or an antigen-binding fragment of an antibody, which is capable of selectively binding to antigens in tumor tissue to subsequently release the drug unit as a free cytotoxic compound.

[0019] The drug linker portion of formula -LU-D' in each ligand drug conjugate compound of the composition has the structure of formula 1A:

[0020]

[0021] Or its salts, especially pharmaceutically acceptable salts,

[0022] The wavy line indicates covalent attachment with L;

[0023] D is the pharmaceutical unit of the cytotoxic compound;

[0024] L B It is the ligand covalently bound part;

[0025] A is the first optional extension subunit;

[0026] The subscript 'a' is 0 or 1, which means that A does not exist or exists, respectively.

[0027] B is an optional branch unit;

[0028] The subscript b is 0 or 1, which means that B does not exist or exists, respectively.

[0029] L O This is the secondary connector portion, wherein the secondary connector has the following formula:

[0030]

[0031] The wavy line adjacent to Y represents L. O The sites covalently attached to the drug unit, and the wavy line adjacent to A' indicates the sites covalently attached to the remaining portion of the drug connector portion;

[0032] A' is the second optional extended subunit, which becomes a subunit of A in the absence of B.

[0033] The subscript a' is 0 or 1, which means A' does not exist or exists, respectively.

[0034] W is a peptide-cleavable unit, wherein the peptide-cleavable unit is a continuous sequence of up to 12 (e.g., 3-12 or 3-10) amino acids, wherein the sequence comprises a tripeptide that imparts selectivity, providing improved selectivity for tumor tissue relative to normal tissue exposure to free cytotoxic compounds released from the ligand-drug conjugate compound of the comparative ligand-drug conjugate composition, the peptide sequence of the peptide-cleavable unit of the comparative ligand-drug conjugate composition being a dipeptide-valine-citrulline- or -valine-alanine-.

[0035] The tumor tissue and normal tissue described herein belong to rodent species, and the composition of Formula 1 provides the enhanced exposure selectivity, as demonstrated by the following:

[0036] When administered at the same effective amount and dosage regimen previously determined for the comparative ligand-drug conjugate composition, the efficacy of the comparative ligand-drug conjugate composition was maintained in a tumor xenograft model, and

[0037] The results showed that when administered to non-tumor-bearing rodents at the same effective dose and dosage regimen as in the tumor xenograft model, compared to equivalent (e.g., identical) administration of the comparative ligand-drug conjugate composition, the plasma concentration of free cytotoxic compounds released from the ligand-drug conjugate compound of the composition was reduced, and / or normal cells were retained in tissues, wherein the ligand units of both conjugate compositions were replaced by non-binding antibodies.

[0038] The toxicity to cells in human tissues of the same type as normal cells in non-tumor-bearing rodent tissues is at least partially responsible for adverse events in human subjects who were given a therapeutically effective amount of the comparative conjugate composition.

[0039] Y is a suicide-type spacer subunit; and

[0040] The subscript y is 0, 1, or 2, which respectively indicate that Y does not exist or that there is 1 or 2 Y;

[0041] The subscript q is an integer ranging from 1 to 4.

[0042] The condition is that when the index b is 0, the index q is 1, and when the index b is 1, the index q is 2, 3, or 4; and

[0043] The ligand drug conjugate compound of the composition has the structure of Formula 1, wherein the subscript p is replaced by the subscript p', wherein the subscript p' is an integer from 1 to 12, 1 to 10 or 1 to 8, or 4 or 8.

[0044] The relevant key implementation schemes provide drug connector compounds of Formula I:

[0045] LU'-(D') (I)

[0046] Or its salts, particularly pharmaceutically acceptable salts, wherein LU' is capable of providing a covalent bond between L and LU in Formula 1, and is therefore sometimes referred to as a linker unit precursor; and D' represents 1 to 4 drug units, wherein the drug linker compound is further defined by the structure of Formula IA:

[0047]

[0048] Among them L B 'L that can be transformed into Equation 1A B This forms a covalent bond with L in Formula 1, and is therefore sometimes referred to as the ligand covalently bonded precursor moiety, and the remaining variable groups of Formula IA are as defined in Formula 1A.

[0049] In some embodiments, this document provides ligand-drug conjugate compositions represented by Formula 1:

[0050] L-[LU-D']p (1)

[0051] Or its pharmaceutically acceptable salt, wherein

[0052] L is the ligand unit;

[0053] LU stands for connector unit;

[0054] D' represents 1 to 4 drug units (D) in each drug junction portion of the formula -LU-D'; and

[0055] The subscript p is a number from 1 to 12, from 1 to 10, or from 1 to 8, or approximately 4 or approximately 8.

[0056] The ligand unit is derived from an antibody or an antigen-binding fragment of an antibody, which is capable of selectively binding to antigens in tumor tissue to subsequently release the one or more drug units as a free drug.

[0057] The drug linker portion of formula -LU-D' in each ligand drug conjugate compound of the composition has the structure of formula 1A:

[0058]

[0059] Or its salts, especially pharmaceutically acceptable salts,

[0060] The wavy line indicates covalent attachment with L;

[0061] D is the drug unit;

[0062] L B It is the ligand covalently bound part;

[0063] A is the first optional extension subunit;

[0064] The subscript 'a' is 0 or 1, which means that A does not exist or exists, respectively.

[0065] B is an optional branch unit;

[0066] The subscript b is 0 or 1, which means that B does not exist or exists, respectively.

[0067] L O This is the secondary connector portion, wherein the secondary connector has the following formula:

[0068]

[0069] The wavy line adjacent to Y represents L. O The sites covalently attached to the drug unit, and the wavy line adjacent to A' indicates the sites covalently attached to the remaining portion of the drug connector portion;

[0070] A' is the second optional extended subunit, which becomes a subunit of A in the absence of B.

[0071] The subscript a' is 0 or 1, which means A' does not exist or exists, respectively.

[0072] W is a peptide-cleavable unit, wherein the peptide-cleavable unit comprises a tripeptide having the sequence -P3-P2-P1-, wherein P1, P2, and P3 are each amino acids, wherein:

[0073] The first of the amino acids P1, P2, or P3 is negatively charged;

[0074] The second of the amino acids P1, P2, or P3 has a hydrophobic aliphatic side chain with a hydrophobicity no greater than that of leucine; and

[0075] The third of the amino acids P1, P2, or P3 is less hydrophobic than leucine.

[0076] The first of amino acids P1, P2, or P3 corresponds to any one of P1, P2, or P3; the second of amino acids P1, P2, or P3 corresponds to one of the two remaining amino acids P1, P2, or P3; and the third of amino acids P1, P2, or P3 corresponds to the last remaining amino acid P1, P2, or P3.

[0077] The condition is that -P3-P2-P1- is not -Glu-Val-Cit- or -Asp-Val-Cit-;

[0078] Y is a suicide-type spacer subunit;

[0079] The subscript y is 0, 1, or 2, representing that Y does not exist, or that one or two Ys exist, respectively; and

[0080] The subscript q is an integer ranging from 1 to 4.

[0081] The condition is that when the index b is 0, the index q is 1, and when the index b is 1, the index q is 2, 3, or 4; and

[0082] The ligand drug conjugate compound of the composition has the structure of Formula 1, wherein the subscript p is replaced by the subscript p', wherein the subscript p' is independently an integer from 1 to 12, 1 to 10 or 1 to 8, or 4 or 8.

[0083] In some embodiments, this document provides ligand-drug conjugate compositions of Formula 1, wherein the ligand-drug conjugate compound in the ligand-drug conjugate composition primarily has a drug linker portion of Formula 1H:

[0084]

[0085] Or a pharmaceutically acceptable salt thereof, and the ligand-drug conjugate composition optionally has a few of the following ligand-drug conjugate compounds, wherein one or more drug linker portions of each such compound have a succinimide ring in a hydrolyzed form, and wherein

[0086] HE is a hydrolysis-enhanced unit;

[0087] A', when present, is a subunit of the first extended subunit (A) shown; the subscript a' is 0 or 1, indicating that A' either does not exist or exists; and

[0088] The wavy line indicates the site where the sulfur atom of the ligand unit is covalently bound.

[0089] In some embodiments that can be combined with any of the foregoing embodiments, this document provides ligand-drug conjugate compositions wherein HE is -(C=O).

[0090] In some embodiments that can be combined with any of the foregoing embodiments, this document provides ligand-drug conjugate compositions wherein -Y y -D has the following structure:

[0091]

[0092] Where -N(R) y D' represents D, where D' is the remainder of D;

[0093] The wavy line indicates the site covalently attached to P1;

[0094] The dashed line represents R. y Optional cyclotization to D';

[0095] R y It is an optionally substituted C1-C6 alkyl group when not cyclized to D', or an optionally substituted C1-C6 alkylene group when cyclized to D';

[0096] Each Q is independently selected from -C1-C8 alkyl, -O-(C1-C8 alkyl), halogen, nitro, and cyano; and

[0097] The subscript m is 0, 1, or 2.

[0098] In some embodiments that can be combined with any of the foregoing embodiments, this document provides ligand-drug conjugate compositions, wherein D is a cytotoxic drug, wherein the cytotoxic drug is an auristatin compound containing a secondary amine, wherein the nitrogen atom of the secondary amine is a site covalently attached to the drug linker portion, and the auristatin compound containing the secondary amine has the formula D. F / E-3 Structure:

[0099]

[0100] The sword symbol indicates the covalent attachment site for the nitrogen atom that provides the urethane functional group;

[0101] R 10 and R 11 One of them is hydrogen and the other is methyl;

[0102] R 13 It is isopropyl or -CH2-CH(CH3)2; and

[0103] R 19B It is -CH(CH3)-CH(OH)-Ph, -CH(CO2H)-CH(OH)-CH3, -CH(CO2H)-CH2Ph, -CH(CH2Ph)-2-thiazolyl, -CH(CH2Ph)-2-pyridyl, -CH(CH2-p-Cl-Ph), -CH(CO2Me)-CH2Ph, -CH(CO2Me)-CH2CH2SCH3, -CH(CH2CH2SCH3)C(=O)NH-quinoline-3-yl, -CH(CH2Ph)C(=O)NH-p-Cl-Ph, or

[0104] R 19B Having structure The wavy line indicates covalent attachment to the remaining portion of the auristatin compound.

[0105] In some embodiments that can be combined with any of the foregoing embodiments, this document provides ligand-drug conjugate compositions, wherein the secondary amine-containing auristatin compound is methylaurestatin E (MMAE) or methylaurestatin F (MMAF).

[0106] In some embodiments that can be combined with any of the foregoing embodiments, this document provides ligand-drug conjugate compositions, wherein the subscript q is 1, and the ligand-drug conjugate compound in the ligand-drug conjugate composition primarily has a drug linker portion of formula 1H-MMAE:

[0107]

[0108] Or a pharmaceutically acceptable salt thereof, and the ligand-drug conjugate composition optionally has a few of the following ligand-drug conjugate compounds, wherein one or more drug linker portions of each such compound have a succinimide ring in a hydrolyzed form, and wherein:

[0109] The subscript a' is 0, and A' does not exist; and

[0110] The wavy line indicates the site where the sulfur atom of the ligand unit is covalently bound.

[0111] In some embodiments that can be combined with any of the foregoing embodiments, this document provides ligand-drug conjugate compositions wherein the peptide cleavable unit is a tripeptide having the sequence -P3-P2-P1-, wherein P1, P2, and P3 are each an amino acid, wherein: the P3 amino acid of the tripeptide is a D amino acid configuration; one of the P2 and P1 amino acids has a hydrophobic aliphatic side chain with less hydrophobicity than leucine; and the other of the P2 and P1 amino acids is negatively charged. In some embodiments, the P3 amino acid is D-Leu or D-Ala. In some embodiments, one of the P2 or P1 amino acids has an aliphatic side chain with less hydrophobicity than valine, and the other of the P2 or P1 amino acids is negatively charged at physiological plasma pH. In some embodiments, the P2 amino acid has an aliphatic side chain with less hydrophobicity than valine, and the P1 amino acid is negatively charged at physiological plasma pH. In some embodiments, -P2-P1- is -Ala-Glu- or -Ala-Asp-. In some embodiments, -P3-P2-P1- is -D-Leu-Ala-Asp-, -D-Leu-Ala-Glu-, -D-Ala-Ala-Asp-, or -D-Ala-Ala-Glu-. In some embodiments, the P3 amino acid is D-Leu or D-Ala, the P2 amino acid is Ala, Glu, or Asp, and the P1 amino acid is Ala, Glu, or Asp.

[0112] In some embodiments, this document provides ligand-drug conjugate compositions, wherein said compound has the following structure:

[0113]

[0114]

[0115] or a pharmaceutically acceptable salt thereof,

[0116] Where L is the ligand unit, and the subscript p' is an integer from 1 to 24.

[0117] In some embodiments that can be combined with any of the foregoing embodiments, this document provides ligand-drug conjugate compositions, wherein L is an antibody-ligand unit of an intact antibody or an antigen-binding fragment thereof. In some embodiments, the intact antibody or fragment thereof is capable of selectively binding cancer cell antigens. In some embodiments, the intact antibody is a chimeric antibody, a humanized antibody, or a human antibody, wherein the antibody is capable of selectively binding cancer cell antigens, or the antibody is a non-binding control antibody, thereby defining a non-binding control conjugate composition.

[0118] In some embodiments that can be combined with any of the foregoing embodiments, this document provides ligand-drug conjugate compositions wherein the subscript p ranges from about 2 to about 12, or from about 2 to about 10, or from about 2 to about 8, and particularly the subscript p is about 2, about 4, or about 8.

[0119] In some embodiments that can be combined with any of the foregoing embodiments, this document provides pharmaceutically acceptable formulations comprising an effective amount of a ligand-drug conjugate composition or an equivalent amount of the non-binding control conjugate described herein, and at least one pharmaceutically acceptable excipient. In some embodiments, the at least one pharmaceutically acceptable excipient is a liquid carrier providing a liquid formulation suitable for lyophilization or administration to a subject in need. In some embodiments, the formulation is a solid or liquid formulation derived from lyophilization, wherein at least one excipient in the solid formulation is a lyophilization protectant.

[0120] In some implementations, this document provides drug connector compounds of formula IA:

[0121]

[0122] or its salt, wherein

[0123] D stands for drug unit;

[0124] L B 'It is the precursor of ligand covalent binding;

[0125] A is the first optional extended subunit;

[0126] The subscript 'a' is 0 or 1, which means that A does not exist or exists, respectively.

[0127] B is an optional branch unit;

[0128] The subscript b is 0 or 1, which means that B does not exist or exists, respectively.

[0129] L O This is the secondary connector portion, wherein the secondary connector has the following formula:

[0130]

[0131] The wavy line adjacent to Y represents L. O The sites covalently attached to the drug unit, and the wavy lines adjacent to A' indicate the sites covalently attached to the remainder of the drug connector compound;

[0132] A' is the second optional extended subunit, which becomes a subunit of A in the absence of B;

[0133] The subscript a' is 0 or 1, which means A' does not exist or exists, respectively.

[0134] W is a peptide-cleavable unit, wherein the peptide-cleavable unit comprises a tripeptide having the sequence -P3-P2-P1-, wherein P1, P2, and P3 are each amino acids, wherein:

[0135] The first of the amino acids P1, P2, or P3 is negatively charged;

[0136] The second of the amino acids P1, P2, or P3 has a hydrophobic aliphatic side chain with a hydrophobicity no greater than that of leucine; and

[0137] The third of the amino acids P1, P2, or P3 is less hydrophobic than leucine.

[0138] The first of amino acids P1, P2, or P3 corresponds to any one of P1, P2, or P3; the second of amino acids P1, P2, or P3 corresponds to one of the two remaining amino acids P1, P2, or P3; and the third of amino acids P1, P2, or P3 corresponds to the last remaining amino acid P1, P2, or P3.

[0139] The condition is that -P3-P2-P1- is not -Glu-Val-Cit- or -Asp-Val-Cit-;

[0140] Y is a suicide-type spacer subunit;

[0141] The subscript y is 0, 1, or 2, representing that Y does not exist, or that one or two Ys exist, respectively; and

[0142] The subscript q is an integer ranging from 1 to 4.

[0143] The condition is that when the index b is 0, the index q is 1, and when the index b is 1, the index q is 2, 3 or 4.

[0144] In some embodiments, this document provides drug connector compounds of formula IA, wherein the drug connector compound has the structure of formula IH:

[0145]

[0146] or a salt thereof, wherein:

[0147] HE is a hydrolysis-enhancing unit; and

[0148] A' is the subunit of the first extended subunit (A) shown when it exists; the subscript a' is 0 or 1, indicating that A' does not exist or exists.

[0149] In some embodiments that can be combined with any of the foregoing embodiments, this document provides a drug connector compound, wherein HE is -(C=O).

[0150] In some embodiments that can be combined with any of the foregoing embodiments, this document provides pharmaceutical connector compounds, wherein -Y y -D has the following structure:

[0151]

[0152] Where -N(R) y D' represents D, where D' is the remainder of D;

[0153] The wavy line indicates the site covalently attached to P1;

[0154] The dashed line represents R. y Optional cyclotization to D';

[0155] R y It is an optionally substituted C1-C6 alkyl group when not cyclized to D', or an optionally substituted C1-C6 alkylene group when cyclized to D';

[0156] Each Q is independently selected from -C1-C8 alkyl, -O-(C1-C8 alkyl), halogen, nitro, and cyano; and

[0157] The subscript m is 0, 1, or 2.

[0158] In some embodiments that can be combined with any of the foregoing embodiments, this document provides a drug connector compound, wherein D is a cytotoxic drug, wherein the cytotoxic drug is a secondary amine-containing aurestatin compound, wherein the nitrogen atom of the secondary amine is a site covalently attached to the drug connector portion, and the secondary amine-containing aurestatin compound has the formula D. F / E-3 Structure:

[0159]

[0160] The sword symbol indicates the covalent attachment site for the nitrogen atom that provides the urethane functional group;

[0161] R 10 and R 11 One of them is hydrogen and the other is methyl;

[0162] R 13 It is isopropyl or -CH2-CH(CH3)2; and

[0163] R 19BIt is -CH(CH3)-CH(OH)-Ph, -CH(CO2H)-CH(OH)-CH3, -CH(CO2H)-CH2Ph, -CH(CH2Ph)-2-thiazolyl, -CH(CH2Ph)-2-pyridyl, -CH(CH2-p-Cl-Ph), -CH(CO2Me)-CH2Ph, -CH(CO2Me)-CH2CH2SCH3, -CH(CH2CH2SCH3)C(=O)NH-quinoline-3-yl, -CH(CH2Ph)C(=O)NH-p-Cl-Ph, or

[0164] R 19B Having structure The wavy line indicates covalent attachment to the remaining portion of the auristatin compound.

[0165] In some embodiments that can be combined with any of the foregoing embodiments, this document provides a pharmaceutical connector compound, wherein the aureatine compound containing a secondary amine is methylaureatine E (MMAE) or methylaureatine F (MMAF).

[0166] In some embodiments that can be combined with any of the foregoing embodiments, this document provides drug connector compounds having the structure of formula IH-MMAE:

[0167]

[0168] or its salt, wherein

[0169] The subscript a' is 0, and A' does not exist.

[0170] In some embodiments that can be combined with any of the foregoing embodiments, this document provides a pharmaceutical linker compound, wherein the peptide cleavable unit is a tripeptide having the sequence -P3-P2-P1-, wherein P1, P2, and P3 are each an amino acid, wherein: the P3 amino acid of the tripeptide is a D amino acid configuration; one of the P2 and P1 amino acids has a hydrophobic aliphatic side chain with a hydrophobicity less than leucine; and the other of the P2 and P1 amino acids is negatively charged. In some embodiments, the P3 amino acid is D-Leu or D-Ala. In some embodiments, one of the P2 or P1 amino acids has an aliphatic side chain with a hydrophobicity no greater than valine, and the other of the P2 or P1 amino acids is negatively charged at physiological plasma pH. In some embodiments, the P2 amino acid has an aliphatic side chain with a hydrophobicity no greater than valine, and the P1 amino acid is negatively charged at physiological plasma pH. In some embodiments, -P2-P1- is -Ala-Glu- or -Ala-Asp-. In some embodiments, -P3-P2-P1- is -D-Leu-Ala-Asp-, -D-Leu-Ala-Glu-, -D-Ala-Ala-Asp-, or -D-Ala-Ala-Glu-. In some embodiments, the P3 amino acid is D-Leu or D-Ala, the P2 amino acid is Ala, Glu, or Asp, and the P1 amino acid is Ala, Glu, or Asp.

[0171] In some embodiments, this document provides drug connector compounds having the following structures:

[0172]

[0173] Or its salt.

[0174] In some implementations, this document provides connector compounds of formula IA-L:

[0175]

[0176] or its salt, wherein

[0177] RG is a reactive group;

[0178] L B 'It is the precursor of ligand covalent binding;

[0179] A is the first optional extension subunit;

[0180] The subscript 'a' is 0 or 1, which means that A does not exist or exists, respectively.

[0181] B is an optional branch unit;

[0182] The subscript b is 0 or 1, which means that B does not exist or exists, respectively.

[0183] L O This is the secondary connector portion, wherein the secondary connector has the following formula:

[0184]

[0185] The wavy line adjacent to Y represents L. O The sites covalently attached to the drug unit, and the wavy lines adjacent to A' indicate the sites covalently attached to the remainder of the drug connector compound;

[0186] A' is the second optional extended subunit, which becomes a subunit of A in the absence of B;

[0187] The subscript a' is 0 or 1, which means A' does not exist or exists, respectively.

[0188] W is a peptide-cleavable unit, wherein the peptide-cleavable unit comprises a tripeptide having the sequence -P3-P2-P1-, wherein P1, P2, and P3 are each amino acids, wherein:

[0189] The first of the amino acids P1, P2, or P3 is negatively charged;

[0190] The second of the amino acids P1, P2, or P3 has a hydrophobic aliphatic side chain with a hydrophobicity no greater than that of leucine; and

[0191] The third of the amino acids P1, P2, or P3 is less hydrophobic than leucine.

[0192] The first of amino acids P1, P2, or P3 corresponds to any one of P1, P2, or P3; the second of amino acids P1, P2, or P3 corresponds to one of the two remaining amino acids P1, P2, or P3; and the third of amino acids P1, P2, or P3 corresponds to the last remaining amino acid P1, P2, or P3.

[0193] The condition is that -P3-P2-P1- is not -Glu-Val-Cit- or -Asp-Val-Cit-;

[0194] Y is a suicide-type spacer subunit;

[0195] The subscript y is 0, 1, or 2, representing that Y does not exist, or that one or two Ys exist, respectively; and

[0196] The subscript q is an integer ranging from 1 to 4.

[0197] The condition is that when the index b is 0, the index q is 1, and when the index b is 1, the index q is 2, 3 or 4.

[0198] In some embodiments, this document provides a linker compound wherein the peptide cleavable unit is a tripeptide having the sequence -P3-P2-P1-, wherein P1, P2, and P3 are each an amino acid, wherein: the P3 amino acid of the tripeptide has a D amino acid configuration; one of the P2 and P1 amino acids has a hydrophobic aliphatic side chain with hydrophobicity less than that of leucine; and the other of the P2 and P1 amino acids is negatively charged.

[0199] In some embodiments, this document provides a connector compound having the structure of formula IA-L-3:

[0200] Or its salt.

[0201] In some embodiments, this document provides a connector compound having the following structure:

[0202]

[0203] Or its salt.

[0204] Those and other embodiments of the present invention are described in more detail in the following “Detailed Description” and “Claims”. Attached Figure Description

[0205] Figure 1A , Figure 1B , Figure 1C and Figure 1D The relationship between tumor volume and days post-implantation in a series of xenograft models treated with subcure doses of 4-loaded ADCs, wherein the ADCs have different tripeptide sequences as peptide-cleavable units and the drug-connector portion is represented by the formula mp-P3-P2-P1-PABC-MMAE, compared with subcure doses of 4-loaded ADCs targeting the same cancer cell antigen and having a drug-connector portion represented by the formula mc-val-cit-PABC-MMAE. Figure 1A The compounds in the test were measured at 4 mg / kg. Figure 1B and Figure 1D The compounds in the test were measured at 3 mg / kg. Figure 1C The compounds were tested at 6 mg / kg.

[0206] Figure 2Neutrophil counts following administration of a series of four loaded unbound control conjugates at 10 mg / kg on day 4, the unbound control conjugates having different tripeptide sequences as peptide cleavable units and drug-linker moieties represented by the formula mp-P3-P2-P1-PABC-MMAE, compared to four loaded unbound conjugates having drug-linker moieties represented by the formula mc-val-cit-PABC-MMAE or mp-val-cit-PABC-MMAE.

[0207] Figure 3 Reticulocyte counts in rat plasma following administration of a series of four loaded unbound conjugates at 10 mg / kg to non-tumor-bearing animals on day 4, the unbound conjugates having different tripeptide sequences as peptide-cleavable units and drug-linker moieties represented by the formula mp-P3-P2-P1-PABC-MMAE, compared to four loaded unbound conjugates having drug-linker moieties represented by the formula mc-val-cit-PABC-MMAE or mp-val-cit-PABC-MMAE.

[0208] Figure 4 Histopathology of rat bone marrow after administration of the mediator or 10 mg / kg of the 4-loaded unbound conjugate to non-tumor-bearing animals on day 4, the unbound conjugate having a different tripeptide sequence as a peptide cleavable unit and a drug-linker moiety represented by the formula mp-P3-P2-P1-PABC-MMAE, compared to the 4-loaded unbound conjugate having a drug-linker moiety represented by the formula mc-val-cit-PABC-MMAE.

[0209] Figure 5A and Figure 5B Free MMAE in rat plasma at different time points following administration of the mediator and 10 mg / kg of four loaded unbound conjugates to non-tumor-bearing animals, the unbound conjugates having different tripeptide sequences as peptide cleavable units and drug-linker moieties represented by the formula mp-P3-P2-P1-PABC-MMAE, compared to four loaded unbound conjugates having drug-linker moieties represented by the formula mc-val-cit-PABC-MMAE.

[0210] Figure 6A and Figure 6B Neutrophil elastase ( Figure 6A ) or cathepsin B ( Figure 6BThe percentage of drug cleaved from the heavy chain of a 4-loaded untargeted conjugate in vitro, the untargeted conjugate having a different tripeptide sequence as a peptide cleavable unit and a drug-linker moiety represented by the formula mp-P3-P2-P1-PABC-MMAE, compared to a 4-loaded untargeted conjugate having a drug-linker moiety represented by the formula mp-val-cit-PABC-MMAE.

[0211] Figure 7 , Figure 8 and Figure 9 In rat plasma ( Figure 7 ), Crab-eating macaque plasma ( Figure 8 ) or human plasma ( Figure 9 After incubation for 96 hours in the medium, a series of 4-loaded untargeted conjugates aggregated, wherein the untargeted conjugates have different tripeptide sequences as peptide cleavable units and the drug-linker portion is represented by the formula mp-P3-P2-P1-PABC-MMAE.

[0212] Figure 10 Accumulation of non-targeted MMAF ADCs in rat plasma at different time points.

[0213] Figure 11 The correlation between reticulocyte depletion caused by non-targeted ADCs and ADC aggregation in rat plasma after 96 h of incubation.

[0214] Figure 12 The correlation between reticulocyte depletion caused by non-targeted ADCs in rats and ADC aggregation in cynomolgus monkey plasma after 96 h of incubation.

[0215] Figure 13 The correlation between reticulocyte depletion caused by non-targeted ADCs in rats and ADC aggregation in human plasma after 96 h of incubation.

[0216] Figure 14 The concentration of antibodies in the extracellular bone marrow compartments of rats administered non-targeted ADCs.

[0217] Figure 15 The amount of free MMAE in bone marrow cells of rats administered with non-targeted ADCs.

[0218] Figure 16 In rats, reticulocyte depletion was induced by non-targeted tripeptide ADC on days 5 and 8 following administration of 20 mg / kg.

[0219] Figure 17 In rats, neutrophil depletion was induced by non-targeted tripeptide ADC on days 5 and 8 following administration of 20 mg / kg.

[0220] Figure 18 Bone histology induced by non-targeted tripeptide ADC on days 5 and 8 after administration of 20 mg / kg in rats.

[0221] Figure 19 The correlation between the cLogP of the linker and the aggregation of the corresponding h00 conjugate in rat plasma after 96 h of incubation (expressed as %HMW = % high molecular weight substance).

[0222] Figure 20 The correlation between reticulocyte depletion caused by non-targeted ADCs in rats and ADC accumulation in rat plasma after 96 h of incubation (expressed as %HMW = % high molecular weight substances).

[0223] Figure 21 The correlation between reticulocyte depletion caused by non-targeted ADCs in rats and ADC aggregation in human plasma after 96 h of incubation (expressed as %HMW = % high molecular weight substances).

[0224] Figure 22 The correlation between reticulocyte depletion caused by non-targeted ADCs in rats and ADC accumulation in cynomolgus monkey plasma (expressed as %HMW = % high molecular weight substances) after 96 h of incubation. Detailed Implementation

[0225] Overview

[0226] This invention is partly based on the unexpected discovery that protease activity in tumor tissue differs significantly from that in untargeted normal tissue, providing additional selectivity for cancer cells targeted by ligand-drug conjugates having protease-activated peptide sequences to conditionally release their conjugated cytotoxic compounds. These sequences utilize this difference when the protease-cleavable peptide sequences disclosed herein are incorporated into the peptide-cleavable linker units of the ligand-drug conjugate compounds. It is believed that, in some cases, sequences possessing this property provide conjugate compounds whose biodistribution and / or sensitivity to proteolytic release of free cytotoxic compounds is more favorable to tumor tissue than to normal tissue.

[0227] 1. Definition

[0228] Unless the context otherwise indicates or implies, the terms used herein have the meanings defined below. Unless otherwise prohibited or implied, for example by including mutually exclusive elements or options, in those definitions and throughout this specification, where the context permits, the terms “a” and “an” mean one or more, and the term “or” means and / or. Thus, as presented in the specification and the appended claims, the singular forms “a”, “an”, and “the” include plural indicators unless the context expressly specifies otherwise.

[0229] At various locations throughout this disclosure, such as in any disclosed embodiment or in the claims, a compound, composition, or method is referred to as "comprise" one or more specified components, elements, or steps. Embodiments of the invention also specifically include compounds, compositions, combinations, or methods that are composed of or substantially composed of those specified components, elements, or steps. The term "comprised of" is used interchangeably with the term "comprising" and is expressed as equivalent terms. For example, a disclosed composition, apparatus, article, or method "comprising" a component or step is open-ended, and they include or are read as those compositions or methods with one or more additional components or steps. However, those terms do not cover unlisted elements that would interfere with the function of the disclosed composition, apparatus, article, or method for its intended purpose. Similarly, the disclosed compositions, apparatus, articles, or methods that "consist of" components or steps are closed in nature, and they do not include or are read as those having a considerable number of one or more additional components or one or more additional steps. Furthermore, the term "consisting essentially of" allows for the inclusion of unlisted elements that do not materially affect the function of the disclosed compositions, apparatus, articles, or methods intended for the purposes further defined herein. Section headings used herein are for organizational purposes only and should not be construed as limiting the subject matter. Unless otherwise stated, conventional methods of mass spectrometry, NMR, HPLC, protein chemistry, biochemistry, recombinant DNA techniques, and pharmacology are employed.

[0230] Unless the context otherwise indicates or implies, when the term "about" is used herein in conjunction with a numerical value or range describing a particular property of a compound or composition, it means that the numerical value or range may deviate from what is reasonably considered by one of ordinary skill in the art, while still describing the particular property. Reasonable deviation includes deviations within the range of accuracy or precision of one or more instruments used to measure, determine, or obtain the particular property. Specifically, the term "about," as used herein, means that a numerical value or range may vary by 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.9%, 0.8%, 0.7%, 0.6%, 0.5%, 0.4%, 0.3%, 0.2%, 0.1%, or 0.01%, typically from 10% to 0.5%, more typically from 5% to 1%, while still describing the particular property.

[0231] The subscript p indicates the average number of drug linker portions in the ligand-drug conjugate composition as further defined herein, and the term “about” reflects the acceptable uncertainty in the art for determining this value from the distribution of the ligand-drug conjugate compound in the composition, as determined by standard methods such as size exclusion, HIC chromatography, or HPLC-MS.

[0232] Unless the context otherwise indicates or implies, the terms “essentially retains,” “essentially retaining,” and similar terms as used herein mean that the properties, characteristics, function, or activity of a compound or composition or part thereof have not been detectably altered or are within the experimental error range that determines the same activity, characteristics, or properties of the compound or composition or part thereof that defines the relevant structure.

[0233] Unless the context otherwise indicates or implies, the terms “substantially retain,” “substantially retain,” and similar terms as used herein mean that a measurement of the physical property or characteristic of a compound or composition or part thereof may differ statistically from the same physical property of another compound or composition or part that determines the relevant structure, but such difference does not translate into a statistically significant or meaningful difference in biological activity or pharmacological property in a suitable bioassay system used to assess that activity or property (i.e., retention or substantial retention of biological activity or property). Therefore, the phrase “substantially retain” refers to the effect of the physical property or characteristic of a compound or composition on the physicochemical, pharmacological, or biological activity that is clearly associated with that physical property or characteristic.

[0234] Unless the context otherwise indicates or implies, terms such as “negligibly,” “negligible,” and similar terms as used herein mean an amount of impurity below the quantification level determined by HPLC analysis. Depending on the context, those terms may alternatively mean that no statistically significant difference was observed between measurements or results, or that the difference is within the experimental error range of the instrument used to obtain those values. A negligible difference in experimentally determined parameter values ​​does not imply the presence of a negligible amount of impurity characteristic of that parameter.

[0235] Unless the context otherwise indicates or implies, terms such as “predominately containing,” “predominately having,” and similar terms as used herein refer to the principal component of a mixture. When a mixture has two components, the principal component constitutes more than 50% of the mixture by weight. For mixtures of three or more components, the principal component is the component present in the largest quantity in the mixture and may or may not represent the majority of the mixture's mass.

[0236] Unless the context otherwise indicates or implies, when the term “electron-withdrawing group” is used herein, it refers to a functional group or negatively charged atom that draws electron density away from atoms bonded in an induced manner and / or through resonance (whichever is more dominant, i.e., the functional group or atom may donate electrons through resonance, but overall it may be electron-withdrawing induced) and has a tendency to stabilize anions or electron-rich moieties. The electron-withdrawing effect is generally transferred in an induced manner (albeit in a weakened form) to other atoms that have been bonded to atoms that have been rendered electron-deficient by the electron-withdrawing group (EWG), thereby reducing the electron density further away from the reaction center.

[0237] Electron-withdrawing groups (EWGs) are typically selected from -C(=O)R', -CN, -NO2, -CX3, -X, -C(=O)OR', -C(=O)NH2, and -C(=O)N(R')R op , -C(=O)R', -C(=O)X, -S(=O)2R op , -S(=O)2OR', -SO3H2, -S(=O)2NH2, -S(=O)2N(R')R op -PO3H2, -P(=O)(OR')(OR op )2、-NO、-NH2、-N(R')(R op ), -N(R op )3 + and its salts, wherein X is -F, -Br, -Cl, or -I, R opEach occurrence is independently selected from the grouping previously described for optional substituents, and R' is -H or R. op (where R) op (As previously defined). In some respects, each R op Independently, it is C1-C 12 Alkyl, C1-C8 alkyl, C1-C6 alkyl, or C1-C4 alkyl, or independently selected from C1-C6 alkyl and optionally substituted phenyl groups, and R' is hydrogen. EWG can also be aryl (e.g., phenyl) or heteroaryl (depending on its substitution) and certain electron-withdrawing heteroaryl groups (e.g., pyridyl). Therefore, in some aspects, "electron-withdrawing group" further encompasses electron-withdrawing C5-C substituents substituted with electron-withdrawing substituents. 24 heteroaryl and C6-C 24 Aryl. More generally, the electron-withdrawing group is independently selected from -C(=O)R', -CN, -NO2, -CX3, and -X, where X is a halogen (usually selected from -F and -Cl), and R' is H, C1-C6 alkyl, or C1-C4 alkyl. Depending on its substituents, the optionally substituted alkyl moiety can also be an electron-withdrawing group, and therefore in such cases, these aspects will be covered in the terminology of electron-withdrawing groups.

[0238] Unless the context otherwise indicates or implies, when the term "electron-donating group" is used herein, it refers to a functional group or positively charged atom that increases the electron density of atoms bonded in an inductive manner and / or through resonance (whichever is more dominant; i.e., the functional group or atom may be electron-withdrawing induced, but generally electron-donating via resonance) and has a tendency to stabilize cations or electron-deficient systems. The electron-donating effect is typically transferred via resonance to other atoms that have been made electron-rich by the electron-donating group (EDG), thereby increasing the electron density further from the reaction center. Typically, electron-donating groups are selected from -OH, -OR', -NH2, -NHR', and N(R')2, where each R' is independently selected from C1-C. 12 Alkyl groups, typically C1-C6 alkyl groups. The C6-C6 alkyl groups are determined by their substituents. 24 Aryl, C5-C 24 heteroaryl or unsaturated C1-C 12 Alkyl moiety can also be an electron-donating group, and in some respects, such moiety is covered in the term electron-donating group.

[0239] Unless the context otherwise indicates or implies, the term "compound" as used herein means and covers the compound itself (whether named or represented by a structure) and one or more salt forms (whether explicitly stated or not, unless the context explicitly excludes such salt forms). Compound salts include zwitterionic salt forms having organic or inorganic counterions, as well as acid addition and base addition salt forms, and salt forms involving two or more counterions that may be the same or different. In some respects, the salt form is a pharmaceutically acceptable salt form of the compound. The term "compound" also covers the solvate form of the compound, in which the solvent non-covalently associates with the compound or reversibly covalently associates with the compound, for example, when the carbonyl group of the compound is hydrated to form a geminal diol. Solvate forms include the compound itself and one or more salt forms, and include hemisolides, monosolvents, disolvents, including hydrates; and when the compound can associate with two or more solvent molecules, said two or more solvent molecules may be the same or different. In some cases, compounds of the present invention will include an explicit reference to one or more of the above-described forms (e.g., salts and solvates), which does not imply any solid form of the compound; however, this reference is for emphasis only and should not be construed as excluding any other forms as identified above. Furthermore, when a salt and / or solvate form of a compound or ligand pharmaceutical conjugate composition is not explicitly mentioned, this omission should not be construed as excluding one or more salt and / or solvate forms of the compound or conjugate unless the context clearly indicates that such salt and / or solvate forms will be excluded.

[0240] Unless the context otherwise indicates or implies, the term “optical isomer” as used herein refers to a related compound that has the same atomic connectivity but is structurally different in that it has one or more chiral centers in one or more opposite stereochemical configurations compared to a reference compound.

[0241] Unless the context otherwise indicates or implies, the term "part" as used herein means a specific segment, fragment, or functional group of a molecule or compound. A chemical part is sometimes referred to as a chemical entity that is embedded in or attached to (i.e., a substituent or variable group) a molecule, compound, or chemical formula.

[0242] Unless the context otherwise indicates or implies, for any substituent group or portion described herein by a given range of carbon atoms, the specified range means describing any number of individual carbon atoms. Thus, references such as “optionally substituted C1-C4 alkyl” or “optionally substituted C2-C6 alkenyl” specifically mean the presence of an alkyl moiety with 1, 2, 3, or 4 carbon atoms as defined herein, or an alkenyl moiety with 2, 3, 4, 5, or 6 carbon atoms as defined herein. All these numerical designations are explicitly intended to disclose all individual carbon atom groups; therefore, “optionally substituted C1-C4 alkyl” includes methyl, ethyl, 3-carbon alkyl, and 4-carbon alkyl, including all their positional isomers, whether substituted or unsubstituted. Thus, when the alkyl moiety is substituted, the numerical designation refers to the unsubstituted base moiety and is not intended to include carbon atoms not directly attached to that base moiety, which may be present in substituents of that base moiety. For esters, carbonates, carbamates, and ureas as defined herein, determined by a given range of carbon atoms, the specified range includes the carbonyl carbon of the corresponding functional group. Therefore, C1 esters are carbamates, and C2 esters are acetates.

[0243] The organic substituents, moieties, and groups described herein, as well as any other parts described herein, will generally not include unstable parts, except that such unstable parts are transient particles that can be used to prepare compounds with sufficient chemical stability for one or more of the uses described herein. Those substituents, moieties, or groups having a pentavalent carbon produced by operations defined herein are explicitly excluded.

[0244] Unless the context otherwise indicates or implies, when the term "alkyl" is used herein, either on its own or as part of another term, it refers to a methyl group or a set of consecutive carbon atoms (one of which is monovalent) wherein one or more carbon atoms are saturated (i.e., containing one or more sp atoms). 3 Alkyl groups are composed of carbon atoms arranged covalently in normal, secondary, tertiary, or cyclic configurations (i.e., linear, branched, cyclic, or some combination thereof). When the continuous saturated carbon atoms are arranged in a ring, such alkyl groups are referred to in some respects as carbocyclic groups as further defined herein.

[0245] When referring to an alkyl moiety or group as an alkyl substituent, the alkyl substituent associated with it in the Markush structure or another organic moiety is a methyl group or a chain of consecutive carbon atoms, the consecutive carbon atoms being sp. 3Carbon is covalently attached to the structure or portion. Therefore, alkyl substituents as used herein contain at least one saturated moiety and may also be substituted by cycloalkyl or aromatic or heteroaromatic moieties or groups, or by alkenyl or ynyl moieties, to produce unsaturated alkyl groups. Thus, optionally substituted alkyl substituents may additionally contain one, two, three, or more independently selected double and / or triple bonds, or may be substituted by alkenyl or ynyl moieties or combinations thereof to define an unsaturated alkyl substituent, and may be substituted by other moieties, including suitable optional substituents as described herein. The number of carbon atoms in a saturated alkyl group can vary and is typically 1-50, 1-30, or 1-20, more typically 1-8, or 1-6; the number of carbon atoms in an unsaturated alkyl moiety or group typically varies between 3-50, 3-30, or 3-20, more typically between 3-8.

[0246] The saturated alkyl moiety contains saturated acyclic carbon atoms (i.e., acyclic sp). 3 Carbon) and free of sp 2 Alternatively, it may be an sp carbon atom, but can be substituted by optional substituents as described herein, provided that such substitution does not occur via the sp of the optional substituent. 3 sp 2 Alternatively, sp carbon atoms may be substituted, as this would affect the identification of the number of carbon atoms in the base alkyl moiety of such substitution, unless the optional substituent is a basic unit as defined herein. Unless the context otherwise indicates or implies, the term "alkyl" will refer to a saturated acyclic hydrocarbon group having a specified number of covalently linked saturated carbon atoms; thus, terms such as "C1-C6 alkyl" or "C1-C6 alkyl" mean an alkyl moiety or group containing one saturated carbon atom (i.e., methyl) or two, three, four, five, or six consecutive acyclic saturated carbon atoms, and "C1-C8 alkyl" means an alkyl moiety or group having one saturated carbon atom or two, three, four, five, six, seven, or eight consecutive saturated acyclic carbon atoms. Generally, a saturated alkyl group is one that does not contain sp carbon atoms in its continuous carbon chain. 2 The C1-C6 or C1-C4 alkyl moiety of the sp carbon atom (the latter sometimes referred to as a lower alkyl group), and in some respects will refer to the alkyl moiety having 1 to 8 consecutive acyclic sp atoms. 3 The saturated C1-C8 alkyl moiety of carbon atoms (when the number of carbon atoms is not specified, it does not contain sp in its continuous carbon chain) 2 (or sp carbon atoms). In other respects, when the range of consecutive carbon atoms defines the term "alkyl" but does not specify it as saturated or unsaturated, the term covers saturated alkyl with a specified range as well as unsaturated alkyl where the lower limit of the range is increased by two carbon atoms. For example, the term "C1-C8 alkyl, but not limited to saturated alkyl" includes saturated C1-C8 alkyl and C3-C8 unsaturated alkyl.

[0247] When a saturated alkyl substituent, part, or group is specified, the class includes those obtained by removing hydrogen atoms from a parent alkane (i.e., the alkyl part is monovalent), and may include methyl, ethyl, 1-propyl (n-propyl), 2-propyl (isopropyl, -CH(CH3)2), 1-butyl (n-butyl), 2-methyl-1-propyl (isobutyl, -CH2CH(CH3)2), 2-butyl (sec-butyl, -CH(CH3)CH2CH3), 2-methyl-2-propyl (tert-butyl, -C(CH3)3), pentyl, isopentyl, sec-pentyl, and other linear and branched alkyl parts.

[0248] Unless the context otherwise indicates or implies, the term "alkylene" as used herein, either on its own or as part of another term, refers to a substituted or unsubstituted saturated branched or straight-chain hydrocarbon dimethyl group having a specified number of carbon atoms and having two radical centers (i.e., divalent), wherein one or more carbon atoms are saturated (i.e., containing one or more sp atoms). 3 The number of carbon atoms ranges from 1 to 50 or 1 to 30, typically from 1 to 20 or 1 to 12 carbon atoms, more typically from 1 to 8, 1 or 6, or 1 to 4 carbon atoms, the radical center being derived from the same or two different saturates (i.e., sp) of the parent alkane. 3 The alkylene moiety is obtained by removing two hydrogen atoms from a carbon atom. In some respects, the alkylene moiety is an alkyl group as described herein, wherein the hydrogen atom has been removed from its other saturated carbon atom or from the carbon radical of the alkyl group to form a die. In other respects, the alkylene moiety is a divalent moiety obtained by removing hydrogen atoms from the saturated carbon atom of the parent alkyl moiety, or is covered by it, and includes, for example but not limited to, methylene (-CH2-), 1,2-ethylene (-CH2CH2-), 1,3-propylene (-CH2CH2CH2-), 1,4-butylene (-CH2CH2CH2CH2-), and similar die groups. Typically, the alkylene moiety contains only sp... 3 The alkylene moiety is a branched or straight-chain hydrocarbon (i.e., fully saturated despite the presence of a free radical carbon atom) and is unsubstituted in some respects. In other respects, the alkylene moiety contains internal unsaturated sites in the form of one or more double and / or triple functional groups (usually one or two such functional groups, more commonly one), such that the terminal carbon of the unsaturated alkylene moiety is monovalent sp. 3Carbon atom. In other respects, the alkylene moiety is substituted at one or more saturated carbon atoms of the saturated alkylene moiety or at one or more saturated and / or unsaturated carbon atoms of the unsaturated alkylene moiety with 1 to 4, usually 1 to 3, or 1 or 2 substituents (as defined herein with respect to optional substituents), excluding alkyl, arylalkyl, alkenyl, ynyl, and any other moiety (when the resulting substituted alkylene moiety differs from the unsubstituted alkylene moiety in the number of consecutive non-aromatic carbon atoms), unless the optional substituent is a basic unit as defined herein.

[0249] Unless the context otherwise indicates or implies, when the term "carbocyclic group" is used herein, either on its own or as part of another term, it refers to a group forming a monocyclic, bicyclic, or tricyclic system, wherein each atom forming the ring system (i.e., the skeleton atom) is a carbon atom, and wherein one or more of these carbon atoms in each ring of the ring system are saturated (i.e., contain one or more sp atoms). 3 (Carbon). Therefore, the carbocyclic group is a cyclic arrangement of saturated carbon atoms, but it may also contain one or more unsaturated carbon atoms, so its carbocyclic ring may be saturated or partially unsaturated or may be fused with an aromatic moiety, wherein the fusion point with the cycloalkyl group is adjacent to the unsaturated carbon of the carbocyclic moiety, and the fusion point with the aromatic ring is adjacent to the aromatic carbon atom of the aromatic moiety.

[0250] Unless otherwise stated, the carbocyclic group may be partially substituted (i.e., optionally substituted) as described for alkyl, alkenyl, ynyl, aryl, arylalkyl, alkylaryl, etc., or may be substituted by another cycloalkyl moiety. The cycloalkyl moiety, group, or substituent includes cyclopropyl, cyclopentyl, cyclohexyl, adamantyl, or other cyclic moiety having only carbon atoms in its ring system.

[0251] When a carbocyclic group is used as a Markush group (i.e., a substituent), the carbocyclic group is attached to a Markush or other organic moiety, the carbocyclic group being associated with the Markush or other organic moiety through the carbon atoms contained in the carbocyclic system of the carbocyclic moiety (provided the carbon is not aromatic). When the unsaturated carbon atom of the olefinic moiety containing the carbocyclic substituent is attached to the Markush moiety associated with it, the carbocyclic group is sometimes referred to as a cycloalkenyl substituent. The number of carbon atoms in a carbocyclic substituent is defined by the total number of skeleton atoms in its carbocyclic system. Unless otherwise stated, this number can vary and is generally in the range of 3 to 50, 1-30, or 1-20, more typically 3-8 or 3-6. For example, C3-C8 carbocyclic groups refer to carbocyclic substituents, portions, or groups containing 3, 4, 5, 6, 7, or 8 carbon atoms in the carbocyclic system, and C3-C6 carbocyclic groups refer to carbocyclic substituents, portions, or groups containing 3, 4, 5, or 6 carbon atoms in the carbocyclic system. Carbocyclic groups can be obtained by removing a hydrogen atom from the ring atom of a parent cycloalkane or cycloalkene. Representative C3-C8 carbocyclic groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclopentadienyl, cyclohexyl, cyclohexenyl, 1,3-cyclohexadienyl, 1,4-cyclohexadienyl, cycloheptyl, 1,3-cycloheptadienyl, 1,3,5-cycloheptanetrienyl, cyclooctyl, and cyclooctadienyl.

[0252] Therefore, carbocyclic substituents, portions, or groups typically have 3, 4, 5, 6, 7, or 8 carbon atoms in their carbocyclic ring system, and may contain exocyclic or intracyclic double bonds or intracyclic triple bonds, or a combination of both, wherein the intracyclic double or triple bonds or a combination of both do not form a cyclic conjugated system with 4n+2 electrons. Bicyclic systems may share two carbon atoms, and tricyclic systems may share a total of 3 or 4 carbon atoms. In some respects, the carbocyclic group is a C3-C8 or C3-C6 carbocyclic group, which may be partially substituted (i.e., optionally substituted) and / or substituted by one or more, 1 to 4, usually 1 to 3, or 1 or 2 of the substituents described herein with respect to alkyl, alkenyl, alkynyl, aryl, arylalkyl, and alkylaryl groups, and in some respects is unsubstituted. In other respects, the cycloalkyl moiety, group, or substituent is a C3-C6 cycloalkyl group selected from cyclopropyl, cyclopentyl, and cyclohexyl, or a C3-C8 cycloalkyl group that covers the group and further covers other ring moieties having no more than 8 carbon atoms in its ring system. When the number of carbon atoms is not specified, the carbocyclic moiety, group, or substituent has 3 to 8 carbon atoms in its carbocyclic system.

[0253] Unless the context otherwise indicates or implies, the term “carbocyclic” as used herein, either on its own or as part of another term, refers to an optionally substituted carbocyclic group as defined above, wherein another hydrogen atom in its cycloalkyl ring system has been removed (i.e., it is divalent) and is C3-C. 50 Or C3-C30 Carbon rings, typically C3-C 20 Or C3-C 12 The carbon ring is more typically a C3-C8 or C3-C6 carbon ring, and in some respects is an unsubstituted or optionally substituted C3, C5, or C6 carbon ring. When the number of carbon atoms is not specified, the carbon ring moiety, group, or substituent has 3 to 8 carbon atoms in its carbon ring system.

[0254] In some respects, additional hydrogen atoms are removed from the monovalent carbon atom of the cycloalkyl group to provide a divalent carbon atom; in some cases, the divalent carbon atom is the spirocarbon atom that interrupts the alkyl moiety with the carbon atom of the carbocyclic moiety. In this case, the spirocarbon atom is attributed to the interrupted alkyl moiety and the carbon atom count of the carbocyclic system, wherein the carbocyclic moiety is indicated as being incorporated into the alkyl moiety. In those respects, the carbocyclic moiety, group, or substituent is a C3-C6 carbocyclic system in the form of a spirocyclic system and selected from cyclopropyl-1,1-diyl, cyclobutyl-1,1-diyl, cyclopentyl-1,1-diyl, and cyclohexyl-1,1-diyl, or a C3-C8 carbocyclic moiety that encompasses the group and is further encompassed by other divalent cyclic moieties having no more than 8 carbon atoms in their ring system. The carbocyclic moiety can be saturated or unsaturated, and / or can be unsubstituted or unreplaced in the same manner as described for the carbocyclic basal moiety. If unsaturated, one or both monovalent carbon atoms of the carbocyclic moiety can be sp atoms from the same or different double bond functional groups. 2 A carbon atom, or two monovalent carbon atoms, can be adjacent or non-adjacent sp atoms. 3 Carbon atom.

[0255] Unless the context otherwise indicates or implies, when the term “alkenyl” is used herein, either on its own or as part of another term, it refers to an organic moiety, substituent, or group that contains one or more double-bonded functional groups (e.g., the -CH=CH- moiety) or 1, 2, 3, 4, 5, or 6 or more, usually 1, 2, or 3 such functional groups, more usually one such functional group, and may in some respects be substituted with an aryl moiety or group (e.g., phenyl) (i.e., optionally substituted), or may contain non-aromatic linked ortho-, secondary, tertiary, or cyclic carbon atoms (i.e., linear, branched, cyclic, or any combination thereof) as part of a base moiety, except that the alkenyl substituent, moiety, or group is a vinyl moiety (e.g., the -CH=CH2 moiety). An alkenyl moiety, group, or substituent having multiple double bonds may have double bonds arranged continuously (i.e., 1,3-butadienyl moiety) or discontinuously with one or more intermediate saturated carbon atoms or combinations thereof, provided that the cyclic continuous arrangement of double bonds does not form a cyclic conjugated system with 4n+2 electrons (i.e., not aromatic).

[0256] The alkenyl moiety, group, or substituent contains at least one sp. 2A carbon atom, wherein the carbon atom is divalent and double-bonded to another organic moiety or Markush structure associated therewith, or contains at least two conjugated sp atoms. 2 carbon atom, where sp 2 One of the carbon atoms is monovalent and single-bonded to another organic moiety or Markush structure associated with it. Typically, when the alkenyl group is used as a Markush group (i.e., a substituent), the alkenyl group is single-bonded to a Markush structure or another organic moiety, and the alkenyl group is sp-bonded through the olefinic functional group of the alkenyl moiety. 2 Carbon associates with a Markush moiety or another organic part. In some respects, when specifying an alkenyl moiety, the class encompasses those corresponding to any optionally substituted alkyl or carbocyclic group, group moiety, or substituent described herein, having one or more internal double bonds (where sp... 2 Carbon atoms are monovalent) and sp from parent olefin compounds 2 The monovalent moiety obtained by removing hydrogen atoms from carbon. Such monovalent moieties include, but are not limited to, vinyl (-CH=CH2), allyl, 1-methylvinyl, butenyl, isobutenyl, 3-methyl-2-butenyl, 1-pentenyl, cyclopentenyl, 1-methyl-cyclopentenyl, 1-hexenyl, 3-hexenyl, and cyclohexenyl. In some aspects, the term alkenyl covers all carbon-containing moieties containing at least one double-bonded functional group and / or other linear, cyclic, and branched moieties, wherein sp... 2 One of the carbon atoms is monovalent.

[0257] The number of carbon atoms in the alkenyl moiety is determined by the sp(s) of one or more alkene functional groups (defined as alkenyl substituents). 2 The number of carbon atoms and the amount of carbon atoms attached to these sp atoms 2 The total number of consecutive non-aromatic carbon atoms in each of the carbon groups (excluding any carbon atoms in other parts where the alkenyl moiety is a variable group or a Markush structure, and carbon atoms from any optional substituents of the alkenyl moiety) is defined. When the double bond functional group is double-bonded with a Markush structure (e.g., =CH2), this number ranges from 1 to 50 or 1 to 30, typically from 1 to 20 or 1 to 12, and more typically from 1 to 8, 1 to 6, or 1 to 4 carbon atoms; or when the double bond functional group is single-bonded with a Markush structure (e.g., -CH=CH2), this number ranges from 2 to 50, typically from 2 to 30, 2 to 20, or 2 to 12, and more typically from 2 to 8, 2 to 6, or 2 to 4 carbon atoms. For example, C2-C8 alkenyl or C2-C8 alkenyl means an alkenyl moiety containing 2, 3, 4, 5, 6, 7, or 8 carbon atoms, wherein at least two carbon atoms are conjugated sp. 2 Carbon atoms, one of which is monovalent; C2-C6 alkenyl or C2-C6 alkenyl means an alkenyl moiety containing 2, 3, 4, 5, or 6 carbon atoms, wherein at least two carbon atoms are conjugated with each other. 2Carbon, in which one of these carbon atoms is monovalent. In some respects, the alkenyl substituent or group is a C2-C6 or C2-C4 alkenyl moiety, which has only two sp atoms conjugated with each other. 2 Carbon, wherein one of these carbon atoms is monovalent, while in other respects, the alkenyl moiety is unsubstituted or substituted by one to four or more, typically one to three, more typically one or two independently selected moieties as disclosed herein, including substituents as defined herein for optional substituents, excluding alkyl, arylalkyl, heteroarylalkyl, alkenyl, ynyl, and any other moiety (when the substituted alkenyl differs from the unsubstituted alkenyl in the number of consecutive non-aromatic carbon atoms), wherein said substitution may be in the number of consecutive sps of the alkenyl moiety. 2 Carbon and sp 3 Any one of the carbon atoms (if present). Typically, alkenyl substituents are those with only two conjugated sp atoms. 2 The C2-C6 or C2-C4 alkenyl moiety of carbon. When the number of carbon atoms is not specified, the alkenyl moiety has 2 to 8 carbon atoms.

[0258] Unless the context otherwise indicates or implies, the term “alkenyl” as used herein, either on its own or as part of another term, refers to an organic moiety, substituent, or group that comprises one or more double-bonded portions (as previously described for alkenyl), has a specified number of carbon atoms, and possesses two radical centers, said radical centers being derived from the same or two different sp[s] of an olefin functional group. 2 The alkenyl group is obtained by removing two hydrogen atoms from a carbon atom or from two separate alkene functional groups in a parent alkene. In some respects, the alkenyl moiety is an alkenyl group as described herein, wherein hydrogen atoms have been removed from the same or different sp groups of the double bond functional groups of the alkenyl group. 2 Carbon atoms removed, or sp from different double bond moieties. 2 Carbon is removed to provide a diol. Typically, the alkenyl moiety encompasses a group containing -C=C- or -C=CX. 1 -C=C- structure of two basis elements, where X 1 The alkylene moiety is either absent or optionally substituted as defined herein, typically C1-C6 alkylene, and more typically unsubstituted. The number of carbon atoms in the alkenyl moiety is determined by the sp(s) of one or more olefinic functional groups (defined as alkenyl moieties). 2 The number of carbon atoms and the amount added to sp 2The total number of consecutive non-aromatic carbon atoms in each of the carbon atoms (excluding other portions where the alkenyl moiety exists as a variable group or any carbon atoms in a Markush structure) is defined. Unless otherwise specified, this number ranges from 2 to 50 or 2 to 30, typically from 2 to 20 or 2 to 12, and more typically from 2 to 8, 2 to 6, or 2 to 4 carbon atoms. For example, C2-C8 alkenyl or C2-C8 alkenyl means an alkenyl moiety containing 2, 3, 4, 5, 6, 7, or 8 carbon atoms, wherein at least two carbon atoms are conjugated with each other. 2 Carbon, one of which is divalent or both are monovalent; C2-C6 alkenyl or C2-C6 alkenyl means an alkenyl moiety containing 2, 3, 4, 5 or 6 carbon atoms, wherein at least two carbon atoms are conjugated with each other. 2 Carbon, of which at least two are sp. 2 Carbon atoms, one of which is divalent or both are monovalent. In some respects, the alkenyl group is composed of two conjugated sp atoms. 2 C2-C6 or C2-C4 alkenyl groups of carbon, where both sp... 2 All carbon atoms are monovalent and are unsubstituted in some respects. When the number of carbon atoms is not specified, the alkenyl moiety has 2 to 8 carbon atoms and is either unsubstituted or substituted in the same manner as described for the alkenyl moiety.

[0259] Unless the context otherwise indicates or implies, the term “alkynyl” as used herein, either by itself or as part of another term, refers to an organic moiety, substituent, or group comprising one or more triple-bonded functional groups (e.g., the -C≡C- moiety) or 1, 2, 3, 4, 5, or 6 or more, usually 1, 2, or 3 such functional groups, more usually one such functional group, and which may in some respects be substituted with an aryl moiety (e.g., phenyl) (i.e., optionally substituted), or substituted with an alkenyl moiety or linked or connected normal, secondary, tertiary, or cyclic carbon atom (i.e., linear, branched, cyclic, or any combination thereof), except that the alkynyl substituent, moiety, or group is -C≡CH. An alkynyl moiety, group, or substituent having multiple triple bonds may have triple bonds arranged continuously or discontinuously with one or more intermediate saturated or unsaturated carbon atoms or combinations thereof, provided that the continuously arranged cyclic triple bonds do not form a cyclic conjugated system with 4n+2 electrons (i.e., not aromatic).

[0260] An alkynyl moiety, group, or substituent contains at least two sp carbon atoms, wherein said carbon atoms are conjugated with each other and wherein one of said sp carbon atoms is single-bonded to another organic moiety or Markush structure associated therewith. When the alkynyl group is used as a Markush group (i.e., is a substituent), the alkynyl group is single-bonded to a Markush or other organic moiety, and the alkynyl group is associated with the Markush or other organic moiety via a triple-bonded carbon (i.e., sp carbon) of the terminal alkyne functional group. In some aspects, when the alkynyl moiety, group, or substituent is specified, the class encompasses any optionally substituted alkyl or carbocyclic group, group moiety, or substituent described herein, having one or more internal triple bonds and a monovalent moiety obtained by removing a hydrogen atom from the sp carbon of the parent alkyne compound. Such monovalent moieties are, for example, but not limited to, -C≡CH and -C≡C-CH3 and -C≡C-Ph.

[0261] The number of carbon atoms in an alkynyl substituent is defined by the number of sp carbon atoms in the alkene functional group (defined as an alkynyl substituent) and the total number of consecutive non-aromatic carbon atoms attached to each of these sp carbons (excluding any carbon atoms in other parts where the alkenyl moiety is a variable group or in a Markush structure). When a triple-bonded functional group is single-bonded to a Markush structure (e.g., -CH≡CH), this number can vary from 2 to 50, typically 2 to 30, 2 to 20, or 2 to 12, and more typically 2 to 8, 2 to 6, or 2 to 4 carbon atoms. For example, C2-C8 ynyl or C2-C8 ynyl means an ynyl moiety containing 2, 3, 4, 5, 6, 7, or 8 carbon atoms, wherein at least two carbon atoms are conjugated sp carbon atoms, and one of these carbon atoms is monovalent; C2-C6 ynyl or C2-C6 ynyl means an ynyl moiety containing 2, 3, 4, 5, or 6 carbon atoms, wherein at least two carbon atoms are conjugated sp carbon atoms, and one of these carbon atoms is monovalent. In some aspects, the ynyl substituent or group is a C2-C6 or C2-C4 ynyl moiety having two conjugated sp carbons, wherein one of these carbon atoms is monovalent, while in other aspects, the ynyl moiety is unsubstituted. When the number of carbon atoms is not specified, the ynyl moiety, group, or substituent has 2 to 8 carbon atoms. The ynyl moiety may be substituted or unsubstituted in the same manner as described with respect to the alkenyl moiety, except that substitution on the monovalent sp carbon is not permitted.

[0262] Unless the context otherwise indicates or implies, the term “aryl” as used herein, either by itself or as part of another term, refers to an organic moiety, substituent, or group having an aromatic ring system or a fused aromatic ring system and free of cyclic heteroatoms, wherein the organic moiety, substituent, or group comprises or consists of one, two, three, or four to six aromatic rings (each aromatic ring being independently and optionally substituted), typically consisting of one to three aromatic rings, more typically consisting of one or two aromatic rings (each aromatic ring being independently and optionally substituted), wherein the rings consist of carbon atoms involved in a cyclic conjugation system (Hückel's rule) involving only 4n+2 electrons (typically 6, 10, or 14 electrons), with some carbon atoms possibly additionally involved in exocyclic conjugation (cross-conjugation, e.g., quinones) with heteroatoms. Aryl substituents, moieties, or groups are typically formed of six, eight, ten, or more up to 24 consecutive aromatic carbon atoms to include C6-C. 24 Aryl, and in some respects C6-C 20 Or C6-C 12 Aryl. An aryl substituent, part, or group is optionally substituted and, in some respects, unsubstituted or substituted by one, two, three, or more, usually one or two independently selected substituents, as defined herein with respect to alkyl, alkenyl, alkynyl, or other portions described herein (including another aryl or heteroaryl to form a biaryl) and other optional substituents as defined herein. In other respects, the aryl group is C6-C. 10 Aryl groups, such as phenyl and naphthyl, as well as phenanthrene. Since the aromaticity of a neutral aryl moiety requires an even number of electrons, it should be understood that a given range of this moiety will not cover species with an odd number of aromatic carbons. When an aryl group is used as a Markush group (i.e., a substituent), the aryl group is attached to a Markush or other organic moiety, associating the aryl group with the Markush or other organic moiety through the aromatic carbons of the aryl group.

[0263] Unless the context otherwise indicates or implies, when the term "heterocyclic group" is used herein, either on its own or as part of another term, it refers to a carbocyclic group in which one or more, but not all, of the skeletal carbon atoms in the carbocyclic system, along with the hydrogen atoms to which they are attached, are replaced (optionally substituted where permitted) by independently selected heteroatoms or heteroatom portions, including but not limited to N / NH, O, S, Se, B, Si, and P, wherein two or more heteroatoms or heteroatom portions (usually two) may be adjacent to each other or separated by one or more carbon atoms in the same ring system, typically separated by one to three carbon atoms. Those heteroatoms or heteroatom portions are typically N / NH, O, and S. Heterocyclic groups typically contain monovalent skeletal carbon atoms or monovalent heteroatoms or heteroatomic moieties and have a total of one to ten heteroatomic and / or heteroatomic moieties, typically a total of 1 to 5, or more typically a total of 1 to 3, or 1 or 2, provided that the skeletal atoms in any one or more heterocycles of the heterocyclic group are not all heteroatoms and / or heteroatomic moieties (i.e., at least one carbon atom in each ring is not replaced by at least one carbon atom in one of these rings that has been substituted), wherein each heteroatom or heteroatomic moiety in said one or more rings (optionally substituted where permitted) is independently selected from N / NH, O, and S, provided that any ring does not contain two adjacent O or S atoms. Exemplary heterocyclic groups and heteroaryl groups are collectively referred to as heterocycles, which are provided by the following literature: Paquette, Leo A.; "Principles of Modern Heterocyclic Chemistry" (WA Benjamin, New York, 1968), especially Chapters 1, 3, 4, 6, 7 and 9; "The Chemistry of Heterocyclic Compounds, A series of Monographs" (John Wiley & Sons, New York, 1950 to present), especially Volumes 13, 14, 16, 19 and 28; and J. Am. Chem. Soc. 1960, 82: 5545-5473, especially 5566-5573).

[0264] When a heterocyclic group is used as a Markush group (i.e., a substituent), the saturated or partially unsaturated heterocycle of the heterocyclic group is attached to the Markush structure or other part. The heterocyclic group associates with the Markush structure or other part through the carbon atom or heteroatom of the heterocycle, wherein such attachment does not result in an unstable or unacceptable oxidation state of the carbon atom or heteroatom. In this context, the heterocyclic group is a monovalent moiety, wherein the heterocycle of the heterocyclic system that defines it as a heterocyclic group is non-aromatic but can be fused with a carbocyclic, aryl, or heteroaryl ring and includes heterocyclic moieties fused with phenyl (i.e., benzo) compounds.

[0265] The heterocyclic group is as follows: C3-C 50 Or C3-C 30 Carbocyclic groups, usually C3-C 20 Or C3-C 12 A carbocyclic group, more typically C3-C8 or C3-C6, wherein one, two, three, or more, but not all, of the carbons in its cycloalkyl ring system, along with their attached hydrogens (usually one, two, three, or four, more typically one or two), are replaced by heteroatoms or heteroatoms independently selected from N / NH, O, and S (optionally substituted where permitted), thus being C3-C6. 50 Or C3-C 30 Heterocyclic groups, usually C3-C 20 Or C3-C 12 Heterocyclic groups, more typically C3-C6 or C5-C6 heterocyclic groups, wherein the subscript indicates the total number of skeletal atoms (including carbon atoms and heteroatoms) in one or more heterocyclic systems of the heterocyclic group. In some aspects, the heterocyclic group contains 0 to 2 N, 0 to 2 O, or 0 to 1 S skeletal heteroatoms (optionally substituted) or some combination thereof, provided that at least one of the said heteroatoms is present in the heterocyclic system of the heterocyclic group. The heterocyclic group can be saturated or partially unsaturated and / or unsubstituted or substituted with an oxo (=O) moiety on the skeletal carbon atom, as in pyrrolidone-2-one, and / or substituted with one or two oxo moiety on the skeletal heteroatoms to contain an oxidized heteroatom, such as, but not limited to, -N (=O), -S (=O)- or -S (=O)2-. A fully saturated or partially unsaturated heterocyclic group may be substituted or further substituted by an alkyl, (hetero)aryl, (hetero)arylalkyl, alkenyl, ynyl, or other moiety as described herein (including optional substituents as defined herein) or by a combination of two, three, or more, typically one or two such substituents. In some respects, the heterocyclic group is selected from pyrrolidinyl, piperidinyl, morpholinyl, and piperazineyl.

[0266] Unless the context otherwise indicates or implies, the term "heterocycle" as used herein, either by itself or as part of another term, refers to a heterocyclic moiety, group, or substituent as defined above, wherein a hydrogen atom from its monovalent carbon atom, a hydrogen atom from a different skeletal atom (carbon or nitrogen, if the latter is present), or an electron (where permitted) from the skeletal nitrogen atom is removed, or an electron from a nitrogen ring atom that is no longer monovalent is removed and replaced by a bond (i.e., it is divalent). In some aspects, the second hydrogen atom being replaced is a hydrogen atom from the monovalent carbon atom of the parent heterocyclic moiety, thus forming a spirocarbon atom, which in some cases can interrupt the alkyl moiety with this carbon ring carbon atom. In this case, the spirocarbon atom is attributed to the carbon atom count of the interrupted alkyl moiety, wherein the heterocycle is indicated as being incorporated into the alkyl moiety.

[0267] Unless the context otherwise indicates or implies, the term "heteroaryl" as used herein, either on its own or as part of another term, refers to an aryl moiety, group, or substituent as defined herein, wherein one or more, but not all, of the aromatic carbons in the aryl ring system are replaced by heteroatoms. Heteroaryls typically contain a total of one to four skeletal heteroatoms in one or more rings of a heteroaryl ring system, provided that not all skeletal atoms in any ring system of the heteroaryl are heteroatoms (optionally substituted where permitted), and the heteroaryl has 0 to 3 N, 1 to 3 N, or 0 to 3 N skeletal heteroatoms, typically 0 to 1 O, and / or 0 to 1 S skeletal heteroatoms, provided that at least one skeletal heteroatomation is present. Heteroaryls can be monocyclic, bicyclic, or polycyclic. Polycyclic heteroaryls are typically C5-C6. 50 Or C5-C 30 heteroaryl, more often C5-C 20 Or C5-C 12 Heteroaryl, bicyclic heteroaryl is usually C5-C 10 Heteroaryl, monocyclic heteroaryl is usually C5-C6 heteroaryl, where the subscript indicates the total number of skeletal atoms (including its carbon atoms and heteroatoms) of one or more aromatic ring systems of heteroaryl. In some aspects, a heteroaryl moiety is a bicyclic aryl moiety in which one, two, three, four, or more, typically one, two, or three carbon atoms and their attached hydrogen atoms of one or more aromatic rings of the parent bicyclic aryl moiety are replaced by independently selected heteroatoms or heteroatom portions; or a monocyclic aryl moiety in which one, two, three, or more, typically one or two carbon atoms and their attached hydrogen atoms of one or more aromatic rings of the parent monocyclic aryl moiety are replaced by independently selected heteroatoms or heteroatom portions, wherein said heteroatoms or heteroatom portions are optionally substituted where permitted, including N / NH, O, and S, provided that the skeletal atoms of any aromatic ring system in the parent aryl moiety are not entirely replaced by heteroatoms and are more typically replaced by oxygen (-O-), sulfur (-S-), nitrogen (=N-), or -NR-, such that the nitrogen heteroatom is optionally substituted, wherein R is -H, a nitrogen protecting group, or an optionally substituted C1-C 20 Alkyl, or optionally substituted C6-C 24 Aryl or C5-C 24 Heteroaryl groups form heterocyclic aryl groups. In other aspects, one, two, or three carbon atoms of one or more aromatic rings of the parent aryl moiety, along with the hydrogen atoms attached to them, are replaced by nitrogen, which is substituted by another organic moiety in a manner that maintains the cyclic conjugated system. In still other aspects, the aromatic carbon groups of the parent aryl moiety are replaced by aromatic nitrogen groups. In any of those aspects, the nitrogen, sulfur, or oxygen heteroatom participates in the conjugated system through π-bonding with adjacent atoms in the ring system or through lone pairs of electrons on the heteroatom. In still other aspects, the heteroaryl group has a heterocyclic structure as defined herein, wherein its ring system has been aromatized.

[0268] Typically, heteroaryl groups are monocyclic, and in some respects have 5- or 6-membered heteroaryl ring systems. A 5-membered heteroaryl is a monocyclic C5-heteroaryl containing 1 to 4 aromatic carbon atoms and the necessary number of aromatic heteroatoms in its heteroaryl ring system. A 6-membered heteroaryl is a monocyclic C6-heteroaryl containing 1 to 5 aromatic carbon atoms and the necessary number of aromatic heteroatoms in its heteroaryl ring system. 5-membered heteroaryl groups have four, three, two, or one aromatic heteroatom, and 6-membered heteroaryl groups include heteroaryl groups having five, four, three, two, or one aromatic heteroatom.

[0269] C5-heteroaryl, also known as 5-membered heteroaryl, is a monovalent moiety obtained by removing a hydrogen atom from a skeletal aromatic carbon or an electron from a skeletal aromatic heteroatom in a parent aromatic heterocyclic compound, where permissible. In some respects, it is selected from pyrrole, furan, thiophene, oxazole, isoxazole, thiazole, isothiazole, imidazole, pyrazole, triazole, and tetraazole. In other respects, the parent heterocycle is selected from thiazole, imidazole, oxazole, and triazole, and is usually thiazole or oxazole, more commonly thiazole.

[0270] C6 heteroaryl is a 6-membered monovalent moiety obtained by removing a hydrogen atom from an aromatic carbon or an electron from an aromatic heteroatom (where permitted), in certain aspects selected from pyridine, pyridazine, pyrimidine, and triazine. The heteroaryl may be substituted or further substituted with an alkyl, (hetero)arylalkyl, alkenyl, or alkynyl group, or substituted or further substituted with an aryl or another heteroaryl group to form a biaryl group, or substituted or further substituted with other moieties as described herein (including optional substituents as defined herein), or substituted or further substituted with combinations of two, three, or more, usually one or two such substituents.

[0271] When the term “arylalkyl” or “heteroarylalkyl” is used herein, either on its own or as part of another term, it refers to an aryl or heteroaryl moiety bonded to an alkyl portion, i.e., (aryl)-alkyl-, wherein the alkyl and aryl groups are as described above. Typically, arylalkyl groups are (C6-C) 24 (aryl)-C1-C 12 Alkyl moiety, group or substituent, heteroarylalkyl is (C5-C 24 (heteroaryl)-C1-C 12 Alkyl moiety, group, or substituent. When a (hetero)arylalkyl group is used as a Markush group (i.e., a substituent), the alkyl moiety of the (hetero)arylalkyl group is attached to the Markush group, and the (hetero)arylalkyl group is sp... 3 Carbon associates with Markush. In some respects, arylalkyl groups are (C6-C6) 24 (aryl)-C1-C 12 Alkyl- or (C6-C) 20(aryl)-C1-C 20 Alkyl-, typically (C6-C) 12 (aryl)-C1-C 12 Alkyl- or (C6-C) 10 (aryl)-C1-C 12 Alkyl-, more typically (C6-C) 10 Aryl)-C1-C6 alkyl, such as, but not limited to, C6H5-CH2-, C6H5-CH(CH3)CH2- and C6H5-CH2-CH(CH2CH2CH3)-. (Hetero)arylalkyl may be unsubstituted or substituted in the same manner as described for the (hetero)aryl and / or alkyl moiety.

[0272] Unless the context otherwise indicates or implies, the term “arylene” or “heteroarylene” as used herein, either by itself or as part of another term, refers to an aromatic or heteroaryl dimethyl moiety (i.e., it is divalent) that forms two covalent bonds in another organic moiety, with respect to the other organic moiety, said bonds being in the ortho, meta, or para configuration. Arylene and some heteroarylene moietyes include divalent types formed by removing a hydrogen atom from a parent aryl or heteroaryl moiety, group, or substituent as defined herein. Other heteroarylene moietyes are divalent types in which a hydrogen atom has been removed from two different aromatic carbon atoms of a parent aromatic heterocycle to form a dimethyl type, or in which a hydrogen atom and another hydrogen atom or an electron has been removed from an aromatic carbon atom or heteroatom of a parent aromatic heterocycle to form a dimethyl type, wherein one aromatic carbon atom and one aromatic heteroatom are monovalent or the two different aromatic heteroatoms are each monovalent. The heteroarylene further includes one or more heteroatoms and / or heteroatoms that partially replace one or more, but not all, of the aromatic carbon atoms of the parent arylene.

[0273] Non-limiting exemplary arylene groups that are optionally substituted at the remaining positions are phenyl-1,2-ethylenedimethylbenzene, phenyl-1,3-ethylenedimethylbenzene, and phenyl-1,4-ethylenedimethylbenzene, as shown in the following structures:

[0274]

[0275] Unless the context otherwise indicates or implies, the term "heteroalkyl" as used herein, either alone or in combination with another term, refers to an optionally substituted straight-chain or branched hydrocarbon that is fully saturated or has 1 to 3 degrees of unsaturation and has 1 to 12 carbon atoms and 1 to 6 heteroatoms, typically 1 to 5 heteroatoms, more typically one or two heteroatoms or heteroatom moieties selected from O, N / NH, Si, and S (optionally substituted where permitted), each nitrogen and sulfur atom being independently and optionally oxidized to an N-oxide, sulfoxide, or sulfone, or one or more of the nitrogen atoms being optionally substituted or quaternized. One or more heteroatoms or heteroatom moieties O, N / NH, S, and / or Si may be located at any internal position of the heteroalkyl group or at the terminal position of an optionally substituted alkyl group of the heteroalkyl group. In some respects, the heteroalkyl group is fully saturated or has 1 degree of unsaturation and contains 1 to 6 carbon atoms and 1 to 2 heteroatoms, while in other respects, the heteroalkyl group is unsubstituted. Non-limiting examples are -CH2-CH2-O-CH3, -CH2-CH2-NH-CH3, -CH2-CH2-N(CH3)-CH3, -CH2-S-CH2-CH3, -CH2-CH2-S(O)-CH3, -NH-CH2-CH2-NH-C(O)-CH2-CH3, -CH2-CH2-S(O)2-CH3, -CH=CH-O-CH3, -Si(CH3)3, -CH2-CH=NO-CH3, and -CH=CH-N(CH3)-CH3. At most two heteroatoms can be consecutive, for example, -CH2-NH-OCH3 and -CH2-O-Si(CH3)3.

[0276] Heteroalkyl groups are generally represented by the number of one or more consecutive heteroatoms and non-aromatic carbon atoms (including those with one or more consecutive carbon atoms attached to one or more heteroatoms), unless otherwise specified (e.g., as described for aminoalkyl groups) or according to the context. Thus, -CH2-CH2-O-CH3 and -CH2-CH2-S(O)-CH3 are both C4-heteroalkyl groups, and -CH2-CH=NO-CH3 and -CH=CH-N(CH3)2 are both C5-heteroalkyl groups. The heteroalkyl group may be unsubstituted, or substituted at its heteroatom or heteroatom component by any portion described herein (including optional substituents as defined herein) (i.e., optionally substituted), and / or substituted at its alkyl component by one to four or more, typically one to three, or one or two independently selected portions described herein (including one or more optional substituents as defined herein) (i.e., optionally substituted), wherein the substituents do not include alkyl, (hetero)arylalkyl, alkenyl, ynyl, another heteroalkyl, or another other portion (when the substituted alkenyl group differs from the unsubstituted aminoalkyl group in the number of consecutive non-aromatic carbon atoms).

[0277] As defined herein, aminoalkyl is an exemplary heteroalkyl group in which the terminal carbon atom of the alkyl moiety, other than its monovalent carbon atom, is replaced by an amino group. When indicated as a substituent for a Markush structure or other organic moiety associated with it, the monovalent carbon atom of the alkyl moiety is attached to another organic moiety to which it will be associated, and the monovalent carbon atom of the alkyl moiety is typically a different carbon atom from the carbon atom attached to the amino group. Aminoalkyl differs from other heteroalkyl groups in that it is numbered solely by indicating the number of consecutive carbon atoms in its alkylene moiety.

[0278] Unless the context otherwise indicates or implies, the term "heteroalkylene" as used herein, either on its own or in combination with another term, means a divalent group obtained from a heteroalkyl group (as discussed above) by removing a hydrogen atom or a heteroatom electron from the parent heteroalkyl group to provide a divalent moiety, such as, but not limited to, -CH2-CH2-S-CH2-CH2- and -CH2-S-CH2-CH2-NH-CH2-. For a heteroalkylene group, one or more heteroatoms may be internal to it or may occupy one or both ends of its optionally substituted alkylene chain, such that one or both of these heteroatoms are monovalent. When a heteroalkylene group is a component of a linker unit, both orientations of that component within the linker unit are permitted, unless the context otherwise indicates or implies. A heteroalkylene group is generally represented by the number of its one or more consecutive heteroatoms and non-aromatic carbon atoms (including those attached to one or more consecutive carbon atoms), unless otherwise indicated or according to the context. Alkylene diamines are heteroalkylene groups in which two monovalent carbon atoms of the alkylene group are replaced by amino groups such that each nitrogen atom is monovalent, and alkylene diamines differ from other heteroalkylene groups in that they are numbered only by indicating the number of adjacent carbon atoms of their alkylene moiety.

[0279] Unless the context otherwise indicates or implies, the term “aminoalkyl” as used herein, either on its own or in combination with another term, refers to a moiety, group, or substituent having a basic nitrogen moiety bonded to a radical terminus of an alkylene moiety as defined above, to provide a primary amine in which the basic nitrogen is not further substituted, or to provide a C1-C amine in which the basic amine is optionally substituted by one or two independently selected substituents as described above. 12 The alkyl moiety is further substituted into a secondary or tertiary amine. In some respects, the optionally substituted alkyl group is a C1-C8 or C1-C6 alkyl group, while in others, the alkyl group is unsubstituted. In still other respects, the basic nitrogen, together with its substituents, defines an optionally substituted C3-C8 heterocyclic group containing a basic nitrogen as a skeletal atom, typically in the form of an optionally substituted nitrogen-containing C3-C6 or C5-C6 heterocyclic group. When an aminoalkyl group is used as a variable group in a Markush structure, the alkylene moiety of the aminoalkyl group is attached to the Markush structure, and the alkylene moiety of the aminoalkyl group is sp... 3Carbon and Markush association, in some respects, sp 3 Carbon is another radical terminus of the aforementioned alkylene group. Aminoalkyl groups are usually represented by the number of consecutive carbon atoms in their alkylene portion. Thus, C1 aminoalkyl groups are, for example, but not limited to -CH2NH2, -CH2NHCH3 and -CH2N(CH3)2, and C2 aminoalkyl groups are, for example, but not limited to -CH2CH2NH2, -CH2CH2NHCH3 and -CH2CH2N(CH3)2.

[0280] Unless the context otherwise indicates or implies, the terms “optionally substituted alkyl,” “optionally substituted alkenyl,” “optionally substituted alkynyl,” “optionally substituted arylalkyl,” “optionally substituted heterocyclic,” “optionally substituted aryl,” “optionally substituted heteroaryl,” “optionally substituted heteroarylalkyl,” etc., as used herein, refer to alkyl, alkenyl, alkynyl, arylalkyl, heterocyclic, aryl, heteroaryl, heteroarylalkyl, or other substituent, part, or group as defined or disclosed herein, wherein one or more hydrogen atoms of the substituent, part, or group have been optionally substituted by one or more different parts or groups, or an alicyclic carbon chain containing one of those substituents, parts, or groups is interrupted by replacing one or more carbon atoms of the chain with one or more different parts or groups. In some aspects, the olefin functional group replaces two consecutive sps of the alkyl substituent. 3 Carbon atom, provided that the radical carbon of the alkyl moiety is not replaced, such that the optionally substituted alkyl group becomes an unsaturated alkyl substituent.

[0281] The optional substituents replacing one or more hydrogens of any of the aforementioned substituents, portions, or groups are independently selected from C6-C6. 24 Aryl, C5-C 24 heteroaryl, hydroxyl, C1-C 20 Alkoxy, C6-C 24 Aryloxy, cyano, halogen, nitro, C1-C 20 Fluoroalkoxy and amino groups (including -NH2 and mono-, di-, and tri-substituted amino groups, and their protected derivatives), or selected from -X, -OR', -SR', -NH2, -N(R')(R op ), -N(R op )3, =NR', -CX3, -CN, -NO2, -NR'C(=O)H, -NR'C(=O)R op -NR'C(=O)R op , -C(=O)R', -C(=O)NH2, -C(=O)N(R')R op -S(=O)2R op , -S(=O)2NH2, -S(=O)2N(R')R op, -S(=O)2NH2, -S(=O)2N(R')R op -S(=O)2OR', -S(=O)R op -OP(=O)(OR')(OR) op ), -OP(OH)3, -P(=O)(OR')(OR op ), -PO3H2, -C(=O)R', -C(=S)R op -CO2R', -C(=S)OR op , -C(=O)SR', -C(=S)SR', -C(=S)NH2, -C(=S)N(R')(R op )2, -C(=NR')NH2, -C(=NR')N(R')R op and its salts, wherein each X is independently selected from halogens: -F, -Cl, -Br, and -I; and wherein each R op Independently selected from C1-C 20 Alkyl, C2-C 20 alkenyl, C2-C 20 alkynyl group, C6-C 24 Aryl, C3-C 24 Heterocyclic group, C5-C 24 Heteroaryl, protecting group and prodrug moiety, or two R op Together with the heteroatoms to which they are attached, C3-C is defined. 24 Heterocyclic group; and R' is hydrogen or R op , where R op Selected from C1-C 20 Alkyl, C6-C 24 Aryl, C3-C 24 Heterocyclic group, C5-C 24 Heteroaryl groups and protecting groups.

[0282] Typically, the optional substituents present are selected from -X, -OH, and -OR. op -SH, -SR op -NH2, -NH(R) op ), -NR'(R op )2、-N(R op 3, =NH, =NR op , -CX3, -CN, -NO2, -NR'C(=O)H, NR'C(=O)R op , -CO2H, -C(=O)H, -C(=O)R op -C(=O)NH2, -C(=O)NR'R op -S(=O)2R op, -S(=O)2NH2, -S(=O)2N(R')R op , -S(=O)2NH2, -S(=O)2N(R')(R op -S(=O)2OR', -S(=O)R op -C(=S)R op , -C(=S)NH2, -C(=S)N(R')R op -C(=NR')N(R) op )2 and its salts, wherein each X is independently selected from -F and -Cl, wherein R op Typically selected from C1-C6 alkyl groups, C6-C 10 Aryl, C3-C 10 Heterocyclic group, C5-C 10 Heteroaryl and protecting group; and R' is usually independently selected from hydrogen, C1-C6 alkyl, C6-C 10 Aryl, C3-C 10 Heterocyclic group, C5-C 10 The heteroaryl group and protecting group are independently selected from R op .

[0283] More typically, the optional substituents present are selected from X, -R op 、-OH、-OR op -NH2, -NH(R) op ), -N(R op )2、-N(R op )3, -CX3, -NO2, -NHC(=O)H, -NHC(=O)R op -C(=O)NH2, -C(=O)NHR op -C(=O)N(R) op )2、-CO2H、-CO2R op -C(=O)H, -C(=O)R op -C(=O)NH2, -C(=O)NH(R) op -C(=O)N(R) op )2, -C(=NR')NH2, -C(=NR')NH(R op -C(=NR')N(R) op 2. Protecting groups and their salts, wherein each X is -F, and R op Independently selected from C1-C6 alkyl, C6-C 10 Aryl, C5-C 10 Heteroaryl and protecting group; and R' is selected from hydrogen, C1-C6 alkyl and protecting group, independently selected from R op .

[0284] In some respects, the optional alkyl substituents are selected from -NH2, -NH(R) op ), -N(R op )2、-N(R op )3, -C(=NR')NH2, -C(=NR')NH(R op ) and -C(=NR')N(R op )2, where R' and R op For example, regarding the above R' or R op Defined by any of the groups. In some of those aspects, R' and / or R op Substituents, along with the nitrogen atoms to which they are attached, provide the basic functional groups for the basic unit (BU), such as when R... op When independently selected from hydrogen and C1-C6 alkyl. Alkylene, carbocyclic, carbocyclic, aryl, arylene, heteroalkyl, heteroalkylene, heterocyclic, heterocyclic, heteroaryl and heteroarylene as described above are similarly substituted or unsubstituted, with exceptions as described in the definition of these moieties (if any).

[0285] Other optional substituents replace carbon atoms in the acyclic carbon chain of the alkyl or alkylene moiety, group, or substituent to provide C3-C. 12 Heteroalkyl or C3-C 12 Heteroalkylene, and for this purpose, other optional substituents are generally selected from -O-, -C(=O)-, -C(=O)O-, -S-, -S(=O)-, -S(=O)2-, -NH-, -NHC(=O)-, -C(=O)NH-, S(=O)2NH-, -NHS(=O)2-, -OC(=O)NH- and -NHC(=O)O (optionally substituted), wherein -NH- is an optional substituted heteroatom moiety whose hydrogen atom is replaced by an independently selected substituent derived from the group previously described with respect to optional substituents of -NH-.

[0286] Unless the context otherwise indicates or implies, as used herein, "optionally substituted heteroatom" on its own or in combination with another term refers to a heteroatom or heteroatom moiety within a functional group or other organic moiety in which the heteroatom is not further substituted or is substituted by any of the aforementioned moiety having a monovalent carbon atom, including but not limited to alkyl, cycloalkyl, alkenyl, aryl, heterocyclic, heteroaryl, heteroalkyl, and (hetero)arylalkyl, or oxidized by substitution with one or two =O substituents. In some aspects, "optionally substituted heteroatom" refers to an aromatic or non-aromatic -NH- moiety that is unsubstituted or in which the hydrogen atom is substituted by any of the aforementioned substituents. In other aspects, "optionally substituted heteroatom" refers to the nitrogen atom of the aromatic skeleton of a heteroaryl group in which the electron of the heteroatom is substituted by any of the aforementioned substituents. To encompass two of those aspects, the nitrogen heteroatom is sometimes referred to as an optionally substituted N / NH.

[0287] Therefore, in some respects, the optional substituents of the nitrogen atom are selected from the optional substituted C1-C. 20 Alkyl, C2-C 20 alkenyl, C2-C 20 alkynyl group, C6-C 24 Aryl, C5-C 24 heteroaryl, (C6-C 24 (aryl)-C1-C 20 Alkyl- and (C5-C) 24 (heteroaryl)-C1-C 20 Alkyl groups, as those terms are defined herein. In other respects, the optional substituents on the nitrogen atom are independently selected from the optional substituted C1-C groups. 12 Alkyl, C2-C 12 alkenyl, C2-C 12 alkynyl group, C6-C 24 Aryl, C5-C 24 heteroaryl, (C6-C 24 (aryl)-C1-C 12 Alkyl- and (C5-C) 24 (heteroaryl)-C1-C 12 Alkyl-, selected from C1-C8 alkyl, C2-C8 alkenyl, C2-C8 alkynyl, C6-C 10 Aryl, C5-C 10 heteroaryl, (C6-C 10 aryl)-C1-C8 alkyl- and (C5-C 10 (heteroaryl)-C1-C8 alkyl, or selected from C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C6-C 10 Aryl, C5-C 10 heteroaryl, (C6-C 10aryl)-C1-C6 alkyl- and (C5-C 10 (heteroaryl)-C1-C6 alkyl-.

[0288] When the optionally substituted nitrogen atom is a covalent attachment site of the PAB or PAB-type portion of the peptide cleavable unit and the suicide spacer unit, it is sometimes designated as J. The optional substituent of the nitrogen atom (if present) is restricted to having a monovalent sp that, compared to the unsubstituted nitrogen atom, does not adversely affect the electron-donating ability of the nitrogen atom to which it is attached. 3 The substituent on the carbon atom allows the nitrogen atom to regain its electron-donating ability when the cleavable unit is cleaved, thus enabling suicide to release the drug unit as a free drug.

[0289] Unless the context otherwise indicates or implies, the term "O-linked portion" as used herein, either alone or in combination with another term, refers to a portion, group, or substituent attached to a Markush structure or another organic part, wherein the portion, group, or substituent is directly associated with the Markush structure or another organic part via the oxygen atom of the O-linked portion. Monovalent O-linked portions are attached via monovalent oxygen and are typically -OH, -OC(=O)R. b (acyloxy group), where R b It is -H, or an optional substituted saturated C1-C 20 Alkyl, optionally substituted unsaturated C1-C 20 Alkyl, optionally substituted C3-C 20 Cycloalkyl groups, wherein the cycloalkyl moiety is optionally substituted C3-C with saturation or partial unsaturation. 20 Alkenyl, optionally substituted C2-C 20 Alkyne group, optionally substituted C6-C 24 Aryl, optionally substituted C5-C 24 heteroaryl or optionally substituted C3-C 24 Heterocyclic group, or R b It is an optional substitution of C1-C 12 Alkyl, optionally substituted C3-C 12 cycloalkyl, optionally substituted C3-C 12 alkenyl or optionally substituted C2-C 12 The alkynyl group, wherein the monovalent O-linked portion further comprises an ether group, said ether group being optionally substituted C1-C. 12 Alkoxy (i.e., C1-C) 12 Aliphatic ether moiety, wherein the alkyl moiety is saturated or unsaturated.

[0290] In other respects, the monovalent O-linked portion is selected from optionally substituted phenoxy groups, optionally substituted C1-C8 alkoxy groups (i.e., C1-C8 aliphatic ethers), and -OC(=O)R. bThe unit price portion, of which R b It is either an optionally substituted C1-C8 alkyl group (which is usually saturated) or an optionally substituted unsaturated C3-C8 alkyl group.

[0291] In other respects, the O-linked portion is selected from -OH, optionally substituted saturated C1-C6 alkyl ethers and unsaturated C3-C6 alkyl ethers, and -OC(=O)R. b The unit price portion, of which R b Typically, it is an optionally substituted C1-C6 saturated alkyl, C3-C6 unsaturated alkyl, C3-C6 cycloalkyl, C2-C6 alkenyl, or phenyl group; or the O-linked portion is selected from groups excluding -OH and / or phenyl groups, or R b The monovalent moiety is selected from optionally substituted C1-C6 saturated alkyl, C3-C6 unsaturated alkyl, and C2-C6 alkenyl groups; or the monovalent O-linked moiety is selected from saturated C1-C6 alkyl ethers, unsaturated C3-C6 alkyl ethers, and -OC(=O)R. b The unsubstituted O-linked substituents, where R b It is an unsubstituted saturated C1-C6 alkyl or an unsubstituted unsaturated C3-C6 alkyl.

[0292] Other exemplary O-linked substituents are provided by the definitions of urethanes, ethers, and carbonates disclosed herein, wherein the monovalent oxygen atom of the urethane, ether, or carbonate functional group is bonded to the Markush structure or other organic moiety associated therewith.

[0293] In other respects, the O-linked portion connected to carbon is divalent and encompasses both =O and -X-(CH2). n -Y-, where X and Y are independently S and O and the subscript n is 2 or 3, to form a spiro-ring system, wherein both X and Y are attached to the carbon.

[0294] Unless the context otherwise indicates or implies, the term “halogen” as used herein, either on its own or in combination with another term, refers to fluorine, chlorine, bromine, or iodine, and is typically -F or -Cl.

[0295] Unless the context otherwise indicates or implies, the term "protecting group" as used herein, either on its own or in combination with another term, refers to a portion that prevents or significantly reduces the ability of the atom or functional group to which it is attached to participate in an undesirable reaction. Typical protecting groups for atoms or functional groups are given in the following literature: Greene (1999), "Protective groups in organic synthesis, 3rd ed.", Wiley Interscience. Protecting groups for heteroatoms (such as oxygen, sulfur, and nitrogen) are sometimes used to minimize or avoid undesirable reactions between heteroatoms and electrophilic compounds. At other times, protecting groups are used to reduce or eliminate the nucleophilicity and / or basicity of unprotected heteroatoms. Non-limiting examples of protected oxygen are given by -OR. PR Given, where R PR The protecting group is the hydroxyl group, which is typically protected as an ester (e.g., acetate, propionate, or benzoate). Other protecting groups on the hydroxyl group prevent nucleophilic interference from organometallic reagents or other strong basic reagents. For this purpose, the hydroxyl group is typically protected as an ether, including but not limited to alkyl ethers or heterocyclic ethers (e.g., methyl ether or tetrahydropyran ether), alkoxymethyl ethers (e.g., methoxymethyl ether or ethoxymethyl ether), optionally substituted aryl ethers, and silyl ethers (e.g., trimethylsilyl ether (TMS), triethylsilyl ether (TES), tert-butyldiphenylsilyl ether (TBDPS), tert-butyldimethylsilyl ether (TBS / TBDMS), triisopropylsilyl ether (TIPS), and [2-(trimethylsilyl)ethoxy]-methylsilyl ether (SEM)). Nitrogen protecting groups include those used for primary or secondary amines, such as those in -NHR. PR or -N(R PR In )2, at least one of R PR It is a nitrogen-protecting group or two R groups. PR Together, we define the nitrogen-protecting group.

[0296] A protecting group is suitable for protection if it prevents or substantially avoids undesirable side reactions or premature loss of the protecting group under the reaction conditions required to achieve one or more desired chemical transformations elsewhere in the molecule and during the purification of the newly formed molecule if necessary, and can be removed without adversely affecting the structural or stereochemical integrity of the newly formed molecule. In some respects, suitable protecting groups are those previously described for protecting functional groups. In other respects, suitable protecting groups are those used in peptide coupling reactions. For example, suitable protecting groups for the basic nitrogen atom of an acyclic or cyclic basic unit are acid-labile ester protecting groups, such as tert-butoxycarbonyl (BOC).

[0297] Unless the context otherwise indicates or implies, the term “ester” as used herein, either on its own or in combination with another term, refers to a substituent, part or group having a -C(=O)-O- structure to define an ester functional group, wherein the carbonyl carbon atom of the structure is not directly attached to another heteroatom, but is directly attached to a hydrogen atom or another carbon atom of an organic part associated with it, and wherein a monovalent oxygen atom is attached to a different carbon atom of the same organic part to provide a lactone or to a Markush structure or some other organic part. Typically, esters, in addition to the ester functional group, contain or are composed of an organic moiety containing 1 to 50 carbon atoms, typically 1 to 20 carbon atoms, or more typically 1 to 8, 1 to 6, or 1 to 4 carbon atoms, and 0 to 10 independently selected heteroatoms (e.g., O, S, N, P, Si, but typically O, S, and N), typically 0 to 2 heteroatoms, wherein the organic moiety is bonded to a -C(=O)-O- structure (i.e., via the ester functional group) to provide a structure of the formula having an organic moiety of -C(=O)-O- or -C(=O)-O- organic moiety.

[0298] When the ester is a substituent or variable group of the Markush structure or other organic moiety associated with it, the substituent is bonded to the structure or other organic moiety via the monovalent oxygen atom of the ester functional group; therefore, it is a monovalent O-linked substituent, sometimes called an acyloxy group. In this case, the organic moiety of the carbonyl carbon attached to the ester functional group is typically C1-C. 20 Alkyl, C2-C 20 alkenyl, C2-C 20 alkynyl group, C6-C 24 Aryl, C5-C 24 heteroaryl, C3-C 24 Heterocyclic groups or substituted derivatives of any of these, for example, having 1, 2, 3, or 4 substituents; more typically C1-C. 12 Alkyl, C2-C 12 alkenyl, C2-C 12 alkynyl group, C6-C 10 Aryl, C5-C 10 heteroaryl, C3-C 10 A heterocyclic group or a substituted derivative thereof, for example having 1, 2 or 3 substituents; or a C1-C8 alkyl, C2-C8 alkenyl, C2-C8 alkynyl or phenyl or a substituted derivative thereof, for example having 1 or 2 substituents; or an unsubstituted C1-C6 alkyl or unsubstituted C2-C6 alkenyl; wherein each individually selected substituent is as defined herein for optional alkyl substituents.

[0299] Exemplary esters, by way of example and not limitation, are acetates, propionates, isopropionates, isobutyrates, butyrates, valerates, isovalerates, caproate, isohexanoate, hexanoate, heptanoate, octanoate, phenylacetate, and benzoates, or those having -OC(=O)R b The structure, where R b As defined for the substituents linked to the acyloxy group O- and generally selected from methyl, ethyl, propyl, isopropyl, 2-methyl-propyl-1-yl, 2,2-dimethyl-propyl-1-yl, propyl-2-en-1-yl, and vinyl.

[0300] Unless the context otherwise indicates or implies, the term "ether" as used herein, either alone or in combination with another term, refers to an organic moiety, group, or substituent comprising one, two, three, four, or more, typically one or two -O- (i.e., oxy) moieties (not bonded to one or more carbonyl moieties), wherein no two -O- moieties are adjacent to each other (i.e., directly attached). Typically, ethers contain the formula of an -O- organic moiety, wherein the organic moiety is as described with respect to an organic moiety bonded to an ester functional group or as described herein with respect to an optionally substituted alkyl group. When an ether is discussed as a substituent or variable group of a Markush structure or other organic moiety associated with it, the oxygen of the ether functional group is attached to the Markush formula associated with it and is sometimes referred to as an "alkoxy" group (which is an exemplary O-linked substituent). In some aspects, the O-linked substituent of an ether is a C1-C group optionally substituted with one, two, three, or four substituents, typically one, two, or three substituents. 20 Alkoxy or C1-C 12 The alkoxy group, and in other respects, is a C1-C8 or C1-C6 alkoxy group optionally substituted with one or two substituents, wherein each independently selected substituent is as defined herein for optional alkyl substituents, and in still other respects, the ether O-linked substituent is an unsubstituted saturated or unsaturated C1-C4 alkoxy group, such as, but not limited to, methoxy, ethoxy, propoxy, isopropoxy, butoxy, and allyloxy (i.e., -OCH2CH=CH2).

[0301] Unless the context otherwise indicates or implies, the term "amide" as used herein, either on its own or in combination with another term, refers to a portion having optionally substituted functional groups having RC(=O)N(R c - or -C(=O)N(R) c The structure of )2, wherein no other heteroatoms are directly attached to the carbonyl carbon of the structure, wherein each R c R is independently hydrogen, a protecting group, or an independently selected organic moiety, and R is hydrogen or an organic moiety wherein R is independently selected from... cThe organic moiety is as described for organic moieties bonded to ester functional groups or as described herein for optionally substituted alkyl groups. When an amide is discussed as a substituent or variable group of a Markush structure or other organic moiety associated with it, the amide nitrogen atom or carbonyl carbon atom of the amide functional group is bonded to that structure or other organic moiety. Amides are typically prepared by condensing an acidic halide (such as an acidic chloride) with a molecule containing a primary or secondary amine. Alternatively, amide coupling reactions well known in the field of peptide synthesis (which in some respects is carried out via activated esters containing carboxylic acid molecules) are used. Exemplary preparations of amide bonds by peptide coupling methods are provided in the following literature: Benoiton (2006) "Chemistry of peptide synthesis", CRC Press; Bodansky (1988) "Peptide synthesis: A practical textbook" Springer-Verlag; Frinkin, M. et al. "Peptide Synthesis" Ann. Rev. Biochem. (1974) 43:419-443. The reagents used in the preparation of activated carboxylic acids are provided in the following literature: Han et al. “Recent development of peptidecoupling agents in organic synthesis” Tet. (2004) 60: 2447-2476.

[0302] Therefore, in some respects, amides are prepared by reacting a carboxylic acid with an amine in the presence of a coupling agent. As used herein, "in the presence of a coupling agent" includes contacting the carboxylic acid with the coupling agent to convert the acid into its activated derivative (such as an activated ester or mixed anhydride), isolating or not isolating the resulting activated derivative of the acid, and then contacting the resulting activated derivative with an amine, either subsequently or simultaneously. In some cases, the activated derivative is prepared in situ. In other cases, the activated derivative can be isolated to remove any undesirable impurities.

[0303] Unless the context otherwise indicates or implies, when the term “carbonate” is used herein, either alone or in combination with another term, it means a substituent, part, or group containing a functional group having the structure -OC(=O)-O- (which defines the carbonate functional group). Generally, as used herein, a carbonate group comprises an organic part bonded to the -OC(=O)-O- structure, said organic part as described herein for organic parts bonded to ester functional groups (e.g., the organic part -OC(=O)-O-). When a carbonate is discussed as a substituent or variable group of a Markush structure or other organic part associated with it, one of the monovalent oxygen atoms of the carbonate functional group is attached to that structure or organic part, while the other monovalent oxygen atom is bonded to a carbon atom of another organic part, as previously described for organic parts bonded to ester functional groups or as described herein for optionally substituted alkyl groups. In this case, the carbonate is an exemplary O-linked substituent.

[0304] Unless the context otherwise indicates or implies, the term "carbamate" as used herein, either on its own or in combination with another term, means containing a compound derived from -OC(=O)N(R). c )-or-OC(=O)N(R c The )2 or -OC(=O)NH (optionally substituted alkyl)- or -OC(=O)N (optionally substituted alkyl)2 represents a substituent, part, or group in an optionally substituted urethane functional group structure, wherein one or more independently selected optional substituted alkyl groups are substituents of exemplary urethane functional groups, and are typically optionally substituted C1-C 12 Alkyl or C1-C8 alkyl, more often optionally substituted C1-C6 alkyl or C1-C4 alkyl, wherein each R c It is independently chosen, where R is independently chosen. c It is hydrogen, a protecting group, or an organic moiety, wherein the organic moiety is as described with respect to organic moiety bonded to ester functional groups or as described herein with respect to optionally substituted alkyl groups. Typically, the urethane group additionally comprises independently selected from R c The organic portion, wherein the organic portion is as described for the organic portion bonded to the ester functional group, the bond being via -OC(=O)-N(R) c The structure proceeds in the form of an organic moiety -OC(=O)-N(R) c )-or-OC(=O)-N(R cThe formula for the organic part is ). When a urethane ester is discussed as a substituent or variable group of a Markush structure or other organic part associated with it, the monovalent oxygen (O-linked) or nitrogen (N-linked) of the urethane functional group is attached to the Markush formula associated with it. The connection of the urethane substituent is explicitly stated (N- or O-linked) or implied in the context in which the substituent is referenced. The O-linked urethane esters described herein are exemplary monovalent O-linked substituents.

[0305] Unless the context otherwise indicates or implies, when the term "ligand-drug conjugate" is used herein, it refers to a construct comprising a ligand unit (L) that binds to or structurally corresponds to a target agent and a drug unit (D) that binds to or structurally corresponds to a free drug, wherein L and D are bonded to each other via a linker unit (LU), and wherein the ligand-drug conjugate is capable of selectively binding to the target portion of a target cell. The term ligand-drug conjugate (LDC) in one aspect refers to a plurality (i.e., a composition) of individual conjugate compounds that are similar or different to some extent in the number of auristatin drug units conjugated to each ligand unit and / or in the position of the ligand units conjugated to the drug units. In some aspects, the term refers to a collection (i.e., a group or plurality) of conjugate compounds having substantially the same ligand units as well as the same drug units and linker units, which in some aspects have a variable loading and / or distribution of auristatin drug linker portions attached to each antibody residue (e.g., when any two ligand-drug conjugate compounds of a plurality of such compounds have the same number of drug units but different positions of the sites attached to the ligand units). In those cases, the ligand-drug conjugate is described by the average drug loading of the conjugate compound.

[0306] The average number of drug units / ligand units in a ligand-drug conjugate composition is the average number of compounds in the ligand-drug conjugate group, sometimes denoted by the subscript p. It reflects the distribution of these compounds in some respects, with the main differences in distribution being the number of drug units conjugated to ligand units and / or their positions on the ligand units they conjugate to.

[0307] The ligand-drug conjugate compounds of the present invention, either on their own or in ligand-drug conjugate compositions, are generally represented by the structure of Formula 1:

[0308] L-[LU-(D')] p’ (1)

[0309] Or a salt thereof, in some respects being a pharmaceutically acceptable salt, wherein L is a ligand unit; LU is a linker unit; the subscript p' is an integer ranging from 1 to 24; and D' represents 1 to 4 drug units. In some respects, the ligand unit binds to or structurally corresponds to an antibody or its antigen-binding fragment, thereby being defined as an antibody ligand unit. In those respects, the antibody ligand unit is capable of selectively binding to an antigen of a target cell to subsequently release a free drug, wherein in one respect, the target antigen is a cancer cell antigen selectively recognized by the antibody ligand unit and is capable of being internalized into the cancer cell along with the bound ADC compound after said binding to initiate intracellular release of the free drug after said internalization. In any of those respects, each drug linker portion in the ligand-drug conjugate compound has the structure of Formula 1A:

[0310]

[0311] Or its salts, which are pharmaceutically acceptable salts in some respects, wherein D in each drug linker portion is a drug unit; the wavy line indicates covalent binding with L; L B This is the ligand covalently bound portion; A is the first optional extension subunit; the subscript a is 0 or 1, indicating that A is absent or present, respectively; B is an optional branch unit; the subscript b is 0 or 1, indicating that B is absent or present, respectively; L O This is the secondary connector portion; D is the drug unit, where the drug unit structurally corresponds to the free drug; and the subscript q is an integer ranging from 1 to 4.

[0312] A ligand-drug conjugate composition containing a distribution or aggregate of ligand-drug conjugate compounds is represented by the structure of Formula 1, wherein the subscript p' is replaced by the subscript p, where the subscript p is a number ranging from about 2 to about 24.

[0313] Unless the context otherwise indicates or implies, the term "ligand unit" as used herein refers to a targeting portion of a ligand-drug conjugate composition or compound that is capable of selectively binding its homologous targeting portion and binding to or corresponding to the structure of a target agent. Ligand units (L) include, but are not limited to, those derived from receptor ligands, antibodies against cell surface antigens, and transporter substrates. In some aspects, receptors, antigens, or transporters bound to the conjugate compound of the ligand-drug conjugate composition are present in greater abundance on abnormal cells compared to normal cells, thereby achieving a desired improvement in tolerability or reducing the potential occurrence or severity of one or more adverse events associated with administration of the unconjugated form of the drug. In other aspects, receptors, antigens, or transporters bound to the ligand unit of the ligand-drug conjugate compound are present in greater abundance on normal cells near the abnormal cells compared to normal cells distant from the abnormal cell site, thereby selectively exposing nearby abnormal cells to the free drug. Embodiments of the invention further describe various aspects of ligand units (including antibody ligand units).

[0314] Unless the context otherwise indicates or implies, as used herein, "target agent" means an agent capable of selectively binding to a target moiety and substantially retaining that capability when bound as a ligand unit to a ligand-drug conjugate. Thus, the ligand unit of a ligand-drug conjugate structurally corresponds to a target agent, and therefore the ligand unit is the target moiety of the conjugate. In some aspects, a target agent is an antibody or fragment thereof that selectively binds to an accessible antigen, which is specific to abnormal cells or present in a higher copy number compared to normal cells, or is an accessible antigen specific to the surrounding environment, in which tolerability is improved to some extent compared to administration of a free drug. In other aspects, a target agent is a receptor ligand that selectively binds to an accessible receptor, which is specific to abnormal cells or present in greater abundance on abnormal cells, or the receptor ligand binds to an accessible receptor on nominally normal cells specific to the environment surrounding the abnormal cells. Typically, a target agent is an antibody as defined herein that selectively binds to a target moiety of abnormal mammalian cells, more commonly, a target moiety of abnormal human cells.

[0315] As defined herein, a “targeting portion” is a portion selectively recognized by a target agent or a targeting portion of a ligand-drug conjugate (which is a ligand unit that binds to or structurally corresponds to the target agent). In some aspects, the targeting portion is present on, within, or near abnormal cells and is typically present on abnormal cells at a greater abundance or copy number than in the environment of normal cells or normal cells located away from the abnormal cell site, to provide improved tolerability relative to administration of free drug or a reduced likelihood of one or more adverse events resulting from such administration. In some aspects, the targeting portion is an antigen that can be selectively bound by an antibody, which is an exemplary target agent that binds to or structurally corresponds to an antibody ligand unit in an antibody-drug conjugate composition or a compound thereof. In other aspects, the targeting portion is a ligand of an extracellularly accessible cell membrane receptor, in some aspects of which the ligand is internalized upon binding of a homologous targeting portion by a ligand unit of the ligand-drug conjugate compound, wherein the ligand unit binds to or structurally corresponds to a receptor ligand, and in other aspects, the receptor is passively or conveniently transported after the ligand-drug conjugate compound binds to a cell surface receptor. In some respects, the target portion is present on abnormal mammalian cells or on mammalian cells in an environment specific to such abnormal cells. In some of those respects, the target portion is an antigen of the abnormal mammalian cell, and more often, a target portion of the abnormal human cell.

[0316] Unless the context otherwise indicates or implies, the term "target cell" as used herein refers to the intended cell with which the ligand-drug conjugate is designed to interact to inhibit the proliferation or other undesirable activity of abnormal cells. In some respects, target cells are overproliferating cells or overactivated immune cells, both of which are exemplary abnormal cells. Typically, those abnormal cells are mammalian cells, more commonly human cells. In other respects, target cells are located in the vicinity of abnormal cells, such that the action of the ligand-drug conjugate on nearby cells has the intended effect on the abnormal cells. For example, nearby cells might be epithelial cells specific to the abnormal vascular system of a tumor. Targeting those vascular cells with the ligand-drug conjugate compound would produce a cytotoxic or cytosuppressive effect on these cells, presumably inhibiting the delivery of nutrients to abnormal cells near the tumor. This inhibition has an indirect cytotoxic or cytosuppressive effect on abnormal cells and may also have a direct cytotoxic or cytosuppressive effect on nearby abnormal cells by releasing its drug payload in the vicinity of these cells.

[0317] Unless the context otherwise indicates or implies, the term “antibody-drug conjugate” as used herein is a subset of the ligand-drug conjugate of Formula 1, and therefore refers to a construct comprising an antibody ligand unit (L) that binds to or corresponds to an antibody or an antigen-binding fragment thereof, and a drug unit (D) that binds to or structurally corresponds to a biologically active compound (commonly referred to as a free drug), wherein L and D are bonded to each other by a linker unit (LU), wherein the antibody-drug conjugate is capable of selectively binding to a target antigen of a target cell via its targeting antibody ligand unit, in some respects said target antigen being an antigen of an abnormal cell (such as a cancer cell).

[0318] The term antibody-drug conjugate (ADC) refers, in one respect, to a plurality of (i.e., compositions of) individual conjugate compounds that are similar or different to some extent in terms of the number of drug units conjugated to each antibody ligand unit and / or the position of the antibody ligand units conjugated to the drug units. In other respects, the term refers to a distribution or collection (i.e., a population or plurality) of conjugate compounds having the same drug-linker motif and antibody ligand unit, which allows for mutant amino acid variations and different glycosylation patterns, as described herein, during antibody production from cell cultures, which in some respects have variable loading and / or distribution of drug-linker motifs attached to each antibody residue (e.g., when any two antibody-drug conjugate compounds among a plurality of such compounds have the same number of drug units but the positions of the drug-linker motifs attached to the targeting antibody ligand units are different). In those cases, the antibody-drug conjugate is described by the average drug loading of the conjugate compound.

[0319] In antibody-drug conjugate compositions having an intact drug linker portion (wherein the linker unit is unbranched), the average number of drug units / antibody ligand units or their antigen-binding fragments is an average of the antibody-drug conjugate compound population and reflects, in some respects, the distribution of these compounds, the main difference being the number of drug units conjugated to the antibody ligand units and / or their positions. When the linker unit is branched, the average reflects the distribution of the drug linker portion over the antibody-drug conjugate compound population. In either case, p is a number ranging from about 2 to about 24 or from about 2 to about 20, typically about 2, about 4, or about 10 or about 8. In other cases, p represents the number of drug units covalently bound to a single antibody ligand unit of the antibody-drug conjugate in the antibody-drug conjugate compound population, where, in some respects, the compounds in the population differ primarily in the number and / or position of drug units or drug linker portions. In this case, p is designated as p' and is an integer ranging from 1 to 24 or from 1 to 20, typically from 1 to 12 or from 1 to 10, more typically from 1 to 8. In other respects, virtually all available reactive functional groups of the antibody target form covalent bonds with the drug linker portion to provide antibody ligand units attached to the maximum number of drug linker portions, so that the p value of the antibody-drug conjugate composition is the same as or nearly the same as the p' value of each antibody-drug conjugate compound in the composition, and thus only a small number of antibody-drug conjugate compounds with lower p' values ​​(if any) are present, as can be detected using appropriate chromatographic methods (such as electrophoresis, HIC, reversed-phase HPLC, or size exclusion chromatography).

[0320] In some aspects, the average number of drug units or drug linker moieties / antibody ligand units in the formulation derived from the conjugation reaction is characterized by conventional chromatographic methods combined with mass spectrometry as described above. In other aspects, the quantitative distribution of the conjugated compound is determined, expressed as a p' value. In those cases, the separation, purification, and characterization of homogeneous antibody-drug conjugated compounds from antibody-drug conjugated compositions where p' is a specific value, and antibody-drug conjugated compounds loaded with other drug units or drug linker moieties, can be achieved, for example, by the chromatographic methods described above.

[0321] Unless the context otherwise indicates or implies, the term "drug linker compound" as used herein refers to a compound having a drug unit covalently attached to a linker unit precursor (LU'), wherein LU' comprises L B The latter is sometimes referred to as a ligand covalently bound precursor (L). B The ') portion contains reactive or activatable functional groups, wherein the activated reactive or activatable functional groups can react with the target agent to covalently bind to the ligand in the ligand covalently bound portion (L). BCovalent bonds are formed between the ligand unit and the ligand conjugate compound of Formula 1, thereby providing a drug linker portion of Formula 1A for the ligand drug conjugate compound of Formula 1, particularly a covalent bond with an antibody ligand unit that binds to or structurally corresponds to an antibody.

[0322] The drug connector compounds of the present invention generally have the general formula of Formula I:

[0323] LU'-(D')(I)

[0324] Or a salt thereof, which in some respects is a pharmaceutically acceptable salt, wherein LU' is a precursor of LU; and D' represents 1 to 4 drug units, wherein the drug linker compound is further defined by the structure of formula IA:

[0325]

[0326] Where L B It contains reactive or activatable functional groups, and the remaining variable groups are as defined in Formula 1A.

[0327] Unless the context otherwise indicates or implies, the term "cytotoxic agent" as used herein is a compound capable of inducing cell death or inhibiting cell proliferation or prolonging cell survival in vitro or in vivo, typically abnormal mammalian cells. Cell inhibitors exert their therapeutic effects primarily by inhibiting the proliferation of abnormal cells rather than by directly killing cells, and are covered within the definition of cytotoxic agents. In some respects, cytotoxic agents are free drugs resulting from the release of pharmaceutical units from antibody-drug conjugates.

[0328] Unless the context otherwise indicates or implies, the phrase “drug unit” as used herein refers to a drug residue covalently attached to a linker unit (LU) in the drug linker portion of a ligand-drug conjugate (LDC), or covalently attached to a linker unit precursor (LU') of a drug linker compound and releasable as a free drug from the drug linker portion or drug linker compound. The free drug may be directly incorporated into the drug unit, or components of the free drug may be covalently attached to the LU or LU' or intermediates thereof, followed by further refinement to complete the structure of the drug unit. When the term “drug” is used alone or in combination with another term (such as “drug unit”) herein, it is not intended to imply that the compound is approved, is eligible for approval, or is expected to be approved for medical or veterinary treatment by a government agency.

[0329] In some respects, the free drug bound to the drug unit is a cytotoxic compound, typically a drug having a secondary aliphatic amine as a conjugate, and includes compounds such as auristatin as defined herein.

[0330] Unless the context otherwise indicates or implies, as used herein, “aurestatin drug,” “aurestatin compound,” and similar terms refer to peptide-based tubulin disruptors having cytotoxic, inhibitory, or anti-inflammatory activity, which contain dolaproline and dolaisoleucine residues or related amino acid residues.

[0331] Some exemplary auristatin has D E Or D F Structure:

[0332]

[0333] Where Z is -O-, -S-, or -N(R) 19 )-, where R 10 -R 21 As defined in the implementation scheme of the auristatin drug unit, and the indicated nitrogen atom It is a secondary amine (e.g., R) 10 and R 11 One of them is hydrogen, and the other is a nitrogen atom (-CH3). In those respects, auristatin is bound to the drug unit via a carbamate functional group containing the nitrogen atom. This carbamate functional group is an exemplary second spacer subunit (Y') and is capable of self-destruction, which is then attached to a PAB or PAB-type spacer subunit (Y) such that the subscript y in any of the drug linker portions described herein is 2.

[0334] Other exemplary auristatins include, but are not limited to, AE, AFP, AEB, AEBV, MMAF, and MMAE, as well as those further described in embodiments of the present invention. The synthesis and structure of auristatins are described in the following documents: U.S. Patent Application Publications Nos. 2003-0083263, 2005-0238649, 2005-0009751, 2009-0111756, and 2011-0020343; International Patent Publications Nos. WO 04 / 010957, WO 02 / 088172, and U.S. Patents Nos. 7,659,241 and 8,343,928. The structures and methods of synthesis disclosed therein are expressly incorporated herein by reference.

[0335] Unless the context otherwise indicates or implies, the phrase "its salt" as used herein refers to the salt form of a compound (e.g., a drug, a drug connector compound, or an LDC compound). The salt form of a compound is one or more internal salt forms and / or involves the inclusion of another molecule, such as an acetate ion, a succinate ion, or other counterions. The counterions in the salt form of a compound are typically organic or inorganic moieties that stabilize the charge on the parent compound. The salt form of a compound has one or more charged atoms in its structure. In cases where multiple charged atoms are part of the salt form, multiple counterions and / or multiple charged counterions are present. Therefore, the salt form of a compound typically has one or more charged atoms corresponding to the non-salt form of the compound, as well as one or more counterions. In some aspects, the non-salt form of a compound contains at least one amino or other basic moiety, thus yielding an acid addition salt with a basic moiety in the presence of an acid. In other aspects, the non-salt form of a compound contains at least one carboxylic acid group or other acidic moiety, thus yielding a carboxylate or other anionic moiety in the presence of a base.

[0336] Exemplary counter anions and counter cations in the form of complex salts include, but are not limited to, sulfates, trifluoroacetates, citrates, acetates, oxalates, chlorides, bromides, iodides, nitrates, bisulfates, phosphates, acid phosphates, isonicotinates, lactates, salicylates, acid citrates, tartrates, oleates, tannates, pantothenates, bitartrates, ascorbic acid salts, succinates, maleates, gentianates, fumarates, gluconates, glucurons, glycosides, formates, benzoates, glutamates, methanesulfonates, ethanesulfonates, benzenesulfonates, p-toluenesulfonates, and bis(hydroxynaphthyl)ates (i.e., 1,1'-methylenebis-(2-hydroxy-3-naphthylcarbamate)).

[0337] The choice of the salt form of the compound depends on the properties that the pharmaceutical product must exhibit (including sufficient water solubility at various pH values), the intended one or more routes of administration, the crystallinity with flow characteristics and low hygroscopicity suitable for handling (i.e., water absorption and relative humidity), and the desired shelf life obtained by determining the chemical stability and solid-state stability under accelerated conditions (i.e., to determine degradation or solid-state changes when stored at 40°C and 75% relative humidity).

[0338] "Pharmaceutically acceptable salt" is a salt form of a compound suitable for administration to a subject as described herein, and in some respects includes countercations or counteranions, as described in PHStahl and CGWermuth, eds., Handbook of Pharmaceutical Salts: Properties, Selection and Use, Weinheim / Zürich: Wiley-VCH / VHCA, 2002.

[0339] Unless the context otherwise indicates or implies, when the term “antibody” is used in its broadest sense herein, it specifically encompasses complete monoclonal antibodies, polyclonal antibodies, monospecific antibodies, multispecific antibodies (e.g., bispecific antibodies), and antibody fragments exhibiting desired biological activity, requiring the antibody fragment to have a desired number of sites for attaching a desired number of drug-linker portions and to be able to specifically and selectively bind to target cancer cell antigens. Antibodies are naturally tetramers and typically consist of two pairs of identical immunoglobulin chains, each pair having one light chain and one heavy chain. In each pair, the variable regions (VL and VH) of the light and heavy chains together are primarily responsible for binding to the antigen. The variable domains of the light and heavy chains consist of a framework region interrupted by three hypervariable regions (also known as “complementarity-determining regions” or “CDRs”). In some respects, the constant regions are recognized by and interact with the immune system (see, for example, Janeway et al., 2001, Immunol. Biology, 5th ed., Garland Publishing, New York) to perform effector functions. Antibodies include any isotype (e.g., IgG, IgE, IgM, IgD, and IgA) or its subclasses (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2). Antibodies can be derived from any suitable species. In some respects, antibodies are human or murine antibodies. Such antibodies include human antibodies, humanized antibodies, or chimeric antibodies.

[0340] In some respects, the antibody is in a reduced form, in which the antibody has undergone reduction of its hinge disulfide bonds. The antibody is then bound to the antibody-drug conjugate as an antibody ligand unit by reacting one or more cysteine ​​thiols obtained from this reduction with a suitable electrophilic reagent of the drug linker compound. This results in the covalent binding of the drug linker moiety to the antibody ligand unit or linker intermediate, which is further refined into the final form of the drug linker moiety.

[0341] As used herein, “monoclonal antibody” refers to an antibody obtained from a substantially homogeneous population of antibodies—that is, a single antibody—that constitutes a population identical in all respects except for naturally occurring mutations (which may be present in small amounts) and / or contains differences in glycosylation patterns. Monoclonal antibodies are highly specific, targeting a single antigenic site. The modifier “monoclonal” indicates the characteristic of antibodies obtained from a substantially homogeneous population of antibodies and should not be interpreted as requiring the production of antibodies through any particular method.

[0342] Unless the context otherwise requires, the terms "selective binding" and "selectively bind" as used herein refer to the ability of an antibody-drug conjugate, its fragments, or antibody ligand units to bind in an immunoselective and specific manner to homologous cancer cell antigens rather than a large number of other antigens. Typically, the antibody or its antigen-binding fragment binds at least about 1 x 10-1 -7 M, preferably about 1x10 -8 M to 1x10 -9 M, 1x10 -10 M or 1x10 - 11 M has an affinity for binding its target cancer cell antigens and binds the predetermined antigen with an affinity at least twice as great as that for binding non-specific antigens (e.g., BSA, casein) rather than closely related antigens, wherein the affinity is substantially preserved when the antibody or its antigen-binding fragment corresponds to or binds to an antibody-drug conjugate as an antibody ligand unit.

[0343] Unless the context otherwise indicates or implies, the term "antigen" as used herein refers to a portion capable of specifically binding to an unconjugated antibody or its antigen-binding fragment, or to an antibody-drug conjugate compound, comprising an antibody ligand unit that binds to or structurally corresponds to the unconjugated antibody. In some aspects, antigens are extracellularly accessible cell surface proteins, glycoproteins, or carbohydrates, particularly proteins or glycoproteins, preferentially displayed by abnormal cells compared to normal cells located away from abnormal cell sites. In those aspects, cell surface antigens are capable of internalization upon selective binding by the conjugate compound of the antibody-drug conjugate composition. Following internalization, intracellular processing of the linker unit of the antibody-drug conjugate compound of the composition releases the drug unit as a free drug. Antigens associated with overproliferating cells (which are cell surface accessible to the antibody-drug conjugate compound) include, for example, but not limited to, cancer-specific antigens as described herein.

[0344] Typically, antigens are associated with cancer. In some aspects, antigens are preferentially displayed by cancer cells compared to normal cells that are not localized to abnormal cells; specifically, the cancer cells displaying antigens are mammalian cancer cells. In other aspects, cancer cell antigens are extracellularly accessible antigens preferentially displayed by nearby normal cells, which are specific to the cancer cell environment, compared to normal cells far from the cancer cell site. For example, nearby cells might be epithelial cells specific to the abnormal vascular system of a tumor. Antibody-drug conjugates targeting those vascular cells will produce cytotoxic or cytosuppressive effects on these cells, presumably inhibiting the delivery of nutrients to cancer cells near the tumor. This inhibition will indirectly produce cytotoxic or cytosuppressive effects on cancer cells, and may also produce direct cytotoxic or cytosuppressive effects on nearby cancer cells after immunoselective binding to the antibody-drug conjugate (ADC) compound and after the release of the drug unit as a free drug. In any of these aspects, cell surface antigens can be internalized to allow the free drug to be delivered intracellularly to the target cell after release from the conjugate.

[0345] Preferred internalizable antigens are those expressed on the surface of cancer cells, with copy numbers of 10,000 or more, 20,000 or more, or 40,000 or more per cell. Cancer cell-associated antigens that are cell surface accessible to and internalizable for ADCs include antigens expressed on certain cancer cells of Hodgkin lymphoma (particularly Reed-Sternberg cells, such as Karpas 299 cells) and advanced lymphomas (sometimes referred to as Ki-1 lymphoma). Other antigens include cancer cells of renal cell adenocarcinoma (e.g., 789-O cells), cancer cells of B-cell lymphomas or leukemias (including non-Hodgkin lymphoma, chronic lymphocytic leukemia (CLL), and acute lymphoblastic leukemia (ALL)) (e.g., CHO cells), cancer cells of acute myeloid leukemia (AML) (e.g., HL-60), and certain transporter receptors commonly expressed on these and other cancer cells.

[0346] Unless the context otherwise indicates or implies, the term "connector unit" as used herein refers to the organic portion in a ligand-drug conjugate that covalently attaches the insert drug unit and the ligand unit (L) (these terms are defined herein), or the organic portion in a drug-connector compound that covalently attaches the drug unit and has reactive functional groups or portions thereof, for interacting with the target agent to form a covalent bond between the L (which binds to or structurally corresponds to the target agent) and the connector unit (LU). Because the connector unit in a drug-connector is capable of forming such a bond, it is considered a precursor to the connector unit in a ligand-drug conjugate and is sometimes denoted as LU'. The connector unit comprises a primary connector (L... R ) and secondary connector (LO The secondary connector is inserted into the drug connector portion of the ligand-drug conjugate compound. R L between and D or inserted into drug connector compounds R Between D and L, in the latter case it can be represented as L. R ', to clearly indicate that it is L in the ligand drug conjugate. R The precursor.

[0347] Unless the context otherwise indicates or implies, the term "primary linker" as used herein refers to the essential component of the linker unit (LU) in a ligand-drug conjugate, which is covalently attached to the ligand unit and the remainder of the LU. R One component of ) is ligand covalently bound (L B The ) section provides self-stabilizing properties in some aspects of the ligand-drug conjugates (LDCs) and drug linker compounds described herein. SS ) connector, thereby defining L SS Primary connector, and provides LDC with other features that can be obtained from L SS The derived self-stabilized (L) S ) connector, thereby defining L S Primary joints, these terms are further described herein. A primary joint optionally contains a branch element (B) and a first optional extension sub-unit (A), depending on the values ​​of the subscripts a and b in Equation 1A, provided that when L R It is L SS or L S A exists in the first-level connector.

[0348] LDC or drug connector compound L SS The primary junction is characterized by succinimide (M) that is close to the basic unit. 2 ) or maleimide (M 1 The L in the LDC composition or its compounds is a portion thereof. S The primary connector is characterized by succinamide (M) which is close to the alkaline unit. 3 ( ) part. The L of the present invention SS or L S The primary connector is further characterized by a first optional extension subunit (A), which exists and includes M. 1 Or M 2 maleimide or succinimide ring system imide nitrogen or M 3 Optional substitution of C1-C amide-nitrogen bond 12The alkylene moiety is substituted in some respects with an acyclic basic unit and may be further substituted by optional substituents, or may be substituted in other respects and combined with an optional substituted cyclic basic unit.

[0349] L in drug linker compounds SS The ligand covalently bound precursor of the primary linker (sometimes shown as L) SS ', to clearly indicate that it is L in the ligand drug conjugate. SS maleimide (M) (precursor) 1 The L group can react with the sulfur atom of the reactive thiol functional group of the target agent to produce a ligand-drug conjugate. SS The sulfur-substituted succinimide moiety (M) in the covalently bound ligand of the primary linker 2 In this context, the thiosubstituent is a ligand unit that binds to or structurally corresponds to the target agent. In cases where the target agent is an antibody or its antigen-binding fragment, the antibody binds to the sulfur atom of a cysteine ​​residue obtained through disulfide bond reduction or introduced through genetic engineering. 2 Bonding. Therefore, the antibody or its antigen-binding fragment binds to L. SS The primary linker is covalently bonded and serves as the antibody ligand unit. L SS M in the primary connector 2 Subsequent hydrolysis produces L S Primary connector, where M 2 Conversion to succinamide moiety (M) 3 This connector portion can be used as two positional isomers (M). 3A and M 3B The presence of a mixture depends on the relative reactivity of the two carbonyl groups in the succinimide ring system to hydrolysis.

[0350] Unless the context otherwise indicates or implies, the term "ligand covalently bound portion" as used herein refers to a portion of the linker unit (LU) in a ligand-drug conjugate that interconnects the ligand unit (L) and the remaining portion of the linker unit, and from the corresponding ligand covalently bound precursor (L') of the linker unit precursor (LU') in the drug linker compound. B It is obtained through the reaction between L and a target agent (such as an antibody or its antigen-binding fragment). For example, when L B 'Contains maleimide moiety (M) 1 When this part reacts with the reactive thiol functional groups of the target agent, L B 'Converted to ligand covalent binding (L BThe sulfur-substituted succinimide moiety is obtained by converting the target into a sulfur-substituted succinimide portion. When the target is an antibody or its antigen-binding fragment, the sulfur substituent contains a sulfur atom from the antibody ligand unit, which is provided in some respects by interchain disulfide bond reduction or by cysteine ​​residues obtained through genetic engineering.

[0351] In another example, when L B When an activated carboxylic acid functional group is present, the reaction between this functional group and a reactive amino group of the target agent (such as the ε-amino group of a lysine residue in an antibody or its antigen-binding fragment) converts the functional group into an amide, wherein the amide functional group generated by this reaction is in L B It is shared between the attached ligand unit (which is an antibody ligand unit in the case of an antibody or antigen-binding fragment). Other L... are described in embodiments of the invention. B Parts and their components containing L B 'Partial transformation.' In yet another example, a target agent with a reactive amino group is derived using a bifunctional molecule to provide an intermediate that, in some cases, generates a reactive thiol functional group, which interacts with L... B 'Partial condensation. As a result of condensation, the resulting L...' B Partially attributed to bifunctional molecules and L B 'atom'.

[0352] The "ligand-covalently binding precursor moiety" is a component of the linker unit of a drug linker compound or its intermediate, containing reactive or activatable functional groups. During the preparation of ligand-drug conjugates (LDCs) (including antibody-drug conjugates (ADCs)), the activated reactive or activatable functional groups can covalently bind to the target agent (such as an antibody or its antigen-binding fragment). Thus, the ligand-binding precursor moiety (L...)... B The portion is converted into ligand covalently bound (L) B ( ) part. In some respects, L B The portion has a functional group capable of reacting with a nucleophilic or electrophilic agent native to the antibody or its antigen-binding fragment, or is introduced into the antibody or antigen-binding fragment by chemical transformation or genetic engineering (see above) to convert it into an antibody ligand unit. In some of those aspects, the nucleophile is the N-terminal amino group of the light or heavy chain of the antibody or its antigen-binding fragment, or the ε-amino group of a lysine residue of the light or heavy chain.

[0353] In other respects, nucleophiles are thiol groups of cysteine ​​residues introduced via genetic engineering into the light or heavy chain of an antibody or its antigen-binding fragment, or from chemically reduced disulfide bonds between the chains of the antibody or antigen-binding fragment. In still other respects, electrophiles are aldehydes introduced via the selective oxidation of the sugar portion of the glycan component of an antibody or its antigen-binding fragment, or ketones derived from non-natural amino acids introduced into the light or heavy chain of an antibody or its antigen-binding fragment using genetically engineered tRNA / tRNA synthetase. Behrens and Liu, “Methods for site-specific drug conjugation to antibodies” mAB (2014) 6(1):46-53, reviewed those and other methods for introducing reactive functional groups into antibodies to provide conjugation sites.

[0354] Unless the context otherwise indicates or implies, as used herein, "secondary joint," "secondary joint portion," and similar terms refer to the organic portion of the joint unit (LU), where the secondary joint (LU) O ) is a component of LU; LU enables the drug unit to connect with the primary linker (L R Interconnected and containing ligand covalently bound (L) B The first optional extended subunit and / or optional branching unit (B) and, in some respects, provide self-stabilization (L) of ligand-drug conjugates (LDCs) (such as antibody-drug conjugates (ADCs)) or drug linker compounds that can be used to prepare the conjugates. SS ) Level 1 connector, or L SS Hydrolysis provides self-stabilized (L) compounds for LDC / ADC. S ) Level 1 connector. In L R For L SS or L S In this case, there exists a first optional extended subunit. In those respects, L R Attached to L by a heteroatom or functional group from the first optional extended subunit (A) O .

[0355] Secondary linkers in ligand-drug conjugates or drug linker compounds typically have the following structures:

[0356]

[0357] When the subscript b is 0, the wavy line adjacent to A' represents L. O The site covalently attached to the primary connector; the wavy line adjacent to Y indicates L. OThe site covalently attached to the drug unit; A' is a second optional spacer subunit, or a subunit of a first optional extension subunit that is present in some respects, with subscript a' being 0 or 1, indicating the absence or presence of A', respectively; Y is a spacer subunit, with subscript y being 0, 1, or 2, indicating the absence or presence of one or both spacer subunits, respectively; and W is a peptide-cleavable unit, wherein the recognition site provided by the peptide-cleavable unit has greater overall selectivity for proteases in tumor tissue homogenates compared to proteases in normal tissue homogenates, wherein the tumor tissue contains target cancer cells and the normal tissue contains non-target normal cells, for which off-target cytotoxicity of the ligand-drug conjugate at least partially results in adverse events typically associated with administration of therapeutically effective doses to mammalian subjects in need. When subscript b is 0, A' becomes a subunit of A when present, in which case the secondary linker has -WY. y - The structure. In any of those aspects, W, Y, and D are arranged in a linear configuration relative to the remainder of LU / LU', such as -WY. y -D is shown, where W is a peptide-cleavable unit, and the subscript y is 0, 1, or 2. When the subscript y is 1 or 2, after protease cleavage, the suicide spacer unit attached to W commits suicide to release D or Y'-D. If a second spacer unit (Y') is present, it is decomposed to complete the release of D as a free drug.

[0358] Secondary linkers (L) that are bonded to D in the linker unit (as illustrated when only one drug unit is attached to LU, where W is a peptide-cleavable unit) O It is usually represented by the following structure:

[0359] When the subscript b is 1; or

[0360] Because A' a’ The subunit is considered as the first optional extended subunit when the subscript b is 0 and the subscript a' is 1;

[0361] Where D is the drug unit, and the remaining variable groups are as described in this paper for L. O Defined;

[0362] Furthermore, the drug connector portion or drug connector compound containing this secondary connector typically has structures of Formula 1B and Formula IB, respectively:

[0363]

[0364]

[0365] Where L BIt is the ligand covalently bound portion as defined in this paper, which is the primary linker (L) of the linker unit (LU) of the drug linker portion of the ligand-drug conjugate compound. R ) components; L B The ' is the ligand covalently bound portion as defined herein, which is the primary linker (L') of the linker unit (LU') in the drug linker compound. R The components of a drug linker compound, when used to prepare a ligand-drug conjugate, are sometimes referred to as L components specifically for the ligand-drug conjugate. R L B The precursors of the ligand covalently bound to LU, the primary linker precursors, and the linker unit precursors; A is the first optional extension subunit; the subscript a is 0 or 1, indicating that A is absent or present, respectively; B is an optional branching unit, the subscript b is 0 or 1, indicating that B is absent or present, where when the subscript b is 0, the subscript a is 1, and the subscript a' is 1, A' is a subunit of A; the subscript q ranges from 1 to 4, where L B / L B 'And A and B (if they exist) are L' R / L R The components are defined as follows: when subscript b is 1, subscript q ranges from 2 to 4; when subscript b is 0, subscript q is 1; and the remaining variable groups are as described in this paper for L. O Defined.

[0366] Unless the context otherwise indicates or implies, "maleimide moiety" as used herein refers to the component of a primary linker in a pharmaceutical linker compound, which in some respects is a component of a self-stabilizing linker, wherein the primary linker is sometimes denoted as L. R 'or L SS 'To clearly indicate that it is L in ligand drug conjugates' R / L SS Precursor. Maleimide moiety (M 1 The sulfur atom of the reactive thiol functional group can participate in Michael addition (i.e., 1,4-conjugated addition) through a target (such as an antibody or its antigen-binding fragment) to provide a sulfur-substituted succinimide (M). 2 The ) part, wherein the thiosubstituent is a ligand unit that binds to or corresponds to the structure of the target agent, as illustrated herein with respect to the antibody ligand unit of an antibody-drug conjugate composition or a compound thereof. The M of the drug linker compound 1 The remainder of the primary linker is attached via its imide nitrogen atom, typically to the first optional extension subunit (A) (when M...). 1 Part is L SS When the component is present, it is attached to the secondary connector (L). O (If neither A nor B exists).

[0367] Besides the imide nitrogen atom, M 1 Partially unsubstituted, but can be asymmetrically substituted at the cyclic double bond in its maleimide ring system. This substitution can lead to a regiochemically preferred conjugated addition of the sulfur atom of the reactive thiol functional group of the target agent to the less hindered or more electron-deficient double-bonded carbon atom of the maleimide ring system (depending on which is more dominant). This conjugated addition produces succinimide (M 2 The part is substituted with sulfur by ligand units through the sulfur atom of the thiol functional group provided by the target agent.

[0368] Unless the context otherwise indicates or implies, "succinimide moiety" as used herein refers to the ligand covalently bound to the primary linker (L... B This is a type of component that is also a component of the linker unit of ligand-drug conjugates (such as antibody-drug conjugates), and is composed of the sulfur atom of the reactive thiol functional group of the antibody or its antigen-binding fragment and the maleimide moiety (M). 1 (It is a drug connector compound or its M-containing compound) 1 The ligand covalently binds to the precursor (L) in the intermediate. B It is produced by Michael addition of a maleimide ring system of a certain type (of the ') portion. Therefore, succinimide (M) 2 The succinimide ring system comprises a sulfur-substituted succinimide ring system having an imide nitrogen atom substituted by a remaining portion of a primary linker, the remaining portion of which is typically a first optional extended subunit (A). In some aspects, the nitrogen atom is substituted by an optional C1-C1 subunit constituting the first optional extended subunit (A). 12 The alkylene moiety is attached to the present unit. When the primary linker is a self-stabilizing linker, the alkylene moiety binds the cyclic basic unit to a first optional extended subunit; the first optional extended subunit is present or substituted with an acyclic basic unit as described elsewhere, and optionally otherwise substituted, and has an M that is optionally substituted with one or more substituents in its succinimide ring system. 2 Part (which may exist in M) 1 (Precursor).

[0369] Therefore, C1-C with optional substitution of A and optional combination with [HE] (which is an optional hydrolysis-enhancing unit) 12 The alkylene moiety is directly covalently attached to an optional secondary connector (L). O (When the subscript b is 0, it exists) or indirectly covalently attached to L via the drug linker portion of Formula 1B or the drug linker compound of Formula IB, -[HE]-B- (when the subscript b is 1). OIn the cases where subscript b is 0, subscript a is 1, and subscript a' is 1, A is represented by the formula -A1[HE]-A2-, where A1 is the first subunit of A and contains C1-C with optional substitutions in optional combinations with HE. 12 The alkylene moiety, and A' (previously denoted as L) O The component) becomes A2 (now the second subunit of A). In those cases where subscript b is 1, subscript a is 1, and subscript a' is 1, A' is a component of the secondary connector, and A is a single unit optionally combined with [HE] or optionally contains two subunits, consisting of -A[HE]-A O - indicates that A O It is an optional subunit of A. When A O When it exists, A is also represented by the formula -A1[HE]-A2-.

[0370] When the self-stabilizing linker (L) in the ligand drug conjugate compound SS When present in ), due to the presence of basic functional groups of acyclic or cyclic basic units nearby, sulfur-substituted succinimides (M) 2 The hydrolysis of the succinimide ring system is pH-controlled, and in some cases, due to asymmetric substitution resulting from thiosubstituents, the hydrolysis provides a self-stabilizing linker (L). S ) in succinic acid-amide (M 3 The positional chemical isomers of M. The relative amounts of those isomers will be at least partially attributed to M. 2 The difference in reactivity between the two carbonyl carbons, at least in part attributable to M 1 Any one or more substituents present in the precursor. Compared to the controlled hydrolysis provided by the basic unit, when L R It has M that is highly variable but does not contain basic units. 2 In some cases, some degree of hydrolysis is also expected.

[0371] In some respects, M 2 The optional substituents on the succinimide ring system are absent, and the first optional extended subunit is present and contains an optionally substituted C1-C at a position away from its attachment site to the imide nitrogen atom, which is optionally attached to [HE] (which is an optional hydrolysis-enhancing unit). 12 Alkylene moiety. In this respect, A is a single unit or further includes A' (which is an optional subunit of A present when subscript b is 0 and subscript a' is 1), and is attached to [HE] which is also present, such that A has the formula -A[HE]-A'-, or when subscript b is 1 and subscript a' is 1, A' is a component present in a secondary linker, and therefore A is given by the formula -A[HE]-A O -express.

[0372] Unless the context otherwise indicates or implies, "succinic acid-amide moiety" as used herein refers to the self-stabilized linker (L) of the linker unit within a ligand-drug conjugate (such as an antibody-drug conjugate). S It is a component of ) and has the structure of succinamide hemiacid residues (whose amide nitrogen is L S Another component is substituted), wherein the component is typically present as a first optional extended subunit (A) or its subunit and contains C1-C optionally attached to [HE]. 12 Alkylene moiety. When subscript b is 0 and subscript a is 0 or 1, the possible structures of A are represented by the formula -A[HE]-A'. a’ - indicates that A', which was previously associated with the secondary connector, does not exist, so the subscript a' is 0, or A' exists as a subunit of A when the subscript a' is 1. When this subunit exists, A is represented by the formula A1[HE]-A2-, where A1 is the first subunit of A and includes optionally substituted C1-C, which is optionally attached to [HE]. 12 The alkylene moiety, and A2 (previously denoted as A') is the second subunit of A. When subscript b is 1 and subscript a is 1, the possible structures of A are given by the formula -A[HE]-A. O - indicates that A O When it exists, it is an optional subunit of A. When this subunit does not exist, A is a single discrete unit, while when A... O When present, A is represented by the formula A1[HE]-A2-, where A1 is the first subunit of A and includes optionally substituted C1-C that is optionally attached to [HE]. 12 The alkylene moiety, and A2 (previously referred to as A) O ) is the second subunit of A.

[0373] In some respects, the alkylene moiety comprises a cyclic basic unit and in others is substituted with an acyclic basic unit, and in either respect is optionally substituted in other ways, wherein succinamide (M 3 The ) portion is further replaced by LS-, where L is a ligand unit (e.g., an antibody ligand unit) that binds to or structurally corresponds to a target agent (e.g., an antibody or its antigen-binding fragment), and S is a sulfur atom from the target agent, antibody, or antigen-binding fragment. M 3 Partially composed of succinimide (M) in the self-stabilizing primary linker 2 The succinimide ring system is generated by partial sulfur substitution, and the self-stabilizing primary linker undergoes the breaking of one of its carbonyl-nitrogen bonds via basic unit-assisted hydrolysis.

[0374] Therefore, M 3Some have free carboxylic acid functional groups and amide functional groups (the latter's nitrogen atom is attached to the remainder of the primary linker), and are LS-substituted on the α-carbon of the carboxylic acid or amide functional group, depending specifically on its M. 2 The hydrolysis site of the precursor. Unbound by theory, it is believed to produce M. 3 The above-mentioned partial hydrolysis provides a linker unit (LU) in the ligand drug conjugate, which is unlikely to cause the conjugate to suffer premature loss of the targeting ligand unit (L) by eliminating thio substituents.

[0375] Unless the context otherwise indicates or implies, as used herein, "self-stabilizing linker" refers to the primary linker of the linker unit (LU) in a ligand-drug conjugate (such as an antibody-drug conjugate), which has M 2 The components; or the primary linker of the linker unit precursor (LU') in a drug linker compound, which has M 1 The component, wherein the component may be specified as L SS ', to indicate that it is L in LDC SS M 2 The precursor of the component. The self-stabilizing junction is then converted into the corresponding self-stabilized junction (L) under controlled hydrolysis conditions. S L SS The basic unit components promote this hydrolysis, therefore containing L SS The LDC / ADC now includes L through its connector unit (LU). S The connector unit is more resistant to premature loss of its ligand unit. In addition to its M... 1 Or M 2 In addition to some, L SS The primary connector further includes a first optional extension subunit (A) that is required to exist, wherein A includes C1-C optionally combined with [HE]. 12 The alkylene moiety, wherein A further comprises an optional subunit (A O When (it exists when the index b is 1), the combination is sometimes designated A1, or when the index b is 0 and the index a' is 1, A further includes A', where, for either of the two values ​​of index b, the additional subunit is designated A2. When A may exist as a single discrete unit or as two discrete units, these two possibilities are given by equation -A[HE]-A O -(when the subscript b is 1) or A[HE]-A' a’ (When the subscript b is 0) means that for any value of the subscript b, it becomes -A[HE]- or -A1[HE]-A2-, depending on whether a second subunit exists. In L SSIn any variant of A within the range, the alkylene portion is either bound to a cyclic basic unit or substituted with an acyclic basic unit, and optionally substituted in other ways.

[0376] Therefore, when the primary linker of a drug linker compound is L SS (Sometimes displayed as L) SS ', to indicate that it is L in the ligand drug conjugate. SS When the precursor is present, the primary connector contains a first optional extension subunit (A) and maleimide (M). 1 In this part, the target agent will attach, and the target agent provides antibody ligand units in the presence of an antibody or its antigen-binding fragment. In those respects, L SS A of C1-C 12 The alkylene portion is attached to M 1 The remaining portion of the maleimide ring system's imide nitrogen and the connector unit, the latter's attachment optionally via [HE]-A O -B- (when the subscript b is 1) or [HE]-A' a’ - (when the index b is 0) occurs, depending on whether A exists. O / A' and [HE]. In some of those aspects, [HE] is the hydrolysis-enhancing moiety and consists of or contains optional substituted electron-withdrawing heteroatoms or functional groups; in some aspects, besides BU, [HE] can enhance the corresponding L in LDC / ADC compounds. SS M in part 2 Partial hydrolysis rate. After binding the drug linker compound to the LDC / ADC compound, L... SS Currently, succinimide (M) containing ligand units that have undergone sulfur substitution is present. 2 The attachment of the ligand unit to its drug linker portion (i.e., the attachment of the ligand unit to its drug linker portion via the sulfur atom of the reactive thiol functional group of the target agent and M) 1 (This occurs via Michael addition to the maleimide ring system).

[0377] In some respects, the cyclic basic unit (cBU) structurally corresponds to the acyclic basic unit by formally cyclizing the basic nitrogen of the unit, such that the cyclic basic unit structure is incorporated into the spiro C4-C, which is optionally substituted. 12 The heterocyclic ring is present in the first optional extended subunit. In such a construct, the spirocarbon is attached to M. 1 The maleimide imide nitrogen, and thus attached to M 2 Nitrogen in the nitrogen, and further attached to L SS The remaining portion of the primary connector, which includes the aforementioned first optional extension subunit (A), optionally via -[HE]-A O -or[HE]-Aa’ - Present in the drug linker portion of Formula 1B or in the drug linker compound of Formula IB. In those respects, cyclic BUs contribute to M 2 The succinimide moiety is hydrolyzed in a manner similar to that of the acyclic basic unit to that of M 3 The corresponding one or more open-loop forms can also be enhanced by [HE].

[0378] In some respects, L SS Primary connector (sometimes shown as L) SS ', to clearly indicate that it is a self-stabilizing (L) compound in a drug linker compound of formula IB. SS L of the primary connector precursor B '-AB b -by general formula M 1 -A(BU)-[HE]-A O -B- (when the subscript b is 1) or M 1 -A(BU)-[HE]-A' a’ - (when the subscript b is 0) indicates that M 1 It is the maleimide moiety, and A is a C1-C that is bound to BU or substituted by BU and otherwise optionally substituted and optionally combined with [HE]. 12 Alkylene, [HE] is an optional hydrolysis-enhancing moiety, where when A is a single discrete unit, the equation becomes M. 1 -A(BU)-[HE]-B-or M 1 -A(BU)[HE]-, or when A has two subunits, the formula becomes M. 1 -A1(BU)-[HE]-A2-B-or M 1 -A1(BU)-[HE]-A2-, where A1 and A2 are both subunits of A.

[0379] In other respects, the L in the drug delivery unit of Formula 1B of the ADC of Formula 1A SS The primary connector is of the general type - M 2 -A(BU)-[HE]-A O -B- (when the subscript b is 1) or M 2 -A(BU)-[HE]-A a’ - (when the subscript b is 0) indicates that M 2 It is the succinimide moiety, where A is the first optional extended subunit and contains C1-C. 12 Alkylene, the C1-C 12 Alkylene-bonded BU or substituted with BU and otherwise optionally substituted and optionally combined with [HE], [HE] being an optional hydrolysis-enhancing moiety, and A O / A' is an optional subunit of A. When A is a single discrete unit, L SS From formula -M 2 -A(BU)-[HE]-B-or-M 2 -A(BU)-[HE]- indicates that when A has two subunits, L SS From formula -M 2 -A1(BU)-[HE]-A2-or-M 2 -A1(BU)-[HE]-A2-B- (when the subscript b is 0 or 1) represents this.

[0380] In other respects, the L in the drug delivery unit of Formula 1B of Formula 1A LDC / ADC S The primary connector is of type M. 3 -A(BU)-[HE]-A O -B- (when the subscript b is 1) or -M 3 -A(BU)-[HE]-A a’ - (when the subscript b is 0) indicates that M 3 It is a succinimide amide moiety, and A is a C1-C moiety that is bound to BU or substituted by BU and otherwise optionally substituted and optionally combined with [HE]. 12 Alkylene, [HE] is an optional hydrolysis-enhancing moiety, and A O / A' is an optional subunit of A, where -A(BU)-[HE]-A is an optional subunit of A when A is a single discrete unit. O -or-A(BU)-[HE]-A a’ - becomes -A(BU)-[HE]-, or when A is or contains two subunits, it becomes -A1(BU)-[HE]-A2-.

[0381] L within the drug linker portion of Formula 1B of some ligand drug conjugates containing Formula 1 SS Exemplary but non-limiting example of a primary connector - L B The -A- structure is represented by the following formula:

[0382]

[0383]

[0384] Where the wavy line represents the site covalently attached to the ligand unit, the well number (#) in the upper structure of 1 with subscript b represents the site covalently attached to the branch unit (B) in Equation 1B, or the well number (#) in the lower structure of 0 with subscript b represents the site covalently attached to the arbitrarily chosen secondary linker (L) in Equation 1B. OThe W covalent attachment sites of ) are shown, where the dashed curves represent optional cyclization, which exists when BU is a cyclic basic unit or does not exist when BU is an acyclic basic unit, where [HE] is an optional hydrolysis-enhancing moiety, and A O / A' is an optional subbase of A, where the subscript z is 0 or an integer in the range 1 to 6; each R d1 Independently selected from hydrogen and optionally substituted C1-C6 alkyl groups, or R d1 The two carbon atoms in the ring, the one or more carbon atoms to which they are attached, and any intermediate carbon atom define the optionally substituted C3-C8 carbon ring, and the remaining R d1 (If any) is independently hydrogen or optionally substituted C1-C6; when BU is an acyclic basic unit, R a2 It is a -H or optionally substituted C1-C8 alkyl group, and when BU is a cyclic basic unit, R a2 It needs to be not -H and include BU and R. a2 The attached carbon atom is defined as a spiro C4-C with an optional substitution of a secondary or tertiary basic nitrogen atom in the skeleton. 12 Heterocyclic rings, such that R in them a2 Compared to corresponding conjugates in which BU is hydrogen-containing and BU is replaced by hydrogen, acyclic or cyclic BU can increase the succinimide (M) content at suitable pH. 2 The hydrolysis rate of the succinamide (M) portion is used to provide the succinamide (M) 3 ) part, and with R in it a2 Compared to the aforementioned conjugates in which hydrogen is present and BU is replaced by hydrogen, the cyclic basic unit essentially retains the characteristics of LDC / ADC (where R is hydrogen). a2 The increase in the hydrolysis rate of the drug junction portion corresponding to the drug junction portion of the acyclic BU (which is hydrogen and BU is acyclic BU).

[0385] Exemplary but non-limiting examples include L SS 'of L B The '-A- structure (which is sometimes found in pharmaceutical linker compounds of formula I used as intermediates in the preparation of ligand-drug conjugate compositions) is represented by the following formula:

[0386]

[0387] Among them, BU and other variable groups are as described above for those with L. SS LDC / ADC of the first-level connector B -A- structure as defined. When a self-stabilizing linker precursor (L) containing a maleimide moiety is present. SS When drug linker compounds are used to prepare LDC / ADCs, the L... SS 'Partially converted into L containing succinimide moiety' SSPrimary linker. Before condensing with the reactive thiol functional group from the target agent (such as an antibody or its antigen-binding fragment), the basic nitrogen atom of BU is usually protonated or protected by an acid-labile protecting group.

[0388] "Self-stabilizing linkers" are derived from self-stabilizing linkers (L...) in ligand-drug conjugates (such as antibody-drug conjugates). SS ) containing M 2 The organic portion undergoes hydrolysis under controlled conditions to provide a self-stabilized linker (L). S The corresponding M) 3 In some cases, the LU component is unlikely to reverse the target portion and the M-containing portion. 1 Part (which provides the original containing M) 2 L SS (Partial) condensation reaction. Except for M 3 In addition to the portion, the self-stabilized connector (LS) also includes a first optional extension subunit (A), which is present and combined with a cyclic basic unit or replaced by an acyclic basic unit, wherein A is covalently attached to M. 3 and L S The remaining portion of the primary connector (i.e., B), or if B is absent, is attached to the secondary connector (L). O M 3 Part of it is L in the ligand drug conjugate SS The succinimide moiety (M) 2 The transformation of M is obtained, where M is the result of the transformation. 2 Partially sulfur-substituted succinimide ring systems, wherein the ring system consists of a sulfur atom in the reactive thiol functional group of the target agent and an L in the drug linker compound. SS 'Part of M' 1 It is obtained by Michael addition to the maleimide ring system, wherein M 2 Compared to the corresponding substituents in M, the derivatives are derived from M. 2 The portion possesses reduced reactivity in eliminating its thiosubstituents. In those respects, those derived from M 2 The part has a corresponding M 2 Succinic acid amide (M 3 The structure of part M, where M 2 The product undergoes hydrolysis of one of the carbonyl-nitrogen bonds in its succinimide ring system, assisted by the basic functional group of BU, due to its appropriate proximity caused by this attachment. Therefore, the product of this hydrolysis possesses [a specific characteristic] in its amide nitrogen atom (which corresponds to L [a specific characteristic]). S M 2 L SSThe precursor contains carboxylic acid and amide functional groups substituted on the imide nitrogen atom by the remaining portion of a primary linker, the remaining portion of which will include at least any optional extended subunit present. In some aspects, the basic functional group is a primary, secondary, or tertiary amine with an acyclic basic unit or a secondary or tertiary amine with a cyclic basic unit. In other aspects, the basic nitrogen of BU is a heteroatom of an optionally substituted basic functional group, such as in the guanidinium moiety. In any aspect, the reactivity of the basic functional group of BU to base-catalyzed hydrolysis is controlled by pH by reducing the protonated state of the basic nitrogen atom.

[0389] Therefore, the self-stabilizing connector (L) S ) usually has M 3 The partial structure, wherein the partial covalently bonded to a first optional extended subunit, the extended subunit being present and combined with a cyclic basic unit or substituted by an acyclic basic unit. In some respects, A is a discrete single unit and in other respects has two or more subunits, and if the two subunits exist as A / A1, optionally combined with [HE], then A is generally represented by A1-A2. The extended subunit A is sequentially coupled with L S The B of the first-level connector or with M 3 L O W covalent bond, A, A' a’ / B and BU components are expressed in general formula -M 3 -A(BU)-[HE]-A' a’ -or M 3 -A(BU)-[HE]-A O The arrangement is represented by -B- (where the subscript b is 0 or 1). When A is a single discrete unit, L S By -M 3 -A(BU)-[HE]-B- (when the subscript b is 1) or -M 3 -A(BU)-[HE]- indicates that when A has two subunits, L S By -M 3 -A1(BU)-A2- or -M 3 -A1(BU)-A2-B- indicates (where the subscript b is 0 or 1 respectively), where BU represents any type of basic unit (cyclic or acyclic).

[0390] LDC / ADC (where L) B It's M 2 Or M 3 In these structures, A(BU) / A1(BU) and [HE] are arranged as described above, where BU is an acyclic basic unit. SS and L S -L in the primary connector BExemplary non-restrictive structures of -A- are shown by way of example, but are not limited to the following structures:

[0391]

[0392] Where -CH(CH2NH2)C(=O)- is A, and when A is a single discrete unit, A O If A' does not exist, or A is A1-A2-, then A O / A' exists as A2, where A / A1 is replaced by BU, where BU is an acyclic basic unit, is -CH2NH2, has an optional geotonic basic nitrogen atom, and the -C(=O)- in this part is an optional hydrolysis-enhancing part [HE], and the well mark in the upper structure indicates covalent attachment with B, and the well mark in the lower structure indicates covalent attachment with L. O The W is covalently attached. Those exemplary structures respectively contain succinimide (M... 2 ) part or succinic acid-amide (M 3 ) part, the latter in L SS Transform into L S The process is assisted by -CH2NH2, M 2 It is produced by the hydrolysis of the succinimide ring.

[0393] LDC / ADC (where L) B It's M 2 Or M 3 ; In these structures, A(BU) / A1(BU), A O / A' and [HE] are arranged in the manner described above, where BU is a cyclic basic unit) of L SS and L S -L in the primary connector B Exemplary non-restrictive structures of -A- are shown by way of example, but are not limited to the following structures:

[0394]

[0395] Where A O When A exists as a single discrete unit, or when it does not exist or the subscript a' is 0, these -M 2 -A(BU)-[HE]-A O / A' a’ -and-M 3 -A(BU)-[HE]-A O / A' a’ -The structure becomes -M 2 -A(BU)-[HE]-and-M 3 -A(BU)-[HE]-, or when A OWhen / A' exists as a subunit of A (denoted as A2), it becomes -M. 2 -A1(BU)-[HE]-A2-and-M 3 -A 1 (BU)-[HE]-A2-, and wherein in either structure BU is a cyclic basic unit in the form of optionally geofused azirmonobutane-3,3-diyl, the structure of which is an exemplary heterocyclic basic unit incorporated into A / A1. This heterocycle corresponds to the aminoalkyl of the acyclic basic unit in the -A1(BU)- or -A(BU)- moiety, wherein the basic nitrogen of the acyclic basic unit is at least partially passed through an R at the α-position of the carbon atom. a2 And the formal circularization back to M 2 The succinimide nitrogen, with an acyclic basic unit attached to M 2 succinimide nitrogen.

[0396] Each of the above -L B The wavy line in the -A- structure represents the M in the structurally corresponding drug connector compound where the sulfur atom is Michaelized to the corresponding drug connector compound. 1 Partial maleimide ring or M-containing ring 1 In the intermediate stage, the sulfur atom of the ligand unit of the reactive thiol functional group of the target agent is covalently attached. The hash mark (#) in the upper structure indicates the site covalently attached to B, where B is L. SS or L S The remaining portion of the primary connector, and the well mark in the lower structure, indicates the connection to L. O The site where W is covalently attached. Due to M 2 The succinimide ring system is asymmetrically substituted due to its thiosubstituents, thus differing in position from the released carboxylic acid group, as defined herein as succinic acid-amide (M). 3 The regional chemical isomers of the region may lead to M 2 Hydrolysis. In the above structure, with A O The adjacent carbonyl functional groups shown are examples of hydrolysis enhancers [HE] as defined herein.

[0397] The above-M 3 -A(BU)-[HE]-A O / A' a’ -、-M 3 -A(BU)-and-M 3 -A1(BU)-[HE]-A2- (where BU is an acyclic or cyclic basic unit) represents the inclusion of a self-stabilized junction (L S Example of a primary connector - L BThe -A- structures are named as such because these structures are unlikely to eliminate the thiosubstituents of the ligand unit, thus leading to the loss of the targeting moiety, unlike the inclusion-M structures from which these structures are derived. 2 -A(BU)-[HE]-A O / A' a’ -、-M 2 -A(BU)-or-M 2 The corresponding L of -A1(BU)-[HE]-A2- SS In comparison to some aspects. Unbound by theory, it is considered to be similar to M. 2 (It no longer restricts the thiosubstituent to conformations favorable for E2 elimination) compared to M 3 Greater conformational flexibility leads to increased stability.

[0398] Unless the context otherwise indicates or implies, "alkaline unit" as used herein refers to a self-stabilizing junction (L) as described herein. SS The organic components within the primary connector are carried to the corresponding L via the BU. S In some parts, BU participates in L SS M 2 The base-catalyzed hydrolysis of the succinimide ring system within the succinimide ring (i.e., catalyzing the addition of a water molecule to one of the carbonyl-nitrogen bonds of the succinimide ring). In some respects, base-catalyzed hydrolysis occurs when attached to L... SS The targeted ligand unit can be tolerated under controlled conditions. In other respects, base-catalyzed hydrolysis is initiated in the presence of L... SS The reaction between the drug linker compound and the target agent is initiated by the Michael addition of the sulfur atom of the reactive thiol functional group of the target agent and the L-type of the drug linker compound. SS 'of M 1 Partial hydrolysis competes with this. Unbound by theory, the following describes various considerations for designing suitable basic units. In one such aspect, regarding BU and M... 2 The ability of the carbonyl group to form hydrogen bonds, the selection of basic functional groups of acyclic basic units and their role in L SS In contrast to M 2 The relative positions of the components effectively increase its electrophilicity, and thus its sensitivity to water attack. In another aspect, those selections that increase the nucleophilicity of water molecules via hydrogen bonding to the basic functional groups of BU are directed towards M. 2 Carbonyl group. In a third aspect, choices are made such that the protonated basic nitrogen does not increase the electrophilicity of the succinimide carbonyl group to a degree that would promote premature hydrolysis (requiring compensation from undesirable excess drug linker compounds) through inductive electron withdrawal. In another aspect, some combination of these mechanistic effects contributes to the catalysis of L... SS Controlled hydrolysis to LS .

[0399] Typically, an acyclic basic unit that functions via any of the mechanisms described above comprises one carbon atom or two to six consecutive carbon atoms, more typically one carbon atom or two or three consecutive carbon atoms, wherein one or more carbon atoms link the basic amino functional group of the acyclic basic unit to the L atom attached thereto. SS The remaining portion of the primary connector. To ensure the basic amine nitrogen atom is at the desired proximity to aid succinimide (M... 2 Partially hydrolyzed into its corresponding ring-opening succinic acid amide (M) 3 ( ) part, relative to A and M 2 The succinimide nitrogen attachment site, the amine-containing carbon chain of the acyclic basic unit is usually attached to L SS -L B -A- Part C1-C 12 The A on the α carbon of the alkylene group (and thus attached to its corresponding M) 1 -A-structured maleimide nitrogen). Typically, the α-carbon in an acyclic basic unit has an (S) stereochemical configuration or a configuration corresponding to the α-carbon of an L-amino acid.

[0400] As mentioned earlier, acyclic or cyclic BUs are typically represented by C1-C, which are optionally replaced in other ways. 12 alkylene moiety and L SS M 1 Or M 2 Or L S M 3 Connection, wherein C1-C 12 The alkylene moiety is either cyclically bonded to a basic unit or substituted with an acyclic basic unit, and is associated with M, respectively. 1 Or M 2 maleimide or succinimide nitrogen, or M 3 The nitrogen atom of the amide is bonded. In some respects, the C1-C atoms of the cyclic basic unit are optionally substituted in other ways. 12 The alkylene moiety is covalently bonded to [HE] and is typically present via an intermediate of ether, ester, carbonate, urea, disulfide, carbamate, or other functional groups, more commonly via an intermediate of an ether, amide, or carbamate functional group. Similarly, the acyclic form of BU is typically present via an intermediate of an L-shaped moiety. B '-A-(where L) B 'Is M 1 ) or -L B -A-(where L) B It's M 2 Or M 3 C1-C, which are arbitrarily substituted by other means for A in ) 12 alkylene moiety and L SSM 1 Or M 2 Or L S M 3 Connection, that is, in relation to M 1 Or M 2 The imino nitrogen atom or M in the maleimide or succinimide ring system 3 (It is composed of M) 2 The amide nitrogen-attached C1-C (generated by hydrolysis of the succinimide ring system) 12 The same carbon atom of the alkylene moiety is replaced by an acyclic basic unit.

[0401] In some respects, the cyclic basic unit is formed by cyclizing the acyclic basic unit into a C1-C structure that is otherwise optionally substituted. 12 Alkyl (R) a2 To combine the structure of the acyclic BU, the C1-C 12 The alkyl group is independently selected from the alkyl group of A / A1, which is bonded to the same α-carbon as the acyclic basic unit, thereby forming a spirocyclic system such that the cyclic basic unit is incorporated into the structure of A / A1, rather than as a substituent of A / A1 when BU is acyclic. In those respects, formal cyclization is directed toward the basic amine nitrogen of the acyclic basic unit, thereby providing the cyclic basic unit as an optionally substituted symmetrical or asymmetric spirocyclic C4-C. 12 Heterocyclic (this depends on the relative carbon chain lengths in the two α-carbon substituents), where the basic nitrogen is now a basic skeleton heteroatom. To ensure that cyclization in the cyclic basic unit substantially preserves the basic properties of the acyclic basic unit, the basic nitrogen atom of the acyclic basic unit should be a nitrogen atom of a primary or secondary amine, not a nitrogen atom of a tertiary amine, as the latter would lead to quaternization of the skeleton nitrogen in the heterocycle of the cyclic basic unit. In this regard, to substantially preserve the basic nitrogen in the L... SS Transform into L S Help M in the process 2 Hydrolysis to M 3 The ability of these primary joints to locate the basic nitrogen in the resulting structure of the cyclic basic unit typically enables the formation of C4-C in the spiral. 12 The number of carbon atoms inserted between the basic nitrogen atom and the spirocarbon in the heterocyclic component does not exceed three, typically one or two. Embodiments of the invention further describe the cyclic basic unit incorporated into A / A1 and L having these as components. SS and L S Primary connector.

[0402] Unless the context otherwise indicates or implies, "hydrolysis-enhanced portion" as used herein refers to the portion optionally present in L SS Primary connector and its hydrolysis product L S L B'-A- or -L B The electron-withdrawing group or portion in the first optional extended subunit (A) of -A-. Hydrolysis-enhancing [HE] moiety (when used as a drug linker portion in LDC / ADCs) SS When the components of A / A1 are present, where A / A1 is similar to M in some respects... 2 Partial imide nitrogen bonding) increases M 2 The electrophilicity of the carbonyl group in succinimide has the least effect on the reaction, which depends on the electron-withdrawing effect of [HE] and M. 2 The degree of proximity of parts to promote M 2 Partially converted to L S M of the first-level connector 3 Partially, A / A1 are respectively bound to or substituted by cyclic basic units or acyclic basic units; for [HE] to M 2 The potential influence of the carbonyl group (by inducing hydrolysis to M) 3 The rate) and one or more of the above-mentioned functions of any type of BU are adjusted to make M 1 From the inclusion of M 1 -A(BU)-[HE]-A O / A' a’ -ofL B Drug linker compounds with an '-A- structure do not undergo significant premature hydrolysis during the preparation of ligand-drug conjugates, two of which are derived from formula M. 1 -A(BU)- and M 1 -A1(BU)-[HE]-A2- indicates that A / A1 is combined with [HE]. Conversely, the combined action of BU and [HE] promotes hydrolysis, which leads to the formation of the general formula -M in the ligand-drug conjugate compound. 2 -A(BU)-[HE]-A O / A' a’ -or a more specific form-M 2 -A(BU)-or-M 2 -A1(BU)-A2-of-L B The -A- structure transforms into its corresponding -M structure under controlled conditions (such as when pH is intentionally increased to reduce the protonation state of the basic unit). 3 -A(BU)-[HE]-A O / A' a’ -、-M 3 -A(BU)- or M 3 -A1(BU)-[HE]-A2-, thus eliminating the need for excessive molar excesses of drug connective compounds to compensate for its M-, thus requiring no excessive molar excess of drug connective compounds to compensate for its M-. 1 Partial hydrolysis. Therefore, the sulfur atom of the reactive thiol functional group of the target agent interacts with M. 1The maleimide ring system (providing attachment to M) 2 Michael addition of the succinimide ring system (targeting ligand unit) typically facilitates effective interaction with M. 1 Hydrolysis occurs at competing rates. Unbound by theory, it is believed that at low pH, for example when the basic amine of BU is in the form of a TFA salt, the M in the drug linker product... 1 Premature hydrolysis occurs much slower than the pH rises to a level suitable for alkaline catalysis using an appropriate buffer, and an acceptable molar excess of the drug linker compound can adequately compensate for the premature M 1 Any loss caused by hydrolysis, premature M 1 Hydrolysis of the sulfur atom of the reactive thiol functional group in the target agent and the M of the drug linker compound 1 Part of the Michael Bonus occurs during the process of completion or near completion.

[0403] As previously discussed, the enhancement of carbonyl hydrolysis by any type of basic unit depends on the basicity of its functional group and the relationship between that basic functional group and M. 1 / M 2 The distance between the carbonyl groups. Typically, [HE] is the C1-C group located at A / A1. 12 The carbonyl moiety distal to the alkylene terminus or other carbonyl-containing functional group, and also providing covalent attachment to A2 or optional secondary linker (which exists when B is absent and A is a single discrete unit), A / A1 with M 2 Or derived from it M 3 Bonding. Carbonyl functional groups other than ketones include esters, carbamates, carbonates, and ureas. When [HE] is an L... SS When the drug junction portion of an ADC with a primary junction contains carbonyl functional groups other than ketones, the carbonyl moiety shared with A / A1 is typically located away from A / A1 and M. 2 The attachment site of the imide nitrogen atom is optionally substituted with the C1-C of A / A1 in other ways. 12 Alkylene bonding, such as when [HE] is -C(=O)-X-, where X is -O- or optionally substituted -NH-. In some respects, the [HE] moiety can be sufficiently distant from the imide nitrogen covalently bonded to A / A1 to observe the effect on M-containing... 2 The hydrolysis sensitivity of some succinimide carbonyl-nitrogen bonds has no significant effect or a slight effect, but the hydrolysis sensitivity is mainly driven by BU.

[0404] Unless the context otherwise indicates or implies, as used herein, "extended subunit" refers to an optional organic portion of the primary or secondary linker in a linker unit of a drug linker compound or drug linker moiety of a ligand-drug conjugate (such as an antibody-drug conjugate), which physically separates the targeting ligand unit (L) from the optional secondary linker (if present). When the linker unit contains L... SS or L S In the case of a primary connector, a first optional extender exists because it provides a basic unit for these types of primary connectors. When the ligand unit lacking this optional extender unit does not have sufficient space to release to allow for efficient treatment of the secondary connector, thereby releasing the drug unit as free drug, L R The presence of the first optional extension subunit (A) may also be required in any type of primary linker. Alternatively, or in addition to steric release, those optional components may be included to provide synthetic convenience in the preparation of pharmaceutical linker compounds. In some respects, when the subscript b is 1, the first or second optional extension subunit (A or A', respectively) is a single unit or may contain multiple subunits (e.g., when A has two subunits denoted by -A1-[HE]-A2-). In other respects, when the subscript b is typically 0, A is a distinct unit or has two distinct subunits (when the subscript is 0 and the subscript a' is 1). In still other respects, B / A' has 2 to 4 independently selected distinct subunits.

[0405] In some respects, when L R It is L SS / L S At that time, in addition to the M of the drug linker compound 1 or the M of the drug linker in LDC / ADC compounds 2 / M 3 In addition to covalent attachment, A can also optionally pass through A O / A' a With branch unit (B) or the existing optional secondary connector (L) O W-bonding, such as in A[HE](A) O / A' does not exist) or A1-[HE]-A2(A O In the context of / A' (which exists), it is generally represented as A-[HE]-A O / A a’ - where A / A1 and A O / A a (When it exists as A2) is also L. SS / L S One of its components.

[0406] In some respects, the subunits of A or A' or any of these extended subunits have the formula -L P(PEG)-, where L P It is a parallel linker unit and PEG is a PEG unit as defined elsewhere. Therefore, in some of those aspects, the linker unit in the drug linker portion of the ligand-drug conjugate, or a drug linker compound in which subscript b is 0 and subscript a' is 1, contains the formula -A1-[HE]-L. P (PEG)-, where A' is -L P (PEG) - and exists as A2. In which the subscript b is 1 and A O Among the other aspects of A2, the linker unit in the drug linker portion of the ligand-drug conjugate or the drug linker compound contains the formula -A1-[HE]-L P (PEG)-B-. In other respects, where subscript b is 1 and subscript a' is 1, the ligand-drug conjugate or drug linker compound contains the formula -A-[HE]-A. O -BL P (PEG), where A' is L P (PEG).

[0407] In some respects, when the subscript a is 1 and thus a first optional extended subunit (A) exists, the unit typically has at least one carbon atom, wherein the atom will L B / L B 'Connected to [HE]. In which L B 'L belongs to drug linker compounds' SS In some aspects of the primary connector, the extended subunit comprises C1-C units that are replaced or combined with basic units. 12 The alkylene moiety, and optionally otherwise substituted, has one radical carbon atom attached to a maleimide nitrogen atom and the other to [HE], wherein [HE] is an optional hydrolysis-enhancing moiety present. In other respects, when L... R 'Not L' SS 'But it still contains maleimide moiety or some other L' B 'Partially, L B 'Attached to an optional first extension subunit (A), the first extension subunit being, in some respects, an optionally substituted C1-C in combination with [HE]. 12 Alkylene. Therefore, in which L R 'Is L SS In some aspects, there exists a first optional extended subunit and it comprises C1-C 12 The alkylene moiety, [HE], and optional subunits (A when the subscript b is 1) O Or, when the subscript b is 0, it is A' a’ When L R 'Is LSS At that time, all of these were L R 'The components, in which the C1-C 12 At the attachment site between the alkylene moiety and the imide nitrogen atom, A is attached to L. R 'The component B or attached as L O The component W. In other respects, when the subscript a is 1 and A exists as a single discrete unit or with two subunits, A has the general formula -A-[HE]-A. O / A a’ -(where A) O / A' a’ (is an optional subunit of A), or more specifically has the formula -A1-[HE]-A2- (when A O A exists as a second subunit of A when the index b is 1, or when the index a' is 1 and the index b is 0, such that A' exists as a second subunit of A. In these respects, A O / A2 or A' / A2 is an α-amino acid, β-amino acid, or other amino acid residue.

[0408] Unless the context otherwise indicates or implies, as used herein, "branching unit" refers to a trifunctional or multifunctional organic portion that is an optional component of a linker unit (LU). A branching unit (B) is present in the primary linker of the drug linker portion of Formula 1A in an LDC / ADC of Formula 1A, when multiple -L... O When the -D portion is present, it is a single drug delivery unit portion. In LDC / ADCs with the above general formula, the absence or presence of branch units is determined by B. b The subscript b indicates that the subscript b is either 0 or 1. The branching unit is at least trifunctional in order to bind to the primary linker. Drug linkers or LDC / ADC compounds with branching units (this is because each drug linker portion of the formula -LU-D has multiple -L...) O The -D part usually contains the formula -A' a’ -WY y - Each secondary connector (L O ), where A' is the second optional extension subunit; the subscript a' is 0 or 1, indicating that A' is absent or present, respectively; W is the peptide cleavable unit; Y is the spacer subunit; and the subscript y is 0, 1 or 2, indicating that one or two spacer subunits are absent or present, respectively.

[0409] In some respects, natural or non-natural amino acid residues, or residues of another amino acid compound with a functionalized side chain, are used to attach two -L... OThe -D portion is a trifunctional branched unit. In some of those aspects, B is an L or D configuration lysine, glutamic acid, or aspartic acid residue, wherein the ε-amino, γ-carboxylic acid, or β-carboxylic acid functional group, along with their amino and carboxylic acid ends, are interconnected within the remainder of LU. This is used to attach 3 or 4 -L... O The more functional branch units of the -D part typically contain the necessary number of trifunctional subunits.

[0410] Unless the context otherwise indicates or implies, “natural amino acid” as used herein means naturally occurring amino acids of the L or D configuration, namely arginine, glutamine, phenylalanine, tyrosine, tryptophan, lysine, glycine, alanine, histidine, serine, proline, glutamic acid, aspartic acid, threonine, cysteine, methionine, leucine, asparagine, isoleucine, and valine, or residues thereof, unless the context otherwise indicates or implies.

[0411] Unless the context otherwise indicates or implies, “non-natural amino acid” as used herein refers to an amino acid or residue thereof containing an α-amino group that has the main chain structure of a natural amino acid but has a side chain group attached to the α-carbon that is not present in the natural amino acid.

[0412] Unless the context otherwise indicates or implies, as used herein, "non-classical amino acid" refers to an amino acid compound whose amine substituent is not bonded to the α-carbon of a carboxylic acid, and therefore is not an α-amino acid. Non-classical amino acids include β-amino acids in which a methylene group is inserted between the carboxylic acid and amino functional groups in natural or non-natural amino acids.

[0413] Unless the context otherwise indicates or implies, as used herein, "peptide" refers to a polymer of two or more amino acids, wherein the carboxylic acid group of one amino acid forms an amide bond with the α-amino group of the next amino acid in the peptide sequence. Methods for preparing amide bonds in polypeptides are also provided in the definition of amides. Peptides may contain naturally occurring amino acids and / or non-natural and / or non-classical amino acids in L- or D-configuration.

[0414] As defined in this article, a "protease" is a protein capable of enzymatically cleaving carbonyl-nitrogen bonds (such as amide bonds, which are commonly found in peptides). Proteases are classified into six major classes: serine proteases, threonine proteases, cysteine ​​proteases, glutamate proteases, aspartic proteases, and metalloproteinases. This naming is attributed to the catalytic residues in their active sites that are primarily responsible for cleaving the carbonyl-nitrogen bonds of their substrates. Proteases are characterized by various specificities, depending on the identity of the residues on the N-terminus and / or C-terminus sides of the carbonyl-nitrogen bond and their diverse distribution (intracellular and extracellular).

[0415] Regulatory proteases are intracellular proteases typically required to regulate cellular activity, which can sometimes become abnormal or disordered in abnormal or otherwise unwanted cells. In some cases, a protease is a regulatory protease involved in cell maintenance or proliferation when its peptide-cleavable unit targets a protease that preferentially distributes within the cell. These proteases include cathepsins. Cathepsins include serine proteases, cathepsin A, cathepsin G, aspartic proteases, cathepsin D, cathepsin E, and cysteine ​​proteases, cathepsin B, cathepsin C, cathepsin F, cathepsin H, cathepsin K, cathepsin L1, cathepsin L2, cathepsin O, cathepsin S, cathepsin W, and cathepsin Z.

[0416] Unless the context otherwise indicates or implies, as used herein, “peptide-cleavable unit” refers to the drug linker portion of a ligand-drug conjugate compound or the organic portion within the secondary linker of a drug linker compound that provides a recognition site for a protease and is capable of enzymatically releasing the conjugated drug unit (D) as a free drug under the enzymatic action of the protease.

[0417] Protease cleavage recognition sites are sometimes limited to those found in abnormal cells (such as cancer cells) or in nominally normal cells targeted by ligand-drug conjugates (which are specific to the environment of nearby abnormal cells but can also be found in normal cells). For this purpose, peptides are typically resistant to circulating proteases to minimize premature release of the free drug or its precursors, which could otherwise lead to off-target adverse events due to systemic exposure. In some aspects, the peptide will have one or more D-amino acids or non-natural or non-classical amino acids to provide this resistance. In some of those aspects, the sequence will comprise a dipeptide or tripeptide, wherein the P2' site contains a D-amino acid and the P1' site contains one of 20 naturally occurring L-amino acids other than L-proline.

[0418] In some respects, the reaction site is more likely to be enzymatically manipulated after the immune selective binding of the target antigen. In some of these respects, the target antigen is located on abnormal cells, making the recognition site more likely to be enzymatically manipulated after the ligand-drug conjugate compound has been internalized into the target abnormal cells. Therefore, those abnormal cells should display the target antigen at a higher copy number compared to normal cells to mitigate target-related adverse events. In other respects, the target antigen is located on normal cells within and specific to the abnormal cellular environment, making the recognition site more likely to be enzymatically manipulated after the ligand-drug conjugate compound has been internalized into these target normal cells. Therefore, those normal cells should display the target antigen at a higher copy number compared to normal cells located far from cancer cell sites to mitigate target-related adverse events.

[0419] In any of the above aspects, proteases in tumor tissue homogenates exhibit greater responsiveness to recognition sites compared to normal tissue homogenates. In some aspects, this greater responsiveness is due to the greater amount of intracellular protease activity in the target cells of tumor tissue compared to the intracellular protease activity in normal cells of normal tissue, and / or the reduced protease activity in the interstitial spaces of normal tissue compared to the activity of the peptide-cleavable units of conventional ligand-drug conjugates. In those aspects, the intracellular proteases are regulatory proteases, and the peptide bonds of the peptide-cleavable units are selectively cleaved not only by proteases in tumor tissue homogenates compared to those in normal tissue homogenates, but also by intracellular regulatory proteases compared to serum proteases.

[0420] Secondary linkers containing peptide-cleavable units typically have the formula -A' a’ -WY y - where A' is the second optional spacer unit when subscript b is 1; subscript a' is 0 or 1; W is a peptide-cleavable unit; Y is an optional spacer unit; and subscript y is 0, 1, or 2. When subscript b is 0 and subscript a' is 1, A' becomes a subunit of A, therefore the secondary linker has the formula -WY. y - For any of the secondary linkers, the protease acts on the peptide sequence containing the peptide-cleavable unit, resulting in the direct release of D. When the subscript y is 0 or 1, the drug-linker fragment of formula YD is produced as a precursor of the free drug, where Y typically undergoes suicide to provide the free drug; or when the subscript y is 2, the first drug-linker fragment of formula Y-Y'-D is produced, where Y is a first spacer subunit undergoing suicide to provide the second drug-linker fragment of formula Y'-D, where Y' is a second spacer subunit that breaks down to complete the release of D as the free drug.

[0421] In some respects, drug linker compounds in which the secondary linker contains a peptide-cleavable unit are represented by the structure of formula IC:

[0422]

[0423] Furthermore, the corresponding drug linker portion of the ligand-drug conjugate is represented by the structure of Formula 1D or Formula 1E:

[0424]

[0425]

[0426] Where W is a peptide cleavable unit and M is of formula IC 1 -A a -B b -、Form 1D of -M 2 -Aa -B b -Japanese 1E of -M 3 -A a -B b - is a primary connector, where M 1 It is the maleimide moiety; M 2 It is the succinimide moiety; M 3 It is the succinamide part; Y is an optional spacer subunit, so the subscript y is 0 or 1, or Y y It is -Y-Y', so the subscript y is 2, and Y and Y' are the first and second spacer subunits, respectively. The remaining variable groups are as defined for the drug linker compound of formula IA and the drug linker portion of formula 1A. The drug linker compound of the present invention contains M 1 Part of L SS 'M' is contained in the primary connector and the drug connector portion of some LDC / ADCs. 2 Part of L SS Primary joints are those in which A or its subunits are replaced by or bonded to a basic unit. Other primary joints are L. S The primary connector, derived from the M-containing formula 1C mentioned above. 2 L SS Primary connectors, by hydrolyzing their succinimide moieties to provide M-containing compounds of formula 1D. 3 part.

[0427] In either of the above aspects, an amide bond, specifically cleaved by a protease produced by or within the target cell, is linked to an amino group of the spacer unit (Y) or the drug unit (if Y is absent). Thus, the protease's action on the peptide sequence in W results in the release of D as the free drug or its precursor Yy-D (spontaneously fragmented to provide the free drug).

[0428] Unless the context otherwise indicates or implies, as used herein, "spacer subunit" refers to a unit of formula -A' in the drug linker portion of a drug linker compound or ligand drug conjugate. a’ -WY y - secondary connector (L O The part in ) where the subscript y is 1 or 2, indicating the presence of 1 or 2 spacer subunits, where A' is a second optional spacer subunit (which becomes part of a primary joint in some aspects as described herein), the primary joint and the secondary joint are covalently attached as subunits of the existing first optional spacer subunit, the subscript a' is 0 or 1, indicating the presence or absence of A'; Y is a spacer subunit, W is equation -P n …[P3]-[P2]-[P1]-or-P n…[P3]-[P2]-[P1]-[P-1]- a peptide-cleavable unit, wherein the subscript n ranges from 0 to 12 (e.g., 0-10, 3-12, or 3-10), and P1, P2, and P3 are amino acid residues that confer selectivity to protease cleavage of tumor tissue homogenate relative to normal tissue homogenate, as described herein. When the subscript y is 1, the spacer subunit is covalently bonded to W and the drug unit (D), or when the subscript y is 2, it is covalently bonded to another such portion (Y') covalently bonded to D. The action of the protease on W initiates the release of D as free drug, as further described in embodiments of the invention.

[0429] As used herein, “suicide moiety” refers to a bifunctional moiety within a suicide spacer subunit (Y), wherein the suicide moiety is covalently attached to a heteroatom of D, or covalently attached to a shared functional group between Y and D (optionally substituted where permitted), and also covalently attached to a peptide-cleavable unit via another optionally substituted heteroatom (J), wherein J is a suitably substituted nitrogen atom in a -NH- or amide functional group, so that the suicide moiety binds these drug linker components to a generally stable trigonal molecule unless activated.

[0430] When the peptide bond between P1 / P-1 and Y is cleaved, D or the first drug linker fragment (i.e., Y'-D) spontaneously separates from the trigonometric molecule through the self-destruction of the suicide portion of the suicide spacer unit. In some respects, the components of the suicide spacer unit inserted between Y'-D or D and the optionally substituted heteroatom J of Y bonded to W have the optionally substituted -C6-C 24 Aromatic-C(R) 8 (R) 9 )-、-C5-C 24 Heteroarylene-C(R) 8 (R) 9 )-、-C6-C 24 Aromatic-C(R) 8 )=C(R 9 )-or-C5-C 24 Heteroaryl-C(R) 8 )-=C(R 9 ), where R 8 and R 9 As described in the embodiments of the present invention, and typically C6-C 10 aryl-CH2- or C5-C 10Heteroarylene-CH2-, wherein the (hetero)arylene group is optionally substituted, wherein the components of the suicide spacer subunit are fragmented by 1,4 or 1,6-elimination to form an imino-quinone methylate or related structure, while releasing D or Y'-D upon cleavage of the protease-cleavable bond between J and W. In some aspects, suicide spacer subunits having the aforementioned components bonded to J include, for example, optionally substituted p-aminobenzyl alcohol (PAB) moiety, or o- or p-aminobenzyl acetal, or other aromatic compounds electronically similar to the PAB group (i.e., PAB type), such as 2-aminoimidazol-5-methanol derivatives (see, for example, Hay et al., 1999, Bioorg. Med. Chem. Lett. 9:2237) or those wherein the phenyl group of the p-aminobenzyl alcohol (PAB) moiety is replaced by a heteroarylene group.

[0431] Unbound by theory, the aromatic carbon of the arylene or heteroarylene portion of the PAB or PAB-type portion of the suicide spacer unit incorporated into the linker unit is substituted with J, wherein the electron-donating heteroatom of J is attached to the cleavage site of W, thereby weakening the electron-donating ability of the heteroatom (i.e., its EDG ability is masked by incorporating the suicide portion of the suicide spacer unit into the linker unit). Another substituent of the hetero(arylene) is a benzyl carbon, which is attached to an optionally substituted heteroatom of D (an optionally substituted functional group shared between Y and D) or a second spacer unit (Y') bonded to the drug unit (D), wherein the benzyl carbon is attached to another aromatic carbon atom of the central arylene or heteroarylene, wherein the aromatic carbon with the weakened electron-donating heteroatom is adjacent (i.e., 1,2-relationship), or two additional positions are removed from the benzyl carbon atom (i.e., 1,4-relationship). Functionalized EDG heteroatoms are selected so that the electron-donating ability of the heteroatoms masked during treatment of the cleavage site of W is restored, thereby initiating 1,4- or 1,6-elimination to expel -D as a free drug from the benzyl substituent, or, upon release of Y'-D, the suicide of Y' provides free drug to induce a therapeutic effect. Exemplary suicide portions and suicidal spacer subunits having those suicide portions are illustrated by embodiments of the present invention.

[0432] Other examples of suicide groups include, but are not limited to, aromatic compounds that are electronically similar to the PAB group, such as 2-aminoimidazolium-5-methanol derivatives (see, for example, Hay et al., 1999, Bioorg. Med. Chem. Lett. 9:2237) and ortho- or para-aminophenylmethyl acetals. Spacers that undergo cyclization upon hydrolysis of the amide bond can be used, such as substituted and unsubstituted 4-aminobutyric amides (see, for example, Rodrigues et al., 1995, Chemistry Biology 2:223), appropriately substituted bicyclic [2.2.1] and bicyclic [2.2.2] ring systems (see, for example, Storm et al., 1972, J. Amer. Chem. Soc. 94:5815), and 2-aminophenylpropionic amides (see, for example, Amsberry et al., 1990, J. Org. Chem. 55:5867). Eliminating the α-substitution of glycine in amine-containing drugs (see, for example, Kingsbury et al., 1984, J. Med. Chem. 27:1447) is also an example of a suicide group. In one embodiment, the spacer unit is a branched bis(hydroxymethyl)styrene (BHMS) unit (as described in WO 2007 / 011968), which can be used to bind and release a variety of drugs. Further suicide spacers are described in WO 2005 / 082023.

[0433] Unless the context otherwise indicates or implies, as used herein, “methylene carbamate unit” refers to an organic portion that is capable of self-destruction and is inserted between a first self-destructive spacer subunit and a drug unit within a linker unit of a ligand-drug conjugate or drug linker compound, and is therefore an exemplary second spacer subunit.

[0434] The methylene carbamate (MAC) unit bonded to the drug unit is represented by Formula III:

[0435]

[0436] Or a pharmaceutically acceptable salt thereof, wherein the wavy line indicates that the methylene carbamate unit is covalently attached to the first suicide spacer unit (Y); D is a pharmaceutical unit having a functional group (e.g., hydroxyl, thiol, amide, or amine functional group) bound to the methylene carbamate unit; T* is a heteroatom from said functional group, including oxygen, sulfur, or nitrogen as an optional substitution -NH-. Upon cleaving of the connector unit containing the MAC unit, the first suicide spacer unit (Y) (as a second suicide spacer unit (Y')) bonded to the MAC unit undergoes fragmentation to release -Y'-D of formula III. The MAC unit then spontaneously decomposes to complete the release of D as a free drug, the assumed mechanism of which is illustrated by embodiments of the present invention.

[0437] As used herein, "PEG unit" refers to a group containing a polyethylene glycol moiety (PEG) having repeating ethylene glycol subunits, wherein the ethylene glycol has the following formula:

[0438]

[0439] PEG includes polydisperse PEG, monodisperse PEG, and discrete PEG. Polydisperse PEG is a heterogeneous mixture with different sizes and molecular weights, while monodisperse PEG is typically purified from heterogeneous mixtures, thus providing a single chain length and molecular weight. Discrete PEG is a compound synthesized stepwise rather than via a polymerization process. Discrete PEG provides a single molecule with a defined and specified chain length.

[0440] PEG units contain at least 2 subunits, at least 3 subunits, at least 4 subunits, at least 5 subunits, at least 6 subunits, at least 7 subunits, at least 8 subunits, at least 9 subunits, at least 10 subunits, at least 11 subunits, at least 12 subunits, at least 13 subunits, at least 14 subunits, at least 15 subunits, at least 16 subunits, at least 17 subunits, at least 18 subunits, at least 19 subunits, at least 20 subunits, at least 21 subunits, at least 22 subunits, at least 23 subunits, or at least 24 subunits. Some PEG units contain up to 72 subunits.

[0441] As used herein, a “PEG capping unit” is a nominally unreactive organic moiety or functional group that caps the free and untethered ends of a PEG unit and is in some respects not hydrogen. In those respects, the PEG capping unit is a methoxy, ethoxy, or other C1-C6 ether, or -CH2-CO2H or other suitable moiety. The ether, -CH2-CO2H, -CH2CH2CO2H, or other suitable organic moiety thus act as the “cap” for the terminal PEG subunit of the PEG unit.

[0442] Unless the context otherwise indicates or implies, as used herein, "parallel linker subunit" refers to the organic portion of the drug linker portion of a drug linker compound or ligand-drug conjugate compound, which typically exists as a subunit of the first or second extended subunit within the linker unit, wherein the parallel linker subunit (L... P This allows the PEG unit to be oriented in a parallel direction toward the hydrophobic drug unit (referred to herein as the hydrophobic drug unit) to at least partially reduce the hydrophobicity of the drug unit. P The structures of the associated PEG units and PEG-capped units are described in WO 2015 / 5057699 (which is expressly incorporated herein by reference), and in some respects, L PIt is a trifunctional α-amino acid, β-amino acid, or other trifunctional amino acid residue.

[0443] As used herein, "intracellularly cleaved," "intracellular cleavage," and similar terms refer to metabolic processes or reactions occurring within target cells against ligand-drug conjugates, thereby disrupting the covalent attachment between the drug unit and ligand unit of the conjugate via a linker unit, resulting in the release of D as a free drug within the target cell. As described herein, in some embodiments, D is initially released as an adduct of the drug unit with one or more suicide spacers, which subsequently spontaneously dissociate from the drug unit to release D as a free drug.

[0444] Unless the context otherwise indicates or implies, as used herein, "hematologic malignancy" refers to a blood cell tumor originating from cells of lymphoid or bone marrow origin and is synonymous with the term "liquid tumor." Hematologic malignancies can be classified as indolent, moderately aggressive, or highly aggressive.

[0445] Unless the context otherwise requires, “lymphoma” as used herein refers to a blood malignancy typically formed by overproliferating cells of lymphoid origin. Lymphomas are sometimes classified into two main types: Hodgkin lymphoma (HL) and non-Hodgkin lymphoma (NHL). Lymphomas can also be classified according to the normal cell type that most closely resembles cancer cells, based on phenotypic markers, molecular markers, or cellularity markers. Subtypes of lymphoma under this classification include, but are not limited to, mature B-cell vegetations, mature T-cell and natural killer (NK) cell vegetations, Hodgkin lymphoma, and immunodeficiency-associated lymphoproliferative disorders. Lymphoma subtypes include precursor T-cell lymphoblastic lymphoma (sometimes called lymphoblastic leukemia because T-cell lymphoblasts are produced in the bone marrow), follicular lymphoma, diffuse large B-cell lymphoma, mantle cell lymphoma, B-cell chronic lymphocytic lymphoma (sometimes called leukemia due to peripheral blood involvement), MALT lymphoma, Burkitt lymphoma, mycosis fungoides and its more aggressive variant Sézary disease, peripheral T-cell lymphoma not otherwise specified, tuberous sclerosis of Hodgkin lymphoma, and mixed-cell subtypes of Hodgkin lymphoma.

[0446] Unless the context otherwise requires, "leukemia" as used herein refers to a blood malignancy typically arising from excessively proliferating cells of bone marrow origin, and includes, but is not limited to, acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL), chronic myeloid leukemia (CML), and acute monocytic leukemia (AMoL). Other leukemias include hairy cell leukemia (HCL), T-cell lymphoblastic leukemia (T-PLL), large granular lymphocytic leukemia, and adult T-cell leukemia.

[0447] Unless the context otherwise indicates or implies, as used herein, “hyperproliferating cell” refers to an abnormal cell characterized by unwanted cell proliferation, or an abnormally high rate or persistent state of cell division, or other cellular activities unrelated to or uncoordinated with the cellular activity of the surrounding normal tissue. In some respects, hyperproliferating cells are hyperproliferating mammalian cells. In other respects, hyperproliferating cells are overstimulated immune cells as defined herein, whose persistent state of cell division or activation occurs after the cessation of the stimulus that may have initially caused the change in cell division. In other respects, hyperproliferating cells are transformed normal or cancerous cells, and their uncontrolled and progressive state of cell proliferation may result in a tumor that is benign, potentially malignant (precancerous), or definitively malignant. Hyperproliferative disorders caused by transformed normal or cancerous cells include, but are not limited to, those characterized by precancerous lesions, hyperplasia, developmental abnormalities, adenomas, sarcomas, blastomas, carcinomas, lymphomas, leukemias, or papillomas. Precancerous lesions are generally defined as lesions exhibiting histological changes and associated with an increased risk of cancer development, and sometimes possessing some, but not all, of the molecular and phenotypic characteristics characterizing cancer. Hormone-related or hormone-sensitive precancerous lesions include, but are not limited to, prostatic intraepithelial neoplasia (PIN) (especially high-grade PIN (HGPIN)), atypical microacinary hyperplasia (ASAP), cervical dysplasia, and ductal carcinoma in situ. Hyperplasia generally refers to the proliferation of cells within an organ or tissue beyond its normally visible extent, which may lead to overall enlargement of the organ or the formation of a benign tumor or growth. Hyperplasia includes, but is not limited to, endometrial hyperplasia (endometriosis), benign prostatic hyperplasia, and ductal hyperplasia.

[0448] Unless the context otherwise requires, "normal cell" as used herein refers to a cell undergoing coordinated cell division in connection with the maintenance of cellular integrity in normal tissues, the replenishment of circulating lymphocytes or blood cells (necessary for regulated cell turnover or tissue repair due to injury), or a regulated immune or inflammatory response induced by pathogen exposure or other cellular damage, wherein the cell division or immune response terminates upon completion of the necessary maintenance, replenishment, or pathogen clearance. Normal cells include normally proliferating cells, normally dormant cells, and normally activated immune cells. Normal cells include normally dormant cells, which are in a resting state... o Non-cancerous cells that are in a state that has not yet been stimulated by stress or mitogens, or immune cells that are normally inactive or have not yet been activated by exposure to pro-inflammatory cytokines.

[0449] Unless the context otherwise indicates or implies, the term "abnormal cell" as used herein refers to a normal cell that is disproportionately responsive to external stimuli or dysfunctional due to its inability to properly regulate its spontaneous intracellular activity (in some cases, with a mutational origin). Abnormal cells include overproliferating cells and overstimulated immune cells, as these terms are defined elsewhere. When present in an organism, these cells typically interfere with the function of other normal cells, thereby causing harm to the organism, and their destructive capacity increases over time. Abnormal cells include cancer cells, overactivated immune cells, and other unwanted cells in the organism. Abnormal cells can also refer to nominally normal cells that exist in an environment of outwardly abnormal cells but still support the proliferation and / or survival of other abnormal cells (such as tumor cells), thus targeting nominally normal cells indirectly inhibits the proliferation and / or survival of tumor cells.

[0450] Unless the context otherwise indicates or implies, "overstimulated immune cells" as used herein refers to cells involved in innate or adaptive immunity characterized by an abnormally persistent state of proliferation or inappropriate stimulation that occurs after the cessation of stimulation (which may initially cause proliferation or changes in stimulation) or in the absence of any external damage. Typically, persistent proliferative or inappropriate stimulation states contribute to the chronic inflammatory state characteristic of disease states or symptoms. In some cases, the stimulus that initially caused the proliferation or changes in stimulation is not attributable to external damage but originates internally, as in autoimmune diseases. In some respects, overstimulated immune cells are pro-inflammatory immune cells that have been overactivated by chronic exposure to pro-inflammatory cytokines.

[0451] In some aspects of the invention, the ligand-drug conjugate composition binds to antigens preferentially displayed by abnormally proliferating, inappropriately or persistently activated pro-inflammatory immune cells. These immune cells include classically activated macrophages or type 1 T helper (Th1) cells that produce interferon-γ (INF-γ), interleukin-2 (IL-2), interleukin-10 (IL-10), and tumor necrosis factor-β (TNF-β), which are involved in macrophage and CD8... + Cytokines that activate T cells.

[0452] Unless the context otherwise indicates or implies, “bioavailability” refers to the systemic utilization (i.e., blood / plasma level) of a given amount of drug administered to a patient. Bioavailability is an absolute term that represents a measure of the time (rate) and total amount (degree) of a drug from the administered dosage form to the total circulation.

[0453] Unless the context otherwise indicates or implies, "subject" refers to a human, non-human primate, or mammal suffering from or susceptible to a hyperproliferative disorder, inflammatory disorder, or immune disorder, or other disorder attributable to abnormal cells, who will benefit from administration of an effective amount of the ligand-drug conjugate. Non-limiting examples of subjects include humans, rats, mice, guinea pigs, monkeys, pigs, goats, cattle, horses, dogs, cats, birds, and poultry. Typically, subjects are humans, non-human primates, rats, mice, or dogs.

[0454] Unless the context otherwise indicates or implies, "carrier" refers to a diluent, adjuvant, or excipient administered with a compound. Such drug carriers can be liquids, such as water and oils, including those of petroleum, animal, plant, or synthetic origin, such as peanut oil, soybean oil, mineral oil, and sesame oil. Carriers can be saline, gum arabic, gelatin, starch paste, talc, keratin, colloidal silica, and urea. Additionally, adjuvants, stabilizers, thickeners, lubricants, and colorants may be used. In one embodiment, the compound or composition and the pharmaceutically acceptable carrier are sterile when administered to a subject. Water is an exemplary carrier when the compound is administered intravenously. Saline solutions and aqueous solutions of dextran and glycerol can also be used as liquid carriers, particularly for injectable solutions. Suitable drug carriers also include excipients such as starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, sodium stearate, glyceryl monostearate, talc, sodium chloride, skim milk powder, glycerol, propylene, glycol, water, and ethanol. If desired, the compositions of the present invention may also contain small amounts of wetting agents, emulsifiers, or pH buffers.

[0455] Unless the context otherwise requires, as used herein, "salt form" refers to a charged compound that ion-associates with one or more counter cations and / or counter anions to form a neutral substance overall. In some respects, the salt form of a compound occurs through the interaction of the basic or acidic functional groups of the parent compound with an external acid or base, respectively. In other respects, the charged atoms of the compound associated with the counter anion are permanent in the sense that they cannot spontaneously dissociate into a neutral substance without altering the structural integrity of the parent compound, such as when a nitrogen atom is quaternized. Thus, the salt form of a compound can comprise the protonated form of the quaternized nitrogen atom and / or the basic functional group within the compound and / or the ionized carboxylic acid of the compound, each of which ion-associates with the counter anion.

[0456] In some respects, the salt form can arise from the interaction between a basic functional group and an ionized acid functional group within the same compound, or involve the inclusion of negatively charged molecules, such as acetate ions, succinate ions, or other counter anions. Therefore, a compound in its salt form may have more than one charged atom in its structure. Where multiple charged atoms of the parent compound are part of the salt form, the salt form can have multiple counter ions, such that the salt form of the compound can have one or more charged atoms and / or one or more counter ions. The counter ions can be any charged organic or inorganic component that stabilizes the parent compound with opposite charges.

[0457] When a basic functional group of a compound (such as a primary, secondary, or tertiary amine or other basic amine functional group) interacts with an organic or inorganic acid having a suitable pKa to protonate the basic functional group, or when a suitable pKa is present... a When the acidic functional group of a compound (such as a carboxylic acid) interacts with a hydroxide salt (such as NaOH or KOH) or an organic base of suitable strength (such as triethylamine) to deprotonate the acidic functional group, the compound is usually obtained in its protonated salt form. In some respects, the salt form of the compound contains at least one basic amine functional group, and thus can form an acid addition salt with that amine group, which includes a basic amine functional group with cyclic or acyclic basic units. In the context of drug linker compounds, a suitable salt form is one that does not unduly interfere with the condensation reaction between the target agent and the drug linker compound (which provides a ligand-drug conjugate).

[0458] Unless the context otherwise requires, as used herein, “pharmaceutically acceptable salt” means a salt form of a compound in which its counterions are acceptable for administration of the salt form to the intended subject and includes both inorganic and organic counterions of cations and anions. Exemplary pharmaceutically acceptable counter anions of basic amine functional groups (such as those in cyclic or acyclic basic units) include, but are not limited to, sulfates, citrates, acetates, oxalates, chlorides, bromides, iodides, nitrates, bisulfates, phosphates, acid phosphates, isonicotinates, lactates, salicylates, acid citrates, tartrates, oleates, tannates, pantothenates, bitartrates, ascorbic acid salts, succinates, maleates, methanesulfonates, benzenesulfonates, gentianates, fumarates, gluconates, glucurons, glycosides, formates, benzoates, glutamates, methanesulfonates, ethanesulfonates, benzenesulfonates, p-toluenesulfonates, and bis(hydroxynaphthyl)ates (i.e., 1,1'-methylene-bis(2-hydroxy-3-naphthyl)ates).

[0459] Generally, pharmaceutically acceptable salts are selected from those described in PHStahl and CGWermuth, eds., Handbook of Pharmaceutical Salts: Properties, Selection and Use, Weinheim / Zürich: Wiley-VCH / VHCA, 2002. The choice of salt depends on the properties the pharmaceutical product must exhibit (including adequate water solubility at various pH values), the intended one or more routes of administration, crystallinity with flow characteristics, and low hygroscopicity suitable for handling (i.e., water absorption relative to relative humidity), and the desired shelf life obtained by determining the chemical and solid-state stability under accelerated conditions when in lyophilized formulations (i.e., for determining degradation or solid-state changes during storage at 40°C and 75% relative humidity).

[0460] Unless the context otherwise indicates or implies, “inhibit,” “inhibition of,” and similar terms mean the reduction of a measurable quantity, or the complete prevention of undesirable activity or outcome. In some respects, undesirable outcomes or activities are associated with abnormal cells and include excessive proliferation, overstimulation, or other dysregulated cellular activity in disease states. The inhibition of such dysregulated cellular activity by ligand-drug conjugates is typically determined relative to untreated cells (dummy cells treated with a medium) in a suitable assay system (e.g., in cell cultures (in vitro) or xenograft models (in vivo)). Typically, ligand-drug conjugates targeting antigens that are absent or have low copy numbers on the target abnormal cells, or genetically engineered to not recognize any known antigens, are used as negative controls.

[0461] Unless the context otherwise requires, “treat,” “treatment,” and similar terms refer to therapeutic treatment that includes preventative measures to prevent recurrence, where the aim is to suppress or slow (relieve) undesirable physiological changes or impairments, such as the development or spread of cancer or tissue damage caused by chronic inflammation. Generally, the beneficial or expected clinical benefits of such therapeutic treatments include, but are not limited to, symptom relief, reduction in disease severity, stabilization of the disease state (i.e., no worsening), delay or slowing of disease progression, improvement or mitigation of the disease state, and remission, whether detectable or undetectable (partial or complete). “Treatment” can also mean extended survival or quality of life compared to expected survival or quality of life without treatment. Patients requiring treatment include those already suffering from a condition or impairment, as well as those susceptible to developing a condition or impairment.

[0462] In the context of cancer, the term "treatment" includes any or all of the following: inhibiting the growth of tumor cells, cancer cells, or tumors; inhibiting the replication of tumor cells or cancer cells; inhibiting the spread of tumor cells or cancer cells; reducing the overall tumor burden or the number of cancer cells; or improving one or more symptoms associated with cancer.

[0463] Unless the context otherwise indicates or implies, the term "therapeuticly effective amount" as used herein refers to the amount of free drug or a ligand-drug conjugate having drug units (which are released as free drug) that is effective in treating a disease or disorder in a mammal. In the case of cancer, a therapeutically effective amount of free drug or a ligand-drug conjugate may reduce the number of cancer cells; reduce tumor size; inhibit (i.e., to some extent slow down and preferably stop) cancer cell infiltration into peripheral organs; inhibit (i.e., to some extent slow down and preferably stop) tumor metastasis; inhibit tumor growth to some extent; and / or alleviate one or more symptoms associated with cancer to some extent. The extent to which the free drug or ligand-drug conjugate can inhibit the growth of existing cancer cells and / or kill existing cancer cells can be cytoseptic and / or cytotoxic. For cancer treatment, efficacy can be measured, for example, by assessing time to disease progression (TTP), determining response rate (RR), and / or overall survival (OS).

[0464] In cases of immune disorders caused by overstimulated immune cells, therapeutically effective doses of a drug can reduce the number of overstimulated immune cells, the degree of stimulation by overstimulated immune cells, and / or the extent of infiltration into other normal tissues, and / or alleviate, to some extent, one or more symptoms associated with immune system dysregulation caused by overstimulated immune cells. Efficacy against immune disorders caused by overstimulated immune cells can be measured, for example, by assessing the levels of one or more inflammatory substitutes (including levels of one or more cytokines such as IL-1β, TNFα, INFγ, and MCP-1) or the number of classically activated macrophages.

[0465] In some aspects of the invention, the ligand-drug conjugate compound associates with an antigen on the surface of a target cell (i.e., an abnormal cell, such as an overproliferating cell or an overstimulated immune cell), and the conjugate compound is then absorbed into the target cell via receptor-mediated endocytosis. Once inside the cell, one or more cleavable units within the linker unit of the conjugate are cleaved, resulting in the release of the drug unit (D) as a free drug. The free drug thus released can then migrate within the cytosol and induce cytotoxic or inhibitory activity, or, in the case of overstimulated immune cells, alternatively inhibit pro-inflammatory signal transduction. In another aspect of the invention, the drug unit (D) is released from the ligand-drug conjugate compound outside but near the target cell, such that the free drug resulting from this release is localized to the desired site of action and can subsequently penetrate the cell rather than be prematurely released at a distal site.

[0466] 2. Implementation Plan

[0467] Various embodiments of the invention are described below, followed by a more detailed discussion of components, such as groups, reagents, and steps, useful in the methods of the invention. Any embodiments selected for the components of the methods may be applied to each aspect of the invention as described herein, or they may relate to a single aspect. In some aspects, the selected embodiments may be combined with any combination suitable for describing auristatin ligand drug conjugates, drug linker compounds, or intermediates thereof having a hydrophobic auristatin F drug unit.

[0468] 2.1 Ligand drug conjugates

[0469] The ligand-drug conjugate (LDC) compounds of the present invention are compounds having a drug unit linked to a ligand unit via an intermediate linker unit (LU), wherein the LU comprises a peptide-cleavable unit that is more readily cleaved by proteolytic hydrolysis of tumor tissue homogenate compared to normal tissue homogenate to release D as a free drug, and the ligand-drug conjugates of the present invention generally have the structure of Formula 1:

[0470] L-[LU-(D')] p’ (1)

[0471] Or its salts, particularly pharmaceutically acceptable salts thereof, wherein L is a ligand unit; LU is a linker unit; D' represents 1 to 4 drug units that bind or structurally correspond to the same free drug for each drug linker portion of formula -LU-(D)'; subscript p' is an integer ranging from 1 to 24, wherein the ligand unit is capable of selectively binding antigens of target aberrant cells, wherein the target antigen is capable of being internalized with the bound conjugate compound to subsequently release the free drug into the cell, wherein each drug linker portion in the ligand-drug conjugate compound has the structure of formula 1A:

[0472]

[0473] Or its salts, particularly pharmaceutically acceptable salts, wherein the -L of the drug connector portion of Formula 1A B -A a -B b - Part typically represents the primary connector (L) of the connector unit (LU) in Equation 1. R ),

[0474] The wavy line indicates covalent attachment to L; L B This is the ligand covalently bound portion; A is the first optional extension subunit; the subscript a is 0 or 1, indicating that A is absent or present, respectively; B is an optional branch unit; the subscript b is 0 or 1, indicating that B is absent or present, respectively; D is the drug unit; the subscript q is an integer ranging from 1 to 4; and L O It is a secondary connector part with the following structure:

[0475]

[0476] The wavy line adjacent to A' represents L. O The site covalently attached to the primary connector; the wavy line adjacent to Y indicates L. O The site covalently attached to the drug unit; A' is the second optional spacer unit, the subscript a' is 0 or 1, indicating that A' is absent or present, respectively; W is the peptide cleavable unit; Y is the spacer unit, and y is 0, 1 or 2, indicating that the spacer unit is absent or that one or two spacer units are present, respectively.

[0477] The ligand-drug conjugate composition comprises a distribution or collection of ligand-drug conjugate compounds and is represented by the structure of Formula 1, wherein the subscript p' is replaced by the subscript p, and the subscript p is a number ranging from about 2 to about 24.

[0478] Conventional ligand-drug conjugates are also represented by Formula 1, but have a peptide-cleavable unit (W) comprising a dipeptide directly covalently attached to D or indirectly via Y, wherein the dipeptide is designed to be selective for specific intracellular proteases (whose activity is upregulated in abnormal cells relative to normal cells). In contrast, the conjugates of the present invention are based on the unexpected discovery that the overall protease activity within tissues containing abnormal cells can be distinguished from that in normal tissues containing normal cells by a suitably designed cleavable unit that simultaneously maintains resistance to cleavage by freely circulating proteases. For the conjugates of the present invention, this distinction is achieved by incorporating a peptide-cleavable unit of certain tripeptides, wherein these tripeptides have been identified by the screening methods described herein, in which the protease activity from tissue homogenates containing abnormal cells is compared with the protease activity from normal tissue homogenates, wherein normal tissue is known to be the origin of one or more on-target and / or off-target adverse events experienced by mammalian subjects when administered a therapeutically effective amount of a conventional ligand-drug conjugate.

[0479] Therefore, in a primary embodiment of the invention, W is a peptide-cleavable unit comprising a tripeptide, which provides a recognition site that is selectively acted upon by one or more intracellular proteases of the target abnormal cells, compared to freely circulating proteases, and is also selectively acted upon by proteases in tumor tissue homogenates, compared to proteases in normal tissue homogenates. For cancer treatment, the tripeptide sequence of the peptide-cleavable unit is selected such that proteases in normal tissues, known to be the origin of on-target and / or off-target adverse events resulting from the administration of therapeutically effective amounts of conventional ligand drug conjugates, are less likely to act on conjugates having a tripeptide-based cleavable unit compared to proteases in tumor tissues, thereby providing greater selectivity for targeting cancer cells. This selection is based on the lower overall protease activity in normal tissue homogenates compared to tumor tissue homogenates of cancer. Compared to the improved conjugates of the present invention, conventional ligand drug conjugates containing dipeptide-cleavable units are designed to be selectively acted upon by cathepsin B and rely primarily on the immunological specificity for selectively targeting cancer cells rather than normal cells; cathepsin B is an intracellular protease whose activity is upregulated in cancer cells. The improved conjugate of the present invention has an additional level of selectivity because it is less susceptible to protease action in normal tissue compared to tumor tissue containing target cancer cells.

[0480] In some embodiments, the drug delivery linker portion of Formula 1A will have a structure represented by Formula 1B:

[0481]

[0482] Where L BIt is a primary connector (L) in a connector unit (LU) for a drug connector portion or drug connector compound, as defined herein. R The ligand covalently bound portion of ); A and B are L R The first optional extension subunit and optional branching unit; the subscript q ranges from 1 to 4; and the remaining variable groups are as described in this paper for L O Defined.

[0483] In some of those implementations, W contains a tripeptide directly attached to the drug unit, so the subscript y is 0. When the subscript y is 1, the tripeptide is attached to a suicide spacer unit, so cleavage of the protease provides a drug linker fragment of formula YD, where Y commits suicide to complete the release of the free drug. When the subscript y is 2, the tripeptide is attached to a first suicide spacer unit (Y), so cleavage of the protease provides a first drug linker fragment of formula Y-Y'-D, where Y' is a second spacer unit and, after the suicide of the first spacer unit, provides a second drug linker fragment of formula Y'-D, which decomposes to complete the release of the free drug.

[0484] An exemplary ligand-drug conjugate compound having a drug linker portion of Formula 1B (where a tripeptide of a peptide-cleavable unit (W) is directly attached to a drug unit or attached to an intermediate spacer subunit) has the structure of Scheme 1a, wherein P1, P2, and P3 are amino acid residues of a tripeptide sequence, and D is attached to a p-aminobenzyl alcohol residue via a carbamate or carbonate functional group, the carbamate or carbonate functional group together with the p-aminobenzyl alcohol residue representing Y. y Where the subscript y is 2. In those exemplary ligand-drug conjugate compounds, the carbonyl functional group of the amide bond adjacent to P1 originates from the C-terminus of the tripeptide sequence, where the amide bond is the site of protease cleavage (indicated by the arrow), while the amino group of the amide bond adjacent to P3 originates from the N-terminus of the tripeptide sequence. Cleavage of the amide functional group to P1 yields a first drug linker fragment having the structure shown in scheme 1a, the first drug linker fragment undergoes suicide to provide a second drug linker fragment, the second drug linker fragment spontaneously decomposes and releases CO2 to complete the release of D as a free drug of formula HT*-D*, which has a hydroxyl or amine group, and its oxygen or nitrogen moiety -NH- is represented by T*, where D* represents the remaining portion of the free drug.

[0485] Option 1a.

[0486]

[0487] In those implementations, one or more amino acids, referred to as P4, P5, etc., may be present in formula -L B -A' a’Between the primary linker and P3, as part of a peptide sequence comprising a tripeptide, the tripeptide imparts selectivity for intracellular proteolysis over proteolysis by freely circulating proteases, and selectivity for proteolysis of tumor tissue homogenates over proteolysis of normal tissue homogenates. The mechanism for releasing free drug from the ligand-drug conjugate having this extended peptide sequence is similar to that of scheme 1a.

[0488] In other embodiments, an amino acid residue called P-1 is inserted into the specific tripeptide W and D or -Y. y Between -D, such that D, or the drug linker fragment initially released by protease action at the specific tripeptide, contains this amino acid, thus requiring further processing by an intracellular endopeptidase to allow the suicide of one or more spacer subunits. For those embodiments, exemplary ligand-drug conjugate compounds having the drug linker portion of Formula 1B (where the specific tripeptide of the peptide-cleavable unit is not directly attached to the drug unit or is attached to an intermediate spacer subunit) have the structure shown in Scheme 1b. Protease cleavage of the sensitive amide bond (indicated by the arrow) between P1 and P-1 provides the drug linker fragment, wherein the first suicide spacer subunit (Y) is present as an amino acid residue that provides the substrate for the endopeptidase to attach to the suicide portion of Y, where Y is a p-aminobenzyl alcohol residue attached to D via a carbamate or carbonate functional group. The amino acid-p-aminobenzyl alcohol residue and the carbamate or carbonate functional group together represent Y. y , where the subscript y is 2. After the endopeptidase removes P-1, suicide occurs as in scheme 1a, releasing D as a free drug of formula HT*-D*.

[0489] Option 1b

[0490]

[0491]

[0492] As before, one or more amino acids, referred to as P4, P5, etc., can be present in formula -L B -A' a’ Between the primary linker and P3, as part of a peptide sequence comprising a tripeptide, the tripeptide imparts selectivity for intracellular proteolysis over that of free-circulating proteases and for proteolysis of tumor tissue homogenates over that of normal tissue homogenates. Although P-1 in scheme 1b is formally part of the first suicide spacer subunit (Y), for convenience it will be associated with the tripeptide sequence, and thus W is a tetrapeptide represented by the SEQ ID describing such a peptide-cleavable unit. Those units and other components of the ligand-drug conjugate of the present invention are further discussed below.

[0493] 2.2.1 ligand unit

[0494] The ligand unit (L) of a ligand-drug conjugate is the targeting portion of the conjugate, selectively binding to the target portion. In some embodiments, the ligand unit selectively binds to a cellular component (cell binder) or other target molecule that serves as the target portion. The role of the ligand unit is to target and present the drug unit of the ligand-drug conjugate to a specific target cell population, whereby the ligand unit interacts with the target cell population to selectively release D as a free drug. Targeting agents providing the ligand unit include, but are not limited to, proteins, peptides, and polypeptides. Exemplary ligand units include, but are not limited to, those provided by proteins, peptides, and polypeptides, such as antibodies (e.g., full-length antibodies and their antigen-binding fragments), interferons, lymphokines, hormones, growth factors, and colony-stimulating factors. Other suitable ligand units are those derived from vitamins, nutrient transport molecules, or any other cell-binding molecule or substance. In some embodiments, the ligand unit is derived from a non-antibody protein targeter. In other embodiments, the ligand unit is derived from a protein targeter, such as an antibody. Preferred targeters are proteins with a large molecular weight, such as cell binders having a molecular weight of at least about 80 kDa.

[0495] Primary linker precursor (L) of targeted drug linker compounds R ') ligand covalently bound precursor (L B Partial reaction, forming a primary connector (L) with the drug-connector portion of Formula 1A. R ) ligand covalently bound (L B Partially covalently attached ligand units. The target agent has, or is modified to have, an appropriate number of attachment sites to accommodate the necessary number of drug-connector moieties as defined by the subscript p, whether the attachment sites are naturally occurring or non-natural (e.g., engineered). For example, to have a value of 6 to 14 for the subscript p, the target agent must be able to form bonds with 6 to 14 drug-connector moieties. Attachment sites can be naturally occurring or engineered into the target agent. The target agent can bond with the connector unit of the drug-connector compound via the reactivity of the target agent or the activation of heteroatoms or heteroatom-containing functional groups. SS Partial bond formation. Reactive or activatable heteroatoms or heteroatom-containing functional groups that may be present on the target include sulfur (in one embodiment, a thiol functional group from the target), C=O (in one embodiment, a carbonyl, carboxyl, or hydroxyl group from the target), and nitrogen (in one embodiment, a primary or secondary amino group from the target). These heteroatoms may be present on the target in their native state (e.g., naturally occurring antibodies) or may be introduced into the target via chemical modification or genetic engineering.

[0496] In one embodiment, the target agent has a thiol functional group (e.g., a thiol functional group of a cysteine ​​residue), and a ligand unit therefrom is attached to the drug linker portion of the ligand-drug conjugate compound via a sulfur atom of the thiol functional group.

[0497] In another embodiment, the target agent has an L-shaped linker unit that can connect with a drug linker compound. R The lysine residues react with the activated esters (including but not limited to N-hydroxysuccinimide ester, pentafluorophenyl ester, and p-nitrophenyl ester), thus generating an amide bond between the nitrogen atom in the ligand unit and the C=O functional group in the linker unit of the drug linker compound.

[0498] In yet another embodiment, the target agent has one or more lysine residues that can be chemically modified to introduce one or more thiol functional groups. The ligand unit of the target agent is attached to the linker unit via the sulfur atom of the introduced thiol functional group. Reagents that can be used to modify lysine include, but are not limited to, N-succinimide-S-acetylthioacetate (SATA) and 2-iminothionane hydrochloride (Traut reagent).

[0499] In another embodiment, the target agent may have one or more carbohydrate groups, which may be chemically modified to have one or more thiol functional groups. A ligand unit from the target agent is attached to a linker unit via a sulfur atom of the introduced thiol functional group, or the target agent may have one or more carbohydrate groups that can be oxidized to provide an aldehyde (-CHO) group (see, for example, Laguzza et al., 1989, J. Med. Chem. 32(3):548-55). The corresponding aldehyde can then be reacted with the L of a drug linker compound having a nucleophilic nitrogen atom. SS Partial reaction. L R Other reaction sites that can react with the carbonyl group on the target agent include, but are not limited to, hydrazine and hydroxylamine. Other protocols for modifying proteins to attach drug linker moieties are described in Coligan et al., Current Protocols in Protein Science, Vol. 2, John Wiley & Sons (2002) (incorporated hereby by reference).

[0500] In a preferred embodiment, the L of the drug connector compound R The reactive group is maleimide (M 1 ) part, and L with L R The covalent attachment is achieved through the thiol functional group of the target agent, therefore the sulfur-substituted succinimide (M 2The thiol functional group is formed via Michael addition. The thiol functional group can exist on the target in its natural state (e.g., naturally occurring residues) or can be introduced into the target via chemical modification and / or genetic engineering.

[0501] For bioconjugates, drug conjugation sites have been observed to influence numerous parameters, including conjugation ease, drug-linker stability, effects on the biophysical properties of the resulting bioconjugate, and in vitro cytotoxicity. Regarding drug-linker stability, the conjugation site between the drug-linker and ligand can affect the ability of the conjugated drug-linker moiety to undergo elimination reactions and the ability of the drug-linker moiety to transfer from the ligand unit of the bioconjugate to alternative reactive thiols present in the bioconjugate environment (e.g., reactive thiols in albumin, free cysteine, or glutathione in plasma). Such sites include, for example, interchain disulfides and selectively engineered cysteine ​​sites. Among other sites, the ligand-drug conjugates described herein can be conjugated to thiol residues at sites less sensitive to elimination reactions (e.g., position 239 according to the EU index described in Kabat).

[0502] In a preferred embodiment, the ligand unit (L) is an antibody or an antigen-binding fragment thereof, thereby defining the antibody ligand unit of an antibody-drug conjugate (ADC), wherein the antibody ligand unit is capable of selectively binding to a target antigen of a cancer cell to subsequently release D as a free drug, wherein the target antigen is capable of being internalized into the cancer cell after binding to initiate intracellular release of the free drug.

[0503] Useful antibodies include polyclonal antibodies, which are heterogeneous groups of antibody molecules derived from the serum of immunized animals. Other useful antibodies are monoclonal antibodies, which are homogeneous groups of antibodies targeting specific antigenic determinants (e.g., cancer cell antigens, viral antigens, microbial antigens, proteins, peptides, carbohydrates, chemicals, nucleic acids, or fragments thereof). Monoclonal antibodies (mAbs) against a target antigen can be prepared using any technique known in the art, which provides the production of antibody molecules through continuous cell lines in culture.

[0504] Useful monoclonal antibodies include, but are not limited to, human monoclonal antibodies, humanized monoclonal antibodies, or chimeric human-mouse (or other species) monoclonal antibodies. Antibodies include full-length antibodies and their antigen-binding fragments. Human monoclonal antibodies can be prepared by any of the many techniques known in the art (e.g., Teng et al., 1983, Proc. Natl. Acad. Sci. USA. 80:7308-7312; Kozbor et al., 1983, Immunology Today 4:72-79; and Olsson et al., 1982, Meth. Enzymol. 92:3-16).

[0505] Antibodies can be immune-specific antibodies that bind to target cells (e.g., cancer cell antigens, viral antigens, or microbial antigens) or functionally active fragments, derivatives, or analogs of other antibodies that bind to tumor cells or the matrix. In this regard, “functionally active” means that the fragment, derivative, or analog can bind to target cells immune-specifically. To determine which CDR sequences bind to antigens, synthetic peptides containing CDR sequences can be used in binding assays of antigens by any binding assay known in the art (e.g., BIA core assay) (see, for example, Kabat et al., 1991, Sequences of Proteins of Immunological Interest, 5th ed., National Institute of Health, Bethesda, MD; Kabat E et al., 1980, J. Immunology 125(3):961-969).

[0506] Other useful antibodies include antibody fragments, such as, but not limited to, F(ab')2 fragments, Fab fragments, Fvs, single-chain antibodies, biantibodies, triantibodies, tetraantibodies, scFv, scFv-FV, or any other molecule with the same specificity as an antibody.

[0507] Additionally, recombinant antibodies comprising both human and non-human portions (such as chimeric and humanized monoclonal antibodies) can be prepared using standard recombinant DNA techniques and are useful antibodies. Chimeric antibodies are molecules in which different portions are derived from different animal species, such as those having a variable region derived from mouse monoclonal antibodies and a constant region from human immunoglobulins. (See, for example, U.S. Patent Nos. 4,816,567 and 4,816,397, which are incorporated herein by reference in their entirety). Humanized antibodies are antibody molecules derived from non-human species that have one or more complementarity-determining regions (CDRs) derived from the non-human species and a framework region derived from human immunoglobulin molecules. (See, for example, U.S. Patent No. 5,585,089, which is incorporated herein by reference in its entirety). Such chimeric and humanized monoclonal antibodies can be produced using recombinant DNA techniques known in the art, for example, using methods described in the following documents (each of which is expressly incorporated herein by reference): International Publication No. WO 87 / 02671; European Patent Publication No. 0 184 187; European Patent Publication No. 0 171 496; European Patent Publication No. 0 173 494; International Publication No. WO 86 / 01533; US Patent No. 4,816,567; European Patent Publication No. 012 023; Berter et al., Science (1988) 240:1041-1043; Liu et al., Proc. Natl. Acad. Sci. (USA) (1987) 84:3439-3443; Liu et al., J. Immunol. (1987) 139:3521-3526; Sun et al., Proc. Natl. Acad. Sci. (USA) (1987) 84:214-218; Nishimura et al., Cancer. Res. (1987) 47:999-1005; Wood et al., Nature (1985) 314:446-449; Shaw et al., J. Natl. Cancer Inst. (1988) 80:1553-1559; Morrison, Science (1985) 229:1202-1207; Oi et al. BioTechniques (1986) 4:214; US Patent No. 5,225,539; Jones et al., Nature (1986) 321:552-525; Verhoeyan et al., Science (1988) 239:1534; and Beidler et al., J. Immunol. (1988) 141:4053-4060.

[0508] Fully human antibodies are particularly preferred and can be produced using transgenic mice that do not express endogenous immunoglobulin heavy and light chain genes but can express human heavy and light chain genes.

[0509] Antibodies include analogs and derivatives that have been modified (i.e., by covalent attachment of any type of molecule if such covalent attachment allows the antibody to maintain its antigen-binding immune specificity). For example, but not limited to, antibody derivatives and analogs include those that have been further modified, for example by glycosylation, acetylation, polyethylene glycolation, phosphorylation, amidation, derivatization by known protecting / blocking groups, proteolytic cleavage, linking to cellular antibody units or other proteins, etc. Any of a number of chemical modifications can be performed using known techniques, including but not limited to specific chemical cleavage, acetylation, formylation, metabolic synthesis in the presence of tunicamycin, etc. Additionally, analogs or derivatives may contain one or more non-natural amino acids.

[0510] Antibodies may have modifications (e.g., substitution, deletion, or addition) in amino acid residues that interact with the Fc receptor. In particular, antibodies may have modifications in amino acid residues identified as being involved in the interaction between the anti-Fc domain and the FcRn receptor (see, for example, International Publication No. WO 97 / 34631, which is incorporated herein by reference in its entirety).

[0511] In specific implementations, known antibodies used for cancer treatment are employed. In some implementations, the antibody will selectively bind to cancer antigens of blood malignancies.

[0512] 2.2.2 Primary connector

[0513] In one set of embodiments, the ligand-drug conjugate comprises one or more of the formula -L R -L O -D drug connector portion, where L O As described in this article -A' a’ -WY y -, where L RA' is a primary linker, a' is a second optional extension subunit, a' is 0 or 1, indicating the absence or presence of A', respectively; Y is a spacer subunit, the subscript y is 0, 1 or 2, indicating the absence or presence of 1 or 2 spacer subunits, respectively; D is a drug unit, and W is a peptide-cleavable unit, wherein the peptide-cleavable unit is a sequence of up to 12 (e.g., 3-12 or 3-10) consecutive amino acids, wherein the sequence comprises a tripeptide, which is more readily cleaved by proteolytic cleavage of tumor tissue homogenate to initiate the release of D as a free drug compared to normal tissue homogenate, wherein cytotoxicity against normal tissue cells (due to the unintended release of the free drug into and / or near these cells) is associated with adverse events resulting from administration of an effective amount of the comparative ligand drug conjugate (whose amino acid sequence of the peptide-cleavable unit is dipeptide-valine-citrulline-) to a subject in need, and / or wherein the tripeptide increases the bioavailability of the ligand drug conjugate compared to the comparative conjugate, but is detrimental to its bioavailability against normal tissue. In some of those implementation schemes, -L R -is-L B -A a -B b -, where L B It is the ligand covalently bound part, A is the first optional extension subunit, the subscript a is 0 or 1, which means A does not exist or exists, respectively, and B is an optional branch unit, the subscript b is 0 or 1, which means B does not exist or exists, respectively.

[0514] In some implementations, the drug connector portion has the following structure:

[0515]

[0516] Or its salts, especially pharmaceutically acceptable salts, wherein L R A', a', Y, y, and D retain their previous meanings, and P1, P2, and P3 are amino acid residues that, compared to the comparative ligand drug conjugate (whose peptide-cleavable unit amino acid sequence is dipeptide-valine-citrulline-), provide, together, the selectivity of proteolysis by tumor tissue homogenate to proteolysis by normal tissue homogenate and / or provide increased bioavailability to tumor tissue but disadvantageous bioavailability to normal tissue, wherein if the subscript y is 1 or 2, proteolytic cleavage occurs at the covalent bond between P1 and Y, or if the subscript y is 0, proteolytic cleavage occurs at the covalent bond between P1 and D, and wherein the tumor tissue and normal tissue belong to the same species.

[0517] As described elsewhere, other embodiments contain an additional amino acid residue, referred to as P-1, between P1 and Y or D (depending on the value of the subscript y), such that the selective endopeptidase action of one or more proteolytic enzymes in the tumor tissue homogenate occurs at the amide bond between P1 and P-1, releasing the -[P-1]-Y y -D drug linker fragment. If the subscript y is 0 (i.e. Y does not exist), the release of free drug from this fragment will occur via the action of an exopeptidase of a proteolytic enzyme to remove the P-1 amino acid residue, thereby directly providing free drug.

[0518] In some embodiments where additional amino acid residues exist between P1 and Y or D, the drug linker portion has the following structure:

[0519]

[0520] Or its salts, especially pharmaceutically acceptable salts, wherein L R A', a', Y, y, and D retain their previous meanings, and P1, P2, and P3 are amino acid residues that, optionally together with P-1, provide selectivity for proteolysis of tumor tissue homogenates over normal tissue homogenates, wherein proteolytic cleavage occurs at the covalent bond between P1 and P-1 to release [P-1]-Y y -D structure connector segment.

[0521] In some of those embodiments, when the subscript y is 0, the [P-1]-D residue resulting from the cleavage of the amide bond between the P1 and P-1 amino acids by the endopeptidase also exerts cytotoxic activity. In other embodiments, the subscript y is 1 or 2, thus the action of the exopeptidase to remove the P-1 amino acid residue provides formula -Y y -D is another drug linker fragment that spontaneously fragments to provide free drug.

[0522] In other embodiments, one or more amino acid residues referred to as P4, P5…Pn (where the subscript n ranges up to 12 (e.g., 3-12 or 3-10)) are located at P3 and L R Or between A' (depending on the value of the subscript a'), in some embodiments, it is also a peptide cleavable unit containing the P-1 amino acid residue. In either case, alternative P4, P5…P are selected. n Amino acid residues, so as not to change the supply of -Y y -D or -[P-1]-Y y -D fragment cleavage sites, but impart the desired physicochemical and / or pharmacokinetic properties to the ligand-drug conjugate, such as increased solubility to reduce aggregation.

[0523] In some embodiments, where one or more additional amino acid residues are present at the N-terminus of P3 or where P-1 is additionally present between P1 and Y or D, the drug linker portion has the following structure:

[0524]

[0525]

[0526] Or its salts, especially pharmaceutically acceptable salts, wherein L R A', a', Y, y, and D retain their previous meanings, and P-1 and P1, P2, P3…P n These are amino acid residues, where the subscript n ranges up to 12 (e.g., 3-12 or 3-10), and P1, P2, and P3, optionally together with P-1, provide selectivity for proteolysis of tumor tissue homogenates over proteolysis of normal tissue homogenates, wherein proteolytic cleavage occurs at P1 and Y. y At the covalent bonds between -D or between P1 and P-1, to release the Y respectively y -D or [P-1]-Y y -D linker fragments, which are subsequently cleaved by exopeptidase to release a Y-type linker fragment. y -D structured drug fragments. In both cases, Y y -D linker fragments will spontaneously decompose to release D as a free drug.

[0527] In any of those implementation schemes, when the subscript b is 0, the L of the drug connector portion R With formula -L B -A a -, where L B A is the ligand covalently bound portion, and A is the first optional extended subunit. In such embodiments, if a is 1 and the subscript a' is 1, then A' exists as a subunit of A and is therefore considered a component of the primary linker.

[0528] In some preferred embodiments where subscript b is 0 and subscript a is 1, equation -L B -A- of L R It is a self-stabilizing connector (L SS ) part or from L SS Succinimide (M 2 Self-stabilizing joints (L) obtained by partially controlled hydrolysis S ) portion. An exemplary L of a ligand-drug conjugate composition or its conjugate compound having a primary linker of any type. SS and L SPrimary connectors are represented by the following structures or their salts, particularly pharmaceutically acceptable salts:

[0529]

[0530] The wavy line indicates a site covalently attached to A' or W, depending on the value of the subscript a'; A' is an optional subunit of A; [HE] is an optional hydrolysis-enhancing unit, a component provided by A; BU is a basic unit; R a2 It is an optional substitution of C1-C 12 Alkyl; the dashed curve represents optional cyclization, thus in the absence of said cyclization, BU is an acyclic basic unit having a primary, secondary, or tertiary amine functional group as the basic functional group of the acyclic basic unit, or in the presence of said cyclization, BU is a cyclized basic unit, wherein R a2 Together with BU and the carbon atoms attached to both, we define the optionally substituted spiro C3-C. 20 Heterocyclic compounds, which contain basic nitrogen atoms in the framework of secondary or tertiary amine functional groups as basic functional groups in the cyclic basic units.

[0531] The basic nitrogen atom of the acyclic basic unit or the cyclic basic unit is optionally protected by a nitrogen protecting group, depending on the degree of substitution of the basic nitrogen atom, or optionally protonated.

[0532] In other preferred embodiments where subscript b is 0 and subscript a is 1, formula -L B The primary connector of the -A- series does not contain a basic unit, and an example of its structure is as follows:

[0533]

[0534] Or its salts, particularly pharmaceutically acceptable salts, wherein variable groups such as those previously targeted at L SS or L S As described in the primary connector.

[0535] Representative LL R - Structure (where L) R The ligand unit (L) covalently attaches to LDC as follows:

[0536]

[0537] And its salts, particularly pharmaceutically acceptable salts, wherein the structure of the succinimide ring system is hydrolyzed into an open-ring form, wherein the (#) sulfur atom shown is derived from the ligand unit; and wherein the wavy line indicates the site covalently attached to the remainder of the conjugate structure.

[0538] Other representative LL R -The structure is as follows:

[0539]

[0540] The nitrogen, carbon, or sulfur atoms indicated by (#) are derived from ligand units; and the wavy lines represent sites covalently attached to the remainder of the conjugate structure.

[0541] In another set of embodiments, the drug linker compound used to prepare the ligand drug conjugate as described in the previous set of embodiments has the formula L as described herein. R '-A' a’ -WY y -D, where L R ' is a primary linker for drug linker compounds. When drug linker compounds are used to prepare conjugates, L R 'Primary linker L of the drug linker portion of the conversion to a ligand-drug conjugate R A' is the second optional extended subunit, and a' is 0 or 1, representing the absence or presence of A', respectively. R When 'a' does not contain a branch unit and the subscript a' is 1, A' is considered as a subunit of A and is thus considered L. R 'part of and as L R The components are present, Y is a spacer unit, the subscript y is 0, 1 or 2, indicating the absence of the spacer unit or the presence of 1 or 2 spacer units, respectively, D is a drug unit, and W is a peptide-cleavable unit containing a tripeptide, which is more susceptible to proteolytic cleavage by tumor tissue homogenate compared to normal tissue homogenate, wherein cytotoxicity against normal tissue cells (due to the unintended release of D as a free drug into and / or near these cells) is associated with adverse events resulting from the administration of a ligand-drug conjugate for targeting cancer cells in tumor tissue. In some embodiments of those embodiments, L R '- is L B '-A a -B b -, where L B ' is the ligand covalently bound portion of the primary linker in a drug linker compound, sometimes referred to as the ligand covalently bound precursor portion, because when a drug linker compound is used to prepare conjugates, it is the primary linker (L) of the drug linker portion of the ligand-drug conjugate. R The ligand covalently bound portion (L) B The precursor of ) is A, which is the first optional extension subunit, with subscript a being 0 or 1, indicating that A does not exist or exists, respectively. B is an optional branch unit, with subscript b being 0 or 1, indicating that B does not exist or exists, respectively.

[0542] In some embodiments, the drug connector compound has the following structure:

[0543] L R′-A′ a′ -[P3]-[P2]-[P1]-Y y -D

[0544] Or its salts, especially pharmaceutically acceptable salts, wherein L R ', A', a', Y, y, and D retain their previous meanings, and P1, P2, and P3 are amino acid residues that together provide selectivity for proteolysis of tumor tissue homogenates over that of normal tissue homogenates, wherein if the subscript y is 1 or 2, proteolytic cleavage occurs at the covalent bond between P1 and Y, or if the subscript y is 0, proteolytic cleavage occurs at the covalent bond between P1 and D.

[0545] As described elsewhere, other embodiments contain an additional amino acid residue, referred to as P-1, between P1 and Y or D (depending on the value of the subscript y), such that the selective endopeptidase action of one or more proteolytic enzymes in the tumor tissue homogenate occurs at the amide bond between P1 and P-1, releasing the -[P-1]-Y y -D drug linker fragment. If the subscript y is 0 (i.e. Y does not exist), the release of free drug from this fragment will occur via the action of an exopeptidase of a proteolytic enzyme to remove the P-1 amino acid residue, thereby directly providing free drug.

[0546] In some embodiments where additional amino acid residues exist between P1 and Y or D, the drug linker compound has the following structure:

[0547] L R ′-A′ a′ -[P3]-[P2]-[P1]-[P-1]-Y y -D

[0548] Or its salts, especially pharmaceutically acceptable salts, wherein L R ', A', a', Y, y, and D retain their previous meanings, and P1, P2, and P3 are amino acid residues that, optionally together with P-1, provide selectivity for proteolysis of tumor tissue homogenates over normal tissue homogenates, wherein proteolytic cleavage occurs at the covalent bond between P1 and P-1 to release [P-1]-Y y -D structure connector segment.

[0549] In some of those embodiments, when the subscript y is 0, the [P-1]-D residue resulting from the cleavage of the amide bond between the P1 and P-1 amino acids by the endopeptidase also exerts cytotoxic activity. In other embodiments, the subscript y is 1 or 2, thus the action of the exopeptidase to remove the P-1 amino acid residue provides formula -Y y-D is another drug linker fragment that spontaneously fragments to provide free drug.

[0550] In other embodiments, one or more amino acid residues referred to as P4, P5…Pn (where the subscript n ranges up to 12 (e.g., 3-12 or 3-10)) are located at P3 and L R Or between A' (depending on the value of the subscript a'), in some embodiments, it is also a peptide cleavable unit containing the P-1 amino acid residue. In either case, alternative P4, P5…P are selected. n Amino acid residues, so as not to change the supply of -Y y -D or -[P-1]-Y y -D fragment cleavage sites, but impart the desired physicochemical and / or pharmacokinetic properties to the ligand-drug conjugate, such as increased solubility to reduce aggregation.

[0551] In some embodiments, where one or more additional amino acid residues are present at the N-terminus of P3 or where P-1 is additionally present between P1 and Y or D, the drug connector compound has the following structure:

[0552] L R′ -A′ a′ -[P n ...[P4]-[P3]-[P2]-[P1]-Y y -D or

[0553] L R′ -A′ a′ -[P n ]...[P4]-[P3]-[P2]-[P1]-[P-1]-Y y -D

[0554] Or its salts, especially pharmaceutically acceptable salts, wherein L R ', A', a', Y, y, and D retain their previous meanings, and P-1 and P1, P2, P3…P n These are amino acid residues, where the subscript n ranges up to 12 (e.g., 3-12 or 3-10), and P1, P2, and P3, optionally together with P-1, provide selectivity for proteolysis of tumor tissue homogenates over proteolysis of normal tissue homogenates, wherein proteolytic cleavage occurs at P1 and Y. y At the covalent bonds between -D or between P1 and P-1, to release the Y respectively y -D or [P-1]-Y y -D linker fragments, which are subsequently cleaved by exopeptidase to release a Y-type linker fragment. y -D structured drug fragments. In both cases, Y y-D connector fragments undergo spontaneous decomposition (also known as suicide) to complete the release of D as a free drug.

[0555] In any of those implementation schemes, when the subscript b is 0, the L of the drug connector compound R 'Having the formula L B '-A a -, where L B A' is the ligand covalently bound precursor portion and A is the first optional extended subunit. In such embodiments, if subscript a is 1 and subscript a' is 1, then A' exists as a subunit of A and is therefore considered a component of the primary linker.

[0556] In some preferred embodiments where subscript b is 0 and subscript a is 1, the pharmaceutical connector compound of formula L B '-A- of L R 'It is a self-stabilizing connector precursor (L) SS The part is named as such because when the drug linker compound is used to prepare conjugates, it transforms into a self-stabilizing linker (L) of the ligand-drug conjugate. SS ( ) section. Exemplary L of drug connector compounds SS 'Primary connectors are represented by the following structures or their salts, particularly pharmaceutically acceptable salts:'

[0557]

[0558] The wavy line indicates a site covalently attached to A' or W, depending on the value of the subscript a'; A' is an optional subunit of A; [HE] is an optional hydrolysis-enhancing unit, a component provided by A; BU is a basic unit; R a2 It is an optional substitution of C1-C 12 Alkyl; the dashed curve represents optional cyclization, thus in the absence of said cyclization, BU is an acyclic basic unit having a primary, secondary, or tertiary amine functional group as the basic functional group of the acyclic basic unit, or in the presence of said cyclization, BU is a cyclized basic unit, wherein R a2 Together with BU and the carbon atoms attached to both, we define the optionally substituted spiro C3-C. 20 Heterocyclic compounds, wherein the basic nitrogen atom of the skeleton containing a secondary or tertiary amine functional group serves as the basic functional group of the cyclic basic unit, wherein the basic nitrogen atom of the acyclic basic unit or the cyclic basic unit is optionally appropriately protected by a nitrogen protecting group, depending on the degree of substitution of the basic nitrogen atom, or optionally protonated.

[0559] In other preferred embodiments where subscript b is 0 and subscript a is 1, formula L B The primary connector of the -A- series does not contain a basic unit, and an example of its structure is as follows:

[0560]

[0561] Or its salts, particularly pharmaceutically acceptable salts, wherein variable groups such as those previously targeted at L SS or L S As described in the primary connector.

[0562] Representative L of drug linker compounds R The structure is as follows:

[0563]

[0564] And its salts, particularly pharmaceutically acceptable salts, wherein the wavy line indicates a site covalently attached to the remainder of LU' in the drug linker compound structure, and wherein the basic nitrogen atom in the second or third structure is optionally protonated to an acid-adding salt or optionally protected. When protected, the protecting group is preferably an acid-unstable protecting group, such as BOC.

[0565] 2.2.3 Peptide cleavable unit

[0566] In some embodiments, the peptide-cleavable unit (W) of the ligand-drug conjugate is a peptide sequence comprising a tripeptide directly attached to D or indirectly attached via one or two suicide spacer subunits, wherein the tripeptide is recognized by at least one, preferably more than one, intracellular protease, wherein at least one protease is upregulated in tumor cells compared to normal cells, and the tripeptide is more susceptible to proteolysis by tumor tissue homogenate containing tumor cells targeted by the ligand-drug conjugate compared to normal tissue homogenate, wherein cytotoxicity to normal tissue is associated with adverse events resulting from administration of a comparative ligand-drug conjugate. In other embodiments, the tripeptide improves the biodistribution of the conjugate to tumor tissue but is detrimental to biodistribution to normal tissue; in some of these embodiments, the tripeptide also improves the selectivity for proteolysis of tumor tissue homogenate compared to proteolysis of normal tissue homogenate. In any of those embodiments, the normal tissue is sometimes bone marrow, and the adverse event to be improved is neutropenia. In another embodiment, the normal tissue is bone marrow, liver, kidney, esophagus, breast, or corneal tissue, and the adverse event to be improved is neutropenia. In some embodiments, the tripeptide is directly attached to D or indirectly attached to D via one or two suicide spacer subunits. In other embodiments, a peptide-cleavable unit (W) comprising a tripeptide as described herein is directly attached to D or indirectly attached to D via one or two suicide spacer subunits via an amino acid that is not part of the tripeptide.

[0567] The peptide cleavable unit (W) of the comparative conjugate is typically a dipeptide that imparts selectivity to a specific intracellular protease upregulated in cancer cells over a freely circulating protease, wherein the specific protease is capable of cleaving the amide bond between the C-terminal amino acid of the dipeptide and the amino group of the suicide spacer unit (Y) to initiate the release of the drug unit as a free drug.

[0568] In some embodiments, ligand-drug conjugates comprising tripeptides as disclosed herein exhibit improved tolerability compared to comparative ligand-drug conjugates, wherein the peptide-cleavable unit of the comparative conjugate is a dipeptide conjugated to a specific intracellular protease upregulated in cancer cells with superior selectivity to freely circulating proteases, wherein the specific protease is capable of cleaving the amide bond between the C-terminal amino acid of the dipeptide and the amino group of the suicide spacer unit (Y) to initiate the release of the drug unit as a free drug. In some embodiments, the dipeptide is known to be selectively cleaved by cathepsin B. In some embodiments, the dipeptide in the...

Claims

1. A ligand-drug conjugate compound or a pharmaceutically acceptable salt thereof, wherein the compound has the following structure: in L is a Ligand unit, wherein the Ligand unit is derived from an antibody or an antigen-binding fragment of an antibody, and the antibody or the antigen-binding fragment of an antibody can selectively bind to an antigen of a tumor tissue; and The subscript p' is an integer from 1 to 12.

2. The ligand drug conjugate compound or a pharmaceutically acceptable salt thereof according to claim 1, wherein L is an antibody ligand unit of a complete antibody or an antigen-binding fragment thereof.

3. The ligand-drug conjugate compound or a pharmaceutically acceptable salt thereof according to claim 2, wherein the intact antibody or fragment thereof is capable of selectively binding to a cancer cell antigen.

4. The ligand drug conjugate compound or a pharmaceutically acceptable salt thereof according to claim 2, wherein the intact antibody is a chimeric antibody, a humanized antibody or a human antibody, wherein the antibody is capable of selectively binding to a cancer cell antigen. 5 . The ligand drug conjugate compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 4 , wherein the subscript p′ is an integer from 1 to 10. 6 . The ligand drug conjugate compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 4 , wherein the subscript p′ is an integer from 1 to 8. 7 . The ligand drug conjugate compound or a pharmaceutically acceptable salt thereof according to claim 6 , wherein subscript p′ is 4. 8 . The ligand drug conjugate compound or a pharmaceutically acceptable salt thereof according to claim 6 , wherein subscript p′ is 8.

9. A pharmaceutically acceptable formulation, wherein the formulation comprises an effective amount of the ligand drug conjugate compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 8 and at least one pharmaceutically acceptable excipient.

10. The pharmaceutically acceptable formulation of claim 9, wherein the at least one pharmaceutically acceptable excipient is a liquid carrier that provides a liquid formulation, wherein the liquid formulation is suitable for lyophilization or administration to a subject in need thereof.

11. The pharmaceutically acceptable formulation according to claim 9, wherein the formulation is a solid resulting from lyophilization or a liquid formulation as defined in claim 10, wherein at least one excipient of the solid is a lyoprotectant.

12. A drug linker compound, wherein the drug linker compound has the following structure: or a salt thereof.

13. A linker compound, wherein the linker compound has the following structure: or a salt thereof, wherein RG is a reactive group.

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