Maytansinoid-based drug delivery systems

Abstract

The present subject matter provides albumin binding prodrugs, maytansinoid-based compounds and uses thereof.

Description

[0001] This application claims priority to U.S. Provisional Application No. 62 / 593,184, filed on November 30, 2017, the disclosure of which is incorporated herein by reference in its entirety. Background Art

[0002] Many drugs, particularly cancer therapeutic agents, have a narrow treatment window, wherein their side effects limit their beneficial effects. The systemic administration of such drugs usually results in limited therapeutic effects, because the dosage required to cause a stronger effect can cause the patient to experience unacceptable side effects. For those drugs with high cytotoxic potential, such as cytostatics, viral inhibitors or immunosuppressants, this is particularly critical. For some cytotoxic agents that inhibit tumor cell growth in the picomolar range, this is even more critical. These agents are usually too toxic to be used as chemotherapeutic agents. For example, tubulin binding maytansine is very effective in inhibiting tumor cell growth, but all fails in various clinical trials due to unacceptable toxicity spectrum.

[0003] Many research efforts have been devoted to the delivery of specific drugs at specific sites of action. Typically, this approach produces higher drug concentrations at the site of action than would be achieved with systemic administration while limiting side effects.

[0004] Drug delivery in oncology is of particular interest due to the narrow therapeutic window of agents used in this indication. Much research effort has focused on conjugating anticancer drugs to a variety of low and high molecular weight carriers, including sugars, growth factors, vitamins, peptides, antibodies, polysaccharides, lectins, serum proteins, and synthetic polymers. In most of these drug delivery systems, the drug is conjugated to the carrier via a spacer bound to a predetermined breakpoint that allows the bound drug to be released at the cellular target site (Kratz et al., ChemMedChem, 3: 20-53 (2008)).

[0005] Conjugates are known in which cytostatics are bound to serum proteins, primarily to specific carrier molecules, such as human serum albumin and human serum transferrin, followed by administration. In other cases, conjugates comprising a therapeutically effective substance, a spacer molecule, and a protein-binding molecule are covalently bound to circulating serum albumin after administration, which results in the transport of the therapeutically effective substance to a target site where it is released (US 7,387,771). In other cases, antibody drug conjugates (ADCs) can transport drugs to target sites for local release (Kratz et al., ChemMedChem, 3: 20-53 (2008); Panowski et al., mAbs, 6, 34-45 (2014); Chari et al., Angewandte Chem. Int. Ed., 53, 3796-3827 (2014)).

[0006] However, when designing a drug delivery system, an appropriate balance should be achieved between maintaining the targeting properties of the drug carrier and controlling the release of the drug. The drug delivery system should have sufficient stability in the bloodstream and allow the drug to be effectively released at the tumor site by enzymatic cleavage, reduction or in a pH-dependent manner (Kratz et al., ChemMedChem, 3: 20-53 (2008)). For highly effective cytotoxic agents from maytansine alkaloids (derived from maytansine), only drug delivery systems in which active species based on maytansine alkaloids are released in a non-specific or reductive manner have been reported. Among these, only those antibodies using monoclonal antibodies as carrier molecules have entered the clinical development stage, and only one antibody-maytansine alkaloid conjugate, i.e. T-DM1, has been reported. , which have gained market approval against certain breast cancer subtypes. Therefore, there remains a need for efficient and less complex drug delivery and release systems that release highly potent cytotoxic agents based on maytansinoids in an effective manner. Summary of the invention

[0007] The present disclosure provides a compound having a structure of formula (I):

[0008]

[0009] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof, wherein:

[0010] R 1 Selected from -H and C1-C4 alkyl;

[0011] The spacer is selected from:

[0012]

[0013] V is absent or selected from -CH2-, -O- and -NR3 -, where R 3 is -H or C1-C4 alkyl;

[0014] Each R 2 are independently selected from -H, a halogen group (e.g., -F, -Cl, -Br or -I) and a C1-C4 alkyl group, or two R 2 Together they form a C3-C6 cycloalkyl group;

[0015] n is 0 to 3;

[0016] X is absent or selected from -CH2-, -O-, -S-, -Se- and -NR 4 -, where R 4 is -H or C1-C4 alkyl;

[0017] Y is selected from ═CH— and ═N—;

[0018] Z 1 , Z 2 , Z 3 and Z 4 Each is independently selected from -H, a halogen group (e.g., -F, -Cl, -Br or -I), -CF3, -OCH3, -CN, -NO2, C1-C4 alkyl and C2-C4 alkoxy;

[0019] AA is an amino acid selected from the group consisting of glycine, D or L proline, sarcosine, N-ethylglycine, D or L alanine, D or L N-methylalanine, β-alanine, N-methyl-β-alanine, α-aminoisobutyric acid, and N-methyl-α-aminoisobutyric acid;

[0020] R' is selected from O and

[0021] Y' is absent or selected from optionally substituted C1-C6 alkyl, -NH-C(O)- and -C(O)-NH-; or Y' is selected from the group consisting of:

[0022] wherein n = 0 to 6;

[0023] R 1’ Not present or selected from the group consisting of:

[0024]

[0025] Among them, M 1 is a pharmaceutically acceptable counterion (e.g., H + 、Na + , K + , Ca 2+ Mg 2+ NR4+ and NHR3 + ; wherein R is H or C1-C4 alkyl);

[0026] R 2’ is an optionally substituted C1-C 18 Alkyl, wherein the C1-C 18 Optionally, up to six carbon atoms in the alkyl group are each independently substituted with -OCH2CH2-;

[0027] Z 1’ , Z 2’ , Z 3’ and Z 4’ Each is independently selected from -H, a halogen group (e.g., -F, -Cl, -Br or -I), -CF3, -OCH3, -CN, -NO2, -SO3M 2 and C1-C4 alkyl, wherein M 2 is a pharmaceutically acceptable counterion (e.g., H + 、Na + , K + , Ca 2 + Mg 2+ 、NR4 + and NHR3 + ; wherein R is H or C1-C4 alkyl);

[0028] TBG is a thiol-binding group selected from the group consisting of an optionally substituted maleimido group, an optionally substituted haloacetamide group, an optionally substituted haloacetate group, an optionally substituted pyridylthio group, an optionally substituted isothiocyanate group, an optionally substituted vinylcarbonyl group, an optionally substituted aziridine group, an optionally substituted disulfide group, an optionally substituted ethynyl group, and an optionally substituted N-hydroxysuccinimide ester group;

[0029] The TBG is optionally combined with a thiol-carrying macromolecular carrier or a thiol-carrying tumor-specific carrier.

[0030] In some embodiments, the present disclosure provides a compound having a structure of Formula (I):

[0031]

[0032] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof, wherein:

[0033] R 1 Selected from -H and C1-C4 alkyl;

[0034] The spacer is selected from:

[0035]

[0036] V is absent or selected from -CH2-, -O- and -NR 3 -, where R 3 is -H or C1-C4 alkyl;

[0037] Each R 2 are independently selected from -H, a halogen group (e.g., -F, -Cl, -Br or -I) and a C1-C4 alkyl group, or two R 2 Together they form a C3-C6 cycloalkyl group;

[0038] n is 0 to 3;

[0039] X is absent or selected from -CH2-, -O-, -S-, -Se- and -NR 4 -, where R 4 is -H or C1-C4 alkyl;

[0040] Y is selected from ═CH— and ═N—;

[0041] Z 1 , Z 2 , Z 3 and Z 4 Each is independently selected from -H, a halogen group (e.g., -F, -Cl, -Br or -I), -CF3, -OCH3, -CN, -NO2, C1-C4 alkyl and C2-C4 alkoxy;

[0042] AA is an amino acid selected from the group consisting of glycine, D or L proline, sarcosine, D or L alanine, D or L N-methylalanine, β-alanine, N-methyl-β-alanine, α-aminoisobutyric acid, and N-methyl-α-aminoisobutyric acid;

[0043] R' is selected from O and

[0044] Y' is absent or selected from optionally substituted C1-C6 alkyl, -NH-C(O)- and -C(O)-NH-; or Y' is selected from the group consisting of:

[0045] Where n = 0 to 6;

[0046] R 1’ Not present or selected from the group consisting of:

[0047]

[0048] Among them, M 1 is a pharmaceutically acceptable counterion (e.g., H + 、Na + , K + , Ca2+ Mg 2+ 、NR4 + and NHR3 + ; wherein R is H or C1-C4 alkyl);

[0049] R 2’ is an optionally substituted C1-C 18 Alkyl, wherein the C1-C 18 Optionally, up to six carbon atoms in the alkyl group are each independently substituted with -OCH2CH2-;

[0050] Z 1’ , Z 2’ , Z 3’ and Z 4’ Each is independently selected from -H, a halogen group (e.g., -F, -Cl, -Br or -I), -CF3, -OCH3, -CN, -NO2, -SO3M 2 and C1-C4 alkyl, wherein M 2 is a pharmaceutically acceptable counterion (e.g., H + 、Na + , K + , Ca 2 + Mg 2+ 、NR4 + and NHR3 + ; wherein R is H or C1-C4 alkyl);

[0051] TBG is a thiol-binding group selected from the group consisting of an optionally substituted maleimido group, an optionally substituted haloacetamide group, an optionally substituted haloacetate group, an optionally substituted pyridylthio group, an optionally substituted isothiocyanate group, an optionally substituted vinylcarbonyl group, an optionally substituted aziridine group, an optionally substituted disulfide group, an optionally substituted ethynyl group, and an optionally substituted N-hydroxysuccinimide ester group;

[0052] The TBG is optionally combined with a thiol-carrying macromolecular carrier or a thiol-carrying tumor-specific carrier.

[0053] In some embodiments, the compound has the structure of Formula (II):

[0054]

[0055] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof,

[0056] in:

[0057] Each R 2 are independently selected from -H and C1-C4 alkyl, or two R 2Together they form a C3-C6 cycloalkyl group;

[0058] X is absent or selected from -CH2-, -O-, -S- and -NR 3 -, where R 3 is -H or C1-C4 alkyl;

[0059] Z 1 , Z 2 , Z 3 and Z 4 Each is independently selected from -H, a halogen group (eg, -F, -Cl, -Br or -I), -CF3, -OCH3, -NO2 and -CH3.

[0060] In some embodiments, the compound has the structure of Formula (III):

[0061]

[0062] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof,

[0063] in:

[0064] Each R 2 are independently selected from -H and C1-C4 alkyl, or two R 2 Together they form a C3-C6 cycloalkyl group;

[0065] X is absent or selected from -CH2-, -O-, -S- and -NR 3 -, where R 3 is -H or C1-C4 alkyl;

[0066] Z 1 , Z 2 , Z 3 and Z 4 Each is independently selected from -H, a halogen group (eg, -F, -Cl, -Br or -I), -CF3, -OCH3, -NO2 and -CH3.

[0067] In some embodiments, the compound has the structure of Formula (IV):

[0068]

[0069] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof,

[0070] in:

[0071] X is absent or selected from -CH2- and -NH-;

[0072] Y is =CH- or =N-;

[0073] Z 1 , Z 2 , Z 3 and Z 3 Each is independently selected from -H, a halogen group (e.g., -F, -Cl, -Br or -I), -CF3, -OCH3, -NO2 and -CH3;

[0074] AA is an amino acid selected from the group consisting of glycine, D or L proline, sarcosine, N-ethylglycine, D or L alanine, D or L N-methylalanine, β-alanine, N-methyl-β-alanine, α-aminoisobutyric acid and N-methyl-α-aminoisobutyric acid.

[0075] In some embodiments, the compound has the structure of Formula (IV):

[0076]

[0077] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof,

[0078] in:

[0079] X is absent or selected from -CH2- and -NH-;

[0080] Y is =CH- or =N-;

[0081] Z 1 , Z 2 , Z 3 and Z 3 Each is independently selected from -H, a halogen group (e.g., -F, -Cl, -Br or -I), -CF3, -OCH3, -NO2 and -CH3;

[0082] AA is an amino acid selected from the group consisting of glycine, D or L proline, sarcosine, D or L alanine, D or L N-methylalanine, β-alanine, N-methyl-β-alanine, α-aminoisobutyric acid and N-methyl-α-aminoisobutyric acid.

[0083] In some embodiments, R 1 is -H. In some embodiments, Z 1 , Z 2 , Z 3 and Z 4 At least one of is not H. In some embodiments, Z 1 , Z 2 , Z 3 and Z 4At least one of is -F or -NO2. In some embodiments, n is 0 and X is absent. In some embodiments, n is 0 and X is -CH2-. In some embodiments, n is 0 and X is -O-, NHMe or -S-. In some embodiments, the compound is selected from:

[0084]

[0085]

[0086]

[0087]

[0088]

[0089] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof.

[0090] In some embodiments, the pharmaceutically acceptable counterion is selected from H + 、Na + , K + , Ca 2+ Mg 2+ NR4 + and NHR3 + ; wherein R is H or C1-C4 alkyl.

[0091] In some embodiments, R' is O. In some embodiments, R' is:

[0092]

[0093] In some embodiments, the compound is not bound to a thiol-bearing macromolecular carrier or a thiol-bearing tumor-specific carrier. In some embodiments, the compound is bound to a thiol-bearing macromolecular carrier or a thiol-bearing tumor-specific carrier. In some embodiments, the thiol-bearing macromolecular carrier or the thiol-bearing tumor-specific carrier is selected from endogenous albumin, exogenous albumin, antibodies, antibody fragments, peptides, natural or synthetic polymers, liposomes, and nanoparticles. In some embodiments, TBG is an optionally substituted maleimide group. In some embodiments, Z 1’ Selected from -NO2 or -SO3M 2 ;

[0094] and Y' is selected from -NHC(O)- or

[0095] In some embodiments, R 1’ yes

[0096]

[0097] In some embodiments, R' is:

[0098]

[0099] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof,

[0100] Where R 2’ Selected from optionally substituted C1-C 18 Alkyl, wherein the C1-C 18 Optionally up to six carbon atoms in the alkyl group are each independently substituted with -OCH2CH2-.

[0101] In some embodiments, R' is:

[0102]

[0103] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof.

[0104] In some embodiments, the compound is selected from:

[0105]

[0106]

[0107]

[0108] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof.

[0109] In some embodiments, R' is:

[0110]

[0111] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof; wherein M 1 It is a pharmaceutically acceptable counter ion.

[0112] In some embodiments, the compound is selected from:

[0113]

[0114] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof.

[0115] In some embodiments, according to any one of claims 14 to 20, wherein R' is:

[0116] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof;

[0117] Among them, M 1 It is a pharmaceutically acceptable counter ion.

[0118] In some embodiments, the compound according to claim 26, wherein the compound is:

[0119] Other embodiments include a pharmaceutical composition comprising a compound disclosed herein and a pharmaceutically acceptable carrier.

[0120] Other embodiments include a method for treating a disease or condition selected from the following: cancer, viral diseases, autoimmune diseases, acute or chronic inflammatory diseases, and diseases caused by bacteria, fungi or other microorganisms, the method comprising administering a therapeutically effective amount of a compound or pharmaceutical composition disclosed herein to a patient in need thereof. In some embodiments, the disease is cancer, for example, cancer selected from adenocarcinoma, uveal melanoma, acute leukemia, acoustic neuroma, ampullary carcinoma, anal cancer, astrocytoma, basal cell carcinoma, pancreatic cancer, connective tissue tumors, bladder cancer, bronchial cancer, non-small cell bronchial cancer, breast cancer, Burkitt's lymphoma, uterine body cancer, CUP syndrome, colon cancer, small intestine cancer, ovarian cancer, endometrial cancer, gallbladder cancer, gallbladder cancer, uterine cancer, cervical cancer, neck, nose, ear tumors, blood tumors, Hairy cell leukemia, urethral cancer, skin cancer, glioma, testicular cancer, Kaposi's sarcoma, laryngeal cancer, bone cancer, colorectal cancer, head and neck cancer, colon cancer, craniopharyngioma, liver cancer, leukemia, lung cancer, non-small cell lung cancer, Hodgkin's lymphoma, non-Hodgkin's lymphoma, stomach cancer, colon cancer, medulloblastoma, melanoma, meningioma, kidney cancer, renal cell carcinoma, oligodendroglioma, esophageal cancer, osteolytic and bone cancer, osteosarcoma, ovarian cancer, pancreatic cancer, penile cancer, prostate cancer, tongue cancer, ovarian cancer, and lymph node cancer.

[0121] Other embodiments include a method of reducing the cytotoxicity of a compound, the method comprising administering to a patient in need thereof a compound or pharmaceutical composition disclosed herein, wherein the administration results in reduced cytotoxicity when compared to an equivalent dose of an unmodified active agent.

[0122] Other embodiments include a method of increasing the concentration of a metabolite of a compound in a tumor, the method comprising administering to a patient in need thereof a compound or pharmaceutical composition disclosed herein, wherein the increase is compared to an equivalent dose of the unmodified active agent.

[0123] Other embodiments include a compound disclosed herein for use as a medicament.

[0124] Other embodiments include a compound disclosed herein for use in treating a disease or condition selected from the group consisting of cancer, viral diseases, autoimmune diseases, acute or chronic inflammatory diseases, and diseases caused by bacteria, fungi or other microorganisms.

[0125] Other embodiments include the use of a compound or pharmaceutical composition disclosed herein in the preparation of a medicament for treating a disease or condition selected from cancer, viral diseases, autoimmune diseases, acute or chronic inflammatory diseases, and diseases caused by bacteria, fungi or other microorganisms. BRIEF DESCRIPTION OF THE DRAWINGS

[0126] Figure 1 The stability of different linkers of 4 in CD1 mouse plasma is shown.

[0127] Figure 2 The geometric mean IC values ​​of a panel of different cell lines are shown. 50 Heatmap of values.

[0128] Figure 3 Tumor growth curves of the control group, maytansine group, and groups treated with compounds 30, 42, 31, and 35 in the RXF631 renal cell tumor model are shown.

[0129] Figure 4 The body weight change curves of the control group, the maytansine group, and the groups treated with compounds 30, 42, 31, and 35 in the RXF631 renal cell tumor model are shown.

[0130] Figure 5 Shown are tumor growth curves for the control group, maytansine group, and groups treated with compounds 32, 30, and 31 in the LXFE 937 squamous cell lung cancer model.

[0131] Figure 6 The body weight change curves of the control group, the maytansine group, and the groups treated with compounds 32, 30, and 31 in the LXFE 937 squamous cell lung cancer model are shown.

[0132] Figure 7 Shown are the tumor growth curves of the control group, the maytansine group, and the groups treated with compounds 30 and 31 in the LXFE 937 squamous cell lung cancer model.

[0133] Figure 8 The body weight change curves of the control group, the maytansine group, and the groups treated with compounds 30 and 31 in the LXFE 937 squamous cell lung cancer model are shown.

[0134] Fig. 9Shown are tumor growth curves of the control group, maytansine group, and groups treated with compounds 30 and 31 in the LXFA 737 lung adenocarcinoma model.

[0135] Fig.10 The body weight change curves of the control group, the maytansine group, and the groups treated with compounds 30 and 31 in the LXFA 737 lung adenocarcinoma model are shown.

[0136] Fig.11 Shown are the tumor growth curves of the control group, the maytansine group, and the groups treated with compounds 32, 30, and 31 in the MDA-MB 231 breast cancer model.

[0137] Fig.12 The body weight change curves of the control group, the maytansine group, and the groups treated with compounds 32, 30, and 31 in the MDA-MB 231 breast cancer model are shown.

[0138] Fig.13 Tumor growth curves of the control group, maytansine group, and groups treated with compounds 30 and 31 in the A2780 ovarian cancer model are shown.

[0139] Fig.14 The body weight change curves of the control group, the maytansine group, and the groups treated with compounds 30 and 31 in the A2780 ovarian cancer model are shown.

[0140] Fig.15 Shown are tumor growth curves of the control group, maytansine group, and groups treated with compounds 30 and 31 in the MDA-MB 468 breast cancer model.

[0141] Fig.16 The body weight change curves of the control group, the maytansine group, and the groups treated with compounds 30 and 31 in the MDA-MB 468 breast cancer model are shown. DETAILED DESCRIPTION

[0142] Unless otherwise defined herein, the scientific and technical terms used in this application shall have the meanings commonly understood by those of ordinary skill in the art. Generally, the nomenclature associated with the techniques of chemistry, molecular biology, cell and cancer biology, immunology, microbiology, pharmacology and protein chemistry described herein is well-known and commonly used in the art.

[0143] All publications, patents and published patent applications mentioned in this application are specifically incorporated herein by reference. In the event of a conflict, this specification (including its specific definitions) shall prevail. Unless otherwise stated, it should be understood that each embodiment disclosed herein can be used alone or in combination with any one or more other embodiments of the present invention.

[0144] definition

[0145] Throughout the specification, the word "comprise" or variations such as "comprises" or "comprising" will be understood to imply the inclusion of a stated integer (or component) or group of integers (or components) but not the exclusion of any other integer (or component) or group of integers (or components).

[0146] Throughout the application, where compounds or compositions are described as having, including, or comprising specific components, it is contemplated that such compounds or compositions may also consist essentially of or consist of the enumerated components. Similarly, where methods or processes are described as having, including, or comprising specific process steps, the processes may also consist essentially of or consist of the enumerated process steps. In addition, it should be understood that the order of steps or the order in which certain actions are performed does not matter as long as the compounds, compositions, and methods described herein remain operable. Moreover, two or more steps or actions may be performed simultaneously.

[0147] The singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise.

[0148] The term "including" is used to mean "including, but not limited to." "Including" and "including, but not limited to" can be used interchangeably.

[0149] As used herein, the term "or" should be understood to mean "and / or" unless the context clearly indicates otherwise.

[0150] The terms "drug", "agent", "therapeutic agent", "therapeutically active agent", "cytotoxic agent or drug", "highly cytotoxic agent or drug" or "therapeutically effective substance" are used to refer to any compound that produces a pharmacological effect by itself or after transformation in the relevant organism, and therefore also includes derivatives from these transformations. The pharmacological effect of the drug of the composition according to the present disclosure can only be a single effect, such as: cell inhibition and / or cytotoxic effect, or a wide spectrum of pharmacological effects, such as having both immunosuppressive and anti-inflammatory effects.

[0151] The terms "patient," "subject," or "individual" are used interchangeably and refer to a human or non-human animal. These terms include mammals, such as humans, primates, livestock (e.g., cattle, pigs), companion animals (e.g., dogs, cats), and rodents (e.g., mice and rats). In certain embodiments, the patient or subject is a human patient or subject, such as a human patient suffering from a condition in need of treatment.

[0152] The term "pharmaceutical composition" refers to a composition suitable for pharmaceutical use for a test animal (including humans and mammals), for example, in combination with one or more pharmaceutically acceptable carriers, excipients or solvents. Such a composition may also include diluents, fillers, salts, buffers, stabilizers, solubilizers, protective agents and other materials well known in the art. In certain embodiments, the pharmaceutical composition encompasses a composition comprising an active ingredient, and an inert ingredient consisting of an excipient, a carrier or a diluent, and any combination, complexation or aggregation of any two or more components, or the decomposition of one or more components, or other types of reactions or interactions of one or more components, and any product produced directly or indirectly. Therefore, the pharmaceutical composition of the present disclosure encompasses any composition made by mixing a compound of the present disclosure with one or more pharmaceutically acceptable excipients, carriers and / or diluents.

[0153] The term "pharmaceutically acceptable carrier" refers to a non-toxic carrier that can be administered to a patient together with the therapeutically effective substances disclosed herein and does not destroy the pharmaceutical activity of the agent. The term "excipient" refers to an additive that is not a pharmaceutically active ingredient in a preparation or composition. In certain embodiments, a "pharmaceutically acceptable" substance is suitable for contact with cells, tissues or organs of animals or humans without excessive toxicity, stimulation, allergic reaction, immunogenicity or other adverse reactions, and its dosage form is: according to the dosing schedule, and has a reasonable benefit / risk ratio. In certain embodiments, a "pharmaceutically acceptable" substance as a component of a pharmaceutical composition is additionally compatible with other ingredients of the composition. In certain embodiments, the terms "pharmaceutically acceptable excipient", "pharmaceutically acceptable carrier" and "pharmaceutically acceptable diluent" include, but are not limited to, pharmaceutically acceptable inactive ingredients, materials, compositions and vehicles, such as liquid fillers, solid fillers, diluents, excipients, carriers, solvents and encapsulation materials. Carriers, diluents and excipients also include all pharmaceutically acceptable dispersion media, coatings, buffers, isotonic agents, stabilizers, absorption delaying agents, antimicrobial agents, antibacterial agents, antifungal agents, adjuvants, etc. The disclosure encompasses the use of conventional excipients, carriers and diluents in pharmaceutical compositions, except where any conventional excipient, carrier or diluent is incompatible with the active ingredient. See, for example, Remington: The Science and Practice of Pharmacy, 21st edition, Lippincott Williams & Wilkins (Philadelphia, Pennsylvania, 2005); Handbook of Pharmaceutical Excipients, 5th edition, Rowe et al., The Pharmaceutical Press and the American Pharmaceutical Association (2005); Handbook of Pharmaceutical Additives, 3rd edition, Ash and Ash, Gower Publishing Co. (2007); and Pharmaceutical Preformulation and Formulation, Gibson, CRC Press LLC (Boca Raton, Florida, 2004).

[0154] The term "pharmaceutically effective amount", "therapeutically effective amount" or "therapeutically effective dose" refers to an amount that effectively treats a patient's disease or condition, for example, the treatment, cure, inhibition or improvement of the overall health, physiological response or condition of a patient suffering from a disease (e.g., cancer) or condition, etc., produces a beneficial and / or desirable change. The complete therapeutic effect does not necessarily occur through a single dose administration, and may only occur after a series of doses. Therefore, a therapeutically effective amount can be administered through one or more administrations. The exact effective amount required for the subject will depend on, for example, the size, health status and age of the subject, the nature and extent of the disease, the combination of the therapy or therapies selected for administration, and the mode of administration. The technician can easily determine the effective amount for a given situation through routine experiments. The technician will recognize that treating cancer includes, but is not limited to, killing cancer cells, preventing the growth of new cancer cells, causing tumor regression (reduced tumor size), causing a reduction in metastasis, improving the patient's vital functions, improving the patient's health, alleviating pain, improving appetite, improving the patient's weight, and any combination thereof. The term "pharmaceutically effective amount", "therapeutically effective amount" or "therapeutically effective dose" also refers to the amount required to improve the patient's clinical symptoms. One or more methods of treating cancer described herein should not be construed or otherwise limited to "curing" cancer.

[0155] As used herein, the term "treating" or "treatment" includes reversing, alleviating or preventing symptoms, clinical signs and potential pathological conditions in some way to improve or stabilize the condition of the subject. As used herein, and as is well known in the art, "treatment" is a method for obtaining beneficial or desired results, including clinical results. Beneficial or desirable clinical results may include, but are not limited to, alleviating, improving or slowing down the progression of one or more symptoms or conditions associated with a condition, for example, whether detectable or undetectable cancer, alleviation of the extent of the disease, stabilization of the disease state (i.e., no deterioration), delay or slowing of disease progression, improvement or alleviation of the disease state and alleviation (whether partial or complete). "Treatment" may also mean an extension of survival compared to the expected survival period without treatment. Exemplary beneficial clinical results are described herein.

[0156] One of a variety of methods known to those skilled in the art can be used to "administer" or "administer" a substance, compound, or agent to a subject. For example, the compound or agent can be administered intravenously, intraarterially, intradermally, intramuscularly, intraperitoneally, subcutaneously, intraocularly, sublingually, orally (by ingestion), intranasally (by inhalation), intraspinal, intracerebrally, and transdermally (by absorption, such as through a skin catheter). The compound or agent can also be appropriately introduced by a rechargeable or biodegradable polymer device or other device (such as a patch and a pump or formulation) that provides a prolonged, slow, or controlled release of the compound or agent. It can also be administered, for example, once, multiple times, and / or over one or more extended periods. In certain aspects, administration includes both direct administration (including self-administration) and indirect administration (including the act of prescribing a drug). For example, as used herein, a physician who instructs a patient to self-administer a drug or is administered a drug by another person, and / or a physician who provides a drug prescription to a patient is administering the drug to the patient. When a method is part of a treatment regimen involving more than one agent or treatment modality, the disclosure contemplates that the agents may be administered at the same or different times and by the same or different routes of administration. The appropriate method for administering a substance, compound, or agent to a subject also depends on, for example, the age of the subject, whether the subject is active or inactive at the time of administration, whether the subject's cognitive abilities are impaired at the time of administration, the extent of the impairment, and the chemical and biological properties (e.g., solubility, digestibility, bioavailability, stability, and toxicity) of the compound or agent.

[0157] The term "substituted" refers to a part having a substituent that replaces hydrogen on one or more carbon atoms of the main chain of a chemical compound. It should be understood that "substituted" or "substituted by..." includes implicit conditions, i.e., such substitution is based on the allowed valence of the substituted atom and the substituent, and the substitution produces a stable compound, for example, it will not spontaneously transform such as by rearrangement, cyclization, elimination, etc. As used herein, the term "substituted" is intended to include all allowed substituents of organic compounds. In a broad sense, allowed substituents include acyclic and cyclic, branched and unbranched, carbocyclic and heterocyclic, aromatic and non-aromatic substituents of organic compounds. For suitable organic compounds, allowed substituents can be one or more and can be the same or different. For the purposes of this disclosure, heteroatoms (such as nitrogen) can have hydrogen substituents, and / or any allowed substituents of organic compounds described herein that meet the valence of heteroatoms. Substituents may include any of the substituents described herein, for example, halogen, hydroxy, carbonyl (such as carboxyl, alkoxycarbonyl, formyl or acyl), thiocarbonyl (such as thioester, thioacetate or thioformate), alkoxy, alkylthio, acyloxy, phosphoryl, phosphate, phosphonate, amino, amide, amine, imino, cyano, nitro, azido, sulfhydryl, alkylthio, sulfate, sulfonate, sulfamoyl, sulfonamido, sulfonyl, heterocyclyl, aralkyl or aromatic (e.g., C6-C 12 aryl) or heteroaromatic (e.g., heteroaryl) moiety.

[0158] "Optional" or "optionally" means that the subsequently described circumstances may or may not occur, and thus the application includes instances where such circumstances occur and instances where such circumstances do not occur. For example, the phrase "optionally substituted" means that non-hydrogen substituents may or may not be present on a given atom, and thus the application includes structures where non-hydrogen substituents are present and structures where they are not present.

[0159] Unless otherwise specified as "unsubstituted", the chemical moieties mentioned herein are understood to include substituted variants. For example, reference to an "alkyl" group or moiety implicitly includes both substituted and unsubstituted variants. Examples of substituents on a chemical moiety include, but are not limited to, halogen, hydroxyl, carbonyl (such as carboxyl, alkoxycarbonyl, formyl or acyl), thiocarbonyl (such as thioester, thioacetate or thioformate), alkoxy, alkylthio, acyloxy, phosphoryl, phosphate, phosphonate, amino, amide, amine, imino, cyano, nitro, azido, sulfhydryl, alkylthio, sulfate, sulfonate, sulfamoyl, sulfonamido, sulfonyl, heterocyclic, aralkyl or aryl or heteroaryl moieties.

[0160] "Aryl" indicates an aromatic carbocyclic ring with a specified number of carbon atoms in the ring, such as 6 to 12 or 6 to 10 carbon atoms.Aryl can be monocyclic or polycyclic (e.g., bicyclic, tricyclic). In some cases, both rings of polycyclic aromatics are aromatic (e.g., naphthyl). In other cases, polycyclic aromatics can include non-aromatic rings (e.g., cycloalkyl, cycloalkenyl, heterocycloalkyl, heterocycloalkenyl) fused to aromatic rings, provided that polycyclic aromatics are combined with the parent structure via the atoms in the aromatic rings. Therefore, 1,2,3,4-tetrahydronaphthalene-5-yl (wherein the part is combined with the parent structure by aromatic carbon atoms) is considered as aryl, while 1,2,3,4-tetrahydronaphthalene-1-yl (wherein the part is combined with the parent structure by non-aromatic carbon atoms) is not considered as aryl. Similarly, 1,2,3,4-tetrahydroquinolin-8-yl (wherein the moiety is attached to the parent structure through an aromatic carbon atom) is considered an aryl group, while 1,2,3,4-tetrahydroquinolin-1-yl (wherein the moiety is attached to the parent structure through a non-aromatic nitrogen atom) is not considered an aryl group. However, the term "aryl" does not encompass or overlap with "heteroaryl" as defined herein, regardless of the point of attachment (e.g., quinolin-5-yl and quinolin-2-yl are both heteroaryl groups).

[0161] "Heteroaryl" indicates an aromatic ring containing a specified number of ring atoms (e.g., 5 to 12 or 5 to 10 heteroaryl groups), the specified number of ring atoms being composed of one or more heteroatoms (e.g., 1, 2, 3 or 4 heteroatoms) selected from N, O and S, and the remaining ring atoms being carbon. A 5-membered heteroaryl is a heteroaryl having 5 ring atoms. A 6-membered heteroaryl is a heteroaryl having 6 ring atoms. The heteroaryl group does not contain adjacent S and O atoms. In some embodiments, the total number of S and O atoms in the heteroaryl group does not exceed 2. In some embodiments, the total number of S and O atoms in the heteroaryl group does not exceed 1. Unless otherwise indicated, a heteroaryl group may be bound to the parent structure through a carbon or nitrogen atom, where valence permits. For example, "pyridyl" includes 2-pyridyl, 3-pyridyl and 4-pyridyl, and "pyrrolyl" includes 1-pyrrolyl, 2-pyrrolyl and 3-pyrrolyl. When nitrogen is present in a heteroaryl ring, the nitrogen may be present in an oxidized state (i.e., N + -O - ) is present. Additionally, when sulfur is present in a heteroaryl ring, it may be in an oxidized state (i.e., S + -O - The heteroaryl group may be monocyclic or polycyclic (eg, bicyclic, tricyclic).

[0162] In some cases, the heteroaryl group is monocyclic. Examples include pyrrole, pyrazole, imidazole, triazole (e.g., 1,2,3-triazole, 1,2,4-triazole, 1,3,4-triazole), tetrazole, furan, isoxazole, oxazole, oxadiazole (e.g., 1,2,3-oxadiazole, 1,2,4-oxadiazole, 1,3,4-oxadiazole), thiophene, isothiazole, thiazole, thiadiazole (e.g., 1,2,3-thiadiazole, 1,2,4-thiadiazole, 1,3,4-thiadiazole), pyridine, pyridazine, pyrimidine, pyrazine, triazine (e.g., 1,2,4-triazine, 1,3,5-triazine) and tetrazine.

[0163] The term "acyl" is art-recognized and refers to a group represented by the general formula hydrocarbyl-C(O)-, for example alkyl-C(O)-.

[0164] The term "alkyl" refers to a saturated aliphatic group, including straight chain alkyl and branched chain alkyl. In some embodiments, a straight chain or branched chain alkyl has 30 or fewer carbon atoms in its backbone (e.g., C1-C2 for a straight chain). 30 , for branched C4-C 30 ), and in other embodiments, have 20 or less. In certain embodiments, the alkyl group is a lower alkyl group, for example, methyl, ethyl, n-propyl, isopropyl, n-butyl, and n-pentyl. Moreover, the term "alkyl" used throughout the specification, examples, and claims is intended to include both "unsubstituted alkyl" and "substituted alkyl", the latter referring to an alkyl portion having a substituent replacing hydrogen on one or more carbon atoms of the hydrocarbon backbone. In certain embodiments, a straight chain or branched chain alkyl group has 30 or fewer carbon atoms in its backbone (e.g., for a straight chain, C1-C 30 , for branched C3-C 30 In some embodiments, the chain has ten or fewer carbon atoms (C1-C 10 ). In other embodiments, the chain has six or fewer carbon atoms (C1-C6) in its backbone.

[0165] The term "hydrazone moiety" or "hydrazone" refers to E and / or Z hydrazones, e.g.,

[0166]

[0167] The stereochemical configuration of the hydrazone moiety can be E or Z. The term hydrazone, as used herein, includes both E and Z isomers. The hydrazone moieties disclosed herein are generally drawn in one configuration, but it is understood that the disclosure may include both E and / or Z.

[0168] At various places in this specification, substituents of compounds of the present disclosure are disclosed in groups or ranges. It is specifically intended that the present disclosure includes each and every individual subcombination of the members of such groups and ranges. For example, the term "C1-C6 alkyl" is specifically intended to disclose individually methyl, ethyl, propyl, isopropyl, n-butyl, sec-butyl, isobutyl, etc.

[0169] "Pharmaceutically acceptable salts" are salts of compounds suitable for pharmaceutical use, including but not limited to metal salts (e.g., sodium, potassium, magnesium, calcium, etc.), acid addition salts (e.g., inorganic acids, carboxylic acids, etc.), and base addition salts (e.g., ammonia, organic amines, etc.). Acid addition salt forms of compounds that exist in free form as bases can be obtained by treating the free base form with an appropriate acid, such as an inorganic acid, for example a hydrohalic acid, such as hydrochloric acid or hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, etc.; or an organic acid, such as, for example, acetic acid, glycolic acid, propionic acid, lactic acid, pyruvic acid, malonic acid, succinic acid, maleic acid, fumaric acid, malic acid, tartaric acid, citric acid, methanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid, cyclopentane, salicylic acid, p-aminosalicylic acid, palmitic acid, etc. (see, for example, WO 01 / 062726. Some pharmaceutically acceptable salts listed by Berge et al., Journal of Pharmaceutical Sciences, 66: 1-19 (1977), which is incorporated herein by reference in its entirety). Compounds containing acidic protons can be converted into their therapeutically active non-toxic base addition salt forms, such as metal salts or amine salts, by treatment with appropriate organic and inorganic bases. Suitable basic salt forms include, for example, ammonium salts, alkali metal and alkaline earth metal salts or ions, for example, lithium, sodium, potassium, magnesium, calcium salts, etc., salts with organic bases, for example, N-methyl-d-glucamine, hybamine salts, and salts with amino acids, such as arginine, lysine, etc. Conversely, the salt forms can be converted into free forms by treatment with an appropriate base or acid. The compounds and their salts can be in the form of solvates, which are included within the scope of the present disclosure. Such solvates include, for example, hydrates, alcoholates, etc. (see, for example, WO 01 / 062726).

[0170] The present disclosure also provides pharmaceutical compositions comprising one or more compounds of the present disclosure and a pharmaceutically acceptable carrier or excipient.The compounds or pharmaceutical compositions of the present disclosure can be used in vitro or in vivo.

[0171] The term "isomer" as used herein includes, but is not limited to, tautomers, cis and trans isomers (E (hetero), Z (co-), R- and S-enantiomers (using R and S notation in accordance with the rules described in Pure Appl. Chem. (1976), 45, 11-30), diastereomers, (D)-isomers, (L)-isomers, stereoisomers, racemic mixtures thereof, and other mixtures thereof. All such isomers and mixtures thereof are intended to be included in the present disclosure. Tautomers, although not explicitly indicated in the formulas described herein, are intended to be included within the scope of the present disclosure.

[0172] The present disclosure also includes isotopically labeled or enriched compounds of the present disclosure. An "isotopically labeled" or "radiolabeled" compound is a compound of the present disclosure in which one or more atoms are replaced or substituted by an atom with an atomic mass or mass number different from the atomic mass or mass number normally found in nature (naturally occurring). Suitable radionuclides that can be incorporated into the compounds of the present disclosure include, but are not limited to 2 H (also written as D for deuterium), 3 H (also written as T for tritium), 11 C. 13 C. 14 C. 13 N. 15 N. 15 O. 17 O. 18 O. 18 F. 35 S. 36 Cl, 82 Br, 75 Br, 76 Br and 77 Br. The radionuclide incorporated into the radiolabeled compounds of the invention will depend on the specific application of the radiolabeled compound. For example, for in vitro metalloproteinase labeling and competition assays, it is generally incorporated 3 H. 14 C. 82 Br, 35 For radio imaging applications, 11 C. 18 F. 75 Br, 76 Br or 77 Br is usually the most useful. Tritium ( 3 H) and 14 C may be useful for ADME studies. In some embodiments, each alkyl, cycloalkyl, alkene, alkylene, and alkoxy is optionally substituted with one or more -D or -F.

[0173] Compounds of the Disclosure

[0174] Embodiments of the present disclosure provide a compound having a structure represented by formula (I):

[0175]

[0176] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof,

[0177] in:

[0178] R 1 Selected from -H and C1-C4 alkyl;

[0179] The spacer is selected from:

[0180]

[0181] V is absent or selected from -CH2-, -O- and -NR 3 -, where R 3 is -H or C1-C4 alkyl;

[0182] Each R 2 are independently selected from -H, a halogen group (e.g., -F, -Cl, -Br or -I) and a C1-C4 alkyl group, or two R 2 Together they form a C3-C6 cycloalkyl group;

[0183] n is 0 to 3;

[0184] X is absent or selected from -CH2-, -O-, -S-, -Se- and -NR 4 -, where R 4 is -H or C1-C4 alkyl;

[0185] Y is selected from ═CH— and ═N—;

[0186] Z 1 , Z 2 , Z 3 and Z 4 Each is independently selected from -H, a halogen group (e.g., -F, -Cl, -Br or -I), -CF3, -OCH3, -CN, -NO2, C1-C4 alkyl and C2-C4 alkoxy;

[0187] AA is an amino acid selected from the group consisting of glycine, D or L proline, sarcosine, N-ethylglycine, D or L alanine, D or L N-methylalanine, β-alanine, N-methyl-β-alanine, α-aminoisobutyric acid, and N-methyl-α-aminoisobutyric acid;

[0188] R' is selected from O and

[0189] Y' is absent or selected from optionally substituted C1-C6 alkyl, -NH-C(O)- and -C(O)-NH-; or Y' is selected from the group consisting of:

[0190] Where n = 0 to 6;

[0191] R 1’ Not present or selected from the group consisting of:

[0192]

[0193] Among them, M 1 is a pharmaceutically acceptable counterion (e.g., H + 、Na + , K + , Ca 2+ Mg 2+ 、NR4 + and NHR3 + ; wherein R is H or C1-C4 alkyl);

[0194] R 2’ is an optionally substituted C1-C 18 Alkyl, wherein the C1-C 18 Optionally, up to six carbon atoms in the alkyl group are each independently substituted with -OCH2CH2-;

[0195] Z 1’ , Z 2’ , Z 3’ and Z 4’ Each is independently selected from -H, a halogen group (e.g., -F, -Cl, -Br or -I), -CF3, -OCH3, -CN, -NO2, -SO3M 2 and C1-C4 alkyl, wherein M 2 is a pharmaceutically acceptable counterion (e.g., H + 、Na + , K + , Ca 2 + Mg 2+ 、NR4 + and NHR3 + ; wherein R is H or C1-C4 alkyl);

[0196] TBG is a thiol-binding group selected from the group consisting of an optionally substituted maleimido group, an optionally substituted haloacetamide group, an optionally substituted haloacetate group, an optionally substituted pyridylthio group, an optionally substituted isothiocyanate group, an optionally substituted vinylcarbonyl group, an optionally substituted aziridine group, an optionally substituted disulfide group, an optionally substituted ethynyl group, and an optionally substituted N-hydroxysuccinimide ester group;

[0197] The TBG is optionally combined with a thiol-carrying macromolecular carrier or a thiol-carrying tumor-specific carrier.

[0198] In some embodiments, in the compounds of formula (I), R 1 Selected from -H and C1-C4 alkyl;

[0199] The spacer is selected from:

[0200]

[0201] V is absent or selected from -CH2-, -O- and -NR 3 -, where R 3 is -H or C1-C4 alkyl;

[0202] Each R 2 are independently selected from -H, a halogen group (e.g., -F, -Cl, -Br or -I) and a C1-C4 alkyl group, or two R 2 Together they form a C3-C6 cycloalkyl group;

[0203] n is 0 to 3;

[0204] X is absent or selected from -CH2-, -O-, -S-, -Se- and -NR 4 -, where R 4 is -H or C1-C4 alkyl;

[0205] Y is selected from ═CH— and ═N—;

[0206] Z 1 , Z 2 , Z 3 and Z 4 Each is independently selected from -H, a halogen group (e.g., -F, -Cl, -Br or -I), -CF3, -OCH3, -CN, -NO2, C1-C4 alkyl and C2-C4 alkoxy;

[0207] AA is an amino acid selected from the group consisting of glycine, D or L proline, sarcosine, D or L alanine, D or L N-methylalanine, β-alanine, N-methyl-β-alanine, α-aminoisobutyric acid, and N-methyl-α-aminoisobutyric acid;

[0208] R' is selected from O and

[0209] Y' is absent or selected from optionally substituted C1-C6 alkyl, -NH-C(O)- and -C(O)-NH-; or Y' is selected from the group consisting of:

[0210] Where n = 0 to 6;

[0211] R 1’ Not present or selected from the group consisting of:

[0212]

[0213] Among them, M 1 is a pharmaceutically acceptable counterion (e.g., H + 、Na + , K + , Ca 2+ Mg 2+ 、NR4 + and NHR3 + ; wherein R is H or C1-C4 alkyl);

[0214] R 2’ is an optionally substituted C1-C 18 Alkyl, wherein the C1-C 18 Optionally, up to six carbon atoms in the alkyl group are each independently substituted with -OCH2CH2-;

[0215] Z 1’ , Z 2’ , Z 3’ and Z 4’ Each is independently selected from -H, a halogen group (e.g., -F, -Cl, -Br or -I), -CF3, -OCH3, -CN, -NO2, -SO3M 2 and C1-C4 alkyl, wherein M 2 is a pharmaceutically acceptable counterion (e.g., H + 、Na + , K + , Ca 2 + Mg 2+ 、NR4 + and NHR3 + ; wherein R is H or C1-C4 alkyl);

[0216] TBG is a thiol-binding group selected from the group consisting of an optionally substituted maleimido group, an optionally substituted haloacetamide group, an optionally substituted haloacetate group, an optionally substituted pyridylthio group, an optionally substituted isothiocyanate group, an optionally substituted vinylcarbonyl group, an optionally substituted aziridine group, an optionally substituted disulfide group, an optionally substituted ethynyl group, and an optionally substituted N-hydroxysuccinimide ester group;

[0217] The TBG is optionally combined with a thiol-carrying macromolecular carrier or a thiol-carrying tumor-specific carrier.

[0218] In some embodiments, R' is O. These novel compounds may represent active substances, and may be, for example, active components of a drug delivery system or active metabolites released from a drug delivery system.

[0219] In certain embodiments, the compound of formula (I) where R' is O has the structure of any one of formula (II'), (III') and (IV'):

[0220]

[0221] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof,

[0222] in:

[0223] Each R 2 are independently selected from -H and C1-C4 alkyl, or two R 2 Together they form a C3-C6 cycloalkyl group;

[0224] X is absent or selected from -CH2-, -O-, -S- and -NR 3 -, where R 3 is -H or C1-C4 alkyl;

[0225] Z 1 , Z 2 , Z 3 and Z 4 Each is independently selected from -H, a halogen group (e.g., -F, -Cl, -Br or -I), -CF3, -OCH3, -NO2 and -CH3;

[0226]

[0227] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof,

[0228] in:

[0229] X is absent or selected from -CH2- and -NH-;

[0230] Y is =CH- or =N-;

[0231] Z 1 , Z 2 , Z 3 and Z 3 Each is independently selected from -H, a halogen group (e.g., -F, -Cl, -Br or -I), -CF3, -OCH3, -NO2 and -CH3;

[0232] AA is an amino acid selected from the group consisting of glycine, D or L proline, sarcosine, N-ethylglycine, D or L alanine, D or L N-methylalanine, β-alanine, N-methyl-β-alanine, α-aminoisobutyric acid and N-methyl-α-aminoisobutyric acid.

[0233] In yet other embodiments, in the compound of formula (IV'):

[0234] X is absent or selected from -CH2- and -NH-; Y is =CH- or =N-;

[0235] Z 1 , Z 2 , Z 3 and Z 3 Each is independently selected from -H, a halogen group (e.g., -F, -Cl, -Br or -I), -CF3, -OCH3, -NO2 and -CH3;

[0236] AA is an amino acid selected from the group consisting of glycine, D or L proline, sarcosine, D or L alanine, D or L N-methylalanine, β-alanine, N-methyl-β-alanine, α-aminoisobutyric acid and N-methyl-α-aminoisobutyric acid.

[0237] In some embodiments, the compound is selected from the following specific compounds:

[0238]

[0239]

[0240]

[0241]

[0242] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof.

[0243] Other embodiments include prodrugs, such as those represented by formula (I), wherein R' is:

[0244] These novel compounds may represent drug delivery systems whereby active metabolites are selectively released from the drug delivery system. These prodrugs include, for example, albumin binding prodrugs.

[0245] In certain embodiments, these compounds have the structure of any one of Formula (II), (III), and (IV):

[0246] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof,

[0247] in:

[0248] Each R 2 are independently selected from -H and C1-C4 alkyl, or two R 2 Together they form a C3-C6 cycloalkyl group;

[0249] X is absent or selected from -CH2-, -O-, -S- and -NR 3 -, where R 3 is -H or C1-C4 alkyl;

[0250] Z 1 , Z 2 , Z 3 and Z 4 Each is independently selected from -H, a halogen group (e.g., -F, -Cl, -Br or -I), -CF3, -OCH3, -NO2 and -CH3;

[0251]

[0252] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof,

[0253] in:

[0254] X is absent or selected from -CH2- and -NH-;

[0255] Y is =CH- or =N-;

[0256] Z 1 , Z 2 , Z 3 and Z 3 Each is independently selected from -H, a halogen group (e.g., -F, -Cl, -Br or -I), -CF3, -OCH3, -NO2 and -CH3;

[0257] AA is an amino acid selected from the group consisting of glycine, D or L proline, sarcosine, N-ethylglycine, D or L alanine, D or L N-methylalanine, β-alanine, N-methyl-β-alanine, α-aminoisobutyric acid, and N-methyl-α-aminoisobutyric acid; and

[0258] where R' is:

[0259]

[0260] In yet other embodiments, in the compound of formula (IV'):

[0261] X is absent or selected from -CH2- and -NH-;

[0262] Y is =CH- or =N-;

[0263] Z 1 , Z 2 , Z 3 and Z 3 Each is independently selected from -H, a halogen group (e.g., -F, -Cl, -Br or -I), -CF3, -OCH3, -NO2 and -CH3;

[0264] AA is an amino acid selected from the group consisting of glycine, D or L proline, sarcosine, D or L alanine, D or L N-methylalanine, β-alanine, N-methyl-β-alanine, α-aminoisobutyric acid, and N-methyl-α-aminoisobutyric acid; and

[0265] where R' is:

[0266]

[0267] In some embodiments, R 1 is -H. In other embodiments, Z 1 , Z 2 , Z 3 and Z 4 At least one of them is not H and / or Z 1 , Z 2 , Z 3 and Z 4 At least one of is -F or -NO2. In some embodiments, when n is 0, X is absent. In other embodiments, when n is 0, X is -CH2-. In some embodiments, n is 0 and X is -O- or -S-. Additional embodiments include pharmaceutically acceptable salts, solvates, hydrates, tautomers, and solid forms of the disclosed compounds.

[0268] In some embodiments, the compound is selected from the following specific compounds:

[0269]

[0270]

[0271]

[0272]

[0273] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof.

[0274] In certain embodiments, the compound is not bound to a thiol-bearing macromolecular carrier or a thiol-bearing tumor-specific carrier. In other embodiments, the compound is bound to a thiol-bearing macromolecular carrier or a thiol-bearing tumor-specific carrier. For example, the thiol-bearing macromolecular carrier or the thiol-bearing tumor-specific carrier is selected from endogenous albumin, exogenous albumin, an antibody, an antibody fragment, a peptide, a natural or synthetic polymer, a liposome, and a nanoparticle.

[0275] In some embodiments, TBG is an optionally substituted maleimide group, such as an unsubstituted maleimide group. In some embodiments, following administration to a subject, such as a human, the maleimide group rapidly and selectively binds to cysteine ​​34 of albumin.

[0276] In some embodiments, Z 1’ Selected from -NO2 or -SO3M 2 and / or Y' is selected from -NHC(O)- or

[0277] In some embodiments, R 1’ yes In some embodiments, R 2’ Selected from optionally substituted C1-C 18 Alkyl, wherein the C1-C 18 Optionally up to six carbon atoms in the alkyl group are each independently substituted with -OCH2CH2- (eg, 1, 2, 3, 4, 5, or 6 carbon atoms are substituted with -OCH2CH2-).

[0278] In some embodiments, R' is:

[0279]

[0280] For example, R' can be:

[0281]

[0282] Specific compounds within the present disclosure include the following:

[0283]

[0284]

[0285]

[0286]

[0287]

[0288] or a pharmaceutically acceptable salt, hydrate, solvate or isomer thereof.

[0289] Pharmaceutical composition

[0290] In some embodiments, the present disclosure provides pharmaceutical compositions comprising the compounds described herein. In some embodiments, the composition comprises a compound of formula (I), wherein R' is:

[0291] Or the various embodiments disclosed herein.

[0292] The total amount of the compound in the composition administered to the patient is the amount that is most suitable for the patient. It will be appreciated by those skilled in the art that different individuals may require different total amounts of therapeutically effective substances. In certain embodiments, the amount of the compound is a pharmaceutically effective amount. The technician will be able to determine the amount of the compound in the composition required for the treatment of the patient based on factors such as the patient's age, weight, and physical condition. The concentration of the compound depends on its solubility in the intravenous administration solution and the volume of liquid that can be administered. For example, the concentration of the compound in the injectable composition can be from about 0.1 mg / ml to about 50 mg / ml. In certain embodiments, the concentration of the compound can be in the range of from about 0.1 mg / ml to about 40 mg / mL.

[0293] The pharmaceutical compositions and kits disclosed herein may also include diluents, fillers, salts, buffers, stabilizers, solubilizers, protective agents, and other materials well known in the art. The term "pharmaceutically acceptable" refers to non-toxic materials that do not interfere with the biological activity effectiveness of the active ingredient. The characteristics of the carrier will depend on the route of administration.

[0294] The composition can be administered in a variety of conventional ways. Exemplary routes of administration that can be used include oral, parenteral, intravenous, intraarterial, cutaneous, subcutaneous, intramuscular, topical, intracranial, orbital, ophthalmic, intravitreal, intraventricular, intracapsular, intraspinal, intracisternal, intraperitoneal, intranasal, aerosol, central nervous system (CNS) administration or administration by suppository. In some embodiments, the composition is suitable for parenteral administration. These compositions can be administered, for example, intraperitoneally, intravenously or intrathecally. In some embodiments, the composition is injected intravenously. In some embodiments, the reconstructed preparation can be prepared by reconstructing a lyophilized compound composition in a reconstructed solution including, for example, alcohol, DMSO and / or polyethylene glycol and water and / or a salt buffer. This reconstruction can include adding a reconstructed solution and, for example, mixing by swirling or vortexing the mixture. Then, by mixing, for example, lactated Ringer's solution, 5% glucose solution, isotonic saline or a suitable salt buffer with the preparation to make an injectable composition, the reconstructed preparation can be made suitable for injection. Those skilled in the art will appreciate that the method of administering a therapeutically effective material preparation or composition will depend on factors such as the age, weight and physical condition of the patient being treated and the disease or condition being treated. Therefore, the technician will be able to select the best method of administration for the patient based on the specific circumstances.

[0295] In some embodiments, the compounds and compositions disclosed herein are used to treat cancer, viral diseases, autoimmune diseases, acute or chronic inflammatory diseases, and diseases caused by bacteria, fungi, or other microorganisms.

[0296] In some embodiments, the compounds disclosed herein can be used in the manufacture of a medicament for treating a disease selected from cancer, viral diseases, autoimmune diseases, acute or chronic inflammatory diseases, and diseases caused by bacteria, fungi or other microorganisms.

[0297] In some embodiments, the cancer is a blood cancer or a solid tumor cancer. In some embodiments, the cancer is selected from carcinoma, sarcoma, leukemia, lymphoma, multiple myeloma and melanoma.

[0298] In some embodiments, the cancer is adenocarcinoma, uveal melanoma, acute leukemia, acoustic neuroma, ampullary cancer, anal cancer, astrocytoma, basal cell carcinoma, pancreatic cancer, connective tissue tumors, bladder cancer, bronchial cancer, non-small cell bronchial cancer, breast cancer, Burkitt's lymphoma, uterine corpus cancer, CUP syndrome, colon cancer, small intestine cancer, ovarian cancer, endometrial cancer, gallbladder cancer, gallbladder cancer, uterine cancer, cervical cancer, neck, nose, ear tumors, blood tumors, hairy cell leukemia disease, urethral cancer, skin cancer, glioma, testicular cancer, Kaposi's sarcoma, laryngeal cancer, bone cancer, colorectal cancer, head and neck cancer, colon cancer, craniopharyngioma, liver cancer, leukemia, lung cancer, non-small cell lung cancer, Hodgkin's lymphoma, non-Hodgkin's lymphoma, stomach cancer, colon cancer, medulloblastoma, melanoma, meningioma, kidney cancer, renal cell carcinoma, oligodendroglioma, esophageal cancer, osteolytic and bone cancer, osteosarcoma, ovarian cancer, pancreatic cancer, penis cancer, prostate cancer, tongue cancer, ovarian cancer and lymph node cancer.

[0299] In some embodiments, the present disclosure provides a kit comprising a compound as described herein and a pharmaceutically acceptable excipient, carrier, and / or diluent.

[0300] In certain embodiments, one or more excipients may be included in the composition. It will be appreciated by those skilled in the art that the selection of any excipient may affect the selection of any other excipient. For example, the selection of excipient may exclude the use of one or more other excipients, because the combination of excipients may produce undesirable effects. Those skilled in the art will be able to empirically determine which excipients (if any) are included in the composition. Excipients may include, but are not limited to, cosolvents, solubilizing agents, buffers, pH adjusting agents, swelling agents, surfactants, encapsulating agents, tonicity regulators, stabilizers, protective agents, and viscosity modifiers. In certain embodiments, it may be beneficial to include a pharmaceutically acceptable carrier in the composition.

[0301] In certain embodiments, solubilizing agent can be included in the composition.Solubilizing agent can be used to increase the solubility of any component (including compound or excipient) of composition.Solubilizing agent as described herein is not intended to constitute an exhaustive list, but is only provided as an exemplary solubilizing agent that can be used in the composition.In certain embodiments, solubilizing agent includes but is not limited to ethanol, tert-butyl alcohol, polyethylene glycol, glycerine, methyl parahydroxybenzoate, propyl parahydroxybenzoate, polyethylene glycol, polyvinyl pyrrolidone, cyclodextrin, such as dimethyl-β-cyclodextrin, hydroxyethyl-β-cyclodextrin, hydroxypropyl-β-cyclodextrin and trimethyl-β-cyclodextrin and combination thereof, and any pharmaceutically acceptable salt and / or combination thereof.

[0302] The pH of the composition can be any pH that provides the desired properties for the preparation or composition. The desired properties can include, for example, compound stability, the retention rate of the compound increased compared to the composition at other pH values, and the filtration efficiency improved. In certain embodiments, the pH value of the composition can be about 3.0 to about 9.0, for example, about 5.0 to about 7.0. In a particular embodiment, the pH value of the composition can be 5.5 ± 0.1, 5.6 ± 0.1, 5.7 ± 0.1, 5.8 ± 0.1, 5.9 ± 0.1, 6.0 ± 0.1, 6.1 ± 0.1, 6.2 ± 0.1, 6.3 ± 0.1, 6.4 ± 0.1, 6.5 ± 0.1, 6.6 ± 0.1, 6.7 ± 0.1, 6.8 ± 0.1, 6.9 ± 0.1, 7.0 ± 0.1, 7.1 ± 0.1 and 7.2 ± 0.1.

[0303] In some embodiments, it may be beneficial to buffer pH by including one or more buffers in the composition. In certain embodiments, the buffer may have, for example, a pKa of about 5.5, about 6.0 or about 6.5. It will be appreciated by those skilled in the art that appropriate buffers may be selected to be included in the composition based on its pKa and other characteristics. Buffers are well known in the art. Therefore, the buffers described herein are not intended to constitute an exhaustive list, but are merely provided as exemplary buffers that may be used in the preparations or compositions of the present disclosure. In certain embodiments, buffers include, but are not limited to, Tris, Tris-HCl, potassium phosphate, sodium phosphate, sodium citrate, sodium ascorbate, a combination of sodium phosphate and potassium phosphate, Tris / Tris-HCl, sodium bicarbonate, arginine phosphate, arginine hydrochloride, histidine hydrochloride, cocoate, succinate, 2-(N-morpholino)ethanesulfonic acid (MES), maleate, bis-tris, phosphate, carbonate and any pharmaceutically acceptable salt and / or combination thereof.

[0304] In some embodiments, a pH adjusting agent may be included in the composition. Changing the pH of the composition may have a beneficial effect on, for example, the stability or solubility of the compound, or may be useful in preparing a composition suitable for parenteral administration. pH adjusting agents are well known in the art. Therefore, the pH adjusting agents described herein are not intended to constitute an exhaustive list, but are merely provided as exemplary pH adjusting agents that can be used in the composition. pH adjusting agents may include, for example, acids and bases. In some embodiments, pH adjusting agents include, but are not limited to, acetic acid, hydrochloric acid, phosphoric acid, sodium hydroxide, sodium carbonate, and combinations thereof.

[0305] In some embodiments, a swelling agent may be included in the composition. Swelling agents are commonly used in freeze-dried compositions to increase the volume of the composition and to aid in visualization of the composition, particularly where freeze-dried particles are difficult to see. Swelling agents may also help prevent the ejection of active ingredients of the pharmaceutical composition and / or aid in cryoprotection of the composition. Swelling agents are well known in the art. Therefore, the swelling agents described herein are not intended to constitute an exhaustive list, but are merely provided as exemplary swelling agents that may be used in the composition.

[0306] Exemplary bulking agents may include carbohydrates, monosaccharides, disaccharides, polysaccharides, sugar alcohols, amino acids and sugar acids and combinations thereof. Carbohydrate bulking agents include, but are not limited to, monosaccharides, disaccharides, polysaccharides, carbohydrates, starches, aldoses, ketoses, amino sugars, glyceraldehydes, arabinose, lyxose, pentoses, ribose, xylose, galactose, glucose, hexose, idose, mannose, talose, heptose, glucose, fructose, methyl α-D-pyranoglucoside, maltose, lactones, sorbose, erythrose, threose, arabinose, allose, altrose, gulose, idose, talose, erythrose, ribose, xylose, Sugar, allulose, tagatose, tagatose glucosamine, galactosamine, arabinan, fructan, fucoidan, galactan, galacturonic acid, glucan, mannan, xylan, inulin, fructan, fucoidan, carrageenan, galactose aldosterone, pectin, amylose, pullulan, glycogen, amylopectin, cellulose, pyrifos polysaccharide, chitin, agarose, keratin, chondroitin, dermatan, hyaluronic acid, xanthan gum, sucrose, trehalose, dextran and lactose. Sugar alcohol bulking agents include but are not limited to sugar alcohols, inositol, sorbitol and mannitol. Sugar acid bulking agents include but are not limited to aldonic acid, uronic acid, aldonic acid, gluconic acid, isoascorbic acid, ascorbic acid, saccharic acid, glucuronic acid, gluconic acid, glucuronic acid, galacturonic acid, mannuronic acid, neuraminic acid, pectic acid and alginic acid. Amino acid bulking agents include, but are not limited to, glycine, histidine, and proline.

[0307] In certain embodiments, surfactants may be included in the composition. Surfactants generally reduce the surface tension of liquid compositions. This may provide beneficial properties, such as improving the convenience of filtration. Surfactants may also be used as emulsifiers and / or solubilizers. Surfactants are well known in the art. Therefore, the surfactants described herein are not intended to constitute an exhaustive list, but are merely provided as exemplary surfactants that may be used in the preparations or compositions of the present disclosure. Surfactants that may be included include, but are not limited to, sorbitol esters, such as polysorbates (e.g., polysorbate 20 and polysorbate 80), lipopolysaccharides, polyethylene glycols (e.g., PEG 400 and PEG 3000), poloxamers (i.e., pluronics), ethylene oxide and polyethylene oxide (e.g., Triton X-100), saponins, phospholipids (e.g., lecithin) and combinations thereof.

[0308] In certain embodiments, encapsulating agent can be included in the composition. Encapsulating agent can isolate molecules and help stabilize or solubilize molecules. Encapsulating agent is well known in the art. Therefore, encapsulating agent as described herein is not intended to constitute an exhaustive list, but is only provided as an exemplary encapsulating agent that can be used in the composition. The encapsulating agent that can be included in the composition includes but is not limited to α-cyclodextrin, β-cyclodextrin, γ-cyclodextrin and combinations thereof (for example, α-cyclodextrin, dimethyl-α-cyclodextrin, hydroxyethyl-α-cyclodextrin, hydroxypropyl-α-cyclodextrin, trimethyl-α-cyclodextrin, β-cyclodextrin, dimethyl-β-cyclodextrin, hydroxyethyl-β-cyclodextrin, hydroxypropyl-β-cyclodextrin, trimethyl-β-cyclodextrin, γ-cyclodextrin, dimethyl-γ-cyclodextrin, hydroxyethyl-γ-cyclodextrin, hydroxypropyl-γ-cyclodextrin, trimethyl-γ-cyclodextrin and combinations thereof).

[0309] In certain embodiments, a tonicity regulator may be included in the composition. When, for example, a composition is administered to a patient by parenteral administration, the tension of the liquid composition is an important consideration. Therefore, a tonicity regulator may be used to help make the composition suitable for administration. Tonicity regulators are well known in the art. Therefore, the tonicity regulators described herein are not intended to constitute an exhaustive list, but are merely provided as exemplary tonicity regulators that may be used in the composition. Tonicity regulators may be ionic or nonionic, including but not limited to inorganic salts, amino acids, carbohydrates, sugars, sugar alcohols, and carbohydrates. Exemplary inorganic salts may include sodium chloride, potassium chloride, sodium sulfate, and potassium sulfate. Exemplary amino acids are glycine. Exemplary sugars may include sugar alcohols, such as glycerol, propylene glycol, glucose, sucrose, lactose, dextrose, and mannitol.

[0310] In some embodiments, stabilizers may be included in the composition. Stabilizers help increase the stability of the compound in the composition. For example, this may occur by reducing the degradation of the compound or preventing it from aggregating. Without wishing to be bound by theory, mechanisms for enhancing stability may include isolating the compound from the solvent or inhibiting free radical oxidation of the therapeutically effective substance. Stabilizers are well known in the art. Therefore, the stabilizers described herein are not intended to constitute an exhaustive list, but are merely provided as exemplary stabilizers that can be used in the composition. Stabilizers may include, but are not limited to, emulsifiers and surfactants.

[0311] In some embodiments, a protective agent may be included in the composition. A protective agent is an agent that protects pharmaceutically active ingredients (e.g., therapeutically effective substances or compounds) from adverse conditions (e.g., instability caused by freezing or lyophilization or oxidation). Protective agents may include, for example, cryoprotectants, lyoprotectants, and antioxidants. When the composition is exposed to a temperature below its freezing point, cryoprotectants may be used to prevent the loss of efficacy of active pharmaceutical ingredients (e.g., anthracycline compounds). For example, a cryoprotectant may be included in a reconstructed lyophilized formulation so that the formulation may be frozen before dilution for intravenous administration. Cryoprotectants are well known in the art. Therefore, the cryoprotectants described herein are not intended to constitute an exhaustive list, but are merely provided as exemplary cryoprotectants that can be used in the composition. Cryoprotectants include, but are not limited to, solvents, surfactants, encapsulating agents, stabilizers, viscosity modifiers, and combinations thereof. Cryoprotectants may include, for example, disaccharides (e.g., sucrose, lactose, maltose, and trehalose), polyols (e.g., glycerol, mannitol, sorbitol, and diethylene glycol), glycols (e.g., ethylene glycol, polyethylene glycol, and propylene glycol).

[0312] Lyoprotectants can be used to stabilize the components of the composition subjected to lyophilization. For example, a therapeutically effective substance can be lyophilized with a lyoprotectant before reconstitution. Lyoprotectants are well known in the art. Therefore, the lyoprotectants described herein are not intended to constitute an exhaustive list, but are merely provided as exemplary lyoprotectants that can be used in the composition. Lyoprotectants include, but are not limited to, solvents, surfactants, encapsulating agents, stabilizers, viscosity modifiers, and combinations thereof. Exemplary lyoprotectants can be, for example, sugars and polyols. Trehalose, sucrose, dextran, and hydroxypropyl-β-cyclodextrin are non-limiting examples of lyoprotectants.

[0313] Antioxidants can be used to prevent oxidation of the components of the composition. Oxidation may lead to aggregation of the drug or other harmful effects on the purity of the drug or its effectiveness. Antioxidants are well known in the art. Therefore, the antioxidants described herein are not intended to constitute an exhaustive list, but are merely provided as exemplary antioxidants that can be used in the composition. The antioxidant can be, for example, sodium ascorbate, citrate, mercaptan, metabisulfite and combinations thereof.

[0314] In some embodiments, a viscosity modifier may be included in the composition. A viscosity modifier changes the viscosity of a liquid composition. This may be beneficial because viscosity plays an important role in making it easy to filter a liquid composition. The composition may be filtered before or after freeze drying and reconstitution. Viscosity modifiers are well known in the art. Therefore, the viscosity modifiers described herein are not intended to constitute an exhaustive list, but are merely provided as exemplary viscosity modifiers that can be used in the composition. Viscosity modifiers include solvents, solubilizers, surfactants, and encapsulating agents. Exemplary viscosity modifiers that may be included in the composition include, but are not limited to, N-acetyl-DL-tryptophan and N-acetyl-cysteine.

[0315] Antitumor activity in human tumor xenograft mouse models

[0316] Albumin-bound maytansinoids 30 and 31 exhibited excellent antitumor activity in five nude mouse human tumor xenograft models, causing partial and complete tumor regression in all human tumor xenografts evaluated (see Figures 3 to 16 ). This includes approximately 80 to 110 mm 3 Starting tumor volumes ranged from 400 mm to 400 mm. 3 In addition, in most cases, treatment with albumin-bound maytansine alkaloids 30 and 31 induced long-term remissions and reduced relative tumor volume (RTV). The parent compound maytansine was mainly inactive in the tested models or showed only minor tumor inhibition. Examples 11 to 19 and Figures 3 to 16 The experimental procedures and results of the tumor-bearing mouse model are described in detail.

[0317] Treatment

[0318] The compounds and compositions described herein can be used in a variety of clinical applications. In embodiments, the method of treatment utilizes a compound of a composition comprising a compound of formula (I), wherein R' is:

[0319] Or the various embodiments disclosed herein.

[0320] The compounds and compositions described herein can induce prolonged or long-term inhibition of tumor growth. In certain embodiments, the prolonged or long-term inhibition of tumor growth is without any weight loss or any or only minimal bone marrow toxicity.

[0321] In some embodiments, the present disclosure provides a method for treating a malignant disease, the method comprising administering to a patient in need thereof a therapeutically effective amount of a pharmaceutical composition comprising a compound described herein. For example, some embodiments include a method for treating a patient suffering from a disease or condition selected from the following: cancer, viral disease, autoimmune disease, acute or chronic inflammatory disease, and disease caused by bacteria, fungi or other microorganisms, the method comprising administering to a patient in need thereof a therapeutically effective amount of a compound according to the present disclosure.

[0322] The present disclosure provides methods for treating a condition or disease in a patient selected from cancer, viral diseases, autoimmune diseases, acute or chronic inflammatory diseases, and diseases caused by bacteria, fungi or other microorganisms, comprising administering to the patient a compound or pharmaceutical composition disclosed herein.

[0323] In some embodiments, the cancer comprises a vascularized tumor. In some embodiments, the cancer is a blood cancer or a solid tumor cancer. In some embodiments, the cancer is selected from carcinoma, sarcoma, leukemia, lymphoma, multiple myeloma and melanoma.

[0324] In some embodiments, the cancer is selected from adenocarcinoma, uveal melanoma, acute leukemia, acoustic neuroma, ampullary cancer, anal cancer, astrocytoma, basal cell carcinoma, pancreatic cancer, connective tissue tumors, bladder cancer, bronchial cancer, non-small cell bronchial cancer, breast cancer, Burkitt's lymphoma, uterine corpus cancer, CUP syndrome, colon cancer, small intestine cancer, ovarian cancer, endometrial cancer, gallbladder cancer, uterine cancer, cervical cancer, neck, nose, ear tumors, blood tumors, hairy cell leukemia, Urethral cancer, skin cancer, glioma, testicular cancer, Kaposi's sarcoma, laryngeal cancer, bone cancer, colorectal cancer, head and neck cancer, colon cancer, craniopharyngioma, liver cancer, leukemia, lung cancer, non-small cell lung cancer, Hodgkin's lymphoma, non-Hodgkin's lymphoma, stomach cancer, colon cancer, medulloblastoma, melanoma, meningioma, kidney cancer, renal cell carcinoma, oligodendroglioma, esophageal cancer, osteolytic and bone cancer, osteosarcoma, ovarian cancer, pancreatic cancer, penis cancer, prostate cancer, tongue cancer, ovarian cancer and lymph node cancer

[0325] Some embodiments include methods of increasing the concentration of a metabolite of a compound in a tumor, the method comprising administering a compound according to the present disclosure. In embodiments, the compound is a compound of formula (I), wherein R' is:

[0326] Or various embodiments disclosed herein. In some embodiments, the increase is compared to an equivalent dose of an unmodified active agent, for example, the "unmodified active agent" can be the same compound of formula (I), wherein R' is O.

[0327] Some embodiments include a method of reducing the cytotoxicity of a compound, the method comprising administering a compound or pharmaceutical composition of the present disclosure to a patient in need thereof, wherein the administration results in reduced cytotoxicity when compared to an equivalent dose of an unmodified active agent. For example, in some embodiments, the method of reducing cytotoxicity comprises administering a compound of formula (I), wherein R' is:

[0328] or various embodiments disclosed herein, and the “unmodified active agent” is the same compound of formula (I), wherein R′ is O.

[0329] illustration

[0330] Having now generally described various aspects of the disclosure, these aspects will be more readily understood by reference to the following examples, which are included merely for the purpose of illustrating certain features and embodiments of the disclosure and are not intended to be limiting.

[0331] Equivalent

[0332] Those skilled in the art will recognize or be able to ascertain, using no more than routine experimentation, numerous equivalents to the compounds, compositions, and methods of use described herein. Such equivalents are considered to be within the scope of the present disclosure.

[0333] Examples

[0334] The following examples demonstrate various embodiments and aspects of the invention.

[0335] abbreviation

[0336] The following is a list of abbreviations used in the examples, along with their full chemical names. If not defined, these terms have their generally accepted meanings.

[0337] aq.=water

[0338] Boc=N-tert-butyloxycarbonyl

[0339] calcd.=calculated value

[0340] DCM = dichloromethane

[0341] DIC = N,N'-diisopropylcarbodiimide

[0342] DMAP = N,N-dimethyl-4-aminopyridine

[0343] DMF=N,N-dimethylformamide

[0344] DMSO = dimethyl sulfoxide

[0345] CV = column volume

[0346] EDC = 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide

[0347] eq = equivalent

[0348] ESI = Electrospray Ionization

[0349] Fmoc = fluorenylmethoxycarbonyl

[0350] HATU = (1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium 3-oxide hexafluorophosphate)

[0351] HOAt = 1-hydroxy-7-azabenzotriazole

[0352] HOBt = N-hydroxybenzotriazole

[0353] HPLC = High Performance Liquid Chromatography

[0354] HRMS = High Resolution Mass Spectrometry

[0355] HSA = human serum albumin

[0356] IC = Ion Chromatography

[0357] LC-MS = Liquid Chromatography Mass Spectrometry

[0358] LRMS = Low Resolution Mass Spectrometry

[0359] MeCN = acetonitrile

[0360] MeOH = methanol

[0361] NMR = Nuclear Magnetic Resonance Spectroscopy

[0362] NP=Normal Phase

[0363] Np=4-nitrophenoxycarbonyl

[0364] nd = Not Detectable

[0365] na = not available

[0366] PBS = phosphate buffered saline, pH 7

[0367] RP = Reverse

[0368] TFA = trifluoroacetic acid

[0369] TRIS = 2-amino-2-(hydroxymethyl)-1,3-propanediol

[0370] Example 1

[0371] Preparation of Linker 1

[0372] Linker 1 can be prepared as described below and is shown in Scheme 1.

[0373] Solution 1

[0374]

[0375] Synthesis of 4-nitro-2-sulfobenzoic acid (A)

[0376]

[0377] To a stirred solution of potassium permanganate (72 g, 460 mmol, 4.5 eq.) in water (450 mL) was added a solution of 4-nitro-2-sulfonic acid hydrate (26 g, 102 mmol, 1.0 eq.) in Millipore water (100 mL) over 10 seconds. The resulting purple mixture was stirred at 115° C. for 5 hours, after which time it turned brown. HPLC analysis (PDA 220 nm) confirmed that the reaction was complete after 5 hours (>90% conversion). The reaction mixture was cooled to room temperature. The brown solid formed during the reaction was removed by suction filtration over a pad of celite, washed with Millipore water (300 mL), and the brown / yellow filtrate solution was concentrated to approximately 125 mL at 40° C. on a rotary evaporator and slowly acidified with a 5M HCl (approximately 2 mL) solution until a white suspension (approximately pH 1.0) was formed. The white suspension is then heated at 100°C until a clear solution is obtained, which is allowed to stand in an ice bath for 10 minutes until a white solid is formed. The white solid is obtained by suction filtration using a sintered filter. The white solid is then dried under high vacuum to give 4-nitro-2-sulfobenzoic acid. Yield: 18 g (72%). Purity by RP-HPLC, 220 nm, >95%. C7H4NO7S[MH] - LRMS-ESI (m / z) calculated value: 245.98. Found value: 245.83.

[0378] Synthesis of 4-amino-2-sulfobenzoic acid (B)

[0379]

[0380] A stirred suspension of 4-nitro-2-sulfobenzoic acid (13 g, 51 mmol, 1.0 eq) in water (75 mL) was heated at reflux until the 4-nitro-2-sulfobenzoic acid was completely dissolved. Acetic acid (7.2 mL) was then added at this temperature, followed by iron powder (9.5 g, 180 mmol, 3.5 eq) in portions over 10 minutes (about 1 g / min) to avoid exothermic reaction. The reaction mixture was then stirred at reflux for 1 hour. During this period, a brown solid was formed and HPLC analysis (PDA 220 nm) confirmed that the reaction was complete (>95% conversion). The brown solid was removed by direct suction filtration on a celite pad (while still hot) and further washed with hot water. The filtrate was refiltered. The resulting filtrate was concentrated to a final volume of 100 mL at 40° C. using a rotary evaporator. Concentrated HCl was added dropwise until pH 1 was reached and a white / yellow solid was precipitated. The suspension was left at 4°C for 1 hour. The solid was collected by suction filtration using a sintered filter and dried under high vacuum to give 4-amino-2-sulfobenzoic acid as a white solid. Yield: 9 g (81%). Purity by RP-HPLC, 220 nm, >95%. C7H4NO5S[MH] - LRMS-ESI (m / z) calculated: 216.00. Found: 216.16.

[0381] Synthesis of 6-Maleimidocaproyl chloride or EMC-Cl (C)

[0382]

[0383] At room temperature and under N2 atmosphere, oxalyl chloride (15mL, 171mmol, 1.1 equivalents) was added to a yellow stirred solution of 6-maleimidocaproic acid (EMC) (33g, 156mmol, 1.0 equivalents) in anhydrous DCM (150mL) using a dropping funnel within 30 minutes (about 0.5mL / min). The reactants were stirred at room temperature for 5 hours. During the reaction time, the color of the reaction solution turned dark yellow, and HPLC analysis (PDA 220nm) confirmed that the reaction was complete after 5 hours (>95% conversion). The solvent was removed at 40°C with a rotary evaporator to obtain an oil. The residual oil was dried overnight under high vacuum (solidified overnight). The light brown solid obtained was crushed and further dried under high vacuum for 20 hours to obtain 6-maleimidocaproyl chloride as a yellow microcrystalline solid. The compound can be used for the next step reaction without further purification. Yield: 34g (95%). Purity by RP-HPLC, 220 nm, >95% as methyl ester. 11 H 16 NO4 (as methyl ester) [M+H] +LRMS-ESI (m / z) calculated: 226.10. Found: 225.97.

[0384] Synthesis of 4-(6-maleimidohexamido)-2-sulfobenzoic acid (D)

[0385]

[0386] Under N2 atmosphere, 4-amino-2-sulfobenzoic acid (18.5 g, 85.0 mmol, 1.0 equivalent) was dissolved in anhydrous DMF (300 mL). The solution was cooled to 4 ° C and stirred for 10 minutes. Then, a dropping funnel was used to drop (about 0.3 mL / min) 4-N-methylmorpholine (18.7 mL, 170 mmol, 2.0 equivalent) into the cooled solution over 1 hour. A dropping funnel was used to drop (about 0.5 g / min) EMC-Cl (29.3 g, 127 mmol, 1.5 equivalent) in anhydrous DMF (200 mL) into the dark brown mixture over 1 hour. The reaction mixture was stirred overnight and then allowed to reach room temperature over 10 hours. After the reaction was completed by HPLC analysis (PDA 220 nm, > 95% conversion), the reaction solution was distributed in 8 × 50 mL falcon tubes.

[0387] The sample was centrifuged at 10°C and 4.000 rpm for 20 minutes. The supernatant was removed by decantation, and the solid was resuspended in 10 mL DMF per tube and centrifuged again at 10°C and 4.000 rpm for 20 minutes. All DMF supernatants were combined and concentrated at 50°C under reduced pressure for 3 hours to obtain a light orange solid. The solid was resuspended in methanol (250 mL) and transferred to 8×50 mL falcon tubes. The sample was centrifuged at 10°C and 4.000 rpm for 20 minutes. The supernatant was removed by decantation, the solid was resuspended in 5 mL methanol per tube and centrifuged again at 10°C and 4.000 rpm for 20 minutes. All solids were combined and dried under high vacuum for 24 hours to obtain a crystalline yellow solid. Yield: 17 g (48%). Purity obtained by RP-HPLC reverse, 220 nm, >80%. C 17 H 17 N2O8S[MH] - LRMS-ESI (m / z) calculated value: 409.08. Found value: 409.13.

[0388] Synthesis of 2-(2-(tert-butoxycarbonyl)hydrazine-1-carbonyl)-5-(6-maleimidohexaamido)benzenesulfonic acid or Boc-protected linker 1 (E)

[0389]

[0390] To a solution of 4-(6-maleimidohexacarboxamido)-2-sulfobenzoic acid (17.0 g, 41.4 mmol, 1.0 equiv) in anhydrous DMF (350 mL) was added EDC-HCl (8.72 g, 45.5 mmol, 1.1 equiv) and 1HOBt (6.15 g, 45.5 mmol, 1.1 equiv) under N2 atmosphere. The reaction mixture was stirred at room temperature for 30 minutes, followed by the addition of tert-butyl carbazate (7.12 g, 53.9 mmol, 1.3 equiv) and the solution turned from clear yellow to slightly reddish. The reaction mixture was stirred at room temperature overnight. After this time, the reaction was confirmed to be complete by HPLC (PDA 220 nm, >95% conversion). The solvent was removed by rotary evaporator at 40°C under high vacuum for 1 hour to give a purple-brown oil, which was equipped with two pre-packed SNAP ULTRA 340 g cartridges and HP-Sphere TM Purification was performed using the Biotage Isolera One rapid purification system with spherical silica. The tubes containing the desired product were combined and dried on a rotary evaporator under high vacuum for 10 hours to give 2-(2-(tert-butoxycarbonyl)hydrazine-1-carbonyl)-5-(6-maleimidohexamino)benzenesulfonic acid as a foamy yellow solid. Yield: 9 g (42%). RP-HPLC (220 nm) > 95%. C 22 H 27 N4O9S[MH] - LRMS-ESI (m / z) calculated: 523.16. Found: 523.15.

[0391] Synthesis of Linker 1

[0392]

[0393] To a cooling (4°C to 5°C) solution of Boc-protected linker 1 (10.2 g, 19.4 mmol, 1.0 equiv.) in anhydrous DCM (30 mL) was added dropwise (about 0.5 mL / min) of TFA (15 mL) over 30 minutes. After addition, the cold bath was removed and the reaction mixture was stirred at room temperature for 3 hours. After this time, the reaction was confirmed to be complete by HPLC analysis (PDA 220 nm). The reaction mixture was poured dropwise into six falcon tubes, and approximately 35 mL of cold diethyl ether was added to each falcon tube. A white precipitate was formed immediately. The test tube was placed at 4°C for 3 hours. After centrifugation of the falcon tube (4000 rpm, 20 minutes, 10°C), the supernatant was removed by decantation, and the solid was resuspended in 5 mL of ether per tube and centrifuged again (4000 rpm, 20 minutes, 10°C). The supernatant was again removed by decantation and the solid was collected and dried under high vacuum to provide Linker 1 as a white microcrystalline solid as a TFA salt. Yield: 10 g (96%). RP-HPLC (220 nm) > 95%. C 17 H 21 N4O7S[M+H] + LRMS-ESI (m / z) calculated value: 425.11. Found value: 425.07. 17 H 19 N4O7S[MH] - LRMS-ESI (m / z) calculated value: 423.11. Found value: 423.12. C 17 H 21 N4O7S[M+H] + HRMS-ESI (m / z) calculated value: 425.1125. Found value: 425.1125. C 17 H 19 N4O7S[MH] - HRMS-ESI (m / z) calculated value: 423.0978. Found value: 423.0980.

[0394] pass 1 H NMR and 13 C NMR confirmed the structure: 1H NMR (400MHz, DMSO-d6) δ11.94 (s, 1H; C1-NH), 10.27 (s, 1H; C8-NH), 8.01 (d, J=2.2Hz, 1H ; C4-CH), 7.93 (dd, J=8.5, 2.2Hz, 1H; C6-CH), 7.68 (d, J=8.4Hz, 1H; C7-CH), 7.00 (s, 2H; C 15-CH, C16-CH), 3.40 (t, J=7.0Hz, 2H; C13-CH2), 2.32 (t, J=7.4Hz, 2H; C9-CH2), 1.60 (p, J=7.5Hz, 2H; C10-CH2), 1.52 (p, J=7.2Hz, 2H; C12-CH2), 1.26 (q, J=8.8Hz, 2H; C11-CH2); 13 C NMR (101MHz, DMSO-d6) δ172.20(C8), 171.54(C14, C17), 167.59(C1), 145.73(C5), 142.09(C3), 134.90(C15, C16), 1 32.09(C7), 123.79(C2), 119.44(C6), 117.47(C4), 37.42(C13), 36.65(C9), 28.22(C12), 26.21(C11), 24.90(C10). C 17 H 21 Calculated value for N4O7S·1 / 2: TFA C, 45.71; H, 4.26; N, 11.85; S, 6.78. Found: C, 46.2917; H, 4.4836; N, 12.8879; S, 6.7886. TFA content 0.51%.

[0395] Linker 1 can also be prepared as described below and shown in Scheme 2.

[0396] Solution 2

[0397]

[0398] Synthesis of 5-amino-2-(2-(tert-butoxycarbonyl)hydrazine-1-carbonyl)benzenesulfonic acid (F)

[0399]

[0400] To a suspension of B (30.00 g, 138.12 mmol, 1.00 equiv) in anhydrous acetonitrile (600 mL) was added triethylamine (41.93 g, 57.76 mL, 414.37 mmol, 3.00 equiv) and the mixture was stirred for 10 minutes. After that, tert-butyl carbazate (27.38 g, 207.19 mmol, 1.50 equiv) was added and the mixture was cooled to -35 °C. At this temperature, propylphosphonic anhydride solution T3P (114.27 g, 106.79 mL, 179.56 mmol, 50% ethyl acetate solution, 1.3 equiv) was added dropwise over 1 hour. The reactants were stirred at -35 °C for 2 h. The mixture was allowed to warm to room temperature and filtered by 545 (100 g) was filtered. Washed with acetonitrile (500 mL) The two filtrates were combined and concentrated to 250 mL. The solution was divided equally into 6 portions and the solvent was removed under reduced pressure. Each portion was dissolved in 1% Et3N in dichloromethane (50 mL) and purified by NP flash chromatography on a Biotage Isolera equipped with a pre-packed SNAP Ultra 340 g cartridge. TM Purification was performed on a TRIAZINE One flash purification system using a step gradient from 2% to 12% methanol (containing 1% NEt3) in DCM (containing 1% NEt3) over 7 column volumes. The purified fractions were then combined, the solvent removed under reduced pressure, and the solid dried under high vacuum to give the title compound F as an off-white solid. Yield: 53.25 g, 108.0 mmol, 78.2% (NMR in DMSO-d6 showed the presence of 1.6 equivalents of triethylamine). HPLC (Method 9, 220 nm) > 99%. C 12 H 16 N3O6S[MH] - LRMS-ESI (m / z) calculated value: 330.08. Found value: 330.08.

[0401] Synthesis of N-ethyl-N-isopropylpropan-2-ammonium 2-(2-(tert-butoxycarbonyl)hydrazine-1-carbonyl)-5-(6-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-)hexamino)benzenesulfonate (E)

[0402]

[0403] To a mixture of F (45.00 g, 91.24 mmol, 1.00 equiv) and 6-maleimidocaproic acid (19.27 g, 91.24 mmol, 1.00 equiv) was added acetonitrile (450 mL), triethylamine (13.85 g, 19.08 mL, 136.86 mmol) and T3P (43.55 g, 40.70 mL, 136.86 mmol, 50% ethyl acetate solution) in batches at room temperature. The solution was stirred at room temperature for 24 hours. The solvent was removed under reduced pressure. The crude material was then purified by a flash purification system using seven pre-packed SNAP Ultra 340g cartridges with a linear gradient running from 2% methanol in dichloromethane to 15% methanol to give the title compound E as an off-white solid. Yield: 30.55 g, 53.5% (NMR in DMSO-d6 showed the presence of 1.1 equivalents of triethylamine). HPLC (Method 9, 220 nm) > 99%. 22 H 27 N4O9S[MH] - LRMS-ESI (m / z) calculated: 523.15. Found: 523.26.

[0404] Synthesis of 5-(6-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)hexanamido)-2-(hydrazinocarbonyl)benzenesulfonic acid (Linker 1)

[0405]

[0406] To a cold suspension (4°C) of E (10.00 g, 15.98 mmol, 1.00 equiv) in dichloromethane (50 mL) was added trifluoroacetic acid (18.22 g, 12.31 mL, 159.81 mmol, 10.17 equiv) over 15 min. The mixture was further stirred at 4°C for 15 min, then allowed to gradually warm to room temperature and stirred for 150 min. The reaction mixture was added dropwise to a stirred solution of methyl tert-butyl ether, MTBE (400 mL) and dichloromethane (200 mL) via a separatory funnel. The resulting white solid was purified by The mixture was filtered through a porosity sinter funnel and washed sequentially with dichloromethane (2×150 mL) and MTBE (1×50 mL), MeOH (1×50 mL) and (again) MTBE (2×150 mL). The solid was dried overnight at room temperature on the sinter funnel for 10 minutes. It was further dried under high vacuum at 25° C. for 18 hours. The final product linker 1 was obtained as a yellow solid. Yield: 5.786 g, 13.63 mmol, 98.7%, HPLC (Method 9, 220 nm)>96%. 17 H 19 N4O7S[MH] -LRMS-ESI (m / z) calculated: 423.10. Found: 422.95.

[0407] Examples 2

[0408] Preparation of keto-maytansinoids by direct esterification of maytansinol with keto acids

[0409] General method A: Maytansinol (500 mg, 0.88 mmol, 1 eq.) and the corresponding keto acid (3.52 mmol, 4 eq.) were dissolved in anhydrous DCM (40 mL) in the presence of activated molecular sieves and cooled to 4 to 5 ° C. with an ice bath. A solution of zinc (II) chloride in ether (2.64 mL, 2.64 mmol, 3.0 eq., 1 M solution) was then added to the solution within 10 seconds. The resulting solution was stirred at 4 to 5 ° C. for 40 minutes, and then N, N'-diisopropylcarbodiimide (0.55 mL, 3.52 mmol, 4 eq.) was added. The mixture was stirred at 4 ° C. and then immersed in an ice bath overnight to allow it to slowly reach room temperature. The conversion was monitored by LC-MS, and after reaching 60%, the reaction mixture was concentrated to half the volume under reduced pressure at 40 ° C. and filtered through a 0.45 μm syringe filter (Macherey-Nagel, PTFE-O-45 / 25) was filtered and the filtrate was evaporated. The final product was prepared using a pre-packed SNAP ULTRA 50g cartridge and HP-Sphere TM Purification was performed using a Biotage Isolera One flash purification system on spherical silica (linear gradient from 100% DCM to 90 / 10 DCM / methanol in 25 CV). The tubes containing the product were combined and dried on a rotary evaporator under high vacuum for 1 hour to afford the corresponding keto maytansinoid.

[0410] General method B: Over activated molecular sieves (0.8 g, 325 mesh particle size, Sigma Aldrich) in the presence of maytansinol (758mg, 1.34mmol, 1.0 equivalent), the corresponding keto acid (1.47mmol, 1.1 equivalent) and DMAP (181mg, 1.47mmol, 1.1 equivalent) were dissolved in anhydrous DCM (30mL) under N2 atmosphere. The mixture was cooled to 4°C in 10 minutes using an ice / water bath. In 30 minutes (about 10mg / min), a solution of EDC-HCl (283mg, 1.47mmol, 1.1 equivalent) in anhydrous DCM (15mL) was added to the cooled mixture, and the reaction mixture was stirred at 4°C for 2 hours. After this time, another portion of the keto acid (1.47 mmol, 1.1 eq) was added, and then a solution of EDC-HCl (283 mg, 1.47 mmol, 1.1 eq) in anhydrous DCM (10 mL) was added to the cooled mixture over 30 min (about 10 mg / min), and the reaction mixture was stirred at 4 ° C. for 2 h under an inert atmosphere. After this time, the addition of reagents was repeated again, and the reaction mixture was stirred at 4 ° C. overnight, then allowed to gradually reach room temperature during this time. The mixture was filtered (Macherey-Nagel, PTFE-O-45 / 25) and the solvent was removed by rotary evaporation at 40°C to a final volume of approximately 10 mL. The crude material was purified using a pre-packed SNAP ULTRA 100 g cartridge and HP-Sphere TM Purification was performed using the Biotage IsoleraOne flash purification system on spherical silica (linear gradient from 100% DCM to 90 / 10 DCM / methanol in 25 CV). The tubes containing the product were combined and the solvent was removed using a rotary evaporator to give a solid. The solid was dried under high vacuum to give the corresponding keto maytansinoid.

[0411] Preparation of Maytansinoid 2

[0412]

[0413] From the reaction of maytansinol with 2-(4-acetyl-2-fluorophenyl)acetic acid, using method A: maytansinoid 2 was obtained as a pale yellow solid. Yield: 47%. Purity by RP-HPLC, 220 nm, 96%. C 38 H 45 ClFN2O 10 [M+H] + LRMS-ESI (m / z) calculated value: 743.22. Found value: 743.25. C 38 H 43 ClFN2O 10[MH] - LRMS-ESI (m / z) calculated: 741.22. Found: 741.41.

[0414] pass 1 H NMR and 13 C NMR confirmed the structure: 1 H NMR (400MHz, CDCl3) δ7.73 (dd, J=7.9, 1.6Hz, 1H; C31-CH), 7.66 (dd, J=10.5, 1.6Hz, 1H; C27-CH), 7.46 (t, J=7.6Hz , 1H; C32-CH), 6.82 (d, J=1.8Hz, 1H; C17-CH), 6.59 (d, J=1.8Hz, 1H; C21-CH), 6.48 (dd, J=15.5, 11.0Hz, 1H; C12-CH ), 6.43 (s, 1H; C9-NH), 6.25 (d, J=10.9Hz, 1H; C13-CH), 5.63 (dd, J=15.4, 8.8Hz, 1H; C11-CH), 4.99 (dd, J=11.8, 2. 7Hz, 1H; C3-CH), 4.27 (td, J=11.2, 10.4, 1.8Hz, 1H; C7-CH), 3.97 (s, 3H; C20-OCH3), 3.90 (d, J=15.7Hz, 1H; C24-CH2 ), 3.76 (d, J=15.5Hz, 1H; C24-CH2), 3.54 (d, J=8.8Hz, 1H; C10-CH), 3.46 (d, J=12.8Hz, 1H; C15-CH2), 3.38 (s, 3H; C 10-OCH3), 3.19 (d, J=12.8Hz, 1H; C15-CH2), 3.00 (s, 3H; C1-NCH3), 2.87 (d, J=9.7Hz, 1H; C5-CH), 2.57 (s, 3H; C29-C H3), 2.51 (dd, J=14.1, 11.9Hz, 1H; C2-CH2), 2.17 (dd, J=13.9, 2.6Hz, 1H; C2-CH2), 1.72 (d, J=13.6Hz, 1H; C8-CH2) , 1.68 (s, 3H; C14-CH3), 1.50 (m, 1H; C6-CH), 1.28 (d, J=6.3Hz, 4H; C6-CH3, C8-CH2), 0.87 (d, J=1.4Hz, 3H; C4-CH3); 13 C NMR (101MHz, CDCl3) δ196.50(C28), 168.86(C23), 168.30(C1), 160.78(d, 1 JC-F =247.0Hz; C26), 156.13(C20), 152.46(C22), 142.53(C18), 140.30(C19), 140.24(C14), 138.49(d, 3 J C-F =6.4Hz; C30), 132.58(C12), 131.68(d, 3 J C-F =3.6Hz; C32), 128.32(C11), 126.38(d, 2 J C-F =16.1Hz; C25), 124.82(d, 4 J C-F =3.3Hz; C31), 124.56(C13), 122.04(C21), 119.52(C16), 114.73(d, 2 J C-F =23.2Hz; C27), 113.13(C17), 88.23(C10), 81.25(C9), 77.95(C3), 74. 37(C7), 66.24(C5), 60.39(C4), 56.91(C20-OCH3), 56.71(C10-OCH3), 4 7.26(C15), 38.37(C6), 36.01(C8), 35.46(C1-NCH3), 33.86(C24), 32.7 6(C2), 26.78(C29), 15.88(C14-CH3), 14.60(C6-CH3), 12.35(C4-CH3).

[0415] Preparation of maytansinoid 3

[0416]

[0417] From the reaction of maytansinol with 2-(3-acetylphenoxy)acetic acid, using Method B: Maytansinoid 3 was obtained as a pale yellow solid. Yield: 49%. Purity by RP-HPLC, 220 nm, 98%. C 38 H 46 ClN2O 11 [M+H] + LRMS-ESI (m / z) calculated: 741.23. Found: 741.23.

[0418] pass 1 H NMR and 13 C NMR confirmed the structure: 1H NMR(400MHz,CDCl3)δ7.58(dt,J=7.8,1.1Hz,1H;C30-CH),7.55(s,1H;C26-CH),7.44(d,J=7.9Hz,1H;C31-CH),7.12(dd,J=8.3,2.8Hz,1H;C32-CH),6.79(d,J=1.8Hz,1H;C17-CH),6.65(d,J=1.8Hz,1H;C21-CH),6.46(dd,J=15.4,11.0Hz,1H;C12-CH),6.37(s,1H;C9-NH),6.26(d,J=11.0Hz,1H;C13-CH),5.62(dd,J=15.4,8.9Hz,1H;C11-CH),5.08(dd,J=11.9,2.7Hz,1H;C3-CH),4.90(d,J=15.9Hz,1H;C24-CH2),4.66(d,J=15.9Hz,1H;C24-CH2),4.28(t,J=10.6Hz,1H;C7-CH),3.96(s,3H;C20-OCH3),3.68(s,1H;C9-OH),3.53(d,J=8.9Hz,1H;C10-CH),3.47(d,J=12.9Hz,1H;C15-CH2),3.36(s,3H;C10-OCH3),3.15(d,J=12.8Hz,1H;C15-CH2),2.92(d,J=9.7Hz,1H;C5-CH),2.87(s,3H;C1-NCH3),2.59(s,3H;C29-CH3),2.55(d,J=11.9Hz,1H;C2-CH2),2.22-2.14(m,1H;C2-CH2),1.71(d,J=1.8Hz,1H;C8-CH2),1.67(s,3H;C14-CH3),1.55-1.45(m,1H;C6-CH),1.28(d,J=6.4Hz,3H;C6-CH3),1.28-1.25(m,1H;C8-CH2),0.86(s,3H;C4-CH3); 13C NMR (101MHz, CDCl3) δ197.99(C28), 168.09(C1), 168.07(C23), 158.27(C25), 156.09(C20), 152.36(C22), 142.46(C18), 140.38(C19), 139. 83(C14), 138.84(C27), 132.85(C12), 130.48(C31), 128.22(C11), 124 .95(C13), 122.41(C30), 122.12(C21), 119.67(C32), 119.25(C16), 11 3.73(C26), 113.02(C17), 88.18(C10), 81.16(C9), 78.13(C3), 74.18( C7), 66.37(C24), 66.26(C5), 60.26(C4), 56.87(C20-OCH3), 56.68(C1 0-OCH3), 47.05(C15), 38.41(C6), 36.34(C8), 35.31(C1-NCH3), 32.67(C2), 26.88(C29), 15.86(C14-CH3), 14.58(C6-CH3), 12.50(C4-CH3).

[0419] Table 1: Maytansinoids synthesized using Method A or B

[0420]

[0421]

[0422]

[0423]

[0424] Example 3

[0425] Keto-maytansinoids can be synthesized in three steps via esterification with Fmoc-protected amino acids, cleavage of the Fmoc group and condensation with keto acids.

[0426] General Method C - Step 1: Reaction of maytansinol with Fmoc-protected amino acids. , 325 mesh particle size, Sigma Aldrich), under N2 atmosphere, maytansinol (565mg, 1.00mmol, 1.0 equivalent), amino acid (3.00mmol, 3.0 equivalents) protected by Fmoc, DMAP (982mg, 8.00mmol, 8.0 equivalents) and scandium (III) trifluoromethanesulfonate (541mg, 1.1mmol, 1.1 equivalents) are dissolved in anhydrous DCM (10mL). The mixture is cooled to 4 DEG C using ice / water bath, and stirred for 30 minutes to reach the temperature. After this time, DIC (2.47mL, 16.0mmol, 16.0 equivalents) is added in 10 minutes (about 0.25mL / min), and the reaction mixture is stirred for 2 hours at 4 DEG C, and it is gradually brought to room temperature during this period. The mixture is filtered by gravity. The filtrate was diluted with DCM (50 mL) and then washed with sodium phosphate buffer (50 mL x 3, pH 7.5) and brine (50 mL). The organic layer was then dried over anhydrous sodium sulfate, filtered by gravity, and the solvent was removed using a rotary evaporator at 40°C to a final volume of approximately 10 mL. The crude material was concentrated on a pre-packed SNAP ULTRA 50 g cartridge and HP-Sphere TM Purification was performed on a Biotage Isolera One flash purification system on spherical silica (linear gradient from 100% DCM to 90 / 10 DCM / methanol in 25 CV). The solvent was removed on a rotary evaporator at 40°C for 2 hours to afford the corresponding product as a pale yellow to yellow solid.

[0427] General Procedure C - Step 2: Deprotection of the Fmoc group. The Fmoc protected intermediate (0.53 mmol, 1.0 eq) was dissolved in DCM (5 mL) and to this solution was added tris-(2-aminoethyl)amine (0.320 mL, 2.13 mmol, 4.0 eq) within 10 seconds. The reaction mixture was stirred at room temperature for 1 hour. The white precipitate formed was filtered off through a pad of celite, washed with DCM (20 mL) and the yellow filtrate was evaporated to dryness on a rotary evaporator at 40 °C for 2 hours and further dried under high vacuum for 4 hours to give the free amine which was used directly in the next step without further purification.

[0428] General Procedure C - Step 3: React the amine-maytansinoid with the keto acid. The free amine intermediate (77.0 μmol, 1.0 eq), keto acid (150 μmol, 2.0 eq), HATU (35 mg, 94.0 μmol, 1.2 eq), HOAt (13 mg, 94.0 μmol, 1.2 eq) and N-methylmorpholine (17 μL, 150 μmol, 2.0 eq) were dissolved in anhydrous DMF (1 mL) under N2 atmosphere. The mixture was stirred at room temperature overnight. The reaction mixture was diluted with DCM (5 mL) and washed with saturated ammonium chloride solution (5 mL×5) and brine (5 mL). The organic phase was then dried over anhydrous sodium sulfate, filtered by gravity, and the solvent was removed on a rotary evaporator at 40°C. The crude material was concentrated on a SNAP ULTRAC-18 12 g cartridge equipped with a pre-packed column and HP-Sphere TM Purification was performed on a Biotage Isolera One fast purification system with C18 25 μm spherical silica (linear gradient from 80 / 20 water / MeCN to 100% MeCN in 20 CV). Product-containing fractions were pooled, frozen in liquid nitrogen, and lyophilized for 24 hours to afford the corresponding ketomaytansinoid.

[0429] Table 2: Maytansinoids synthesized using Method C (Steps 1 to 3)

[0430]

[0431]

[0432] Example 4

[0433] Preparation of Maytansinoid 28

[0434]

[0435] Maytansinol (56.5 mg, 0.10 mmol, 1.0 eq) was dissolved in anhydrous DCM (10 mL), and a solution of zinc (II) chloride in ether (0.3 mL, 0.30 mmol, 3.0 eq, 1 M solution) was added at room temperature under N2 atmosphere, and stirring was maintained for 10 minutes. 4-Acetylphenyl isocyanate (48.3 mg, 0.30 mmol, 3.0 eq) was added within 10 seconds, and the resulting solution was stirred at room temperature for 5 hours. After this time, the reaction was confirmed to be complete by HPLC analysis (PDA 220 nm). Volatiles were removed with a rotary evaporator at 40 ° C. The crude material was purified by NP chromatography using a SNAP ULTRA 10 g cartridge equipped with a pre-loaded SNAP ULTRA 10 g cartridge and HP-Sphere TMPurification was performed by Biotage Isolera One flash purification system using spherical silica (linear gradient from 100% DCM to 90 / 10 DCM / methanol in 13 CV) followed by RP chromatography using prepacked SNAP ULTRA C-18 12 g cartridges and HP-Sphere TM Purification was performed on C18 25 μm spherical silica (linear gradient from 80 / 20 water / MeCN to 100% MeCN in 30 CV). The fractions containing the product were combined, frozen in liquid nitrogen, and lyophilized for 24 hours to give compound 28 as a white solid. Yield: 20 mg (28%). Purity by RP-HPLC (220 nm) > 95%. C 37 H 45 ClN3O 10 [M+H] + LRMS-ESI (m / z) calculated value: 726.27. Found value: 726.25. C 37 H 43 ClN3O 10 [MH] - LRMS-ESI (m / z) calculated: 724.27. Found: 724.43.

[0436] Example 5

[0437] Preparation of Maytansinoid 29

[0438] Synthesis of May-ONp: Pyridine (25 μL, 310 μmol, 2.0 equiv) was added to a solution of maytansinol (88 mg, 155 μmol, 1.0 equiv) in DCM (8 mL) at room temperature. The resulting clear solution was stirred for 15 minutes, then cooled to 4 ° C, and p-nitrophenyl chloroformate (219 mg, 1.08 mmol, 7.0 equiv) was added to DCM (4 mL), after which a white precipitate was formed immediately. The reaction mixture was stirred at room temperature for 24 hours. The crude material was concentrated using a rotary evaporator at 40 ° C for 1 hour and purified by the Biotage Isolera One rapid purification system using a pre-packed SNAP ULTRA 25g cartridge and HP-Sphere TM Purification was performed on spherical silica (linear gradient from 100% ethyl acetate to 40 / 60 ethyl acetate / DCM in 50CV). The product-containing fractions were dried on a rotary evaporator at 40°C for 30 minutes and dried under high vacuum for another 30 minutes to afford the intermediate May-ONp as a white solid. Yield: 102 mg (90%). 35 H 41 ClN3O12 [M+H] + LRMS-ESI (m / z) calculated value: 730.23. Found value: 730.01.

[0439]

[0440] Synthesis of Maytansinoid 29: A solution of compound May-ONp (3.7 mg, 5.00 μmol, 1.0 equiv) in DCM (1 mL) was added at room temperature to a solution of 4-(aminomethyl)acetophenone (1.1 mg, 7.50 μmol, 1.5 equiv) in DCM / DMF (2:0.1 v / v) and stirred for 5 minutes, followed by the addition of triethylamine (2 μL, 10.0 μmol, 2.0 equiv). The reaction mixture was stirred at room temperature for 2 hours and at 60°C overnight. The crude material was concentrated at 40°C for 1 hour and purified by the Biotage Isolera One rapid purification system using a pre-packed SNAP ULTRA 10 g cartridge and HP-Sphere TM Purification was performed on spherical silica (linear gradient from 100% chloroform to 90 / 10 chloroform / methanol in 20CV). The product-containing fractions were dried on a rotary evaporator at 40°C for 30 minutes and dried under high vacuum for another 30 minutes to give compound 29 as a white solid. Yield: 1 mg (27%). Purity by RP-HPLC (220 nm) > 95%. C 38 H 46 ClN3O 10 [M+H] + LRMS-ESI (m / z) calculated value: 739.28. Found value: 739.96.

[0441] Example 6

[0442] Preparation of albumin-bound maytansinoids

[0443] General Method D for the synthesis of albumin-bound maytansinoids from keto-maytansinoids and maleimide-hydrazide linkers: To a stirred solution of keto-maytansinoid (1.0 equiv) in anhydrous solvent (suitable solvents are DCM, DMSO, dioxane, 2-methyltetrahydrofuran) at room temperature under N2 atmosphere, molecular sieves (1:1 to 5:1 w / w) and catalyst (TFA, p-toluenesulfonic acid, amberlyst-H form, amberlite-Na form) were added, followed by a solution of hydrazide linker (1.0 to 5.0 equiv) in anhydrous DMSO. The reaction mixture was stirred at room temperature and the conversion was confirmed using HPLC analysis (PDA 220 nm) (>90% conversion). The reaction mixture was filtered, concentrated on a rotary evaporator at 30°C, and purified by Biotage Isolera One Rapid Purification System using a pre-packed SNAP ULTRA HP-Sphere TM The free acid is obtained by chromatography on spherical silica (linear gradient from 100% DCM to 90 / 10 DCM / methanol), drying on a rotary evaporator at 30°C for 30 minutes, and then drying under high vacuum for another 30 minutes. The product is redissolved in a suitable solvent (methanol, acetone, 2-methyltetrahydrofuran or other organic polar solvents) and then neutralized with a salt solution (sodium salt, potassium salt, triethylammonium salt) until the pH range is 5.5 to 7.5. The solution is frozen in liquid nitrogen and lyophilized for 24 hours to obtain the compound as a salt. The content of counterions is determined by IC. The sodium ion content ranges from 0.2 to 1.0 equivalents (about 0.2% to 0.9%), while the triethylammonium ion content ranges from 0.8 to 2.0 equivalents (about 6% to 16%).

[0444] Preparation of albumin-bound maytansinoid 30

[0445]

[0446] Reaction of maytansinoid 2 with linker 1: Maytansinoid 2 (170 mg, 0.23 mmol, 1.0 eq), molecular sieves (0.2 g, powder, activated, 325 mesh particle size) and To a solution of 1,4-dimethoxy-1,4-dextrin (20 mg, macropore, 30 to 60 mesh) in anhydrous DCM (2 mL) was added a solution of linker 1 (51 mg, 0.12 mmol, 2.0 equiv) in anhydrous DMSO (0.6 mL). The reaction mixture was stirred at room temperature, and after 3 hours, HPLC analysis (PDA 220 nm) confirmed the completion of the reaction (>98% conversion). The reaction mixture was concentrated on a rotary evaporator at 30° C. and filtered through a 0.45 μm syringe filter (Macherey-Nagel, The filtrate was diluted with anhydrous DCM (8 mL) and purified by Biotage Isolera One rapid purification system using a pre-packed SNAP ULTRA 10 g cartridge and HP-Sphere TM Spherical silica was purified (linear gradient from 100% DCM to 90 / 10 DCM / MeOH in 22CV). The combined fractions containing the product were dried at 30°C on a rotary evaporator for 30 minutes and dried under high vacuum for another 30 minutes to obtain the free acid form 30 as a white-yellow solid. The solid was dissolved in MeOH / acetone (50:50 v / v, about 4 mL in total), which was then neutralized with a 5 mM solution of NaHCO3 in millipore water until pH 6.8 value 7.1 (pH measured using a pH meter). The solution was frozen in liquid nitrogen and lyophilized for 24 hours to provide maytansinoid prodrug 30 as a yellow-white foam. Yield: 161 mg (60%). 55 H 61 ClFN6O 16 S[MH] - LRMS-ESI (m / z) calculated value: 1147.36. Found value: 1147.84. Na + The content range is between 0.2 and 0.8%.

[0447] Preparation of albumin-bound maytansinoid 31

[0448]

[0449] From the reaction of maytansinoid 3 with linker 1, it was found that maytansinoid 3 (186 mg, 0.25 mmol, 1.0 equiv), molecular sieves (0.4 g, powder, activated, 325 mesh particle size, SigmaAldrich) and To a stirred solution of (IR120 Na form, 484 mg, 2.1 mmol / mL, 4.0 equiv) in anhydrous dichloromethane (4 mL) was added a solution of linker 1 (329 mg, 0.77 mmol, 2.5 equiv) in anhydrous DMSO (4 mL). The reaction mixture was stirred at room temperature and after 6 hours, HPLC analysis (PDA 220 nm) confirmed the completion of the reaction (>98% conversion). The reaction mixture was concentrated on a rotary evaporator at 30°C and filtered through a 0.45 μm syringe filter (Macherey-Nagel, PTFE-O-45 / 25). The filtrate was neutralized with sodium hydroxide (318 μL of 1 M NaOH solution, 1.3 equivalents). The neutralized mixture was added dropwise to a 50 mL falcon tube containing a cooled mixture of methyl tert-butyl ether (27 mL) and isopropanol (14 mL) (4°C ice bath) and centrifuged at 4°C for 5 minutes. The supernatant was decanted and the pellet was resuspended in dichloromethane (10 mL) and purified by Biotage Isolera One Rapid Purification System using a pre-packed SNAP ULTRA 10g cartridge and HP-Sphere TM Purification was performed on spherical silica (linear gradient from 100% DCM to 90 / 10 DCM / MeOH in 22 CV). The combined product-containing fractions were dried on a rotary evaporator at 30°C for 30 minutes and dried under high vacuum for an additional 10 hours to afford 31 as a white-yellow solid. Yield: 156 mg (52%). Purity by RP-HPLC (220 nm) > 95%. C 55 H 62 ClN6O 17 S[MH] - LRMS-ESI (m / z) calculated value: 1145.36. Found value: 1145.87. The content of Na has a window of 0.2 to 0.8%.

[0450] Table 3: Albumin-bound maytansinoids synthesized using General Method D

[0451]

[0452]

[0453]

[0454] Examples 7

[0455] Stability and release kinetics of maytansinoid-HSA conjugates in pH 4.0 and pH 7.4 buffer solutions

[0456] To prepare albumin-bound maytansinoid HSA conjugates, HSA (200 μM: 361.8 μL, 1078 μM free Cys34; 100 μM: 180.9 μL, 1078 μM free Cys34) was diluted with PBS buffer (4 mM sodium phosphate, 150 mM NaCl, pH 7.4) (200 μM: 678.2 μL; 100 μM: 859.1 μL) and DMSO (200 μM: 130.0 μL; 100 μM: 195.0 μL) and incubated in a heating block at 37°C for 30 min. Albumin-bound maytansinoids were added to the pre-incubated HSA samples as 2 mM stock solutions in DMSO (200 μM: 130.0 μL; 100 μM: 65.0 μL) to produce 200 μM or 100 μM maytansinoid prodrug solutions and 300 μM or 150 μM free Cys 34. The mixture at pH 7.4 was reacted at 37°C for 10 minutes and then analyzed by RP-HPLC (Phenomenex Aeris WP XB-C18, 3.6 μm, 250×4.6 mm) every hour for 24 hours.

[0457] To study the release kinetics at pH 4.0, the mixture was acidified with 50 mM sodium acetate buffer pH 3.0 (200 μM: 119.3 μL; 100 μM: 125.2 μL) and 1 M HCl (200 μM: 12.7 μL; 100 μM: 6.8 μL) to reach pH 4.0. The mixture was then analyzed once an hour for 24 hours by RP-HPLC (Phenomenex Aeris WP XB-C18, 3.6 μm, 250×4.6 mm).

[0458] The following RP-HPLC conditions were used: Phenomenex Aeris WP XB-C18 3.6 μm, 250×4.6 mm; eluent A (100% 20 mM Tris buffer pH 8.0) and eluent B (90:10, MeCN: water), with gradient elution of eluent B (25% 0 to 0.5 min, 25 to 35% 0.5 to 2.5 min, 35 to 85% 2.5 to 16 min, 85 to 95% 16 to 17 min, 95% 17 to 20 min, 95 to 25% 20 to 25 min, 25% 25 to 30 min, flow rate 1.0 mL / min). Column oven temperature 37° C.; autosampler temperature 37° C.; injection volume 20 μL.

[0459] To quantify the percentage of free drug released, a standard curve of free maytansinoid was prepared at different concentrations (200 μM, 100 μM, 50 μM, 25 μM and 12.5 μM). The area under the curve (AUC) was determined at 250 nm.

[0460] Table 4: Release kinetics of maytansinoid-HSA conjugates at pH 4.0 and pH 7.4

[0461]

[0462]

[0463] *Measured value is 200μM

[0464] **Measured at 100 μM

[0465] Example 8

[0466] Stability of maytansinoids and maytansinoid-HSA conjugates in CD1 mouse and human plasma: To investigate the stability of maytansinoids and maytansinoid-HSA conjugates in CD1 and human plasma, the compounds were incubated for 24 hours at 37° C. The remaining free maytansinoid or the release of the corresponding maytansinoid was quantified by LC-MS / MS (or HPLC) at certain time points.

[0467] LC-MS quantitative procedure: The combined CD1 mouse or human plasma (K2EDTA, innovative research) was centrifuged (1 minute, 12,044g). The supernatant was first filtered through a filter needle (5μm, sterile, Becton Dickinson) and then through a cellulose acetate membrane (0.45μm, sterile, Carl Roth). The filtered plasma (1710μL) was incubated at 37°C in a heating block for 30 minutes. Free maytansinoids or albumin-bound maytansinoids were added to the pre-incubated plasma samples in the form of a 300μM stock solution of DMSO (190μL) to obtain a 30μM solution of the corresponding maytansinoid or maytansinoid-HSA conjugate. The mixture was reacted at 37°C for 10 minutes, and then three samples (70 μL) were placed in a 96-well plate at each time point (0, 1, 2, 3, 4, 5, 21, and 24 hours), sealed with a plastic mat, immediately frozen with liquid nitrogen and stored at -20°C. At the end of the experiment, the 96-well plate was thawed at room temperature. The samples (30 μL) were then transferred directly to the 96-well Impact plate using a multichannel pipette. TM Protein precipitation plate ( ), washed once with MeCN (150 μL), and then loaded with internal standard (120 μL, 5 μg / mL maytansine in MeCN). TM The precipitation plate was sealed with a silica gel pad and shaken at 420 rpm for 2 minutes. TM The precipitation plate is placed on a 96-well sample manifold and the filtrate is collected on another 96-well plate by applying a moderate vacuum. The 96-well plate is sealed with a silicone pad and kept at room temperature until LC-MS / MS quantification. The filtrate is injected into the LC-MS to be quantified using the MRM mode.

[0468] LC-MS Method: Phenomenex Omega Polar C18, 1.6 μm, , 50×2.1 mm, column; eluent A (0.1% formic acid in water) and eluent B (0.1% formic acid in MeCN), gradient elution with eluent B (20% 0 to 0.5 min, flow rate 0.4 mL / min; 20 to 60% 0.5 to 9.0 min, 0.4 mL / min; 60 to 100% 9.0 to 9.5 min, flow rate 0.4 mL / min; 100 to 20% 9.5 to 10.5 min, flow rate 0.4 mL / min; 20% 10.5 to 12 min, 0.6 mL / min). Column box temperature 25°C; autosampler temperature room temperature; 10 μL injection volume.

[0469] The area under the curve (AUC) of each free drug at time point 0 was set to 100% of the value of the corresponding prodrug. The AUC of each sample was normalized based on the AUC of maytansine.

[0470] HPLC quantitative procedure: Samples were prepared as described above before using 2 mM stock solutions in DMSO of free drug and prodrug. After incubation at 37 ° C, samples were injected directly into HPLC every hour for 24 hours. The area under the curve (AUC) of each free maytansinoid (200 μM in 10% DMSO in PBS buffer) was used as the 100% value of the corresponding albumin-bound maytansinoid.

[0471] HPLC method: Phenomenex Aeris WP XB-C18, 3.6 μm, 250×4.6 mm, column; eluent A (20 mM Tris buffer pH 8.0) and eluent B (90:10, MeCN: water), gradient elution with eluent B (25% 0 to 0.5 min, 25 to 35% 0.5 to 2.5 min, 35 to 85% 2.5 to 16 min, 85 to 95% 16 to 17 min, 95% 17 to 20 min, 95 to 25% 20 to 25 min, 25% 25 to 30 min, flow rate = 1.0 mL / min). Column box temperature 37° C.; autosampler temperature 37° C.; injection volume 20 μL.

[0472] Table 5 below lists the relative release of free maytansinoids and the conversion rate to maytansinol.

[0473] Table 5: Relative release of free maytansinoid and / or maytansinol in CD1 mouse and human plasma for maytansinoid-HSA conjugates

[0474]

[0475]

[0476] *Data obtained by HPLC

[0477] Table 6 below lists the amount of free maytansinoid remaining and the conversion rate to maytansinol.

[0478] Table 6: Remaining amounts of free maytansinoids and maytansinol in CD1 mouse and human serum albumin after 24 hours

[0479]

[0480] Figure 1 The stability of different linkers (maytansinoid 4) in CD1 mouse plasma is depicted in FIG.

[0481] Example 9

[0482] In vitro binding kinetics of albumin-bound maytansinoids in pooled human whole blood and plasma: To study the binding kinetics of albumin-bound maytansinoids in pooled human plasma and pooled human whole blood, compounds were incubated with pooled human plasma at 37°C and samples were taken at designated time points. The remaining albumin-bound maytansinoids were quantified by HPLC.

[0483] HPLC quantification procedure: To study binding kinetics in human whole blood, blood (K2EDTA, 36 donors, Zen-Bio; 900 μL) was pre-incubated in a heating block at 37° C. for 30 minutes. To study binding kinetics in pooled human plasma, pooled whole blood was centrifuged (10 minutes, 1811 g), supernatant plasma was collected, and then pre-incubated at 37° C. for 30 minutes.

[0484] Albumin-bound maytansinoids were added to pre-incubated whole blood / plasma samples in a 10-fold dilution of 1.2 mM stock solution and 1.48 mM citrate (100 μL) in 2.5% propylene glycol in 10 mM sodium phosphate buffer, producing a 12.0 μM solution of corresponding albumin-bound maytansinoids. Sampling (190 μL) was performed after 15 seconds, 2 minutes, 4 minutes, 8 minutes and 15 minutes. Samples were added directly to 760 μL MeCN containing maytansine (5 μg / mL) as internal standards, and vortexed for 1 minute. After centrifugation (12,044 g for 1 minute), supernatant (850 μL) was concentrated under high vacuum. Residue was dissolved in DMSO / water (1: 1 v / v; 95 μL), and then analyzed by RP-HPLC. Percent binding was determined by comparing the area under the curve (AUC) at 220 nm for albumin-bound maytansinoids relative to a control sample in PBS buffer (100% value).All experiments were performed in triplicate.

[0485] HPLC method: Phenomenex Kinetex Polar C18, 2.6 μm, , 150×4.6 mm; eluent A (95:5 5 mM ammonium acetate: MeCN) and eluent B (5:95 5 mM ammonium acetate: MeCN), gradient elution with eluent B (30% 0 to 0.5 min, 30 to 95% 0.5 to 9.0 min, 95% 11.0 min, 95 to 30% 11.0 to 14.5 min, 30% 15.0 min, flow rate = 1.0 mL / min). Column box temperature ambient temperature; autosampler temperature 4°C; 50 μL injection volume.

[0486] The following table lists the relative amounts of albumin-bound maytansinoids at 0 and 8 minutes:

[0487] Table 7: Amount of Bound Albumin-Bound Maytansinoids in Pooled Human Whole Blood and Plasma after 15 Seconds and 8 Minutes

[0488]

[0489] Example 10

[0490] use Cell viability assay, in vitro cytotoxicity of maytansine, DM1, DM1-SMe, maytansinol and maytansinoids in different cell lines

[0491] This study used Promega Cell viability assay was used to test the in vitro efficacy of all compounds. The cancer cell lines tested were: LXFL 529 (large cell lung cancer), RKO (colon cancer), SW-620 (colon cancer), CAL-27 (head and neck cancer), LXFL 1674L (large cell lung cancer), MDA-MB 468 (breast cancer), SK-OV-3 (ovarian cancer), PAXF1657 (pancreatic cancer), MCF7 (breast cancer), COLO 205 (colon cancer), MDA-MB 231 (breast cancer), BT-474 (breast cancer) and Hep G2 (liver cancer).

[0492] Cells were harvested from exponential phase cultures, counted and plated in 96-well flat-bottom microtiter plates at cell densities depending on the cell line growth rate (4,000 and 60,000 cells). After a 24-hour recovery period, cells were allowed to resume exponential growth before the addition of 10 μL of culture medium (four control wells / plate) or culture medium containing test compounds.

[0493] Compounds were administered in duplicate at 10 concentrations in half-log dilution steps, and cells were treated for 96 hours. After treatment and incubation of cells, 20 μL / well Reagents. After incubation for up to 4 hours, fluorescence (FU) was measured using an EnSpire multilabel reader (excitation λ = 531 nm, emission λ = 615 nm). Sigmoidal concentration response curves were fit to the data points (T / C values) obtained for each cell line using a 4-parameter nonlinear curve fit (Oncotest Warehouse Software). IC 50 Values ​​are reported as relative IC 50 Values, which are the concentrations of test compound that produce a response (inhibition of colony formation / viability) between the top plateau and the bottom plateau of the sigmoidal concentration-response curve (inflection point of the curve), or reported as absolute IC 50 values, which are the concentrations of the test compound at the intersection of the concentration-response curves where T / C = 50%. To calculate the average IC 50 The results are presented as mean plots or heat maps of all cell lines tested (relative to the geometric mean IC 50 Value of a single IC 50 See Table 8 and Figure 2 . Figure 2 The IC values ​​of all tested compounds are shown. 50 Heat map.

[0494] Table 8

[0495]

[0496]

[0497] Example 11

[0498] General Procedure for Evaluating Maytansine and Albumin-Binding Maytansine Albumin-Binding Compounds in Patient-Derived Tumor Xenograft Models

[0499] Female immunodeficient NMRI nude mice from Charles River received unilateral tumor implantation with human cancer tumors subcutaneously in the left flank under isoflurane anesthesia until the cancer tumor became palpable and reached a size of 80 to 200 mm. 3 until (unless otherwise stated).

[0500] The animals were housed in cages maintained at a temperature of 25 ± 1 ° C, a relative humidity of 45 to 65%, and a ventilation rate of 60 times per hour. The animals were placed on a 14-hour light: 10-hour dark artificial light cycle. The animals were fed with autoclaved Teklad Global 19% protein extruded feed (T.2019S.12) from Envigo RMSSARL and had access to sterile filtered and acidified (pH 2.5) tap water, which was changed twice a week. Feed and water were provided ad libitum. Prior to treatment, animals were randomized into groups (7 mice per group, unless otherwise stated), considering comparable median and mean values ​​of group tumor volume. Animals were routinely monitored twice a day on weekdays and once a day on Saturdays and Sundays. Starting from day 0, animals were weighed twice a week. The relative body weight (RBW) of individual animals was calculated by dividing the individual absolute body weight (BW) on day X by the mean value of the individual body weight on day X. x ) divided by the individual body weight on the day of randomization multiplied by 100%. On the day of randomization (day 0) and then twice a week, tumor volume was determined by two-dimensional measurement with calipers. Tumor volume was calculated according to the following equation: Tumor volume [mm 3 ]=1[mm]×w 2 [mm 2 ]×0.5, where "1" is the length of the tumor and "w" is the width of the tumor. The absolute individual tumor volume [mm 3 The relative volume of a single tumor on day X (RTVx) was calculated by dividing the absolute single tumor volume of the same tumor on the day of randomization by 100%. The schedule was applied within the limits allowed by the animal welfare policy. 3Individual mice were terminated at tumor volumes (unilateral). All test compounds were administered on days 1, 8, 15, and 22 and were provided as lyophilized solids containing sucrose. On the day of treatment, the lyophilized samples were dissolved in 10 mM sodium phosphate buffer, pH 6.8, containing 20% ​​propylene glycol, and injected intravenously (100 μL / 20-g mouse) together with vehicle (10 mM sodium phosphate buffer, 20% propylene glycol, and 5% sucrose-pH 6.8).

[0501] Example 12

[0502] General Procedure for Evaluating Maytansine and Albumin-Binding Maytansine Albumin-Binding Compounds in Cancer Cell Line-Derived Xenograft Models

[0503] Female immunodeficient NMRI nude mice from Janvier, France, received 5×10 6 Cultured cancer cells (unless otherwise stated) until tumors were palpable and reached a size of 80 to 200 mm 3 (unless otherwise specified). The animals were housed in cages maintained at 22 ± 1°C, 50 ± 10% relative humidity, and 60 ventilation times per hour. The animals were placed in a 12 h light: 12 h dark artificial light cycle. Animals were fed autoclaved SsniffNM and had access to sterile filtered and acidified (pH 4.0) tap water, which was changed twice a week. Feed and water were provided ad libitum. Before treatment, animals were randomized into groups (7 mice per group, unless otherwise stated), taking into account comparable median and mean values ​​of group tumor volume. Animals were routinely monitored twice a day on weekdays and once a day on Saturdays and Sundays. Starting from day 0, the body weight of mice was determined twice a week, and the average body weight of each group was related to the initial value (in percentage) to calculate body weight change (BWC). On the day of randomization (day 0), and then twice or three times a week, two-dimensional measurements were performed with calipers to determine tumor volume. Tumor volume was calculated according to the following equation:

[0504] Tumor volume [mm 3 ]=1[mm]×w 2 [mm 2 ]×0.5, where "1" is the length of the tumor and "w" is the width of the tumor. The absolute individual tumor volume [mm 3 The relative volume of a single tumor on day X (RTV) was calculated by dividing the absolute single tumor volume of the same tumor on the day of randomization and multiplying by 100%. x ). Apply schedule within the limits allowed by animal welfare policy. 3Individual mice were terminated when the tumor volume (unilateral) was less than 10%, or ulcers were visible. All test compounds were administered on days 1, 8, 15, and 22 and were provided as lyophilized solids containing sucrose. On the day of treatment, the lyophilized samples were dissolved in 10 mM sodium phosphate buffer, pH 6.8, containing 20% ​​propylene glycol, and injected intravenously (100 μL / 20-g mouse) with vehicle (10 mM sodium phosphate buffer, 20% propylene glycol, and 5% sucrose-pH 6.8).

[0505] Example 13

[0506] Evaluation of maytansine and albumin-binding maytansine alkaloids 30, 42, 31, and 35 in the human PDX renal cell carcinoma model RXF 631

[0507] Maytansine and albumin-binding compounds 30, 42, 31, and 35 were evaluated in the renal cancer cell RXF 631 model following the general procedure for patient-derived xenograft models. 3 of average size after starting treatment. Figure 3 Tumor growth curves of the control group, the maytansine group, and the groups treated with compounds 30, 42, 31, and 35 are shown. Figure 4 The body weight change curves of the control group, the maytansine group, and the groups treated with compounds 30, 42, 31, and 35 are shown.

[0508] Example 14

[0509] Evaluation of maytansine and albumin-binding maytansine alkaloids 32, 30, and 31 in the human PDX squamous cell lung cancer model LXFE 937

[0510] Maytansine and albumin-binding compounds 32, 30, and 31 were evaluated in the squamous cell lung cancer LXFE 937 model following the general procedure for patient-derived xenograft models. 3 of average size after starting treatment. Figure 5 Tumor growth curves of the control group, the maytansine group, and the groups treated with compounds 32, 30, and 31 are shown. Figure 6 The body weight change curves of the control group, the maytansine group, and the groups treated with compounds 32, 30, and 31 are shown.

[0511] Example 15

[0512] Evaluation of maytansine and albumin-binding maytansine alkaloids 30 and 31 in the human PDX squamous cell lung cancer model LXFE 937 (large tumors)

[0513] Maytansine and albumin-binding compounds 30 and 31 were evaluated in the squamous cell lung cancer LXFE 937 model following the general procedure for patient-derived xenograft models. 3 of average size after starting treatment. Figure 7 Tumor growth curves of the control group, the maytansine group, and the groups treated with compounds 30 and 31 are shown. Figure 8 The body weight change curves of the control group, the maytansine group, and the groups treated with compounds 30 and 31 are shown.

[0514] Example 16

[0515] Evaluation of maytansine and albumin-binding maytansine alkaloids 30 and 31 in the human PDX lung adenocarcinoma model LXFA 737

[0516] Maytansine and albumin-binding compounds 30 and 31 were evaluated in the LXFA 737 model of lung adenocarcinoma following the general procedure for patient-derived xenograft models. 3 of average size after starting treatment. Fig. 9 Tumor growth curves of the control group, the maytansine group, and the groups treated with compounds 30 and 31 are shown. Fig.10 The body weight change curves of the control group, the maytansine group, and the groups treated with compounds 30 and 31 are shown.

[0517] Example 17

[0518] Evaluation of maytansine and albumin-binding maytansine alkaloids 32, 30, and 31 in the human xenograft breast cancer model MDA-MB-231

[0519] Maytansine and albumin-binding compounds 32, 30, and 31 were evaluated in the MDA-MB 231 breast cancer model following the general procedure for cancer cell line-derived xenograft models. 3 of average size after starting treatment. Fig.11 Tumor growth curves of the control group, the maytansine group, and the groups treated with compounds 32, 30, and 31 are shown. Fig.12 The body weight change curves of the control group, the maytansine group, and the groups treated with compounds 32, 30, and 31 are shown.

[0520] Example 18

[0521] Evaluation of maytansine and albumin-binding maytansine alkaloids 30 and 31 in the human xenograft ovarian cancer model A2780

[0522] Maytansine and albumin-binding compounds 30 and 31 were evaluated in the A2780 ovarian cancer model according to the general procedure for ovarian cancer cell line xenograft models. 3 of average size after starting treatment. Fig.13 Tumor growth curves of the control group, the maytansine group, and the groups treated with compounds 30 and 31 are shown. Fig.14 The body weight change curves of the control group, the maytansine group, and the groups treated with compounds 30 and 31 are shown.

[0523] Example 19

[0524] Evaluation of maytansine and albumin-binding maytansine alkaloids 30 and 31 in the human xenograft breast cancer model MDA-MB 468

[0525] Maytansine and albumin-binding compounds 30 and 31 were evaluated in breast cancer MDA-MB 468 according to the general procedure for cancer cell line-derived xenograft models. 3 of average size after starting treatment. Fig.15 Tumor growth curves of the control group, the maytansine group, and the groups treated with compounds 30 and 31 are shown. Fig.16 The body weight change curves of the control group, the maytansine group, and the groups treated with compounds 30 and 31 are shown.

Claims

1. A compound having a structure of formula (II): or a pharmaceutically acceptable salt or hydrate thereof, in: Each R 2 independently -H; X is absent or selected from -CH2-, -O- and -S; Z 1 is selected from -H, -F, -Cl, -Br and -I; Z 2 , Z 3 and Z 4 each independently is -H; R' is Z 1’ Selected from -H, -F, -Cl, -Br, -I, -CF3, -OCH3, -CN, -NO2, -SO3M 2 and C1-C4 alkyl, wherein M 2 is a pharmaceutically acceptable counterion selected from H + 、Na + , K + , Ca 2+ Mg 2+ NR4 + and NHR3 + ; wherein R is H or C1-C4 alkyl; Z 2’ , Z 3’ and Z 4’ each independently is -H; Y' is -NH-C(O)-, where n Y '= 0 to 6; R 1’ Not present or selected from the group consisting of: Among them, M 1 is a pharmaceutically acceptable counterion selected from H + 、Na + , K + , Ca 2+ Mg 2+ NR4 + and NHR3 + ; wherein R is H or C1-C4 alkyl; R 2’ is an optionally substituted C1-C 18 alkyl; TBG is a maleimide group; The TBG is optionally combined with a thiol-bearing macromolecular carrier or a thiol-bearing tumor-specific carrier, wherein the thiol-bearing macromolecular carrier or the thiol-bearing tumor-specific carrier is selected from endogenous albumin, exogenous albumin, antibodies, antibody fragments and peptides.

2. The compound according to claim 1, or a pharmaceutically acceptable salt or hydrate thereof, wherein Z 1 Yes -F.

3. The compound according to claim 1, or a pharmaceutically acceptable salt or hydrate thereof, wherein X is absent.

4. The compound according to claim 1, or a pharmaceutically acceptable salt or hydrate thereof, wherein X is -CH2-.

5. The compound according to claim 1, or a pharmaceutically acceptable salt or hydrate thereof, wherein X is -O- or -S-.

6. The compound according to claim 1, wherein the compound is selected from: or a pharmaceutically acceptable salt or hydrate thereof.

7. The compound according to claim 1, or a pharmaceutically acceptable salt or hydrate thereof, wherein the TBG is not bound to a thiol-bearing macromolecular carrier or a thiol-bearing tumor-specific carrier.

8. The compound according to claim 1, or a pharmaceutically acceptable salt or hydrate thereof, wherein the TBG is bound to a thiol-bearing macromolecular carrier or a thiol-bearing tumor-specific carrier.

9. The compound according to claim 1, or a pharmaceutically acceptable salt or hydrate thereof, wherein Z 1’ Selected from -NO2 or -SO3M 2 ; Y' is selected from -NHC(O)- or and M 2 is a pharmaceutically acceptable counterion selected from H + 、Na + , K + , Ca 2+ Mg 2+ NR4 + and NHR3 + ; wherein R is H or C1-C4 alkyl.

10. The compound according to claim 1, or a pharmaceutically acceptable salt or hydrate thereof, wherein R 1’ yes:

11. The compound according to claim 1, wherein R' is: or a pharmaceutically acceptable salt or hydrate thereof.

12. A compound selected from the group consisting of: or a pharmaceutically acceptable salt or hydrate thereof.

13. The compound according to claim 1, wherein R' is: or a pharmaceutically acceptable salt or hydrate thereof; and Among them, M 1 is a pharmaceutically acceptable counterion selected from H + 、Na + , K + , Ca 2+ Mg 2+ NR4 + and NHR3 + ; wherein R is H or C1-C4 alkyl.

14. The compound according to claim 13, wherein the compound is selected from: or a pharmaceutically acceptable salt or hydrate thereof.

15. A compound according to any one of claims 1 to 4, wherein R' is: or a pharmaceutically acceptable salt or hydrate thereof; and Among them, M 1 is a pharmaceutically acceptable counterion selected from H + 、Na + , K + , Ca 2+ Mg 2+ NR4 + and NHR3 + ; wherein R is H or C1-C4 alkyl.

16. The compound according to claim 15, wherein the compound is: or a pharmaceutically acceptable salt or hydrate thereof.

17. A pharmaceutical composition comprising the compound according to any one of claims 1 to 16 or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.

18. A compound according to any one of claims 1 to 16 or a pharmaceutically acceptable salt thereof for use as a medicament.

19. A compound according to any one of claims 1 to 16 or a pharmaceutically acceptable salt thereof for use in treating cancer.

20. The compound for use according to claim 19, wherein the cancer is selected from adenocarcinoma, uveal melanoma, acoustic neuroma, ampullary carcinoma, anal cancer, astrocytoma, basal cell carcinoma, pancreatic cancer, connective tissue tumors, bladder cancer, bronchial cancer, breast cancer, Burkitt's lymphoma, uterine corpus cancer, CUP syndrome, colon cancer, small intestine cancer, ovarian cancer, endometrial cancer, gallbladder cancer, uterine cancer, cervical cancer, neck, nose, ear tumors, blood tumors, urethral cancer, skin cancer, glioma, testicular cancer, Kaposi's sarcoma, laryngeal cancer, bone cancer, colorectal cancer, head and neck cancer, craniopharyngioma, liver cancer, lung cancer, Hodgkin's lymphoma, non-Hodgkin's lymphoma, gastric cancer, medulloblastoma, melanoma, meningioma, kidney cancer, oligodendroglioma, esophageal cancer, penile cancer, prostate cancer, tongue cancer and lymph node cancer.

21. The compound for use according to claim 19, wherein the cancer is selected from the group consisting of non-small cell bronchogenic carcinoma, cholecystoma carcinoma, hairy cell leukemia, non-small cell lung cancer, renal cell carcinoma, osteolytic carcinoma, osteoblastic carcinoma and osteosarcoma.

22. Use of a compound according to any one of claims 1 to 16 or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition according to claim 17, in the preparation of a medicament for treating cancer.

23. The method according to claim 22, wherein the cancer is selected from the group consisting of adenocarcinoma, uveal melanoma, acoustic neuroma, ampullary carcinoma, anal cancer, astrocytoma, basal cell carcinoma, pancreatic cancer, connective tissue tumors, bladder cancer, bronchial cancer, breast cancer, Burkitt's lymphoma, uterine corpus cancer, CUP syndrome, colon cancer, small intestine cancer, ovarian cancer, endometrial cancer, gallbladder cancer, uterine cancer, cervical cancer, neck, nose, ear tumors, blood tumors, urethral cancer, skin cancer, glioma, testicular cancer, Kaposi's sarcoma, laryngeal cancer, bone cancer, colorectal cancer, head and neck cancer, craniopharyngioma, liver cancer, lung cancer, Hodgkin's lymphoma, non-Hodgkin's lymphoma, gastric cancer, medulloblastoma, melanoma, meningioma, kidney cancer, renal cell carcinoma, oligodendroglioma, esophageal cancer, penile cancer, prostate cancer, tongue cancer and lymph node cancer.

24. The use according to claim 22, wherein the cancer is selected from the group consisting of non-small cell bronchogenic carcinoma, cholecystokinin carcinoma, hairy cell leukemia, non-small cell lung cancer, renal cell carcinoma, osteolytic carcinoma, osteoblastic carcinoma and osteosarcoma.

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