Bioactive conjugates, preparation methods, and use thereof

Abstract

A compound of formula (I) is provided (wherein the formula is a conjugator which is a group capable of binding BA and connecting BA to the remainder of the compound, a cleavable group which comprises at least one site which can be cleaved by a β-glucuronidase enzyme, a spacer which provides the distance between the conjugator and the payload, the payload which is a cytotoxic agent, e.g., MMAE, and BA is selected from a humanized, monoclonal, chimeric, or human antibody or its antigen-binding fragment). JPEG2026521949000480.jpg25165

Description

[Technical Field]

[0001] Cross-reference of related applications This application claims priority to International Application No. PCT / CN2023 / 104052, filed on 29 June 2023, the disclosure of which is incorporated herein by reference in its entirety.

[0002] This specification provides an antibody-drug conjugate platform and an antibody-drug conjugate (ADC) comprising the platform and an antibody, or an antigen-binding fragment thereof, as well as an ADC platform and the use of the ADC.

[0003] Sequence List This application includes a sequence listing submitted electronically in XML format. The XML file is named "01368-0074-00PCT-ST26.xml", was created on June 20, 2024, and has a size of 8,226 bytes. The sequence listing is incorporated herein by reference in its entirety. [Background technology]

[0004] Antibody-drug conjugates (ADCs) contain antibodies against tumor antigens conjugated to a biologically active small molecule, such as a toxin or payload (i.e., a drug). ADCs selectively deliver the payload to cells expressing the tumor antigen. The payload, monomethyl auristatin E (MMAE), is an antimitotic agent that inhibits cell division. ADCs containing MMAE can be highly hydrophobic, leading to aggregation and nonspecific uptake of the ADC, resulting in a high drug-to-antibody ratio (DAR). ADCs containing MMAE can be unstable, leading to deconjugation, premature payload release, an inadequate pharmacokinetic profile, and off-target effects.

[0005] The linker portion of an ADC, which contains a moiety that directly conjugates with the antibody, is a key feature in developing highly active, optimized therapeutic agents at well-tolerated doses. Electrophilic maleimide functional groups have been used to link the linker to the free thiol of the antibody. In vivo, the conjugation product is subjected to slow elimination, thus reversing the conjugation reaction and freeing the maleimide of the ADC, which then migrates to any other available thiol, including those from serum albumin in plasma.

[0006] There is a continuing need for the development of novel linkers for use with MMAEs in ADCs that would resist deconjugation and premature payload release, enabling increased DAR, increased circulatory stability, improved pharmacokinetics, and improved efficacy. This disclosure addresses these needs. [Overview of the Initiative] [Means for solving the problem]

[0007] This specification provides an antibody-drug conjugate platform and antibody-drug conjugates (ADCs). It also provides the use of the ADC platform for preparing ADCs.

[0008] In some embodiments, the ADC compound of formula (I) is used herein: [ka] or a pharmaceutically acceptable salt, tautomer, solvate, or stereoisomer thereof (in the formula, BA is a binder selected from humanized, monoclonal, chimeric, or human antibodies or their antigen-binding fragments; The conjugator is given by equations (II) and (III): [ka] Selected from the group consisting of; U is a bond, heteroarylene, or arylene; V is a bond or -C≡C-(CH2) n -; n is an integer from 0 to 10 (including both ends); W2 is -C(=O)-, -NH-, or -O-; RG is

Chem.

Chem.

Chem.

[0009] In some embodiments, as used herein, the ADC compound of formula (XI): [ka] or a pharmaceutically acceptable salt, tautomer, solvate, or stereoisomer thereof (in the formula, BA is a binder selected from humanized, monoclonal, chimeric, or human antibodies or their antigen-binding fragments; The conjugator is given by formula (II) or (III): [ka] Having; U is a bond, heteroarylene, or arylene; V represents a bond or -C≡C-(CH2) n -and; n is an integer between 0 and 10 (including both ends); W2 is -C(=O)-, -NH-, or -O-; RG is [ka] , -(succinimido-3-yl-N)-, or [ka] and; RS is -NR 1a R 1b and; R 1a and R 1b Each of these independently represents H or substituted or unsubstituted C. 1-4 It is alkyl; RE stands for bond, -O-, -OC(=O)-, -OC(=O)NR 6 -, -NHC(=O)NR 6 -, -OS(=O)2NR 6 -, -NHS(=O)2NR 6 -, or -OC(=O)NHS(=O)2NR 6 -and; R 6 is H or substituted or unsubstituted C 1-4 It is alkyl; W3 is -C(=O)-, -NH-, or -O-; Each of s and t is independently 0, 1, or 2; [ka] This indicates a covalent bond site within the compound. * indicates a connection where the conjugator connects to the BA; Spacers are used for bonding, ** -NH-(CH2CH2O) m -CH2CH2-C(=O)-, ** -C(=O)-(CH2CH2O) m -CH2CH2-NH-, ** -(CH2) m -C(=O)-, ** -CH2-C(=O)-NH-(CH2) m -C(=O)-, ** -(CH2CH2O) m -CH2CH2-C(=O)-, ** -C(=O)-(CH2CH2O) m -CH2CH2-NH-, ** -CH[-(CH2) m -COOH]-C(=O)-, ** -CH2-C(=O)-NH-(CH2) m -C(=O)-NH-(CH2) m -C(=O)-, ** -C(=O)-(CH2) m -C(=O)-, ** -NH-(CH2) m -C(=O)-, or ** -NH-(CH2) m -O- is; m is an integer between 1 and 12 (including both ends); ** indicates a coupling where the spacer connects to the conjugator; The cleavable bodies are given by formulas (IVa'), (IVb'), (IVc'), (Va'), (Vb'), (VIa'), (VIb'), (VIIa'), or (VIIb'): [ka] Having; Su is the sugar portion; Each R 2 These are, independently, hydrogen, halogen, substituted or unsubstituted C 1-4 It is alkyl, -CN, or -NO2; # indicates a connection where the severable part is attached to the branched part; The branched fields are given by equations (XIIa), (XIIb), (XIIc), and (XIId): [ka] Selected from; Each of p and q is independently 1, 2, 3, or 4; The hydrophilic material is -NH-(CH2CH2O) a -(CH2) b CH3, -N(CH3)-((CH2) c C(=O)N(CH3)) a -(CH2) c C(=O)NH2, -C(=O)-(CH2CH2O) a -(CH2) b CH3, -C(=O)-(CH2CH2O) a -(CH2) b NHC(=O)(CH2)3N(CH3)2(CH2)3S(=O)2OH, -NH-(CH2CH2O) a -(CH2) b NHC(=O)(CH2)3N(CH3)2(CH2)3S(=O)2OH, [ka] and; a is an integer between 1 and 18 (including both ends); b is 0, 1, or 2; c is 1, 2, 3, or 4; The payload is a payload residue; x is between 1 and 15 (including both ends). It will be provided.

[0010] In some embodiments, the platform is a conjugator-linker-payload compound of formula (I-I):

Chemical formula

Chemical formula

Chemical formula

[0011] In some embodiments, the platform is a conjugator-linker-payload compound of formula (XV):

Chemical formula

Chemical formula

Chemical formula

[0012] Additional objectives and benefits are partially shown in the following description, partially understood from the description, or learned through practice. These objectives and benefits are realized and achieved by the elements and combinations specifically indicated in the attached claims.

[0013] Please understand that the general explanation above and the detailed explanation below are merely illustrative and descriptive, and do not constitute limitations on the scope of the claims.

[0014] The accompanying drawings incorporated herein, and which constitute part thereof, serve to illustrate embodiments and, together with the specification, illustrate the principles described herein. [Brief explanation of the drawing]

[0015] [Figure 1A] This shows data on ADC cell killing against U937 cells. [Figure 1B] This shows data on ADC cell killing against U937 cells. [Figure 1C] This shows data on ADC cell killing against U937 cells. [Figure 1D] This shows data on ADC cell killing against U937 cells. [Figure 1E] This shows data on ADC cell killing against U937 cells. [Figure 1F] This shows data on ADC cell killing against U937 cells. [Figure 1G] This shows data on ADC cell killing against U937 cells. [Figure 1H] This shows data on ADC cell killing against U937 cells. [Figure 1I] This shows data on ADC cell killing against U937 cells. [Figure 2A] This shows data on ADC cell killing against HL60 cells. [Figure 2B] This shows data on ADC cell killing against HL60 cells. [Figure 2C] This shows data on ADC cell killing against HL60 cells. [Figure 2D] This shows data on ADC cell killing against HL60 cells. [Figure 2E] This shows data on ADC cell killing against HL60 cells. [Figure 2F] This shows data on ADC cell killing against HL60 cells. [Figure 2G] This shows data on ADC cell killing against HL60 cells. [Figure 2H]This shows data on ADC cell killing against HL60 cells. [Figure 2I] This shows data on ADC cell killing against HL60 cells. (Figure 2J) This shows data on ADC cell killing against HL60 cells. [Figure 3A] This shows data on ADC cell killing against TF1 cells. [Figure 3B] This shows data on ADC cell killing against TF1 cells. [Figure 3C] This shows data on ADC cell killing against TF1 cells. [Figure 3D] This shows data on ADC cell killing against TF1 cells. [Figure 3E] This shows data on ADC cell killing against TF1 cells. [Figure 3F] This shows data on ADC cell killing against TF1 cells. [Figure 3G] This shows data on ADC cell killing against TF1 cells. [Figure 3H] This shows data on ADC cell killing against TF1 cells. [Figure 3I] This shows data on ADC cell killing against TF1 cells. [Figure 4A] A and B show data on ADC cell killing against NCI-H358 cells. [Figure 4B] A and B show data on ADC cell killing against NCI-H358 cells. [Figure 5A] A and B show data on ADC cell killing against NCI-H1048 cells. [Figure 5B] A and B show data on ADC cell killing against NCI-H1048 cells. [Figure 6A] A and B show data on ADC cell killing against MDA-MB-453 cells. [Figure 6B] A and B show data on ADC cell killing against MDA-MB-453 cells. [Figure 7]The ADC, payload, and total antibody ("TAB") PK profiles in mice are shown. [Figure 8] This study demonstrates the antitumor efficacy of ADC in the NCI-H1650 xenograft model. [Modes for carrying out the invention]

[0016] This specification provides monomethyl auristatin E (MMAE)-based antibody-drug conjugates (ADCs), as well as covalent linkers and conjugator-linker-payload (platforms) for constructing such ADCs. ADCs may be used to treat diseases or disorders, such as cancer, by providing compositions containing ADCs. The MMAE ADCs of this disclosure may better resist deconjugation and premature payload release, enabling increased DAR, increased circulatory stability, improved pharmacokinetics, and improved efficacy compared to known MMAE ADCs. The conjugate includes a β-glucuronide-based linker containing a site that can be cleaved by an enzyme having β-glucuronidase activity.

[0017] definition In this disclosure, the following terms have the meanings set forth below unless otherwise indicated. Unless otherwise defined, all technical and scientific terms used herein have the same meanings as those commonly understood by those skilled in the art in which this disclosure relates. If there are multiple definitions of a term provided herein, these definitions shall prevail unless otherwise noted.

[0018] Where a trade name is used herein, references to a trade name also refer to the product formulation, generic drugs, and active pharmaceutical ingredients of the trade name product, unless otherwise indicated by the context.

[0019] In this specification, the term “antibody” is used in its broadest sense and particularly includes intact monoclonal antibodies, polyclonal antibodies, monospecific antibodies, multispecific antibodies (e.g., bispecific antibodies), and antibody fragments exhibiting desired biological activity. Intact antibodies primarily have two regions: a variable region and a constant region. The variable region binds to and interacts with the target antigen. The variable region includes a complementation-determining region (CDR) that recognizes and binds to a specific binding site on a particular antigen. The constant region is recognized by the immune system and can interact with it (e.g., Janeway et al., 2001, Immuno. Biology, 5). th See Ed., Garland Publishing, New York. Antibodies can be of any type (e.g., IgG, IgE, IgM, IgD, and IgA), class (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2), or subclass. Antibodies can originate from any suitable species. In some embodiments, antibodies are of human or mouse origin. Antibodies can be, for example, human, humanized, or chimeric.

[0020] The term "humanized" or "humanized antibody" refers to a form of antibody that contains sequences from non-human (e.g., mouse) antibodies and human antibodies. Such antibodies contain minimal sequences derived from non-human immunoglobulins. Typically, humanized antibodies contain substantially all of at least one, and usually two, variable domains, all or substantially all of the hypervariable loops correspond to those of non-human immunoglobulins, and all or substantially all of the FR region are from human immunoglobulin sequences. Humanized antibodies also optionally contain at least a portion of the immunoglobulin constant region (Fc), typically those of human immunoglobulins. If necessary to distinguish humanized antibodies from rodent antibodies, the prefixes "hum," "hu," "Hu," or "h" are added to the antibody clone name. Humanized forms of rodent antibodies typically contain the same CDR sequence as the rodent antibody, but may include certain amino acid substitutions to increase affinity, increase the stability of the humanized antibody, remove post-translational modifications, or for other reasons.

[0021] As used herein, the term "monoclonal antibody" refers to an antibody obtained from a substantially homogeneous population of antibodies; that is, the individual antibodies constituting the population are identical except for naturally occurring mutations that may exist in small amounts. Monoclonal antibodies are highly specific and directed against a single antigenic site. The modifier "monoclonal" should not be interpreted as requiring the production of the antibody by any particular method.

[0022] An "intact antibody" includes an antigen-binding variable region, as well as a light chain constant domain (CL) and a heavy chain constant domain, CH1, CH2, CH3, and CH4 (where appropriate for the antibody class). The constant domains may be native sequence constant domains (e.g., human native sequence constant domains) or amino acid sequence variants thereof.

[0023] An "antibody fragment" includes a portion of an intact antibody that contains its antigen-binding or variable region. Examples of antibody fragments include Fab, Fab', F(ab')2, and Fv fragments, diabodies, triabodies, tetrabodies, linear antibodies, single-chain antibody molecules, scFv, scFv-Fc, multiple-specific antibody fragments formed from antibody fragments, fragments generated by Fab expression libraries, or any of the above epitope-binding fragments that specifically bind to a target antigen (e.g., cancer cell antigen, viral antigen, or microbial antigen).

[0024] An "antigen" is the substance to which an antibody specifically binds.

[0025] The terms "specific binding" and "specifically binding" mean that an antibody or antibody derivative binds to its corresponding target antigen in a highly selective manner and does not bind to a large number of other antigens. Typically, an antibody or antibody derivative binds to at least approximately 1 × 10⁻¹⁶ antigens. -7 M, 10 -8 M, 10 -9 M, 10 -10 M, 10 -11M, or 10 -12 It binds with affinity M and binds to the default antigen with an affinity at least twice as high as the affinity it has for binding to nonspecific antigens other than the default antigen or closely related antigens (e.g., BSA, casein).

[0026] The term “inhibit” or “inhibit” means to reduce by a measurable amount or to prevent overall.

[0027] The term “therapeutic effective dose” refers to the amount of a drug effective in treating a disease or disorder in a mammal. In the case of cancer, a therapeutic effective dose of a drug may reduce the number of cancer cells; reduce tumor size; inhibit (i.e., slow or stop to some extent) cancer cell invasion into peripheral organs; inhibit (i.e., slow or stop to some extent) tumor metastasis; inhibit tumor growth to some extent; and / or alleviate to some extent one or more of the symptoms associated with cancer. To the extent that a drug can inhibit growth and / or kill existing cancer cells, it may be inhibitory and / or cytotoxic. For cancer therapy, effectiveness may be measured, for example, by assessing time to disease progression (TTP) and / or determining the rate of response (RR).

[0028] The term “substantial” or “substantially” refers to the majority of a mixture or sample, i.e., >50% of the population, preferably more than 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% of the population.

[0029] The terms "intracellularly cleaved" and "intracellular cleavage" refer to intracellular metabolic processes or reactions to ligand-drug conjugates (e.g., antibody-drug conjugates (ADCs)) where the covalent bond between the drug site (D) and the ligand unit (e.g., antibody (BA or Ab)), such as the linker, is broken, resulting in the release of the drug from the antibody inside the cell, or another metabolite of the conjugate. Therefore, the cleavage sites of drug-linker-ligand conjugates are intracellular metabolites.

[0030] The term "cytotoxic activity" refers to the cytotoxic, cell proliferation-inhibiting, or antiproliferative effect of a drug-linker-ligand conjugate compound, or the intracellular metabolites of a drug-linker-ligand conjugate. Cytotoxic activity can be expressed as the IC50 value, which is the concentration (molar concentration or mass) per unit volume at which half of the cells survive.

[0031] The term “cytotoxic agent,” as used herein, refers to a substance that inhibits cellular function and / or causes cellular destruction. The term is intended to include radioisotopes (e.g., radioisotopes of 211At, 131I, 125I, 90Y, 186Re, 188Re, 153Sm, 212Bi, 32P, 60C, and Lu), chemotherapeutic agents, and toxins such as small molecule toxins or enzymatically active toxins of bacterial, fungal, plant, or animal origin (including synthetic analogs and their derivatives).

[0032] The terms “cancer” and “malignant” refer to or represent a physiological condition or disorder in mammals typically characterized by uncontrolled cell growth. “Tumor” includes one or more cancer cells.

[0033] In this specification, “autoimmune disease” refers to a disease or disorder that originates from and is directed against the tissues or proteins of an individual.

[0034] Examples of “patient” or “subject” include, but are not limited to, mammals such as humans, rats, mice, guinea pigs, monkeys, pigs, goats, cattle, horses, dogs, or cats, and birds or poultry. In one embodiment, the patient is a human.

[0035] The term “to treat” or “treatment,” unless otherwise indicated by context, refers to therapeutic treatment and preventive measures to prevent relapse, the purpose of which is to inhibit or slow (weaken) undesirable physiological changes or impairments, such as the progression or development of cancer. For the purposes of this disclosure, beneficial or desired clinical outcomes include, but are not limited to, symptom relief, reduction of disease severity, stabilization (i.e., non-worsening) of the disease, delay or slowing of disease progression, improvement or mitigation of the disease state, and remission (whether partial or total), whether detectable or undetectable. “Treatment” may also mean extending survival compared to the survival expected without treatment. Persons requiring treatment include those who already have a condition or impairment and those who are prone to developing a condition or impairment.

[0036] In relation to cancer, the term “to treat” includes any or all of the following: inhibiting tumor cells, cancer cells, or tumor growth; inhibiting the replication of tumor cells or cancer cells; reducing the overall tumor burden or decreasing the number of cancer cells; and improving one or more symptoms associated with the disease.

[0037] In relation to autoimmune diseases, the term “treating” includes any or all of the following: inhibiting the replication of cells associated with an autoimmune disease state, including but not limited to cells that produce autoimmune antibodies; reducing the autoimmune antibody load; and improving one or more symptoms of an autoimmune disease.

[0038] As used in this application, the specification, and the attached claims, the indefinite articles "a" and "an" and the definite article "the" refer to multiple referents as well as a single referent, unless the context otherwise makes clear.

[0039] As used herein, and unless otherwise specified, the terms “about” and “approximately” mean, when used in relation to the amount or weight percentage of a component of a composition, an amount or weight percentage that would be recognized by those skilled in the art as providing an equivalent pharmacological effect to that obtained from a specified amount or weight percentage. In a given embodiment, when used in this context, the terms “about” and “approximately” mean an amount or weight percentage within 30%, 20%, 15%, 10%, or 5% of the specified amount or weight percentage.

[0040] As used herein, and unless otherwise specified, the terms “about” and “approximately” indicate that the value or range of value may deviate to an extent that is reasonable to a person skilled in the art while still describing the solid form, when used in relation to a range of numerical values ​​or values ​​provided to characterize a particular solid form, such as a specific temperature or temperature range representing melting, dehydration, desolvation, or glass transition temperature; a change in mass, such as a change in mass depending on temperature or humidity; solvent or water content, such as in terms of mass or percentage; or, when used in relation to a peak position, such as in analysis by IR or Raman spectroscopy or XRPD. Techniques for characterizing crystalline and amorphous solids include, but are not limited to, thermogravimetric analysis (TGA), differential scanning calorimetry (DSC), X-ray powder diffraction (XRPD), single-crystal X-ray diffraction, vibrational spectroscopy, such as infrared (IR) and Raman spectroscopy, solid-state and solution nuclear magnetic resonance (NMR) spectroscopy, optical microscopy, hot-stage optical microscopy, scanning electron microscopy (SEM), electron crystallography and quantitative analysis, particle size analysis (PSA), surface area analysis, solubility studies, and dissolution studies. In certain embodiments, the terms “about” and “approximately” indicate, when used in this context, that the numerical value or range of values ​​may vary within 30%, 20%, 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1.5%, 1%, 0.5%, or 0.25% of the described value or range of values. For example, in some embodiments, the value of the XRPD peak position can vary by up to ±0.2°²θ while still representing a particular XRPD peak.

[0041] As used herein, the term “including both ends” means, when used in relation to a range, that the endpoints of the range are included. For example, if n is an integer between 0 and 4, then n could be 0, 1, 2, 3, or 4.

[0042] The "alkyl" group is a saturated, partially saturated, or unsaturated linear or branched acyclic hydrocarbon having 1 to 10 carbon atoms, typically 1 to 8 carbon atoms, or in some embodiments, 1 to 6, 1 to 4, or 2 to 6 carbon atoms. Typical alkyl groups include -methyl, -ethyl, -n-propyl, -n-butyl, -n-pentyl, and n-hexyl; saturated branched alkyl groups include -isopropyl, -sec-butyl, -isobutyl, -tert-butyl, -isopentyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2,3-dimethylbutyl, etc. Examples of unsaturated alkyl groups include, but are not limited to, vinyl, allyl, CH=CH(CH3), -CH=C(CH3)2, -C(CH3)=CH2, -C(CH3)=CH(CH3), C(CH2CH3)=CH2, C≡CH, -C≡C(CH3), -C≡C(CH2CH3), -CH2C≡CH, -CH2C≡C(CH3), and CH2C≡C(CH2CH3). Alkyl groups can be substituted or unsubstituted. In certain embodiments, where alkyl groups are referred to as "substituted" in this specification, they may be substituted with any substituent(s) as those found in the compounds and embodiments disclosed herein, as well as halogens (chloro, iodine, bromo, or fluoro); hydroxyl; alkoxy; alkoxyalkyl; amino; alkylamino; carboxy; nitro; cyano; thiol; thioether; imine; imide; amidine; guanidine; enamine; aminocarbonyl; acylamino; phosphonato; phosphine; thiocarbonyl; sulfonyl; sulfone; sulfonamide; ketone; aldehyde; ester; urea; urethane; oxime; hydroxylamine; alkoxyamine; aralkoxyamine; N-oxide; hydrazine; hydrazide; hydrazone; azide; isocyanate; isothiocyanate; cyanate; thiocyanate; B(OH)2; or O(alkyl)aminocarbonyl.

[0043] The "alkenyl" group is a linear or branched acyclic hydrocarbon having 2 to 10 carbon atoms, typically 2 to 8 carbon atoms, and containing at least one carbon-carbon double bond. Representative linear and branched (C2-C8) alkenyls include -vinyl, -allyl, -1-butenyl, -2-butenyl, -isobutyrenyl, -1-pentenyl, -2-pentenyl, -3-methyl-1-butenyl, -2-methyl-2-butenyl, -2,3-dimethyl-2-butenyl, -1-hexenyl, 2-hexenyl, -3-hexenyl, -1-heptenyl, -2-heptenyl, -3-heptenyl, -1-octenyl, -2-octenyl, and 3-octenyl. The double bond of the alkenyl group is either non-conjugated or conjugated to other unsaturated groups. The alkenyl group can be unsubstituted or substituted.

[0044] A "cycloalkyl" group is a saturated or partially saturated cyclic alkyl group of 3 to 10 carbon atoms having a monocyclic ring or multiple condensed or crosslinked rings, which can be optionally substituted with 1 to 3 alkyl groups. In some embodiments, the cycloalkyl group has 3 to 8 ring members, while in other embodiments, the number of ring carbon atoms is in the range of 3 to 5, 3 to 6, or 3 to 7. Examples of such cycloalkyl groups include monocyclic structures such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, 1-methylcyclopropyl, 2-methylcyclopentyl, 2-methylcyclooctyl, etc., or polycyclic or crosslinked ring structures such as adamantyl, etc. Examples of unsaturated cycloalkyl groups include, among others, cyclohexenyl, cyclopentenyl, cyclohexadienyl, butadienyl, pentadienyl, and hexadienyl. Cycloalkyl groups can be substituted or unsubstituted. Examples of such substituted cycloalkyl groups include cyclohexanone, etc.

[0045] An "aryl" group is an aromatic carbocyclic group of 6 to 14 carbon atoms having a single ring (e.g., phenyl) or multiple fused rings (e.g., naphthyl or anthryl). In some embodiments, the aryl group contains 6 to 14 carbon atoms in its ring portion and 6 to 12 or further 6 to 10 carbon atoms in the other portion. Specific aryls include phenyl, biphenyl, naphthyl, etc. The aryl group can be substituted or unsubstituted. The term "aryl group" also includes groups containing fused rings, such as fused aromatic-aliphatic ring systems (e.g., indanyl, tetrahydronaphthyl, etc.).

[0046] The "arylene" group is a divalent aryl group as defined herein.

[0047] A "heteroaryl" group is an aryl ring system having 1 to 4 heteroatoms as ring atoms in a heteroaromatic ring system, with the remaining atoms being carbon atoms. In some embodiments, the heteroaryl group contains 5 to 6 ring atoms in the ring portion of the group, and 6 to 9 or 6 to 10 atoms on the other side. Suitable heteroatoms include oxygen, sulfur, and nitrogen. In certain embodiments, the heteroaryl ring system is monocyclic or bicyclic. Non-restrictive examples include pyrrolyl, pyrazolyl, imidazolyl, triazolyl, tetrazolyl, oxazolyl, isoxazolyl, thiazolyl, pyrrolyl, pyridyl, pyridadinyl, pyrimidinyl, pyrazinyl, thiophenyl, benzothiophenyl, furanyl, benzofuranyl (e.g., isobenzofuran-1,3-diimine), indolyl, azaindolyl (e.g., pyrrolopyridyl or 1H-pyrrolo[2,3-b]pyridyl), indazolyl, benzimidazolyl (e.g., 1H-benzo[d]imidazolyl), This includes, but is not limited to, groups such as imidazopyridyl (e.g., azabenzimidazolyl, 3H-imidazo[4,5-b]pyridyl, or 1H-imidazo[4,5-b]pyridyl), pyrazolopyridyl, triazolopyridyl, benzotriazolyl, benzoxazolyl, benzothiazolyl, benzothiadiazolyl, isoxazolopyridyl, thianaphthalenyl, purinyl, xanthinyl, adeninyl, guaninyl, quinolinyl, isoquinolinyl, tetrahydroquinolinyl, quinoxalinyl, and quinazolinyl groups.

[0048] The "heteroarylene" group is a divalent heteroaryl group as defined herein.

[0049] A "heterocyclyl" is an aromatic (also called heteroaryl) or non-aromatic cycloalkyl group in which 1 to 4 of the ring carbon atoms are independently replaced by heteroatoms from the group consisting of O, S, and N. In some embodiments, the heterocyclyl group contains 3 to 10 ring members, while other such groups have 3 to 5, 3 to 6, or 3 to 8 ring members. Heterocyclyls can also be bonded to other groups at any ring atom (i.e., at any carbon or heteroatom of the heterocyclic ring). Heterocyclyl groups can be substituted or unsubstituted. Heterocyclyl groups include unsaturated, partially saturated, and saturated ring systems, such as imidazolyl, imidazolinyl, and imidazolidinyl groups. The term "heterocyclyl" includes fused ring species, such as those containing fused aromatic and non-aromatic groups, such as benzotriazolyl, 2,3-dihydrobenzo[l,4]dioxynyl, and benzo[l,3]dioxolyl. The term also includes bridged polycyclic ring systems containing heteroatoms, such as quinuclidyls, but are not limited to these.Typical examples of heterocyclyl groups include azilidinyl, azetidinyl, pyrrolidyl, imidazolidinyl, pyrazolidinyl, thiazolidinyl, tetrahydrothiophenyl, tetrahydrofuranil, dioxolyl, furanil, thiophenyl, pyrrolyl, pyrrolidyl, imidazolyl, imidazolinil, pyrazolyl, pyrazolinil, triazolyl, tetrazolyl, oxazolyl, isoxazolyl, thiazolyl, thiazolinil, isothiazolyl, thiadiazolyl, oxadiazolyl, piperidyl, piperazinyl, morpholinil, thiomorpholinil, tetra Lahydrothiopyranil (e.g., tetrahydro-2H-pyranil), tetrahydrothiopyranil, oxatian, dioxyl, dithianil, pyranil, pyridyl, pyrimidinil, pyridazinil, pyrazinil, triazinil, dihydropyridyl, dihydrodithinyl, dihydrodithionyl, homopiperazinil, quinuclidyl, indolyl, indolinyl, isoindolyl, azaindolyl (pyrrolopyridyl), indazolyl, indolidinil, benzotriazolyl, benzimidazolyl, benzofuranil, benzothiophenyl, benzthiazolyl Benzoxadiazolyl, benzoxadinyl, benzodithinyl, benzoxadinyl, benzothiadinyl, benzoxazolyl, benzothiazolyl, benzothiadiazolyl, benzo[l,3]dioxolyl, pyrazolopyridyl, imidazopyridyl (azabenzimidazolyl; e.g., 1H-imidazo[4,5-b]pyridyl, or 1H-imidazo[4,5-b]pyridyl), triazolopyridyl, isoxazolopyridyl, prinyl, xanthinyl, adeninyl, guaninyl, quinolinyl, isoquinolinyl, quinolidinyl, quinoxalini This includes, but is not limited to, quinazolinyl, cinolinyl, phthalazinyl, naphthilidinyl, pteridinyl, thianaphthalenyl, dihydrobenzothiadinyl, dihydrobenzofuranyl, dihydroindolyl, dihydrobenzodioxynyl, tetrahydroindolyl, tetrahydroindazolyl, tetrahydrobenzimidazolyl, tetrahydrobenzotriazolyl, tetrahydropyrrolopyridyl, tetrahydropyrazolopyridyl, tetrahydroimidazopyridyl, tetrahydrotriazolopyridyl, and tetrahydroquinolinyl groups.Typical substituted heterocyclyl groups can be monosubstituted or polysubstituted, for example, pyridyl or morpholinyl groups that are 2, 3, 4, 5, or 6-substituted, or disubstituted with various substituents such as those listed below.

[0050] The "cycloalkylalkyl" group is a radical of the formula -alkyl-cycloalkyl (wherein alkyl and cycloalkyl are defined above). A substituted cycloalkylalkyl group may be substituted with the alkyl, cycloalkyl, or both alkyl and cycloalkyl parts of its group. Representative cycloalkylalkyl groups include, but are not limited to, cyclopentylmethyl, cyclopentylethyl, cyclohexylmethyl, cyclohexylethyl, and cyclohexylpropyl. Representative substituted cycloalkylalkyl groups may be monosubstituted or multiple substituted.

[0051] An "aralkyl" group is a radical of the formula -alkyl-aryl (wherein alkyl and aryl are defined above). A substituted aralkyl group can be substituted with the alkyl, aryl, or both alkyl and aryl parts of the group. Typical aralkyl groups include, but are not limited to, benzyl and phenethyl groups and condensed (cycloalkylaryl)alkyl groups, such as 4-ethyl-indanyl.

[0052] The "heterocyclylalkyl" group is a radical of the formula -alkyl-heterocyclyl (wherein alkyl and heterocyclyl are defined above). A substituted heterocyclylalkyl group may be substituted with the alkyl, heterocyclyl, or both alkyl and heterocyclyl portions of its group. Representative heterocyclylalkyl groups include, but are not limited to, 4-ethyl-morpholinyl, 4-propylmorpholinyl, furan-2-ylmethyl, furan-3-ylmethyl, pyrzin-3-ylmethyl, (tetrahydro-2H-pyran-4-yl)methyl, (tetrahydro-2H-pyran-4-yl)ethyl, tetrahydrofuran-2-ylmethyl, tetrahydrofuran-2-ylethyl, and indole-2-ylpropyl.

[0053] "Halogen" refers to chloro, iodine, bromo, or fluoro compounds.

[0054] A "hydroxyalkyl" group is the alkyl group described above that is substituted with one or more hydroxyl groups.

[0055] The "alkoxy" group is O(alkyl) (where alkyl is defined above).

[0056] The "alkoxyalkyl" group is (alkyl)O(alkyl) (where alkyl is defined above).

[0057] As used herein, “alkynyl” refers to a monovalent hydrocarbon radical moiety containing at least two carbon atoms and one or more carbon-carbon triple bonds. Alkynnyls are optionally substituted and may be linear, branched, or cyclic. Alkynnyls are radicals having 2 to 20 carbon atoms, i.e., C 2-20 Alkynnyl; a radical having 2 to 12 carbon atoms, i.e., C 2-12 Alkynnyl; radicals having 2 to 8 carbon atoms, i.e., C 2-8 Alkynnyl; radicals having 2 to 6 carbon atoms, i.e., C 2-6Alkynnyls; and their radicals having 2 to 4 carbon atoms, i.e., C 2-4 This includes, but is not limited to, alkynyl moieties. Examples of alkynyl moieties include, but are not limited to, ethinyl, propynyl, and butynyl.

[0058] As used herein, “haloalkyl” refers to an alkyl group as defined above, wherein the alkyl group comprises at least one substituent selected from halogens, such as fluorine (F), chlorine (Cl), bromine (Br), or iodine (I). Examples of haloalkyl groups include, but are not limited to, -CF3, -CH2CF3, -CCl2F, and -CCl3.

[0059] As used herein, “haloalkoxy” refers to an alkoxy as defined above, wherein the alkoxy comprises at least one substituent selected from halogens, e.g., F, Cl, Br, or I.

[0060] As used herein, “arylalkyl” refers to a monovalent radical of an alkyl compound in which the alkyl compound is substituted with an aromatic substituent, i.e., the aromatic compound contains a single bond to the alkyl group, and the radical is localized to the alkyl group. Arylalkyls are bonded to the exemplified chemical structures via the alkyl group. Arylalkyls may be represented by structures such as B-CH2-, B-CH2-CH2-, B-CH2-CH2-CH2-, B-CH2-CH2-CH2-CH2-, B-CH(CH3)-CH2-CH2-, B-CH2-CH(CH3)-CH2- (wherein B is an aromatic moiety, e.g., phenyl). Arylalkyls are optionally substituted, i.e., the aryl group and / or alkyl group may be substituted as disclosed herein. Examples of arylalkyls include, but are not limited to, benzyl.

[0061] As used herein, “alkylaryl” refers to a monovalent radical of an aryl compound in which the aryl compound is substituted with an alkyl substituent, i.e., the aryl compound contains a single bond to the alkyl group, and the radical is localized to the aryl group. The alkylaryl group is bonded to the exemplified chemical structure via the aryl group. Alkylaryls can be represented by structures such as -B-CH3, -B-CH2-CH3, -B-CH2-CH2-CH3, -B-CH2-CH2-CH2-CH3, -B-CH(CH3)-CH2-CH3, and -B-CH2-CH(CH3)-CH3 (wherein B is an aromatic moiety, e.g., phenyl). Alkylaryls are optionally substituted, i.e., the aryl group and / or alkyl group can be substituted as disclosed herein. Examples of alkylaryls include, but are not limited to, toluyl.

[0062] As used herein, “aryloxy” refers to a monovalent radical of an aromatic compound in which the ring atom is a carbon atom and the ring is substituted with an oxygen radical, i.e., the aromatic compound contains a single bond to the oxygen atom and the radical is localized on the oxygen atom, for example, in the case of phenoxy, C6H5-O-. Aryloxy substituents are bonded to the compound they substitute via this oxygen atom. Aryloxys are optionally substituted. Aryloxys are their radicals having 6 to 20 ring carbon atoms, i.e., C 6-20 Aryloxy; radicals having 6 to 15 ring carbon atoms, i.e., C 6-15 Aryloxys, and their radicals having 6 to 10 ring carbon atoms, i.e., C 6-10 This includes, but is not limited to, aryloxy groups. Examples of aryloxy groups include, but are not limited to, phenoxy, naphthoxy, and anthroxy.

[0063] The "amino" group is a radical with the formula NH2.

[0064] The "hydroxylamine" group is represented by formula N(R# )OH or NHOH (wherein R in the formula) # (is a radical of a substituted or unsubstituted alkyl, cycloalkyl, cycloalkylalkyl, aryl, aralkyl, heterocyclyl, or heterocyclylalkyl group as defined herein.)

[0065] The "alkoxyamine" group is represented by formula -N(R # )O-alkyl or -NHO-alkyl (wherein R # It is the radical of (as defined above).

[0066] The "aralcoxyamine" group is represented by formula N(R # )O-aryl or NHOaryl (wherein R # It is the radical of (as defined above).

[0067] The "alkylamine" group is a radical of the formula NH alkyl or N(alkyl)2 (wherein each alkyl is independently as defined above).

[0068] The "aminocarbonyl" group is represented by the formula -C(=O)N(R # )2, -C(=O)NH(R # ), or C(=O)NH2(wherein each R # It is the radical of (as defined above).

[0069] The "acylamino" group is represented by the formula NHC(=O)(R # ) or N(alkyl)C(=O)(R # )(wherein each alkyl and R # It is an independent radical of (as defined above).

[0070] The "O(alkyl)aminocarbonyl" group is represented by the formula -O(alkyl)C(=O)N(R # )2,-O(alkyl)C(=O)NH(R # ), or -O(alkyl)C(=O)NH2 (wherein each R #It is an independent radical of (as defined above).

[0071] The "N-oxide" group is represented by the formula -N + -O - It is radical.

[0072] The "carboxyl" group is a radical with the formula C(=O)OH.

[0073] The "ketone" group is represented by the formula C(=O)(R # )(wherein, R # It is the radical of (as defined above).

[0074] The "aldehyde" group is a radical with the formula -CH(=O).

[0075] The "ester" group is represented by the formula C(=O)O(R # ) or OC (=O) (R # )(wherein, R # It is the radical of (as defined above).

[0076] The "urea" group is represented by the formula -N(alkyl)C(=O)N(R # )2,-N(alkyl)C(=O)NH(R # ), -N(alkyl)C(=O)NH2, -NHC(=O)N(R # )2, -NHC(=O)NH(R # ), or NHC(=O)NH2 # (In the formula, each alkyl and R # It is an independent radical of (as defined above).

[0077] The "imine" group is represented by the formula -N=C(R # )2 or -C(R # )=N(R # )(In the formula, each R # It is an independent radical of (as defined above).

[0078] The "imide" group is represented by the formula -C(=O)N(R#)C(=O)(R #) or N((C=O)(R # ))2(In the formula, each R # It is an independent radical of (as defined above).

[0079] The "urethane" group is represented by the formula -OC(=O)N(R # )2, -OC(=O)NH(R # ), -N(R # )C(=O)O(R # ), or -NHC(=O)O(R # )(In the formula, each R # It is an independent radical of (as defined above).

[0080] The "amidine" group is represented by the formula -C(=N(R # ))N(R # )2, -C(=N(R # ))NH(R # ), -C(=N(R # ))NH2, -C(=NH)N(R # )2, -C(=NH)NH(R # ), -C(=NH)NH2, -N=C(R # )N(R # )2, -N=C(R # )NH(R # ), -N=C(R # )NH2, -N(R # )C(R # )=N(R # ), -NHC(R # )=N(R # ), -N(R # )C(R # )=NH, or -NHC(R # )=NH(in the formula, each R # It is an independent radical of (as defined above).

[0081] The "guanidine" group is represented by formula -N(R # )C(=N(R # ))N(R # )2, -NHC(=N(R # ))N(R # )2, -N(R #)C(=NH)N(R # )2, -N(R # )C(=N(R # ))NH(R # ), -N(R # )C(=N(R # ))NH2, -NHC(=NH)N(R # )2, -NHC(=N(R # ))NH(R # ), -NHC(=N(R # ))NH2, -NHC(=NH)NH(R # ), -NHC(=NH)NH2, -N=C(N(R # )2)2, -N=C(NH(R # ))², or -N=C(NH²)² (where each R # It is an independent radical of (as defined above).

[0082] The "enamine" group is represented by formula -N(R # )C(R # )=C(R # )2, -NHC(R # )=C(R # )2, -C(N(R # )2)=C(R # )2, -C(NH(R # ))=C(R # )2, -C(NH2)=C(R # )2, -C(R # )=C(R # )(N(R # )2), C(R # )=C(R # )(NH(R # )) or -C(R # )=C(R # )(NH2)(wherein, each R # It is an independent radical of (as defined above).

[0083] The "oxime" group is represented by the formula -C(=NO(R # ))(R # ), -C(=NOH)(R # ), -CH(=NO(R # )), or -CH(=NOH)(in the formula, each R# It is an independent radical of (as defined above).

[0084] The "hydrazide" group is represented by the formula -C(=O)N(R # )N(R # )2, -C(=O)NHN(R # )2, -C(=O)N(R # )NH(R # ), -C(=O)N(R # )NH2, -C(=O)NHNH(R # )2, or -C(=O)NHNH2(where R # It is an independent radical of (as defined above).

[0085] The "hydrazine" group is represented by formula -N(R # )N(R # )2, -NHN(R # )2, -N(R # )NH(R # ), -N(R # )NH2, -NHNH(R # )2, or -NHNH2(wherein each R # It is an independent radical of (as defined above).

[0086] The "hydrazone" group is represented by the formula -C(=NN(R # )2)(R # )2, -C(=NNH(R # ))(R # )2, -C(=N-NH2)(R # )2, -N(R # )(N=C(R # )2), or -NH(N=C(R # )2)(In the formula, each R # It is an independent radical of (as defined above).

[0087] The "azide" group is a radical of formula -N3.

[0088] The "isocyanate" group is a radical with the formula N=C=O.

[0089] The "isothiocyanate" group is a radical with the formula N=C=S.

[0090] The "cyanate" group is the radical of formula OCN.

[0091] The "thiocyanate" group is a radical of formula SCN.

[0092] The "thioether" group is represented by the formula -S(R # )(wherein, R # It is the radical of (as defined above).

[0093] The "thiocarbonyl" group is represented by the formula -C(=S)(R # )(wherein, R # It is the radical of (as defined above).

[0094] The "sulfinyl" group is represented by the formula -S(=O)(R # )(wherein, R # It is the radical of (as defined above).

[0095] The "sulfone" group is represented by the formula -S(=O)2(R # )(wherein, R # It is the radical of (as defined above).

[0096] The "sulfonylamino" group is represented by the formula -NHSO2(R # ) or -N(alkyl)SO2(R # )(wherein each alkyl and R # It is a radical of (as defined above).

[0097] The "sulfonamide" group is represented by the formula -S(=O)2N(R # )2, or -S(=O)2NH(R # ), or -S(=O)2NH2(wherein each R # It is an independent radical of (as defined above).

[0098] The "phosphonate" group is represented by the formula -P(=O)(O(R # ))2, -P(=O)(OH)2, -OP(=O)(O(R # ))(R # ), or -OP(=O)(OH)(R # )(In the formula, each R # It is an independent radical of (as defined above).

[0099] The "phosphine" group is represented by formula -P(R # )2(in the formula, each R # It is an independent radical of (as defined above).

[0100] Where the groups described herein, except for alkyl groups, are referred to as “substituted,” they may be substituted with any suitable substituent(s). Exemplary examples of substituents are those found in the compounds and embodiments disclosed herein, as well as halogens (chloro, iodo, bromo, or fluoro); alkyl; hydroxyl; alkoxy; alkoxyalkyl; amino; alkylamino; carboxy; nitro; cyano; thiol; thioether; imine; imide; amidine; guanidine; enamine; aminocarbonyl; acylamino; phosphonate; phosphine; thiocarbonyl; sulfinyl; sulfone; sulfonamide; ketone; aldehyde; ester; urea; urethane; oxime; hydroxylamine; alkoxyamine; aralkoxyamine; N-oxide; hydrazine; hydrazide; hydrazone; azide; isocyanate; isothiocyanate; cyanate; thiocyanate; oxygen (=O); B(OH)2, O(alkyl)aminocarbonyl; monocyclic or condensed or These are cycloalkyls that may be non-condensed polycyclic (e.g., cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl), or heterocyclyls that may be monocyclic or condensed or non-condensed polycyclic (e.g., pyrrolidyl, piperidyl, piperazinyl, morpholinyl, or thiadinyl); monocyclic or condensed or non-condensed polycyclic aryl or heteroaryls (e.g., phenyl, naphthyl, pyrrolyl, indolyl, furanyl, thiophenyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, triazolyl, tetrazolyl, pyrazolyl, pyridinyl, quinolinyl, isoquinolinyl, acridinyl, pyrazinyl, pyridadinyl, pyrimidinyl, benzimidazolyl, benzothiophenyl, or benzofuranyl) aryloxys; aralkyloxys; heterocyclyloxys; and heterocyclylalkoxys.

[0101] As used herein, the term “pharmaceutically acceptable salt” refers to a salt prepared from a pharmaceutically acceptable nontoxic acid or base, including inorganic acids or bases and organic acids or bases.

[0102] As used herein, and unless otherwise indicated, the term “solvate” means a compound or salt thereof that further comprises a stoichiometric or nonstoichiometric amount of solvent bonded by non-covalent intermolecular forces. In one embodiment, the solvate is a hydrate.

[0103] As used herein, and unless otherwise indicated, the term “hydrate” means a compound or salt thereof that further contains stoichiometric or nonstoichiometric amounts of water bonded by non-covalent intermolecular forces.

[0104] As used herein, and unless otherwise indicated, the term “prodrug” means a compound derivative that can be hydrolyzed, oxidized, or otherwise reacted under biological conditions (in vitro or in vivo) to provide an active compound. Examples of prodrugs include, but are not limited to, derivatives and metabolites of compounds containing biohydrolyzable sites, such as biohydrolyzable amides, biohydrolyzable esters, biohydrolyzable carbamates, biohydrolyzable carbonates, biohydrolyzable ureids, and biohydrolyzable phosphate analogs. In certain embodiments, a prodrug of a compound having a carboxyl functional group is a lower alkyl ester of a carboxylic acid. Carboxylate esters can be formed by esterifying any of the carboxylic acid sites present in the molecule. Prodrugs are typically described in Burger's Medicinal Chemistry and Drug Discovery 6. th It can be prepared using well-known methods, such as those described in (Donald J. Abraham ed., 2001, Wiley) and Design and Application of Prodrugs (H. Bundgaard ed., 1985, Harwood Academic Publishers GmbH).

[0105] As used herein, and unless otherwise indicated, the terms “stereoisomer” or “stereoisomerically pure” mean one stereoisomer of a compound that is substantially free of other stereoisomers of that compound. For example, a stereoisomerically pure compound having one chiral center is substantially free of the opposite enantiomer of the compound. A stereoisomerically pure compound having two chiral centers is substantially free of the other diastereomer of the compound. Typical stereoisomerically pure compounds include those containing more than about 80 wt% of one stereoisomer of the compound and less than about 20 wt% of the other stereoisomer of the compound, more than about 90 wt% of one stereoisomer of the compound and less than about 10 wt% of the other stereoisomer of the compound, more than about 95 wt% of one stereoisomer of the compound and less than about 5 wt% of the other stereoisomer of the compound, or more than about 97 wt% of one stereoisomer of the compound and less than about 3 wt% of the other stereoisomer of the compound. Compounds may have chiral centers and may arise as racemates, individual enantiomers or diastereomers, and mixtures thereof. All such isomeric forms, including mixtures thereof, are included in the embodiments disclosed herein. The use of stereoisomerically pure forms of such compounds, and the use of mixtures of those forms, are encompassed by the embodiments disclosed herein. For example, mixtures containing equimolar or unequal amounts of enantiomers of a particular compound may be used in the methods and compositions disclosed herein. These isomers may be synthesized asymmetrically or divided using standard techniques, such as chiral columns or chiral resolving agents.For example, see Jacques, J., et al., Enantiomers, Racemates and Resolutions (Wiley Interscience, New York, 1981); Wilen, SH, et al., Tetrahedron 33:2725 (1977); Eliel, EL, Stereochemistry of Carbon Compounds (McGraw Hill, NY, 1962); and Wilen, SH, Tables of Resolving Agents and Optical Resolutions p.268 (EL Eliel, Ed., Univ. of Notre Dame Press, Notre Dame, IN, 1972).

[0106] It should be noted that the compound may include E and Z isomers, or mixtures thereof, as well as cis and trans isomers, or mixtures thereof. In a given embodiment, the compound is isolated as either a cis or trans isomer. In other embodiments, the compound is a mixture of cis and trans isomers.

[0107] A "tautomer" refers to an isomer of a compound that is in equilibrium with it. The concentrations of these isomers depend on the environment in which the compound is found, for example, whether the compound is in a solid state or in an organic solution or aqueous solution. For example, in aqueous solution, pyrazole may exhibit the following isomers, which are called tautomers: [ka]

[0108] As will be readily apparent to those skilled in the art, diverse functional groups and other structures can exhibit tautomerism, and all tautomers of a compound are within the scope of this disclosure.

[0109] It should also be noted that compounds may contain one or more atomic isotopes in a non-natural proportion. For example, compounds may contain radioactive isotopes, such as tritium. 3 H), Iodine-125( 125 I), Sulfur-35( 35 S), or carbon-14 ( 14 C) may be radioactively labeled, or deuterium ( 2 H), carbon-13 ( 13 C), or nitrogen-15( 15 They can be isotopedally enriched with N), etc. As used herein, “isotopololog” is an isotopedally enriched compound. The term “isotopically enriched” refers to an atom having an isotope composition other than the natural isotope composition of that atom. “Isotopedally enriched” may also refer to a compound containing at least one atom having an isotope composition other than the natural isotope composition of that atom. The term “isotope composition” refers to the amount of each isotope present for a given atom. Radiolabeled compounds and isotopedally enriched compounds are useful as therapeutic agents, e.g., cancer and inflammation treatments, research reagents, e.g., binding assay reagents, and diagnostic agents, e.g., in vivo contrast agents. All isotope variations of the compounds described herein, whether radioactive or not, are intended to be included within the scope of the embodiments provided herein. In some embodiments, isotopologs of compounds are provided, for example, isotopologs are compounds enriched with deuterium, carbon-13, or nitrogen-15.

[0110] It should be noted that if there is a contradiction between the structure being shown and the name of that structure, the structure being shown should be given more weight.

[0111] As used herein, the term “residue” refers to a chemical site within a compound that is retained after a chemical reaction. For example, the term “amino acid residue” or “N-alkyl amino acid residue” refers to the product of an amide coupling or peptide coupling of an amino acid or N-alkyl amino acid to a suitable coupling partner, in which case, for example, a water molecule is expelled after the amide or peptide coupling of the amino acid or N-alkyl amino acid, resulting in a product having an amino acid residue or N-alkyl amino acid residue incorporated into the product.

[0112] As used herein, “sugar,” “sugar group,” or “sugar residue” refers to a carbohydrate moiety that may contain 3-carbon (their) units, 4-carbon (tetrose) units, 5-carbon (pentose) units, 6-carbon (hexose) units, 7-carbon (heptose) units, or combinations thereof, and may be monosaccharides, disaccharides, trisaccharides, tetrasaccharides, pentasaccharides, oligosaccharides, or any other polysaccharides. In some examples, “sugar,” “sugar group,” or “sugar residue” may include furanoses (e.g., ribofuranose, fructofuranose) or pyranoses (e.g., glucopyranose, galactopyranose), or combinations thereof. In some examples, “sugar,” “sugar group,” or “sugar residue” may include aldoses or ketoses, or combinations thereof. Non-limiting examples of monosaccharides include ribose, deoxyribose, xylose, arabinose, glucose, mannose, galactose, and fructose. Non-exclusive examples of disaccharides include sucrose, maltose, lactose, lactulose, and trehalose. Other “sugars” or “sugar groups” or “sugar residues” include, but are not limited to, amylose, amylopectin, glycogen, inulin, and cellulose, as well as polysaccharides and / or oligosaccharides. In some examples, the “sugar” or “sugar group” or “sugar residue” is an amino sugar. In some examples, the “sugar” or “sugar group” or “sugar residue” is a glucamine residue (1-amino-1-deoxy-D-glucitol) linked to the remainder of the molecule (i.e., glucamide) via its amino group to form an amide bond with the remainder of the molecule.

[0113] As used herein, “binding agent” refers to a given binding partner, for example, any molecule capable of binding with specificity to an antigen, such as an antibody.

[0114] As used herein, the term “amino acid” refers to an organic compound containing an amino (-NH2) and a carboxyl (-COOH) functional group, along with a side chain (R group) specific to each amino acid. Amino acids can be protein-constitutive or non-protein-constitutive. “Protein-constitutive” means that the amino acid is one of the 20 naturally occurring amino acids found in proteins. Protein-constitutive amino acids include alanine, arginine, asparagine, aspartic acid, cysteine, glutamine, glutamic acid, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine. “Non-protein-constitutive” means that any of the amino acids is not naturally found in proteins or is not directly produced by cellular mechanisms (e.g., is a product of post-translational modification). Non-protein-constitutive amino acids include, but are not limited to, gamma-aminobutyric acid (GABA), taurine (2-aminoethanesulfonic acid), theanine (L-γ-glutamylethylamide), hydroxyproline, beta-alanine, ornithine, and citrulline.

[0115] As used herein, “peptide” (in its various grammatical forms) is defined in its broadest sense to refer to compounds of two or more subunit amino acids, amino acid analogs, or other peptidomimetics. Subunits may be linked by peptide bonds or other bonds, e.g., esters, ethers, etc. As used herein, the term “amino acid” refers to any natural and / or unnatural, protein-constitutive or non-protein-constitutive, or synthetic amino acid, including glycine and both D and L optical isomers, as well as amino acid analogs and peptidomimetics. When the peptide chain is short, for example, two, three or more amino acids, it is generally called an oligopeptide. When the peptide chain is longer, the peptide is typically called a polypeptide or protein. Full-length proteins, analogs, variants, and fragments thereof are encompassed by definition. The term also includes post-expression modifications of polypeptides, e.g., glycosylation, acetylation, phosphorylation, etc. Furthermore, since ionizable amino and carboxyl groups are present in the molecule, certain peptides may be obtained as acidic or basic salts or in neutral forms. Peptides can be obtained directly from source organisms, or they can be produced recombinantly or synthetically.

[0116] The amino acid sequences of antibodies may be numbered using any known numbering scheme, including those described by Kabat et al. ("Kabat" numbering scheme); Al-Lazikani et al., 1997, J.Mol.Biol., 273:927-948 ("Chothia" numbering scheme); MacCallum et al., 1996, J.Mol.Biol., 262:732-745 ("Contact" numbering scheme); Lefranc et al., Dev.Comp.Immunol., 2003, 27:55-77 ("IMGT" numbering scheme); and Honegge and Pluckthun, J.Mol.Biol., 2001, 309:657-70 ("AHo" numbering scheme). Unless otherwise specified, the numbering scheme used herein is the Kabat numbering scheme. However, the selection of numbering schemes is not intended to distinguish between sequences where they do not exist, and those skilled in the art can easily determine the sequence locations by examining the amino acid sequences of one or more antibodies. Unless otherwise stated, the “EU numbering scheme” is typically used when referring to residues in the constant region of the antibody heavy chain (as reported, for example, in Kabat et al., (cited above)).

[0117] As used herein, the term “anti-HER2 antibody” refers to an antibody that selectively binds to the HER2 receptor, such as trastuzumab. In one embodiment, trastuzumab may be produced and used as described in US6407213 and US5821337 (the entire disclosure of which is incorporated herein by reference).

[0118] As used herein, the term “anti-HER3 antibody” refers to an antibody that selectively binds to the HER3 receptor, such as patritumab. In one embodiment, patritumab may be produced and used as described in US7705130 (the entire disclosure of which is incorporated herein by reference).

[0119] As used herein, the term “anti-PTK7 antibody” refers to an antibody that selectively binds to the PTK7 receptor, such as cofetuzumab. In one embodiment, cofetuzumab may be produced and used as described in US9777070 (the entire disclosure of which is incorporated herein by reference).

[0120] As used herein, the term "ifinatamab" refers to an antibody that selectively binds to the B7H3 receptor, i.e., an anti-human B7H3 antibody. In one embodiment, ifinatamab may be produced and used as described in US10117952 or WO2022102695 (the entire disclosure of which is incorporated herein by reference).

[0121] As used herein, the term "6E7" refers to the CLL1 monoclonal antibody.

[0122] As used herein, the term “cytotoxic activity” refers to activity that reduces or diminishes the cellular viability of the cell line being tested.

[0123] In the following claims and prior descriptions, unless the context requires other meanings through explicit expression or necessary implication, the word “comprise” or variations such as “comprises” or “comprising” are used in a comprehensive sense, that is, to specify the presence of the described features, but not to exclude the presence or addition of further features in various embodiments.

[0124] Conjugate In embodiments, the conjugate, or its pharmaceutically acceptable salt, tautomer, solvate, or stereoisomer, includes a protein linked to at least one payload or payload residue (also referred herein as a drug unit) and linked via a covalent linker to at least one hydrophilic site (also referred herein as a cleavable unit or cleavable element). The covalent linker is directly or indirectly bound to each of the protein, payload residue, and hydrophilic site. In some embodiments, the protein is a binder, such as an antibody or its antigen-binding fragment.

[0125] In some embodiments, the protein is directly bound to a covalent linker, for example, a linker as shown herein. In such cases, the binder is one binding site away from the covalent linker. The covalent linker may also be directly bound to a payload residue such that the covalent linker is one binding site away from the payload residue. The payload may be any payload as shown herein, for example, MMAE. In some embodiments, the covalent linker may also be directly bound to a hydrophilic site such that the covalent linker is one binding site away from the hydrophilic site. The hydrophilic site may be any hydrophilic site as shown herein.

[0126] In some embodiments, the binder is indirectly bonded to the covalent linker such that the binder is separated from the covalent linker by more than one bond site. In such cases, the binder is bonded to the covalent linker via another site. For example, the binder may be bonded to a maleimide group, which may be bonded to a polyethylene glycol group, which may be bonded to the covalent linker.

[0127] In some cases, covalent linkers are also indirectly bound to payload residues such that the covalent linker is located more than one binding site away from the payload residue. Covalent linkers are bound to the payload via another site. For example, a covalent linker may be bound to a dipeptide, e.g., Val-Ala or Val-Cit, which may be bound to a p-aminobenzyl alcohol (PAB)-derived unit, which may be bound to a payload residue. An example of a PAB-derived unit is -NH-(C6H4)-CH2-OC(=O)-, where the phenylene moiety of the PAB unit may be substituted with -C1-C8 alkyl, -O-(C1-C8 alkyl), -halogen, -nitro, or -cyano.

[0128] In some embodiments, the covalent linker is indirectly bonded to the hydrophilic site such that the covalent linker is located at a distance of more than one bond site from the hydrophilic site. The covalent linker is bonded to the hydrophilic site via another site.

[0129] Compounds disclosed herein include at least a conjugator, a spacer, a cleavable, and a payload. In some embodiments, the compound includes a branch. In some embodiments, such as a conjugate, the compound includes a binder (BA). The conjugator is a group capable of binding to the BA and connecting the BA to the remainder of the compound. The cleavable includes at least one site that can be cleaved by a β-glucuronidase enzyme, e.g., glycosidic linkage or binding. The cleavable may include a hydrophilic site. A spacer is a group that extends the linker and provides more distance between the conjugator and the payload. The spacer can connect the conjugator to a cleavable or to a branch, if present. The branch is a group that extends the linker framework and includes a hydrophilic site. Spacers, cleavables, and branch may form the linker portion of a conjugate. The payload (also referred herein as a drug unit) is a cytotoxic agent or a residue thereof.

[0130] Appearance 1

[0131] This specification refers to compounds having formula (I). [ka] or a pharmaceutically acceptable salt, tautomer, solvate, or stereoisomer thereof (in the formula, BA is a binder selected from humanized, monoclonal, chimeric, or human antibodies or their antigen-binding fragments; The conjugator is given by equations (II) and (III): [ka] Selected from the group consisting of; U is a bond, heteroarylene, or arylene; V represents a bond or -C≡C-(CH2) n -and; n is an integer between 0 and 10 (including both ends); W2 is -C(=O)-, -NH-, or -O-; RG is [ka] , succinimido-3-yl-N)-, or [ka] and; RS is -NR 1a R 1b and; R 1a and R 1b Each of these independently represents H or substituted or unsubstituted C. 1-4 It is alkyl; RE stands for bond, -O-, -OC(=O)-, -OC(=O)NR 6 -, -NHC(=O)NR 6 -, -OS(=O)2NR 6 -, -NHS(=O)2NR 6 -, or -OC(=O)NHS(=O)2NR6 -and; R 6 is H or substituted or unsubstituted C 1-4 It is alkyl; W3 is -C(=O)-, -NH-, or -O-; Each of s and t is independently 0, 1, or 2; [ka] This indicates a covalent bond site within the compound. * indicates a connection where the conjugator connects to the BA; Spacers are used for bonding, ** -NH-(CH2CH2O) m -CH2CH2-C(=O)-, ** -(CH2) m -C(=O)-, ** -CH2-C(=O)-NH-(CH2) m -C(=O)-, ** -(CH2CH2O) m -CH2CH2-C(=O)-, ** -CH[-(CH2) m -COOH]-C(=O)-, ** -CH2-C(=O)-NH-(CH2) m -C(=O)-NH-(CH2) m -C(=O)-, ** -C(=O)-(CH2) m -C(=O)-, ** -C(=O)-(CH2CH2O) m -CH2CH2-NH-, ** -NH-(CH2) m -C(=O)-, or ** -NH-(CH2) m -O- is; m is an integer between 1 and 12 (including both ends); ** indicates a coupling where the spacer connects to the conjugator; The cleavable bodies are given by formulas (IVa), (IVb), (IVc), (Va), (Vb), (VIa), (VIb), (VIIa), or (VIIb): [ka] Having; Su is the sugar portion; Each R 2 These are, independently, hydrogen, halogen, substituted or unsubstituted C 1-4 It is alkyl, -CN, or -NO2; *** indicates a connection where the severable body connects to the spacer; The payload is a payload residue; x is between 1 and 15 (including both ends). It will be provided.

[0132] In some embodiments, the conjugator has formula (II).

[0133] In some embodiments, U is arylene.

[0134] In some embodiments, U is phenylene.

[0135] In some embodiments, U is [ka] That is the case.

[0136] In some embodiments, U is an arylene and V is a bond.

[0137] In some embodiments, U is arylene and V is -C≡C-(CH2) n - is

[0138] In some embodiments, U is arylene and V is -C≡C-(CH2)3-.

[0139] In some embodiments, U is a heteroarylene.

[0140] In some embodiments, U is a divalent pyrimidine ring.

[0141] In some embodiments, U is [ka] That is the case.

[0142] In some embodiments, U is a heteroarylene and V is a bond.

[0143] In some embodiments, U is a heteroarylene and V is -C≡C-(CH2) n - is

[0144] In some embodiments, U is a heteroarylene and V is -C≡C-(CH2)3-.

[0145] In some embodiments, U is a bond.

[0146] In some embodiments, U is a bond and V is a bond.

[0147] In some embodiments, U is a bond and V is -C≡C-(CH2) n - is

[0148] In some embodiments, U is a bond and V is -C≡C-(CH2)3-.

[0149] In some embodiments, W2 is -C(=O)-.

[0150] In some embodiments, the conjugator has formula (III).

[0151] In some embodiments, RS is -NH2 or -N(CH3)2.

[0152] In some embodiments, RS is -NH2.

[0153] In some embodiments, RE is -OC(=O)NH-.

[0154]

[0155] In some embodiments, RG is [ka] That is the case.

[0156] In some embodiments, RG is [ka] , -(succinimido-3-yl-N)-, or [ka] That is the case.

[0157] In some embodiments, RG is [ka] That is the case.

[0158] A specified RG as described herein, for example, [ka] It can be induced to undergo a ring-opening process by conjugation with BA. During such a process, the maleimide structure formed by the conjugation of RG and BA, for example, [ka] It undergoes ring opening. It is understood that there are two possible positional isomers that may result from such ring opening of the maleimide structure. For example, [ka] It undergoes ring opening. [ka] It may form, [ka] It undergoes ring opening. [ka] This can form. For simplicity, unless otherwise indicated, both possible ring-opening positional isomers are encompassed by the illustration of a single ring-opening positional isomer herein. Thus, unless otherwise indicated, the illustration of a single ring-opening positional isomer [ka] teeth, [ka] This includes, and similarly, illustrations of ring-opening positional isomers. [ka] teeth, [ka] This includes. In some embodiments, a mixture of ring-opening positional isomers exists. In some embodiments, a single ring-opening positional isomer exists.

[0159] In some embodiments, each of s and t is 2.

[0160] In some embodiments, the spacer is connected, ** -NH-(CH2CH2O) m -CH2CH2-C(=O)-, ** -C(=O)-(CH2CH2O) m -CH2CH2-NH-, or ** -NH-(CH2) m It is -O-.

[0161] In some embodiments, the spacer is connected, ** -NH-(CH2CH2O)m -CH2CH2-C(=O)-, or ** -NH-(CH2) m It is -O-.

[0162] In some embodiments, m is 2, 4, 6, or 8.

[0163] In some embodiments, the conjugator is formula (IIa1), (IIa2), (IIa3), or (IIIa) [ka] It holds.

[0164] In some embodiments, the severable body is given by the following formula: [ka] Take one of them.

[0165] In some embodiments, the payload is expressed by the following formula: [ka] It is one of the residues.

[0166] In some embodiments, the payload is [ka] That is the case.

[0167] In some embodiments, the compound is [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (wherein Ab is 6E7 or its antigen-binding fragment); or [ka] (In the formula, Ab is ifinatamab or its antigen-binding fragment.) Alternatively, it may be any of the pharmaceutically acceptable salts, tautomers, solvates, or stereoisomers mentioned above.

[0168] In some embodiments, the compound is [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (wherein Ab is 6E7 or its antigen-binding fragment); or [ka] (In the formula, Ab is ifinatamab or its antigen-binding fragment.) Alternatively, it may be any of the pharmaceutically acceptable salts, tautomers, solvates, or stereoisomers mentioned above.

[0169] In one embodiment, x is 1 to 15 (inclusive). In one embodiment, x is 1 to 12. In one embodiment, x is 1 to 10. In one embodiment, x is 2 to 10. In one embodiment, x is 3.5 to 10. In one embodiment, x is 3.5 to 8. In one embodiment, x is 3.5 to 6. In one embodiment, x is 3.5 to 4.5.

[0170] Binder This specification provides, for example, binders (BA or Ab) for use in the ADC described herein.

[0171] The compound of formula (I) may include any of the BAs described herein.

[0172] In some embodiments, BA is an antibody or its antigen-binding fragment, for example, a humanized, chimeric, or human antibody or its antigen-binding fragment.

[0173] In some embodiments, the antibody or its antigen-binding fragment is a monoclonal antibody, a chimeric antibody, a humanized antibody, a manipulated human antibody, a single-chain antibody (scFv), a Fab fragment, a Fab' fragment, or an F(ab')2 fragment.

[0174] In some embodiments, BA is B7H3, cytokeratin 15, PTK7, HER3, HER2, CD7, CD19, CD20, CD22, CD25, CD27, CD30, CD33, CD37, CD38, CD46, CD70, CD71, CD74, CD79b, CD123, CD138, CD142, CD166, CD205, CD228, CCR2, CA6, p-cadherin, CEA, CEACAM5, It is an antibody or antigen-binding fragment that binds to one or more of the following: C4.4a, DLL3, EGFR, EGFRVIII, ENPP3, EphA2, EphrinA, FLOR1, FGFR2, GCC, cKIT, LIV1, LY6E, MSLN, MUC16, NaPi2b, Nectin4, gpNMB, PSMA, SLITRK6, STEAP1, TROP2, 5T4, SSEA4, GloboH, Gb5, STn, Tn, B7H3, BCMA, MUC1, cMet, ROR1, MSLn, FRa, CLDN18.2, CLDN6, PTK7, Axl, FGFR2b, CLL1, CCR7, GPC1, GPC3, ISAC, CDCP1, ITGB6, ADAM9, or CD45-iADC.

[0175] In some embodiments, BA is an anti-human B7H3 antibody, an anti-CLL1 antibody, an anti-PTK7 antibody, an anti-HER3 antibody, an anti-HER2 antibody, or any of the aforementioned antigen-binding fragments. In some embodiments, the anti-human B7H3 antibody is ifinatamab. In some embodiments, the anti-CLL1 antibody is 6E7. In some embodiments, the anti-PTK7 antibody is cofetuzumab. In some embodiments, the anti-HER3 antibody is patrizumab. In some embodiments, the anti-HER2 antibody is trastuzumab.

[0176] In some embodiments, the antibody or its antigen-binding fragment specifically binds to human B7H3. In some embodiments, the antibody or its antigen-binding fragment is ifinatamab.

[0177] In some embodiments, the antibody or its antigen-binding fragment specifically binds to CLL1. In some embodiments, the antibody or its antigen-binding fragment is 6E7.

[0178] In some embodiments, the antibody or its antigen-binding fragment specifically binds to PTK7. In some embodiments, the antibody or its antigen-binding fragment is cofetuzumab.

[0179] Appearance 2

[0180] In this specification, compounds of formula (XI): [ka] or a pharmaceutically acceptable salt, tautomer, solvate, or stereoisomer thereof (in the formula, BA is a binder selected from humanized, monoclonal, chimeric, or human antibodies or their antigen-binding fragments; The conjugator is given by formula (II) or (III): [ka] Having; U is a bond, heteroarylene, or arylene; V represents a bond or -C≡C-(CH2) n -and; n is an integer between 0 and 10 (including both ends); W2 is -C(=O)-, -NH-, or -O-; RG is [ka] , succinimido-3-yl-N)-, or [ka] and; RS is -NR 1a R 1b and; R 1a and R1b Each of these independently represents H or substituted or unsubstituted C. 1-4 It is alkyl; RE stands for bond, -O-, -OC(=O)-, -OC(=O)NR 6 -, -NHC(=O)NR 6 -, -OS(=O)2NR 6 -, -NHS(=O)2NR 6 -, or -OC(=O)NHS(=O)2NR 6 -and; R 6 is H or substituted or unsubstituted C 1-4 It is alkyl; W3 is -C(=O)-, -NH-, or -O-; Each of s and t is independently 0, 1, or 2; [ka] This indicates a covalent bond site within the compound. * indicates a connection where the conjugator connects to the BA; Spacers are used for bonding, ** -NH-(CH2CH2O) m -CH2CH2-C(=O)-, ** -C(=O)-(CH2CH2O) m -CH2CH2-NH-, ** -(CH2) m -C(=O)-, ** -CH2-C(=O)-NH-(CH2) m -C(=O)-, ** -(CH2CH2O) m -CH2CH2-C(=O)-, ** -C(=O)-(CH2CH2O) m -CH2CH2-NH-, ** -CH[-(CH2) m -COOH]-C(=O)-, ** -CH2-C(=O)-NH-(CH2) m -C(=O)-NH-(CH2) m -C(=O)-, ** -C(=O)-(CH2) m -C(=O)-,** -NH-(CH2) m -C(=O)-, or ** -NH-(CH2) m -O- is; m is an integer between 1 and 12 (including both ends); ** indicates a coupling where the spacer connects to the conjugator; The cleavable bodies are given by formulas (IVa'), (IVb'), (IVc'), (Va'), (Vb'), (VIa'), (VIb'), (VIIa'), or (VIIb'): [ka] Having; Su is the sugar portion; Each R 2 These are, independently, hydrogen, halogen, substituted or unsubstituted C 1-4 It is alkyl, -CN, or -NO2; # indicates a connection where the severable part is attached to the branched part; The branched fields are given by equations (XIIa), (XIIb), (XIIc), and (XIId): [ka] Selected from; Each of p and q is independently 1, 2, 3, or 4; The hydrophilic material is -NH-(CH2CH2O) a -(CH2) b CH3, -N(CH3)-((CH2) c C(=O)N(CH3)) a -(CH2) c C(=O)NH2, -C(=O)-(CH2CH2O) a -(CH2) b CH3, -C(=O)-(CH2CH2O) a -(CH2) b NHC(=O)(CH2)3N(CH3)2(CH2)3S(=O)2OH, -NH-(CH2CH2O) a -(CH2) b NHC(=O)(CH2)3N(CH3)2(CH2)3S(=O)2OH, [ka] and; a is an integer between 1 and 18 (including both ends); b is 0, 1, or 2; c is 1, 2, 3, or 4; The payload is a payload residue; x is between 1 and 15 (including both ends). It will be provided.

[0181] In some embodiments, the conjugator has formula (II).

[0182] In some embodiments, U is arylene.

[0183] In some embodiments, U is phenylene.

[0184] In some embodiments, U is [ka] That is the case.

[0185] In some embodiments, U is an arylene and V is a bond.

[0186] In some embodiments, U is arylene and V is -C≡C-(CH2) n - is

[0187] In some embodiments, U is arylene and V is -C≡C-(CH2)3-.

[0188] In some embodiments, U is a heteroarylene.

[0189] In some embodiments, U is a divalent pyrimidine ring.

[0190] In some embodiments, U is [ka] That is the case.

[0191] In some embodiments, U is a heteroarylene and V is a bond.

[0192] In some embodiments, U is a heteroarylene and V is -C≡C-(CH2) n - is

[0193] In some embodiments, U is a heteroarylene and V is -C≡C-(CH2)3-.

[0194] In some embodiments, U is a bond.

[0195] In some embodiments, U is a bond and V is a bond.

[0196] In some embodiments, U is a bond and V is -C≡C-(CH2) n - is

[0197] In some embodiments, U is a bond and V is -C≡C-(CH2)3-.

[0198] In some embodiments, W2 is -C(=O)-.

[0199] In some embodiments, the conjugator has formula (III).

[0200] In some embodiments, RS is -NH2 or -N(CH3)2.

[0201] In some embodiments, RS is -NH2.

[0202] In some embodiments, RE is -OC(=O)NH-.

[0203] In some embodiments, RG is [ka] That is the case.

[0204] In some embodiments, RG is [ka] , -(succinimido-3-yl-N)-, or [ka] That is the case.

[0205] In some embodiments, RG is [ka] That is the case.

[0206] A specified RG as described herein, for example, [ka] It can be induced to undergo a ring-opening process by conjugation with BA. During such a process, the maleimide structure formed by the conjugation of RG and BA, for example, [ka] It undergoes ring opening. It is understood that there are two possible positional isomers that may result from such ring opening of the maleimide structure. For example, [ka] It undergoes ring opening. [ka] It may form, [ka] It undergoes ring opening. [ka] This can form. For simplicity, unless otherwise indicated, both possible ring-opening positional isomers are encompassed by the illustration of a single ring-opening positional isomer herein. Thus, unless otherwise indicated, the illustration of a single ring-opening positional isomer [ka] teeth, [ka] This includes, and similarly, illustrations of ring-opening positional isomers. [ka] teeth, [ka] This includes. In some embodiments, a mixture of ring-opening positional isomers exists. In some embodiments, a single ring-opening positional isomer exists.

[0207] In some embodiments, s and t are independently 0 or 2.

[0208] In some embodiments, W3 is -C(=O)-.

[0209] In some embodiments, the spacer is connected, ** -NH-(CH2CH2O) m -CH2CH2-C(=O)-, or ** -C(=O)-(CH2CH2O) m It is -CH2CH2-NH-. In some embodiments, the spacer is a bond.

[0210] In some embodiments, m is 2, 4, 6, or 8.

[0211] In some embodiments, the conjugator is given by formula (IIa1), (IIa2), (IIa3), (IIa4), (IIa5), or (IIIa): [ka] It holds.

[0212] In some embodiments, the severable body is given by the following formula: [ka] Take one of them.

[0213] In some embodiments, the severable body is given by formula (IVa'1), (IVc'1), or (Va'1): [ka] It holds.

[0214] In some embodiments, the hydrophilic material is -NH-(CH2CH2O) a -(CH2) b CH3, -N(CH3)-((CH2) c C(=O)N(CH3)) a -(CH2) c C(=O)NH2, -C(=O)-(CH2CH2O) a -(CH2) b CH3, -C(=O)-(CH2CH2O) a -(CH2) b NHC(=O)(CH2)3N(CH3)2(CH2)3S(=O)2OH, [ka] That is the case.

[0215] In some embodiments, a is 4, 8, 11, or 12.

[0216] In some embodiments, b is 0 or 2.

[0217] In some embodiments, c is 1.

[0218] In some embodiments, a is 12 and b is 0.

[0219] In some embodiments, the hydrophilic material is -NH-(CH2CH2O) 12 -CH3, -N(CH3)-((CH2)C(=O)N(CH3)) 11 -(CH2) c C(=O)NH2, -C(=O)-(CH2CH2O) 12 -CH3, -C(=O)-(CH2CH2O)8-(CH2)2NHC(=O)(CH2)3N(CH3)2(CH2)3S(=O)2OH, [ka] That is the case.

[0220] In some embodiments, each of p and q is independently 2 or 4.

[0221] In some embodiments, the payload is [ka] It is a residue.

[0222] In some embodiments, the payload is [ka] That is the case.

[0223] In some embodiments, the compound is [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is ifinatamab or its antigen-binding fragment); [ka] (wherein Ab is cofetuzumab or its antigen-binding fragment); or [ka] (In the formula, Ab is cofetuzumab or its antigen-binding fragment.) Alternatively, it may be any of the pharmaceutically acceptable salts, tautomers, solvates, or stereoisomers mentioned above.

[0224] In some embodiments, the compound is [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (In the formula, Ab is 6E7 or its antigen-binding fragment); [ka] (wherein Ab is ifinatamab or its antigen-binding fragment); [ka] (wherein Ab is cofetuzumab or its antigen-binding fragment); or [ka] (In the formula, Ab is cofetuzumab or its antigen-binding fragment.) Alternatively, it may be any of the pharmaceutically acceptable salts, tautomers, solvates, or stereoisomers mentioned above.

[0225] The binder may be BA or Ab, which may be any BA as described above in Embodiment 1.

[0226] X may be as described above with respect to Embodiment 1.

[0227] Appearance 3

[0228] In this specification, compounds of formula (II): [ka] or a pharmaceutically acceptable salt, tautomer, solvate, or stereoisomer thereof (in the formula, The conjugator is given by formula (I-II) or (I-III): [ka] Having; U is a bond, heteroarylene, or arylene; V represents a bond or -C≡C-(CH2) n -and; n is an integer between 0 and 10 (including both ends); W2 is -C(=O)-, -NH-, or -O-; RG is [ka] and; RS is -NR 1a R 1b and; R 1a and R 1b Each of these independently represents H or substituted or unsubstituted C. 1-4 It is alkyl; RE stands for bond, -O-, -OC(=O)-, -OC(=O)NR 6 -, -NHC(=O)NR 6 -, -OS(=O)2NR 6 -, -NHS(=O)2NR 6 -, or -OC(=O)NHS(=O)2NR 6 -and; R 6 is H or substituted or unsubstituted C 1-4 It is alkyl; W3 is -C(=O)-, -NH-, or -O-; Each of s and t is independently 0, 1, or 2; [ka] This indicates a covalent bond site within the compound. Spacers are used for bonding, ** -NH-(CH2CH2O)m -CH2CH2-C(=O)-, ** -(CH2) m -C(=O)-, ** -CH2-C(=O)-NH-(CH2) m -C(=O)-, ** -(CH2CH2O) m -CH2CH2-C(=O)-, ** -CH[-(CH2) m -COOH]-C(=O)-, ** -CH2-C(=O)-NH-(CH2) m -C(=O)-NH-(CH2) m -C(=O)-, ** -C(=O)-(CH2) m -C(=O)-, ** -NH-(CH2) m -C(=O)-, or ** -NH-(CH2) m -O- is; m is an integer between 1 and 12 (including both ends); ** indicates a coupling where the spacer connects to the conjugator; The cleavable bodies are given by formulas (IVa), (IVb), (IVc), (Va), (Vb), (VIa), (VIb), (VIIa), or (VIIb): [ka] Having; Su is the sugar portion; Each R 2 These are, independently, hydrogen, halogen, substituted or unsubstituted C 1-4 It is alkyl, -CN, or -NO2; *** indicates a connection where the severable body connects to the spacer; The payload is the payload residue. It will be provided.

[0229] In some embodiments, the conjugator has formula (I-II).

[0230] In some embodiments, U is arylene.

[0231] In some embodiments, U is phenylene.

[0232] In some embodiments, U is [ka] That is the case.

[0233] In some embodiments, U is an arylene and V is a bond.

[0234] In some embodiments, U is arylene and V is -C≡C-(CH2) n - is

[0235] In some embodiments, U is arylene and V is -C≡C-(CH2)3-.

[0236] In some embodiments, U is a heteroarylene.

[0237] In some embodiments, U is a divalent pyrimidine ring.

[0238] In some embodiments, U is [ka] That is the case.

[0239] In some embodiments, U is a heteroarylene and V is a bond.

[0240] In some embodiments, U is a heteroarylene and V is -C≡C-(CH2) n - is

[0241] In some embodiments, U is a heteroarylene and V is -C≡C-(CH2)3-.

[0242] In some embodiments, U is a bond.

[0243] In some embodiments, U is a bond and V is a bond.

[0244] In some embodiments, U is a bond and V is -C≡C-(CH2) n - is

[0245] In some embodiments, U is a bond and V is -C≡C-(CH2)3-.

[0246] In some embodiments, W2 is -C(=O)-.

[0247] In some embodiments, the conjugator has formula (I-III).

[0248] In some embodiments, RS is -NH2 or -N(CH3)2.

[0249] In some embodiments, RS is -NH2.

[0250] In some embodiments, RE is -OC(=O)NH-.

[0251] In some embodiments, RG is [ka] That is the case.

[0252] In some embodiments, RG is [ka] That is the case.

[0253] In some embodiments, each of s and t is 2.

[0254] In some embodiments, W3 is -C(=O)- or -NH-.

[0255] In some embodiments, the spacer is connected, ** -NH-(CH2CH2O) m -CH2CH2-C(=O)-, or ** -C(=O)-(CH2CH2O) m It is -CH2CH2-NH-.

[0256] In some embodiments, m is 2, 4, 6, or 8.

[0257] In some embodiments, the conjugator is (I-IIa1) or (I-IIIa1): [ka] It holds.

[0258] In some embodiments, the severable body is given by the following formula: [ka] Take one of them.

[0259] In some embodiments, the payload is [ka] [ka] It is a residue.

[0260] In some embodiments, the payload is [ka] That is the case.

[0261] In some embodiments, the compound is [ka] [ka] [ka] or a pharmaceutically acceptable salt, tautomer, solvate, or stereoisomer thereof.

[0262] Pattern 4

[0263] In this specification, compounds of formula (XV): [ka] or a pharmaceutically acceptable salt, tautomer, solvate, or stereoisomer thereof (in the formula, The conjugator is given by formula (I-II) or (I-III): [ka] It has, U is a bond, heteroarylene, or arylene; V represents a bond or -C≡C-(CH2) n -and; n is an integer between 0 and 10 (including both ends); W2 is -C(=O)-, -NH-, or -O-; RG is [ka] and; RS is -NR 1a R 1b and; R 1a and R 1b Each of these independently represents H or substituted or unsubstituted C. 1-4 It is alkyl; RE stands for bond, -O-, -OC(=O)-, -OC(=O)NR 6 -, -NHC(=O)NR 6 -, -OS(=O)2NR 6 -, -NHS(=O)2NR 6 -, or -OC(=O)NHS(=O)2NR6 -and; R 6 is H or substituted or unsubstituted C 1-4 It is alkyl; W3 is -C(=O)-, -NH-, or -O-; Each of s and t is independently 0, 1, or 2; [ka] This indicates a covalent bond site within the compound. Spacers are used for bonding, ** -NH-(CH2CH2O) m -CH2CH2-C(=O)-, ** -(CH2) m -C(=O)-, ** -CH2-C(=O)-NH-(CH2) m -C(=O)-, ** -(CH2CH2O) m -CH2CH2-C(=O)-, ** -CH[-(CH2) m -COOH]-C(=O)-, ** -CH2-C(=O)-NH-(CH2) m -C(=O)-NH-(CH2) m -C(=O)-, ** -C(=O)-(CH2) m -C(=O)-, ** -NH-(CH2) m -C(=O)-, or ** -NH-(CH2) m -O- is; m is an integer between 1 and 12 (including both ends); ** indicates a coupling where the spacer connects to the conjugator; The cleavable bodies are given by formulas (IVa'), (IVb'), (IVc'), (Va'), (Vb'), (VIa'), (VIb'), (VIIa'), or (VIIb'): [ka] Having; Su is the sugar portion; Each R 2 These are, independently, hydrogen, halogen, substituted or unsubstituted C 1-4 It is alkyl, -CN, or -NO2; # indicates a connection where the severable part is attached to the branched part; The branched fields are given by equations (XIIa), (XIIb), (XIIc), and (XIId): [ka] Selected from; Each of p and q is independently 1, 2, 3, or 4; The hydrophilic material is -NH-(CH2CH2O) a -(CH2) b CH3, -N(CH3)-((CH2) c C(=O)N(CH3)) a -(CH2) c C(=O)NH2, -C(=O)-(CH2CH2O) a -(CH2) b CH3, -C(=O)-(CH2CH2O) a -(CH2) b NHC(=O)(CH2)3N(CH3)2(CH2)3S(=O)2OH, -NH-(CH2CH2O) a -(CH2) b NHC(=O)(CH2)3N(CH3)2(CH2)3S(=O)2OH, [ka] and; a is an integer between 1 and 18 (including both ends); b is 0, 1, or 2; c is 1, 2, 3, or 4; The payload is the payload residue. It will be provided.

[0264] In some embodiments, the conjugator has formula (I-II).

[0265] In some embodiments, U is arylene.

[0266] In some embodiments, U is phenylene.

[0267] In some embodiments, U is [ka] That is the case.

[0268] In some embodiments, U is an arylene and V is a bond.

[0269] In some embodiments, U is arylene and V is -C≡C-(CH2) n - is

[0270] In some embodiments, U is arylene and V is -C≡C-(CH2)3-.

[0271] In some embodiments, U is a heteroarylene.

[0272] In some embodiments, U is a divalent pyrimidine ring.

[0273] In some embodiments, U is [ka] That is the case.

[0274] In some embodiments, U is a heteroarylene and V is a bond.

[0275] In some embodiments, U is a heteroarylene and V is -C≡C-(CH2) n - is

[0276] In some embodiments, U is a heteroarylene and V is -C≡C-(CH2)3-.

[0277] In some embodiments, U is a bond.

[0278] In some embodiments, U is a bond and V is a bond.

[0279] In some embodiments, U is a bond and V is -C≡C-(CH2) n - is

[0280] In some embodiments, U is a bond and V is -C≡C-(CH2)3-.

[0281] In some embodiments, W2 is -C(=O)-.

[0282] In some embodiments, the conjugator has formula (I-III).

[0283] In some embodiments, RS is -NH2 or -N(CH3)2.

[0284] In some embodiments, RS is -NH2.

[0285] In some embodiments, RE is -OC(=O)NH-.

[0286] In some embodiments, RG is [ka] That is the case.

[0287] In some embodiments, RG is [ka] That is the case.

[0288] In some embodiments, each of s and t is 2.

[0289] In some embodiments, the spacer is connected, ** -NH-(CH2CH2O) m-CH2CH2-C(=O)-, ** -C(=O)-(CH2CH2O) m -CH2CH2-NH-, or ** -NH-(CH2) m -O-. In some embodiments, the spacer is bonded, ** -NH-(CH2CH2O) m -CH2CH2-C(=O)-, or ** -NH-(CH2) m It is -O-.

[0290] In some embodiments, m is 2, 4, 6, or 8.

[0291] In some embodiments, the conjugator is given by formula (I-IIa1) or (I-IIIa1): [ka] It holds.

[0292] In some embodiments, the severable body is given by the following formula: [ka] Take one of them.

[0293] In some embodiments, the payload is [ka] It is a residue.

[0294] In some embodiments, the payload is [ka] That is the case.

[0295] In some embodiments, a is 12 and b is 0.

[0296] In some embodiments, the compound is [ka] [ka] [ka] or a pharmaceutically acceptable salt, tautomer, solvate, or stereoisomer thereof.

[0297] How to create a conjugate This specification provides a method for preparing a conjugate by contacting a conjugator (BA) with a conjugator-linker-payload compound under conditions suitable for forming a bond between the conjugator and the conjugator-linker-payload compound. The reaction conditions can be any suitable reaction conditions known in the art. The conjugator is an antibody, and the bond can form an antibody-drug conjugate.

[0298] Examples of such reactions are provided in the following embodiments.

[0299] In some embodiments, a method for preparing a conjugate comprises treating or contacting a compound with a binder under coupling conditions. The compound may include a reactive linker bound to at least one payload. The compound may be either a linker or a platform compound disclosed herein.

[0300] Pharmaceutical composition Furthermore, this specification provides compositions comprising a pharmaceutical composition containing the ADC shown herein. In some embodiments, the composition (e.g., the pharmaceutical composition) further comprises pharmaceutically acceptable excipients.

[0301] The pharmaceutical compositions according to this disclosure may be prepared in the form of lyophilized formulations or aqueous solutions by mixing an antibody-drug conjugate of desired purity with one or more pharmaceutically acceptable carriers (Remington's Pharmaceutical Sciences 16th edition, Osol, A. Ed. (1980)). The pharmaceutically acceptable carriers are typically nontoxic to the recipient at the dosage and concentration used and include buffers, e.g., phosphates, citrates, and other organic acids; antioxidants including ascorbic acid and methionine; preservatives (e.g., octadecyldimethylbenzylammonium chloride; hexamethonium chloride; benzalkonium chloride; benzethonium chloride; phenol, butyl, or benzyl alcohol; alkylparabens, e.g., methyl or propylparaben; catechol; resorcinol; cyclohexanol; 3-pentanol; and m-cresol); low molecular weight (less than about 10 residues) polypeptides; proteins, Examples of pharmaceutically acceptable carriers herein include, but are not limited to, serum albumin, gelatin, or immunoglobulin; hydrophilic polymers, such as polyvinylpyrrolidone; amino acids, such as glycine, glutamine, asparagine, histidine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates (including glucose, mannose, or dextrin); chelating agents, such as EDTA; sugars, such as sucrose, mannitol, trehalose, or sorbitol; salt-forming counterions, such as sodium; metal complexes (e.g., Zn-protein complexes); and / or nonionic surfactants, such as polyethylene glycol (PEG). Exemplary pharmaceutically acceptable carriers herein also include interstitial drug dispersants, such as soluble neutral-active hyaluronidase glycoproteins (sHASEGP), such as human soluble PH-20 hyaluronidase glycoprotein, such as rHuPH20 (HYLENEX®, Baxter International, Inc.). Certain exemplary sHASEGPs and methods of use, including rHuPH20, are described in U.S. Patent Nos. US7,871,607 and 2006 / 0104968.In one embodiment, sHASEGP is combined with one or more additional glycosaminoglycansases, such as chondroitinase.

[0302] How to use conjugates In some embodiments, methods for treating a disease or disorder (e.g., cancer) in an object (e.g., a patient) that requires treatment of the disease or disorder (e.g., cancer) are shown herein, the methods comprising administering a therapeutically effective amount of the conjugate disclosed herein to the patient.

[0303] The conjugates disclosed herein may be administered by any preferred means, including parenteral, intrapulmonary, intranasal, and, if desired for local treatment, intrafocal administration. Parenteral administration includes intramuscular, intravenous, intra-arterial, intraperitoneal, or subcutaneous administration. Dosage may be by any preferred route, e.g., injection, e.g., intravenous or subcutaneous injection, depending in part whether the administration is simple or chronic. Various dosing schedules, including but not limited to single or multiple doses, bolus doses, and pulse infusions over various time points, are contemplated herein.

[0304] The conjugates of this disclosure may be formulated, administered, and given in a manner consistent with good medical practice. Considerations in this context include the specific disorder being treated, the specific mammal being treated, the clinical condition of the individual patient, the cause of the disorder, the site of drug delivery, the method of administration, the scheduling of administration, and other factors known to the medical practitioner. [Examples]

[0305] The following examples are intended to be illustrative and should not be considered limiting in any way. Unless otherwise specified, the experimental methods in the examples described below are conventional methods. Unless otherwise specified, reagents and substances are commercially available. All solvents and chemicals used were analytical grade or of chemical purity. Solvents were redistilled before use. Anhydrous solvents were prepared according to standard or reference methods. Silica gel (100-200 mesh) for column chromatography and silica gel (GF254) for thin-layer chromatography (TLC) are commercially available from Tsingdao Haiyang Chemical Co., Ltd. or Yantai Chemical Co., Ltd. of China; unless otherwise specified, all were eluted with petroleum ether (60-90°C) / ethyl acetate (v / v) and visualized with a solution of molybdenum phosphate in iodine or ethanol. Unless otherwise specified, all extraction solvents were dried with anhydrous Na2SO4. 1 ¹H NMR spectra were recorded using Bruck-400 and Varian 400MR nuclear magnetic resonance spectrometers with TMS (tetramethylsilane) as an internal standard. Coupling constants were given in Hertz. Peaks were reported as singlelines (s), doublelines (d), triplelines (t), quadruplines (q), quintuplines (p), hexatlines (h), heptuplines (hept), multilines (m), or combinations thereof; br represents broadness. LC / MS data were recorded using Agilent 1100, 1200 high-performance liquid chromatography-ion trap mass spectrometers (LC-MSD Trap) equipped with diode array detectors (DAD) detecting at 214 nm and 254 nm, and an ion trap (ESI source). All compound names except reagents were generated using ChemDraw® 18.0.

[0306] For the sake of brevity, certain abbreviations are used herein. One example is the one-letter abbreviation for amino acids. The amino acids and their corresponding three-letter and one-letter abbreviations are as follows: [Table 19]

[0307] In the following examples, the following abbreviations are used: [Table 20]

[0308] UPLC analysis method Method A: Mobile phase A: 0.1% FA in water, B: MeCN; Gradient: Maintain 10% B for 0.2 minutes, 10% to 95% B for 5.8 minutes, maintain 95% B for 0.5 minutes; Flow rate: 0.6 mL / min; Column: ACQUITY UPLC(registered trademark) BEH C18 1.7 μm.

[0309] Method B: Mobile phase A: 0.1% FA in water, B: MeCN; Gradient: Maintain 10% B for 0.5 minutes, 10%~90% B for 2.5 minutes, maintain 90% B for 0.2 minutes; Flow rate: 0.6 mL / min; Column: ACQUITY UPLC(registered trademark) BEH C18 1.7 μm.

[0310] Method C: Mobile phase A: 0.1% FA in water, B: MeCN; Gradient: Maintain 10% B for 0.2 min, 10% to 90% B for 1.3 min, maintain 90% B for 0.3 min; Flow rate: 0.6 mL / min; Column: ACQUITY UPLC(registered trademark) BEH C18 1.7 μm.

[0311] Example Int-1 [ka] Step 1: (2S,3S,4S,5R,6S)-2-(methoxycarbonyl)-6-(2-nitro-4-((((4-nitrophenoxy)carbonyl)oxy)methyl)phenoxy)tetrahydro-2H-pyran-3,4,5-triyltriacetate(Int-1b)

[0312] A solution of Int-1a (3.00 g, 6.18 mmol) (commercially available) in anhydrous DMF (60 mL) was mixed with bis(4-nitrophenyl)carbonate (2.85 g, 9.271 mmol) and DIPEA (2.18 mL, 12.36 mmol) at 0°C and stirred at room temperature for 3 hours. The reaction was quenched with aqueous AcOH, extracted with SiO (100 mL x 3), and washed with water (100 mL) and brine (100 mL). The organic phase was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (SiO / petroleum ether = 30 / 70) to obtain Int-1b (4 g, 99.5% yield) as an off-white solid. MS(ESI) m / z: 673.2[M+Na] + .

[0313] Step 2: (2S,3R,4S,5S,6S)-2-(4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-nitrophenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(Int-1c)

[0314] To a solution of Int-1b (2.00 g, 3.08 mmol) and MMAE (2.45 g, 3.38 mmol) in anhydrous DMF (30 mL), HOBt (126 mg, 0.92 mmol) and DIPEA (1.66 mL, 9.22 mmol) were added and the mixture was stirred at room temperature for 16 hours. The organic phase was concentrated and purified by silica gel column chromatography (CH2Cl2 / MeOH = 90 / 10) to obtain Int-1c (3.50 g, 92.6% yield) as a white solid. MS(ESI) m / z: 1229.7[M+H] + .

[0315] Step 3: (2S,3R,4S,5S,6S)-2-(2-amino-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(Int-1)

[0316] AcOH (0.99 mL, 17.08 mmol) was added at 0°C to a solution of Zn powder (6.96 g, 105.34 mmol) in EtOH (10.0 mL) and water (1 mL), and the mixture was stirred at 0°C for 1 hour. Then, Int-1c (3.50 g, 2.85 mmol) in EtOH (10 mL) was gradually added to the above mixture at 0°C, and the mixture was stirred at room temperature for 4 hours. The resulting solution was filtered and washed with EtOH (100 mL). The filtrate was concentrated to remove EtOH. Water (100 mL) was added to the residue and extracted with ELISA (100 mL × 3). The combined organic layer was dried over Na2SO4 and concentrated. The residue was purified by silica gel column chromatography (CH2Cl2 / MeOH = 90 / 10) to obtain Int-1 (3.30 g, 96.6% yield) as a white solid. MS(ESI)m / z:1199.8[M+H] + .

[0317] Example Int-2 [ka] Int-2 was synthesized following a modified synthesis procedure from the reference (Tetrahedron Letters 54(2013)349-3495).

[0318] Example Int-3 [ka] Step 1: Methyl(R)-3-(((benzyloxy)carbonyl)amino)-4-((tert-butoxycarbonyl)amino)butanoate(Int-3b)

[0319] Int-3a (2.00 g, 5.68 mmol) and K2CO3 (863 mg, 6.24 mmol) were added to DMF (10 mL), followed by the dropwise addition of CH3I (1.61 g, 11.35 mmol) at 0°C. The resulting mixture was stirred at 0°C for 20 minutes, then warmed to 25°C and stirred further at 25°C for 60 minutes. The reaction process was monitored by TLC (petroleum ether / RINKAN) and LC-MS. After complete reaction, the reaction mixture was diluted with RINKAN (80 mL) and washed with brine (30 mL x 3) and H2O (30 mL x 2). The organic layer was dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to obtain the methyl ester of Int-3b (2.08 g, quantitatively) as a pale yellow solid. MS(ESI) m / z: 267.2[M-Boc+H] + .

[0320] Step 2: Benzyl tert-butyl (4-hydroxybutan-1,2-diyl)(R)-dicarbamate(Int-3c)

[0321] Int-3b (1.00 g, 2.73 mmol) was dissolved in MeOH (15 mL), followed by the addition of LiBH4 (2 M stock solution in THF, 6.80 mL) at 0°C. The resulting mixture was stirred at 25°C for 2 hours. The reaction process was monitored by LC-MS and TLC. After complete reaction, the reaction was quenched by adding saturated aqueous solution NH4Cl (10 mL). The reaction mixture was diluted with H2O (80 mL) and extracted with HCl (50 mL x 3). The combined organic layer was washed with brine (40 mL x 2) and water (40 mL x 2), dried over anhydrous Na2SO4, filtered, concentrated under reduced pressure, and further purified by flash column chromatography (petroleum ether / HCl) to obtain Int-3c (760 mg, 82.3% yield) as a white solid. MS(ESI) m / z: 239.2[M-Boc+H] + .

[0322] Step 3: Benzyl tert-butyl (4-(((4-nitrophenoxy)carbonyl)oxy)butane-1,2-diyl)(R)-dicarbamate (Int-3d)

[0323] Int-3c (300 mg, 0.89 mmol) and bis(4-nitrophenyl)carbonate (405 mg, 1.33 mmol) were dissolved in DMF (5 mL), followed by the addition of DIEA (229 mg, 1.77 mmol). The resulting mixture was stirred at 25°C for 1.5 hours. After complete reaction, the reaction mixture was diluted with RINKAN (100 mL) and washed with brine (35 mL x 2) and water (35 mL x 2). The organic layer was dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to obtain Int-3d as a white solid (371 mg, 83.1% yield). MS(ESI)m / z: 404.4[M-Boc+H] + .

[0324] Step 4: (9H-Fluoren-9-yl)methyl tert-butyl (4-(((3-(dimethylamino)-3-oxopropyl)carbamoyl)oxy)butan-1,3-diyl)(S)-dicarbamate(Int-3e)

[0325] Int-3d (420 mg, 0.83 mmol) and 3-aminopropanoic acid (149 mg, 1.67 mmol) were dissolved in DMF (5 mL), followed by the addition of aqueous NaHCO3 (1 M, 5 mL). The resulting mixture was stirred at 25°C for 2.5 hours. After complete reaction, the reaction mixture was concentrated and purified by flash column chromatography (DCM / MeOH) to obtain Int-3e as a pale yellow solid (365 mg, 96.5% yield). MS(ESI) m / z: 354.4 [M-Boc+H] + .

[0326] Step 5: (R)-7-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)-2,2-dimethyl-4,11-dioxo-3,10-dioxa-5,12-diazapentadecane-15-acid(Int-3f)

[0327] Int-3e (360 mg, 0.79 mmol) was dissolved in MeOH (18 mL), followed by the addition of Pd / C (infiltration base, 108 mg). The resulting mixture was stirred at room temperature under H2 (15 psi) for 2 hours. After complete reaction, the reaction mixture was filtered and concentrated under reduced pressure to obtain Int-3f as a clear syrup (252 mg, 99.4% yield). The crude product was used directly in the next step without purification. MS(ESI) m / z: 320.3[M+H] + .

[0328] Step 6: (R)-7-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)-2,2-dimethyl-4,11-dioxo-3,10-dioxa-5,12-diazapentadecane-15-acid(Int-3)

[0329] Int-3f (250 mg, 0.78 mmol) and Int-3g (243 mg, 1.57 mmol) were dissolved in a mixed solvent of ACN (8 mL) and aqueous NaHCO3 (1 M, 16 mL). The resulting mixture was stirred at 0°C for 1 hour, and then stirred further at 25°C until the reaction was complete. The reaction mixture was then acidified with aqueous KHSO4 (20 mL) and extracted with siRNA (35 mL x 3). The combined organic layer was dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to obtain a yellow oil as the crude product. This was purified by flash column chromatography to obtain Int-3 (280 mg, 89.6% yield) as a white solid. MS(ESI)m / z:422.3[M+Na] + .

[0330] 1 H NMR (400 MHz, d6-DMSO) δ 12.48 (s, 1H), 7.03-7.01 (m, 2H), 6.99 (s, 2H), 4.08-4.03 (m, 3H), 3.86-3.83 (m, 2H), 3.14-3.11 (m, 2H), 2.35 (t, J=7.2 Hz, 2H), 2.16-2.09 (m, 1H), 1.91-1.84 (m, 1H), 1.32 (s, 9H).

[0331] Example Int-4 [ka] Step 1: Methyl 4-(5-(methylthio)-1,2,4-thiadiazole-3-yl)benzoate (Int-4c)

[0332] Compound Int-4b (109.72 mg, 0.568 mmol), K2CO3 (168 mg, 0.947 mmol), and Pd(dppf)Cl2.DCM (34.6 mg, 0.047 mmol) were added to a solution of compound Int-4a (100 mg, 0.47 mmol) in toluene (4 mL) and H2O (1 mL). The mixture was prepared by adding 110 ml of compound Int-4b (109.72 mg, 0.568 mmol), K2CO3 (168 mg, 0.947 mmol), and Pd(dppf)Cl2.DCM (34.6 mg, 0.047 mmol). o The mixture was stirred in 1C under an N2 atmosphere for 3 hours. The mixture was filtered through a Celite pad, diluted with EA (100 mL), and washed with brine (50 mL x 4). The organic layer was dried over anhydrous Na2SO4, filtered, and concentrated. The crude product was purified by flash column chromatography (eluted at PE:EA = 0-40%). Compound Int-4c (56 mg, 44.4% yield) was obtained as an off-white solid. MS(ESI) m / z: 267.1[M+H] + .

[0333] Step 2: 4-(5-(methylthio)-1,2,4-thiadiazole-3-yl)benzoic acid (Int-4d)

[0334] A solution of compound Int-4c (54 mg, 0.20 mmol) in MeOH (3 mL) and H2O (1 mL) was mixed with LiOH (17 mg, 0.41 mmol). The mixture was stirred at room temperature for 2 hours. The mixture was adjusted to pH 7 and purified by prep-HPLC (FA conditions) to obtain compound Int-4d (36 mg, 70.3% yield) as a white solid. MS(ESI) m / z: 253.1[M+H] + .

[0335] Step 3: 4-(5-(methylsulfonyl)-1,2,4-thiadiazole-3-yl)benzoic acid (Int-4)

[0336] m-CPBA (96 mg, 0.55 mmol) was added to a solution of compound Int-4d (35 mg, 0.14 mmol) in DCM (3 mL) and THF (3 mL). The mixture was stirred at room temperature for 16 hours. The mixture was concentrated and purified by prep-HPLC (method: column: XBridge Prep C18 OBD 5 μm 19*250 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min). Compound Int-4 (12 mg, 99% purity) was obtained as a white solid. MS(ESI) m / z: 284.8[M+H] + .

[0337] Example 1: Synthesis of conjugator-linker-payload Example 1-1 [ka] 1-1 was synthesized according to PCT Publication WO2017165851.

[0338] Examples 1-2 [ka] Step 1: (R)-38-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-2,2-dimethyl-4,32-dioxo-3,8,11,14,17,20,23,26,29-nonoxa-5,33-diazanonatricontan-39-acid(1-2c) [ka]

[0339] To a solution of 1-2b (350 mg, 0.19 mmol) (commercially available) in anhydrous DMF (5 mL), HATU (236 mg, 0.61 mmol) and DIPEA (0.23 mL, 1.29 mmol) were added and the mixture was stirred at room temperature for 20 minutes. 1-2a (264.5 mg, 0.71 mmol) (commercially available) was added to the mixture and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-2c (345 mg, 59.9% yield) as a colorless oil. MS(ESI) m / z: 892.8[M+H] + .

[0340] Step 2: (R)-33-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-1-amino-27-oxo-3,6,9,12,15,18,21,24-octaoxa-28-azatetratriacontane-34-acid(1-2d)trifluoroacetate [ka]

[0341] To a solution of 1-2c (180.0 mg, 0.20 mmol) in CH2Cl2 (6 mL), TFA (1.56 mL, 20.18 mmol) was added at 0°C and the mixture was stirred at 0°C for 30 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5 μm 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-2d (trifluoroacetate) (150 mg, 82.1% yield) as a white solid. MS(ESI) m / z: 792.8 [M+H] + .

[0342] Step 3: Benzyl 4-(dimethylamino)butanoate (1-2f) [ka]

[0343] TsOH.H2O (1.20 g, 6.26 mmol) was added to a solution of 1-2e (1.0 g, 5.97 mol) and benzyl alcohol (1.11 g, 10.14 mmol) in toluene (8 mL). The mixture was degassed for 10 minutes, N2 was refilled, and toluene (10 mL) was heated and refluxed to remove water using a Dean-Stark trap. The mixture was stirred at 130°C for 4 hours. After cooling the reaction mixture, the toluene phase was extracted four times with water. The pH of the aqueous phase was adjusted to pH 10 with 1N NaOH. The aqueous phase was then extracted with SiO2 (50 mL x 3). The organic layer was evaporated and purified by silica gel column chromatography (CH2Cl2 / MeOH = 90 / 10) to obtain 1-2f (910 mg, 68.9% yield) as a yellow oil. MS(ESI) m / z: 223.2[M+H] + .

[0344] Step 4: 3-((4-(benzyloxy)-4-oxobutyl)dimethylammonio)propane-1-sulfonate (1-2g) [ka]

[0345] 2,6-di-tert-butylpyridine (1.21 g, 6.17 mmol) was added to a solution of 1-2f (910 mg, 4.11 mol) and 1,3-propanesultone (761 mg, 6.17 mmol) in anhydrous DMF (10 mL). The mixture was stirred at 150 °C for 5 hours. After cooling, the reaction mixture was concentrated. The residue was partitioned between ethyl acetate (50 mL) and water (50 mL). The aqueous phase was then extracted three times with ethyl acetate, the pH of the solution was adjusted to 10 with NH3H2O, and further extracted with ethyl acetate (50 mL x 2). The aqueous solution was concentrated under reduced pressure and purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min). The fraction was lyophilized to obtain 1-2 g (510 mg, 36.1% yield) as a white solid. MS(ESI) m / z: 344.4[M+H] +

[0346] Step 5: 3-((3-carboxypropyl)dimethylammonio)propane-1-sulfonate (1-2h) [ka]

[0347] A solution of 1-2 g (510 mg, 1.49 mmol) in MeOH (10 mL) was treated with 100 mg of 10% Pd / C. The suspension was hydrogenated in a parr shaker at 40 psi for 2 hours at room temperature. The reaction mixture was filtered. The methanol solution was concentrated under reduced pressure to obtain 1-2 h (375 mg, crude) as a colorless oil, which was used directly in the next step. MS(ESI) m / z: 254.2 [M+H] + .

[0348] Step 6: 3-(dimethyl(4-oxo-4-(perfluorophenoxy)butyl)ammonio)propane-1-sulfonate(1-2i) [ka]

[0349] To a solution of 1-2h (260 mg, 1.03 mmol) in 0.1N HCl (2 mL) and MeCN (2 mL), pentafluorophenol (382 mg, 2.05 mmol) and EDCI (497 mg, 2.57 mmol) were added and the mixture was stirred at room temperature for 16 hours. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5 μm 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-2i (260 mg, 60.4% yield) as a colorless oil. MS(ESI) m / z: 420.4[M+H] + .

[0350] Step 7: (R)-5-carboxy-1-(9H-fluoren-9-yl)-43,43-dimethyl-3,11,39-trioxo-2,14,17,20,23,24,26,29,32,35-decaxa-4,10,38,43-tetraazahexatetracanthane-43-ium-46-sulfonate(1-2j) [ka]

[0351] To a solution of 1-2i (75 mg, 0.18 mmol) and 1-2d (72 mg, 0.09 mmol) in water (1 mL) and MeCN (1 mL), saturated NaHCO3 (0.5 mL) was added and the mixture was stirred at room temperature for 1 hour. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5 μm 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-2j (75 mg, 40.8% yield) as a white solid. MS(ESI) m / z: 1029.9 [M+H] + .

[0352] Step 8: (S)-5-((3-((5-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl (L)-2-(((2S,3R,4S,5S,6S)-6-carboxy-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)oxy)phenyl)amino)-3-oxopropyl)carbamoyl)-1-(9H-fluoren-9-yl)-43,43-dimethyl-3,11,39-trioxo-2,14,17,20,23,26,29,32,35-nonanoxa-4,10,38,43-tetraazahexatetracanthane-43-ium-46-sulfonate(1-2l) [ka]

[0353] To a solution of 1-2j (73 mg, 0.07 mmol) in anhydrous DMF (2 mL), HATU (33 mg, 0.085 mmol) and DIPEA (0.025 mL, 0.14 mmol) were added and the mixture was stirred at room temperature for 10 minutes. 1-2k (89.3 mg, 0.078 mmol) (synthesized according to PCT publication WO2017165851) was added to the mixture and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-2 L (90 mg, 59.1% yield) as a white solid. MS(ESI) m / z: 2140.5[M+H] + .

[0354] Step 9: (S)-42-amino-47-((5-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13- Dioxa-4,7,10-triazatetradecyl)-2-(((2S,3R,4S,5S,6S)-6-carboxy-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)oxy)phenyl)amino)-4,4-dimethyl-8,36,43,47-tetraoxo-12,15,18,21,24,27,30,33-octaoxa-4,9,37,44-tetraazaheptatetracontan-4-ium-1-sulfonate(1-2m) [ka]

[0355] To a solution of 1-2 L (90 mg, 0.042 mmol) in anhydrous DMF (2 mL), Et2NH (0.088 mL, 0.84 mmol) was added and stirred at room temperature for 30 minutes. 1-2 m (80 mg, 0.042 mmol, crude) was evaporated five times with toluene to remove H2O. MS(ESI)m / z:1918.2[M+H] + .

[0356] Step 10: (7S,10S)-10-((3-((5-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-(((2S ,3R,4S,5S,6S)-6-carboxy-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)oxy)phenyl)amino)-3-oxopropyl)carbamoyl)-7-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)-2,2,48,48-tetramethyl-4,8,16,44-tetraoxo-3,19,22,25,28,31,34,37,40-nonanoxa-5,9,15,43,48-pentazahenpentacontan-48-ium-51-sulfonate(1-2o) [ka]

[0357] To a solution of 1-2n (15 mg, 0.05 mmol) (synthesized according to PCT publication WO2017165851) in anhydrous DMF (2 mL), HATU (26 mg, 0.05 mmol) and DIPEA (0.015 mL, 0.083 mmol) were added and the mixture was stirred at room temperature for 10 minutes. Then, 1-2m (80 mg, 0.042 mmol) was added to the mixture and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-2o (30 mg, 32.9% yield) as a white solid. MS(ESI) m / z: 2184.5[M+H] + .

[0358] Step 11: (S)-42-((S)-3-amino-2-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)propanamide)-47-((5-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-di Methyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-(((2S,3R,4S,5S,6S)-6-carboxy-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)oxy)phenyl)amino)-4,4-dimethyl-8,36,43,47-tetraoxo-12,15,18,21,24,27,30,33-octaoxa-4,9,37,44-tetraazaheptatetracontan-4-ium-1-sulfonate(1-2)trifluoroacetate [ka]

[0359] To a solution of 1-2o (30 mg, 0.014 mmol) in CH2Cl2 (0.4 mL), TFA (0.1 mL, 4.81 mmol) was added at 0°C and the mixture was stirred at 0°C for 30 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-2 (trifluoroacetate) (22.5 mg, 74.5% yield) as a white solid. MS(ESI) m / z: 2084.4[M+H] + .

[0360] Examples 1-3 [ka]

[0361] Step 1: (2S,3R,4S,5S,6S)-2-(2-(3-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)propanamide)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amide (1-1-Methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate (1-3a) [ka]

[0362] To a solution of Int-1 (400 mg, 0.33 mmol) and Fmoc-β-alanine (126 mg, 0.33 mmol) in anhydrous DMF (3.5 mL), HATU (154 mg, 0.40 mmol) and 2,6-lutidine (0.12 mL, 1.00 mmol) were added and the mixture was stirred at room temperature for 20 hours. The resulting solution was quenched with saturated NH4Cl and extracted with CH2Cl2 (50 mL × 3). The organic phase was concentrated and purified by silica gel column chromatography (CH2Cl2 / MeOH = 98 / 2) to obtain 1-3a (398 mg, 79.9% yield) as a colorless oil. MS(ESI) m / z: 1494.3[M+H] + .

[0363] Step 2: (2S,3S,4S,5R,6S)-6-(2-(3-aminopropanamide)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-2k) trifluoroacetate [ka]

[0364] A solution of 1-3a (398 mg, 0.27 mmol) in water (2.5 mL) and THF (2.5 mL) was mixed with 1N LiOH (2.34 mL, 1.87 mmol) at 0°C and stirred at room temperature for 3 hours. The pH was adjusted to approximately 4 by gradually adding FA. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-2k (trifluoroacetate) (275 mg, 82.8% yield) as a white solid. MS(ESI) m / z: 1131.0[M+H] + .

[0365] Step 3: (2S,3S,4S,5R,6S)-6-(4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10- Dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-((R)-7-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)-2,2-dimethyl-4,11-dioxo-3,10-dioxa-5,12-diazapentadecane-15-amide)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-3b) [ka]

[0366] To a solution of 1-2k (trifluoroacetate) (30 mg, 0.024 mmol) in anhydrous DMF (2 mL), Int-2 (13 mg, 0.029 mmol) and DIPEA (0.013 mL, 0.072 mmol) were added at 0°C and the mixture was stirred at room temperature for 1 hour. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-3b (25 mg, 72.0% yield) as a white solid. MS(ESI) m / z: 1440.9[M+H] + .

[0367] Step 4: (2S,3S,4S,5R,6S)-6-(2-(3-((((R)-4-amino-3-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)butoxy)carbonyl)amino)propanamide)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-fu Phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-3)trifluoroacetate [ka]

[0368] A solution of 1-3b (25 mg, 0.017 mmol) in CH2Cl2 (0.5 mL) was mixed with TFA (0.13 mL, 1.76 mmol) at 0°C and stirred at 0°C for 30 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5 μm 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-3 (trifluoroacetate) (19 mg, 75.3% yield) as a white solid. MS(ESI) m / z: 1340.8 [M+H] + .

[0369] Examples 1-4 [ka] Step 1: (2S,3R,4S,5S,6S)-2-(2-amino-4-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate (1-4a) [ka]

[0370] A solution of Int-1a (500 mg, 1.03 mmol) in  (10 mL) was infiltrated with Pd / C (75 mg) and stirred at room temperature under an H2 atmosphere for 16 hours. The solution was filtered and washed with  (30 mL) and CH2Cl2 (30 mL). The filtrate was concentrated to obtain crude product 1-4a (470 mg, crude) as a yellow solid. MS(ESI)m / z:456.4[M+H] + .

[0371] Step 2: (2S,3R,4S,5S,6S)-2-(2-(1-(9H-fluoren-9-yl)-3-oxo-2,7,10,13,16-pentaoxa-4-azanonadecane-19-amide)-4-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-4c) [ka]

[0372] To a solution of 1-4a (95.0 mg, 0.21 mmol) and 1-4b (113 mg, 0.23 mmol) (commercially available) in anhydrous DMF (2 mL), HATU (96 mg, 0.25 mmol) and DIPEA (0.074 mL, 0.42 mmol) were added and stirred at room temperature for 2 hours. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5 μm 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-4c (107 mg, 55.4% yield) as a colorless oil. MS(ESI) m / z: 925.7 [M+H] + .

[0373] Step 3: (2S,3R,4S,5S,6S)-2-(2-(1-(9H-fluoren-9-yl)-3-oxo-2,7,10,13,16-pentaoxa-4-azanonadecane-19-amide)-4-((((4-nitrophenoxy)carbonyl)oxy)methyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-4d) [ka]

[0374] A solution of 1-4c (107 mg, 0.12 mmol) in anhydrous DMF (2 mL) was mixed with bis(4-nitrophenyl)carbonate (53 mg, 0.17 mmol) and DIPEA (0.037 mL, 0.21 mmol) at 0°C and stirred at room temperature for 6 hours. The solution was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (CH2Cl2 / MeOH = 90 / 10) to obtain 1-4d (110 mg, 87.2% yield) as an off-white solid. MS(ESI) m / z: 1090.8[M+H] + .

[0375] Step 4: (2S,3R,4S,5S,6S)-2-(2-(1-(9H-fluoren-9-yl)-3-oxo-2,7,10,13,16-pentaoxa-4-azanonadecane-19-amide)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane (1-4e) [ka]

[0376] To a solution of 1-4d (110 mg, 0.11 mmol) and MMAE (81 mg, 0.11 mmol) in anhydrous DMF (3 mL), HOBt (4.1 mg, 0.03 mmol) and DIPEA (0.033 mL, 0.18 mmol) were added and the mixture was stirred at room temperature for 16 hours. The solution was concentrated under reduced pressure and purified by silica gel column chromatography (CH2Cl2 / MeOH = 90 / 10) to obtain 1-4e (90 mg, 76.3% yield) as a white solid. MS(ESI) m / z: 1670.3[M+H] + .

[0377] Step 5: (2S,3S,4S,5R,6S)-6-(2-(1-amino-3,6,9,12-tetraoxapentadecane-15-amide)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1 -Methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-4f)trifluoroacetate [ka]

[0378] A solution of 1-4e (90 mg, 0.054 mmol) in THF (1 mL) and water (1 mL) was mixed with 1N LiOH (0.38 mL, 0.38 mmol) at 0°C and stirred at room temperature for 3 hours. The pH was adjusted to approximately 6 by gradually adding 20% ​​aqueous AcOH. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-4f (45 mg, 58.8% yield) (trifluoroacetate) as a white solid. MS(ESI) m / z: 1307.3[M+H] + .

[0379] Step 6: (2S,3S,4S,5R,6S)-6-(4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-tri Oxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-((R)-7-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)-2,2-dimethyl-4,11,15-trioxo-3,10,19,22,25,28-hexaoxa-5,12,16-triazahentriacontane-31-amide)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-4g) [ka]

[0380] To a solution of Int-3 (6.8 mg, 0.015 mmol) in anhydrous DMF (0.5 mL), HATU (6.5 mg, 0.017 mmol) and DIPEA (0.005 mL, 0.028 mmol) were added and the mixture was stirred at room temperature for 10 minutes. Then, 1-4f (trifluoroacetate) (20 mg, 0.014 mmol) was added to the mixture and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-4 g (15 mg, 63.1% yield) as a white solid. MS(ESI) m / z: 1689.4[M+H] + .

[0381] Step 7: (2S,3S,4S,5R,6S)-6-(2-((R)-26-amino-25-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)-17,21-dioxo-4,7,10,13,22-pentaoxa-16,20-diazahexacosaneamide)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R) -1-Hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-4)trifluoroacetate [ka]

[0382] To a solution of 1-4 g (15 mg, 0.009 mmol) in CH2Cl2 (0.4 mL), TFA (0.1 mL, 3.10 mmol) was added at 0°C and the mixture was stirred at 0°C for 30 minutes. The resulting solution was concentrated under reduced pressure and purified by prep-HPLC (Method: Column XBridge Prep C18 OBD 5 μm 19*150 mm; Mobile phase: A-Water (0.1% TFA): B-Acetonitrile; Flow rate: 20 mL / min). The fraction was lyophilized to obtain 1-4 (trifluoroacetate) (8.5 mg, 56.2% yield) as a white solid. MS(ESI) m / z: 1589.3 [M+H] + .

[0383] Examples 1-5 [ka] Process 1: (2S,3R,4S,5S,6S)-2-(2-(1-(9H-fluoren-9-yl)-3-oxo-2,7,10,13,16,19,22-heptaoxa-4-azapentacosan-25-amide)-4-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-5b) [ka]

[0384] To a solution of 1-4a (80 mg, 0.18 mmol) and 1-5a (112 mg, 0.19 mmol) in anhydrous DMF (1 mL), HATU (81 mg, 0.21 mmol) and DIPEA (0.062 mL, 0.35 mmol) were added and the mixture was stirred at room temperature for 20 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5 μm 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-5b (80 mg, 44.9% yield) as a colorless oil. MS(ESI) m / z: 1035.7 [M+Na] +

[0385] Step 2: (2S,3R,4S,5S,6S)-2-(2-(1-(9H-fluoren-9-yl)-3-oxo-2,7,10,13,16,19,22-heptaoxa-4-azapentacosan-25-amide)-4-((((4-nitrophenoxy)carbonyl)oxy)methyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-5c) [ka]

[0386] To a solution of 1-5b (80 mg, 0.079 mmol) in anhydrous DMF (2 mL), bis(4-nitrophenyl)carbonate (36 mg, 0.12 mmol) and DIPEA (0.025 mL, 0.14 mmol) were added at 0°C, and the mixture was stirred at room temperature for 6 hours. The solution was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (CH2Cl2 / MeOH = 90 / 10) to obtain 1-5c (80 mg, 86% yield) as an off-white solid. MS(ESI) m / z: 1178.8 [M+H] + .

[0387] Step 3: (2S,3R,4S,5S,6S)-2-(2-(1-(9H-fluoren-9-yl)-3-oxo-2,7,10,13,16,19,22-heptaoxa-4-azapentacosan-25-amide)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phen Lupropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-5d) [ka]

[0388] To a solution of 1-5c (80 mg, 0.068 mmol) and MMAE (59 mg, 0.081 mmol) in anhydrous DMF (2 mL), HOBt (2.8 mg, 0.020 mmol) and DIPEA (0.022 mL, 0.12 mmol) were added and the mixture was stirred at room temperature for 16 hours. The resulting solution was concentrated and purified by silica gel column chromatography (CH2Cl2 / MeOH = 90 / 10) to obtain 1-5d (80 mg, 67.1% yield) as a white solid. MS(ESI) m / z: 1758.3 [M+H] + .

[0389] Step 4: (2S,3S,4S,5R,6S)-6-(2-(1-amino-3,6,9,12,15,18-hexaoxahenicosan-21-amide)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino) -1-Methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-5e)trifluoroacetate [ka]

[0390] A solution of 1-5d (80 mg, 0.046 mmol) in THF (1 mL) and water (1 mL) was mixed with 1N LiOH (0.32 mL, 0.32 mmol) at 0°C and stirred at room temperature for 3 hours. The pH was adjusted to approximately 6 by gradually adding 20% ​​aqueous AcOH. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-5e (trifluoroacetate) (40 mg, 63.0% yield) as a white solid. MS(ESI) m / z: 1395.2[M+H] + .

[0391] Step 5: (2S,3S,4S,5R,6S)-6-(4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo -2,13-Dioxa-4,7,10-Triazatetradecyl)-2-((R)-7-(2,5-Dioxo-2,5-Dihydro-1H-Pyrrole-1-yl)-2,2-dimethyl-4,11,15-Trioxo-3,10,19,22,25,28,31,34-Octaoxa-5,12,16-Triazaheptatriacontan-37-amide)Phenoxy)-3,4,5-Trihydroxytetrahydro-2H-Pyran-2-carboxylic acid (1-5f) [ka]

[0392] To a solution of Int-3 (5.2 mg, 0.013 mmol) in anhydrous DMF (0.5 mL), HATU (5.0 g, 0.013 mmol) and DIPEA (0.004 mL, 0.022 mmol) were added and the mixture was stirred at room temperature for 10 minutes. Then, 1-5e (trifluoroacetate) (15 mg, 0.011 mmol) was added to the mixture and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-5f (9.0 mg, 47.1% yield) as a white solid. MS(ESI) m / z: 1777.4[M+H] + .

[0393] Step 6: (2S,3S,4S,5R,6S)-6-(2-((R)-32-amino-31-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)-23,27-dioxo-4,7,10,13,16,19,28-heptaoxa-22,26-diazadotricontanamid)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S ,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-5)trifluoroacetate [ka]

[0394] A solution of 1-5f (9.0 mg, 0.005 mmol) in CH2Cl2 (0.8 mL) was mixed with TFA (0.2 mL, 1.77 mmol) at 0°C and stirred at 0°C for 30 minutes. The resulting solution was concentrated and purified by prep-HPLC (Method: Column XBridge Prep C18 OBD 5um 19*150 mm; Mobile phase: A-Water (0.1% TFA): B-Acetonitrile; Flow rate: 20 mL / min). The fraction was lyophilized to obtain 1-5 (trifluoroacetate) (6.9 mg, 81.2% yield) as a white solid. MS(ESI) m / z: 1676.7[M+H] + .

[0395] Examples 1-6 [ka] Process 1: (2S,3R,4S,5S,6S)-2-(2-(1-(9H-fluoren-9-yl)-3-oxo-2,7,10,13,16,19,22,25,28-nonoxa-4-azahentriacontane-31-amide)-4-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-6b) [ka]

[0396] To a solution of 1-4a (50 mg, 0.11 mmol) and 1-6a (77 mg, 0.12 mmol) in anhydrous DMF (1 mL), HATU (51 mg, 0.13 mmol) and DIPEA (0.062 mL, 0.35 mmol) were added and the mixture was stirred at room temperature for 20 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5 μm 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-6b (44 mg, 36.4% yield) as a colorless oil. MS(ESI) m / z: 1102.8[M+H] + .

[0397] Step 2: (2S,3R,4S,5S,6S)-2-(2-(1-(9H-fluoren-9-yl)-3-oxo-2,7,10,13,16,19,22,25,28-nonoxa-4-azahentriacontane-31-amide)-4-((((4-nitrophenoxy)carbonyl)oxy)methyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-6c) [ka]

[0398] To a solution of 1-6b (44 mg, 0.04 mmol) in anhydrous DMF (1 mL), bis(4-nitrophenyl)carbonate (18 mg, 0.060 mmol) and DIPEA (0.013 mL, 0.072 mmol) were added at 0°C, and the mixture was stirred at room temperature for 6 hours. The solution was concentrated under reduced pressure. The residue was purified by silica column gel chromatography (CH2Cl2 / MeOH = 90 / 10) to obtain 1-6c (45 mg, 88.9% yield). MS(ESI) m / z: 1266.9 [M+H] + .

[0399] Step 3: (2S,3R,4S,5S,6S)-2-(2-(1-(9H-fluoren-9-yl)-3-oxo-2,7,10,13,16,19,22,25,28-nonoxa-4-azahentriacontane-31-amide)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1 -Phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrroridine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-6d) [ka]

[0400] To a solution of 1-6c (45 mg, 0.036 mmol) and MMAE (34 mg, 0.046 mmol) in anhydrous DMF (2 mL), HOBt (1.5 mg, 0.011 mmol) and DIPEA (0.019 mL, 0.11 mmol) were added and the mixture was stirred at room temperature for 16 hours. The resulting solution was concentrated and purified by silica gel column chromatography (CH2Cl2 / MeOH = 90 / 10) to obtain 1-6d (50 mg, 76.3% yield) as a white solid. MS(ESI) m / z: 1845.9 [M+H] + .

[0401] Step 4: (2S,3S,4S,5R,6S)-6-(2-(1-amino-3,6,9,12,15,18,21,24-octaoxaheptacosan-27-amide)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl) Amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-6e)trifluoroacetate [ka]

[0402] A solution of 1-3d (50 mg, 0.027 mmol) in THF (1 mL) and water (1 mL) was mixed with 1N LiOH (0.19 mL, 0.190 mmol) at 0°C and stirred at room temperature for 3 hours. The pH was adjusted to approximately 6 by gradually adding 20% ​​aqueous AcOH. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-6e (trifluoroacetate) (25 mg, 62.2% yield) as a white solid. MS(ESI) m / z: 1483.6[M+H] + .

[0403] Step 5: (2S,3S,4S,5R,6S)-6-(4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2 ,13-Dioxa-4,7,10-Triazatetradecyl)-2-((R)-7-(2,5-Dioxo-2,5-Dihydro-1H-Pyrrole-1-yl)-2,2-Dimethyl-4,11,15-Trioxo-3,10,19,22,25,28,31,34,37,40-Decaoxa-5,12,16-Triazatritetracontan-43-amide)Phenoxy)-3,4,5-Trihydroxytetrahydro-2H-Pyran-2-carboxylic acid (1-6f) [ka]

[0404] To a solution of Int-3 (6.1 mg, 0.015 mmol) in anhydrous DMF (0.5 mL), HATU (5.8 mg, 0.015 mmol) and DIPEA (0.004 mL, 0.025 mmol) were added and the mixture was stirred at room temperature for 10 minutes. Then, 1-6e (trifluoroacetate) (20 mg, 0.013 mmol) was added to the mixture and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-6f (11 mg, 47.1% yield) as a white solid. MS(ESI) m / z: 1865.4[M+H] + .

[0405] Step 6: (2S,3S,4S,5R,6S)-6-(2-((R)-38-amino-37-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)-29,33-dioxo-4,7,10,13,16,19,22,25,34-nonanoxa-28,32-diazaoctatriacotanamid)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(( (1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-6)trifluoroacetate [ka]

[0406] A solution of 1-6f (11 mg, 0.006 mmol) in CH2Cl2 (0.8 mL) was mixed with TFA (0.2 mL, 1.774 mmol) at 0°C and stirred at 0°C for 30 minutes. The resulting solution was concentrated and purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min). The fraction was lyophilized to obtain 1-6 (trifluoroacetate) (7.8 mg, 70.4% yield) as a white solid. MS(ESI) m / z: 1764.8 [M+H] + .

[0407] Examples 1-7 [ka] 1-7 (7.5 mg) was obtained by following the same procedure as in Examples 1-8. MS(ESI)m / z: 1573.5[M+H] + .

[0408] Examples 1-8 [ka] Step 1: (2S,3R,4S,5S,6S)-2-(2-(1-(9H-fluoren-9-yl)-3-oxo-2,7,10,13,16,19,22-heptaoxa-4-azapentacosan-25-amide)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phen Lupropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-8a) [ka]

[0409] A solution of Int-1 (100 mg, 0.083 mmol), 1-5a (54 mg, 0.092 mmol), and HATU (39 mg, 0.1 mmol) in anhydrous DMF (1 mL) was mixed with 2,6-lutidine (25 μL, 0.21 mmol) and stirred at room temperature for 17 hours. The solution was added to 0.5 N HCl (1 mL) and water (10 mL) and extracted with CH2Cl2 / MeOH (10:1, v / v, 11 mL × 4). The organic phase was concentrated and purified by flash column chromatography (CH2Cl2 / CH2Cl2=50 / 50 with 10% MeOH) to obtain compound 1-8a as a colorless oil (130 mg, 88.7% yield). MS(ESI) m / z: 1756.9[M+H] + .

[0410] Step 2: (2S,3S,4S,5R,6S)-6-(2-(1-amino-3,6,9,12,15,18-hexaoxahenicosan-21-amide)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-amide) (1-8b) [ka]

[0411] Compound 1-8a (120 mg, 0.068 mmol) was mixed with a solution of LiBr (359 mg, 4.1 mmol) in MeCN / H2O (1.7 / 0.17 mL), followed by the addition of TEA (58 μL, 0.41 mmol), and the mixture was stirred at room temperature for 3 hours. The pH was adjusted to 6 by adding HOAc. The solution was concentrated, and the residue was dissolved in anhydrous DMF (2 mL). Et2NH (144 μL, 1.37 mmol) was added, and the mixture was stirred at room temperature for a further 1 hour. The solution was concentrated and purified by prep-HPLC (column: Xbridge Prep C18 OBD (trademark) 5 μm, 19*150 mm; mobile phase A: 0.1% FA in water, B: MeCN; gradient: 20%~60% B; flow rate: 20 mL / min), and lyophilized to obtain compound 1-8b as a white solid (69 mg, 72.4% yield). MS(ESI)m / z:1394.8[M+H] + .

[0412] Step 3: (2S,3S,4S,5R,6S)-6-(4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10- Dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-(1-(4-(5-(methylsulfonyl)-1,2,4-thiadiazole-3-yl)phenyl)-1-oxo-5,8,11,14,17,20-hexaoxa-2-azatricosan-23-amide)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid(1-8) [ka]

[0413] DIEA (3 μL, 0.017 mmol) was added to a solution of Int-4 (4.6 mg, 0.016 mmol) and HATU (6.1 mg, 0.016 mmol) in anhydrous DMF (0.5 mL), and the mixture was stirred at room temperature for 15 minutes. The solution was added to a solution of 1-8b (20 mg, 0.014 mmol) in anhydrous DMF (0.5 mL), and DIEA (2 μL, 0.012 mmol) was added, and the mixture was stirred at room temperature for 10 minutes. The solution was purified by prep-HPLC (column: Xbridge Prep C18 OBD (trademark) 5 μm, 19*150 mm; mobile phase A: 0.1% FA in water, B: MeCN; gradient: 30%~65% B; flow rate: 20 mL / min), and lyophilized to obtain compound 1-8 as a white solid (13.4 mg, 56.3% yield). MS(ESI)m / z:1661.2[M+H] + .

[0414] Examples 1-9 [ka] Step 1: (2S,3R,4S,5S,6S)-2-(2-(1-(9H-fluoren-9-yl)-3-oxo-2,7,10,13,16,19,22,25,28-nonaoxa-4-azahentriacontane-31-amide)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1 -Phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-9a) [ka]

[0415] To a solution of Int-1 (50 mg, 0.042 mmol) and 1-6a (31 mg, 0.047 mmol) in anhydrous DMF (1 mL), HATU (19 mg, 0.050 mmol) and DIPEA (0.015 mL, 0.13 mmol) were added and the mixture was stirred at room temperature for 18 hours. The resulting solution was quenched with saturated NH4Cl and extracted with ethyl acetate. The organic phase was concentrated and purified by silica gel column chromatography (CH2Cl2 / MeOH = 98 / 2) to obtain 1-9a (50 mg, 65.0% yield) as a white solid. MS(ESI) m / z: 1845.4[M+H] + .

[0416] Step 2: (2S,3S,4S,5R,6S)-6-(2-(1-amino-3,6,9,12,15,18,21,24-octaoxaheptacosan-27-amide)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl) Amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-9b)trifluoroacetate [ka]

[0417] A solution of 1-9a (50 mg, 0.027 mmol) in water (0.5 mL) and THF (0.5 mL) was mixed with 1N LiOH (0.19 mL, 0.19 mmol) at 0°C and stirred at room temperature for 3 hours. The pH was adjusted to approximately 4 by gradually adding formic acid. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-9b (trifluoroacetate) (20 mg, 49.8% yield) as a white solid. MS(ESI) m / z: 1483.6 [M+H] + .

[0418] Step 3: (2S,3R,4S,5S,6S)-2-(2-amino-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-9) [ka]

[0419] To a solution of Int-4 (4.3 mg, 0.015 mmol) in anhydrous DMF (0.5 mL), HATU (5.8 mg, 0.015 mmol) and DIPEA (0.0084 mL, 0.025 mmol) were added and the mixture was stirred at room temperature for 15 minutes. Then, 1-9b (trifluoroacetate) (20 mg, 0.013 mmol) was added to the mixture and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-9 (14 mg, 63.9% yield) as a white solid. MS(ESI) m / z: 1750.3[M+H] + .

[0420] Examples 1-10 [ka] Step 1: tert-butyl(S)-42-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-39-oxo-2,5,8,11,14,17,20,23,26,29,32,35-dodecaoxa-38-azatritetracontane-43-oate(1-10b) [ka]

[0421] To a solution of Fmoc-L-glutamic acid 1-tert-butyl ester (69 mg, 0.16 mmol) and 1-10a (90 mg, 0.16 mmol) in anhydrous DMF (2 mL), HATU (74 mg, 0.19 mmol) and DIPEA (0.058 mL, 0.32 mmol) were added and the mixture was stirred at room temperature for 20 minutes. The resulting solution was quenched with saturated NH4Cl and extracted with  (30 mL × 3). The organic phase was concentrated and purified by silica gel column chromatography (CH2Cl2 / MeOH = 98 / 2) to obtain 1-10b (130 mg, 83.6% yield) as a colorless oil. MS(ESI) m / z: 967.9 [M+H] + .

[0422] Step 2: (S)-42-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-39-oxo-2,5,8,11,14,17,20,23,26,29,32,35-dodecaoxa-38-azatritetracontan-43-acid(1-10c) [ka]

[0423] To a solution of 1-10b (60 mg, 0.062 mmol) in CH2Cl2 (0.4 mL), TFA (0.19 mL, 2.48 mmol) was added and the mixture was stirred at room temperature for 3 hours. The resulting solution was concentrated under vacuum and co-evaporated five times with toluene to obtain 1-10c (55 mg, crude) as a colorless oil, which was used directly in the next step. MS(ESI)m / z:911.9[M+H] + .

[0424] Step 3: (2S,3S,4S,5R,6S)-6-(2-((S)-42-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-39,43-dioxo-2,5,8,11,14,17,20,23,26,29,32,35-dodecaoxa-38,44-diazaheptatetracontane-47-amide)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R ,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-10d) [ka]

[0425] To a solution of 1-10c (16 mg, 0.018 mmol) in anhydrous DMF (0.5 mL), HATU (7.4 mg, 0.019 mmol) and DIPEA (0.009 mL, 0.048 mmol) were added and the mixture was stirred at room temperature for 10 minutes. Then, 1-2k (20 mg, 0.016 mmol) was added to the mixture at 0°C and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-10d (13 mg, 40.0% yield) as a white solid. MS(ESI) m / z: 2024.1[M+H] + .

[0426] Step 4: (2S,3S,4S,5R,6S)-6-(2-((S)-42-amino-39,43-dioxo-2,5,8,11,14,17,20,23,26,29,32,35-dodecaoxa-38,44-diazaheptatetracontane-47-amide)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R )-1-Hydroxy-1-phenylpropane-2-yl)amino)-1-Methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-10e) [ka]

[0427] A solution of 1-10d (13 mg, 0.006 mmol) in anhydrous DMF (1 mL) was mixed with Et2NH (0.02 mL, 0.19 mmol) and stirred at room temperature for 20 minutes. 1-10e (7 mg, crude) was evaporated with toluene five times to remove the solvent. MS(ESI) m / z: 1801.9[M+H] + .

[0428] Step 5: (2S,3S,4S,5R,6S)-6-(4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,1 3-Dioxa-4,7,10-Triazatetradecyl)-2-((S)-42-(4-(5-(methylsulfonyl)-1,2,4-thiadiazole-3-yl)benzamide)-39,43-Dioxo-2,5,8,11,14,17,20,23,26,29,32,35-Dodecaoxa-38,44-Diazaheptatetracontan-47-amide)phenoxy)-3,4,5-Trihydroxytetrahydro-2H-pyran-2-carboxylic acid(1-10) [ka]

[0429] To a solution of Int-4 (2.0 mg, 0.007 mmol) in anhydrous DMF (1 mL), HATU (3.0 mg, 0.008 mmol) and DIPEA (0.002 mL, 0.013 mmol) were added and the mixture was stirred at room temperature for 15 minutes. Then, 1-10e (7.0 mg, 0.006 mmol) was added to the mixture and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-10 (5.2 mg, 38.8% yield) as a white solid. MS(ESI) m / z: 2068.1[M+H] + .

[0430] Examples 1-11 [ka] Process 1: (2S,3S,4S,5R,6S)-6-(2-((S)-42-amino-39,43-dioxo-2,5,8,11,14,17,20,23,26,29,32,35-dodecaoxa-38,44-diazaheptatetracontane-47-amide)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R )-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-11a) [ka]

[0431] To a solution of Int-3 (18 mg, 0.007 mmol) in anhydrous DMF (1 mL), HATU (17 mg, 0.044 mmol) and DIPEA (0.013 mL, 0.074 mmol) were added and the mixture was stirred at room temperature for 10 minutes. Then, 1-10e (40 mg, 0.037 mmol) was added to the mixture and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-11a (50 mg, 61.9% yield) as a white solid. MS(ESI) m / z: 2183.1[M+H] + .

[0432] Step 2: (2S,3S,4S,5R,6S)-6-(2-((S)-42-(3-((((R)-4-amino-3-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)butoxy)carbonyl)amino)propanamide)-39,43-dioxo-2,5,8,11,14,17,20,23,26,29,32,35-dodecaoxa-38,44-diazaheptatetracontane-47-amide)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12 -(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-11)trifluoroacetate [ka]

[0433] A solution of 1-11a (50 mg, 0.023 mmol) in CH2Cl2 (2 mL) was mixed with TFA (0.43 mL, 5.73 mmol) at 0°C and stirred at 0°C for 30 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5 μm 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-11 (trifluoroacetate) (36 mg, 71.5% yield) as a white solid. MS(ESI) m / z: 2083.0 [M+H] + .

[0434] Examples 1-12 [ka] Step 1: tert-butyl(S)-40-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-1-amino-3,6,9,12,15,18,21,24,27,30,33,36-dodecamethyl-1,4,7,10,13,16,19,22,25,28,31,34,37-tridecaoxo-3,6,9,12,15,18,21,24,27,30,33,36-dodecazahentetracontane-41-oate(1-12b) [ka]

[0435] To a solution of 1-12a (45 mg, 0.052 mmol) and Fmoc-L-glutamic acid 1-tert-butyl ester (23.3 mg, 0.054 mmol) in anhydrous DMF (1 mL), HATU (23.8 mg, 0.062 mmol) and DIPEA (0.028 mL, 0.155 mmol) were added, and the mixture was stirred at room temperature for 30 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-12b (36 mg, 71.5% yield) as a white solid. MS(ESI) m / z: 1278.3[M+H] + .

[0436] Step 2: (S)-40-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-1-amino-3,6,9,12,15,18,21,24,27,30,33,36-dodecamethyl-1,4,7,10,13,16,19,22,25,28,31,34,37-tridecaoxo-3,6,9,12,15,18,21,24,27,30,33,36-dodecazahentetracontan-41-acid(1-12c) [ka]

[0437] To a solution of 1-12b (45 mg, 0.035 mmol) in CH2Cl2 (0.4 mL), TFA (0.19 mL, 2.48 mmol) was added and the mixture was stirred at room temperature for 3 hours. The resulting solution was concentrated under vacuum and co-evaporated five times with toluene to obtain 1-12c (40 mg, crude) as a colorless oil, which was used directly in the next step. MS(ESI)m / z:1222.0[M+H] + .

[0438] Step 3: (2S,3S,4S,5R,6S)-6-(2-((S)-40-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)-1-amino-3,6,9,12,15,18,21,24,27,30,33,36-Dodecamethyl-1,4,7,10,13,16,19,22,25,28,31,34,37,41-Tetradecaoxo-3,6,9,12,15,18,21,24,27,30,33,36,42-Tridecazapentatetracontane-45-amide)-4-((5S,8S,11S,1 2R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-12d) [ka]

[0439] To a solution of 1-12c (40 mg, 0.032 mmol) in anhydrous DMF (1 mL), HATU (15 mg, 0.039 mmol) and DIPEA (0.012 mL, 0.064 mmol) were added and the mixture was stirred at room temperature for 10 minutes. Then, 1-2k (trifluoroacetate) (40 mg, 0.032 mmol) was added to the mixture and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-12d (25 mg, 33.3% yield) as a white solid. MS(ESI) m / z: 2333.5[M+H] + .

[0440] Step 4: (2S,3S,4S,5R,6S)-6-(4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-trioxa (Thetetradecyl)-2-((S)-1,40-diamino-3,6,9,12,15,18,21,24,27,30,33,36-dodecamethyl-1,4,7,10,13,16,19,22,25,28,31,34,37,41-tetradekaoxo-3,6,9,12,15,18,21,24,27,30,33,36,42-tridecazapentatetracontan-45-amide)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-12e) [ka]

[0441] A solution of 1-12d (25 mg, 0.011 mmol) in anhydrous DMF (1 mL) was mixed with Et2NH (0.034 mL, 0.32 mmol) and stirred at room temperature for 20 minutes. 1-12e (22 mg, crude) was evaporated five times with toluene to remove the solvent, and this was used directly in the next step. MS(ESI)m / z: 2111.5[M+H]

[0442] Step 5: (2S,3S,4S,5R,6S)-6-(2-((S)-1-amino-40-((R)-7-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)-2,2-dimethyl-4,11-dioxo-3,10-dioxa-5,12-diazapentadecane-15-amide)-3,6,9,12,15,18,21,24,27,30,33,36-dodecamethyl-1,4,7,10,13,16,19,22,25,28,31,34,37,41-tetradecaoxo-3,6,9,12,15,18,21,24,27,30,33,36,42-tridecazapentate Lacontan-45-amide)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-12f) [ka]

[0443] To a solution of Int-3 (5 mg, 0.013 mmol) in anhydrous DMF (1 mL), HATU (4.8 mg, 0.013 mmol) and DIPEA (0.004 mL, 0.021 mmol) were added and the mixture was stirred at room temperature for 10 minutes. Then, 1-12e (22 mg, 0.01 mmol) was added to the mixture and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-12f (13 mg, 50.1% yield) as a white solid. MS(ESI) m / z: 2491.8[M+H] + .

[0444] Step 6: (2S,3S,4S,5R,6S)-6-(2-((S)-1-amino-40-(3-((((R)-4-amino-3-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)butoxy)carbonyl)amino)propanamide)-3,6,9,12,15,18,21,24,27,30,33,36-dodecamethyl-1,4,7,10,13,16,19,22,25,28,31,34,37,41-tetradecaoxo-3,6,9,12,15,18,21,24,27,30,33,36,42-tridecazapentatetracontane-45-amide)- 4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-12)trifluoroacetate [ka]

[0445] A solution of 1-12f (13 mg, 0.005 mmol) in CH2Cl2 (0.5 mL) was mixed with TFA (0.1 mL, 1.30 mmol) and stirred at 0°C for 30 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5 μm 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-12 (trifluoroacetate) (10 mg, 76.5% yield) as a white solid. MS(ESI) m / z: 2392.3 [M+H] + .

[0446] Examples 1-13 [ka] Step 1: Methyl 3-fluoro-4-hydroxy-5-nitrobenzoate (1-13b)

[0447] A 65% aqueous nitric acid solution (1.4 mL, 30.43 mmol) and fuming nitric acid (2.1 mL, 21.43 mmol) were added to a solution of 1-13a (1.80 g, 10.6 mmol) in diethyl ether (50 mL) at -10°C. The ice bath was removed, and the reaction mixture was stirred at 30°C for 2 hours, then partitioned between water and ethyl acetate. The organic layer was concentrated and purified by flash column chromatography (petroleum ether / ethyl ethyl acetate = 66 / 34) to obtain 1-13b as a yellow solid (600 mg, 26.3% yield). MS(ESI) m / z: 214.0 [MH] + ]

[0448] Step 2: 2-Fluoro-4-(hydroxymethyl)-6-nitrophenol(1-13c)

[0449] 1-13b (1.20 g, 5.58 mmol) was dissolved in anhydrous CH2Cl2 (100 mL), and diisobutylaluminum hydride (1 N, 16 mL) was added under a nitrogen atmosphere at -78 °C. The mixture was stirred at -78 °C for 1 hour, then quenched with MeOH (4 mL), followed by 1 N HCl (4 mL). The mixture was purified by flash column chromatography (petroleum ether / Â=50 / 50) to obtain 1-13c as a yellow solid (830 mg, 80.6% yield). MS(ESI) m / z: 186.1 [MH] + ]

[0450] Step 3: 3-Fluoro-4-hydroxy-5-nitrobenzaldehyde (1-13d)

[0451] Dess-Martin periodinane (2.82 g, 6.65 mmol) was added to a solution of 1-13c (830 mg, 4.44 mmol) in anhydrous CH2Cl2 (20 mL). The mixture was then allowed to rise to room temperature and stirred for 2 hours. The mixture was quenched with aqueous Na2S2O3 and Na2CO3, and purified by flash column chromatography (petroleum ether / siRNA = 50 / 50) to obtain 1-13d as a yellow solid (300 mg, 36.5% yield). MS(ESI) m / z: 184.1 [MH] + ]

[0452] Step 4: (2S,3R,4S,5S,6S)-2-(2-fluoro-4-formyl-6-nitrophenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-13e)

[0453] 1-13d (300 mg, 1.62 mmol) was added to a solution of acetobrom-α-D-glucuronide methyl ester (579 mg, 1.46 mmol) and Ag2O (751 mg, 3.24 mmol) in CH3CN (10 mL). The mixture was stirred in the dark at room temperature for 4 hours. The suspension was filtered through Celite, and the filtrate was partitioned between ethyl acetate (50 mL) and water (50 mL). The organic layer was washed three times with aqueous NaHCO3 (50 mL), dried over Na2SO4, and concentrated to obtain 1-13e as a yellow solid (648 mg, 79.8% yield) without further purification. MS(ESI)m / z: 524.4[M+H] +

[0454] Step 5: (2S,3R,4S,5S,6S)-2-(2-fluoro-4-(hydroxymethyl)-6-nitrophenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-13f)

[0455] NaBH4 (48.90 mg, 1.29 mmol) was added at 0°C to a solution of 1-13e (648 mg, 1.29 mmol) in IPA / CH2Cl2 (3 / 12 mL). The mixture was allowed to rise to room temperature, stirred for 1 hour, and then quenched with aqueous NH4Cl. The mixture was partitioned between water and dichloromethane. The organic layer was collected, concentrated, and purified by flash column chromatography (petroleum ether / Â=50 / 50) to obtain 1-13d as a yellow solid (350 mg, 53.8% yield). MS(ESI) m / z: 526.5[M+H] +

[0456] Step 6: (2S,3R,4S,5S,6S)-2-(2-amino-6-fluoro-4-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate (1-13g)

[0457] To a solution of 1-13f (350 mg, 0.70 mmol) in THF / MeOH / H2O (5 / 1 / 1 mL), Fe (388 mg, 6.95 mmol) and NH4Cl (186 mg, 3.48 mmol) were added. The mixture was refluxed at 70°C for 4 hours. The mixture was partitioned between water and dichloromethane. The organic layer was collected and concentrated to obtain 1-13 g of crude oil (330 mg) without further purification.

[0458] Step 7: (2S,3R,4S,5S,6S)-2-(2-(3-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)propanamide)-6-fluoro-4-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-13h)

[0459] EEDQ (345 mg, 1.39 mmol) was added to a solution of 1-13 g (330 mg, 0.70 mmol) and Fmoc-β-alanine (260 mg, 0.84 mmol) in CH2Cl2 / DMF (5 / 1 mL). The mixture was stirred overnight at room temperature and purified by flash column chromatography (MeOH / CH2Cl2 = 5 / 95) to obtain 1-13d as a white solid (200 mg, 37.4% yield in 2 steps). MS(ESI) m / z: 767.6 [M+H] +

[0460] Step 8: (2S,3R,4S,5S,6S)-2-(2-(3-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)propanamide)-6-fluoro-4-((((4-nitrophenoxy)carbonyl)oxy)methyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-13i)

[0461] DIPEA (136 μL, 0.78 mmol) was added to a solution of 1-13h (200 mg, 0.26 mmol) and 4-nitrophenyl chloroformate (105 mg, 0.52 mmol) in anhydrous CH2Cl2 (4 mL). The mixture was stirred overnight at room temperature and purified by flash column chromatography (MeOH / CH2Cl2 = 5 / 95) to obtain 1-13i as a white solid (100 mg, 41.2% yield). MS(ESI) m / z: 932.6 [M+H] +

[0462] Step 9: (2S,3R,4S,5S,6S)-2-(2-(3-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)propanamide)-4-((5S,8S,11R,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amino)-1 -Methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-6-fluorophenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-13j) [ka]

[0463] To a solution of 1-13i (100 mg, 0.11 mmol) in 2 mL of DMF, MMAE (77 mg, 0.11 mmol) and DIPEA (56.1 μL, 0.32 mmol) were added, followed by the addition of HOBt (4.4 mg, 0.027 mmol). The mixture was stirred at room temperature for 24 hours and then partitioned between water and ethyl acetate. The organic layer was purified by flash column chromatography (MeOH / CH2Cl2 = 5 / 95) to obtain 1-13j as a white solid (101 mg, 62.3% yield). MS(ESI) m / z: 1511.5[M+H] +

[0464] Step 10: (2S,3S,4S,5R,6S)-6-(2-(3-aminopropanamide)-4-((5S,8S,11R,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-6-fluorophenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-13k) [ka]

[0465] A solution of 1-13j (101 mg, 0.07 mmol) in THF / MeOH (1 / 1 mL) was mixed with 1N LiOH (400 μL) at 0°C. The mixture was stirred at room temperature for 20 minutes, quenched with HOAc (12 μL), and purified by prep-HPLC (column: Xbridge Prep C18 OBD (trademark) 5 μm, 19*150 mm; mobile phase A: 0.1% FA in water, B: MeCN; gradient: 30%~55% B; flow rate: 20 mL / min). Lyophilization yielded 1-13k (40 mg, 52.1% yield). MS(ESI) m / z: 1149.2[M+H] +

[0466] Step 11: (2S,3S,4S,5R,6S)-6-(4-((5S,8S,11R,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl -3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-((R)-7-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)-2,2-dimethyl-4,11-dioxo-3,10-dioxa-5,12-diazapentadecane-15-amide)-6-fluorophenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-13l) [ka]

[0467] A solution of 1-13k (26.2 mg, 0.023 mmol) and Int-2 (12.3 mg, 0.027 mmol) in DMF (1 mL) was mixed with DIPEA (12 μL, 0.069 mmol). The mixture was stirred overnight at room temperature and purified by prep-HPLC (column: Xbridge Prep C18 OBD® 5 μm, 19*150 mm; mobile phase A: 0.1% FA in water, B: MeCN; gradient: 45%~85% B; flow rate: 20 mL / min). The mixture was lyophilized to obtain 1-13 L (15 mg, 45.1% yield). MS(ESI) m / z: 1459.6 [M+H] +

[0468] Step 12: (2S,3S,4S,5R,6S)-6-(2-(3-((((R)-4-amino-3-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)butoxy)carbonyl)amino)propanamide)-4-((5S,8S,11R,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1 -phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-6-fluorophenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid(1-13) [ka]

[0469] 1-13L (15 mg, 10.28 μmol) was dissolved in CH2Cl2 / TFA (1 / 0.2 mL) and stirred at room temperature for 30 minutes. The mixture was purified by prep-HPLC (column: Xbridge Prep C18 OBD (trademark) 5 μm, 19*150 mm; mobile phase A: 0.1% TFA in water, B: MeCN; gradient: 40%~70% B; flow rate: 20 mL / min) and lyophilized to obtain 1-13 (5 mg, 35.8% yield). MS(ESI) m / z: 1359.3 [M+H] +

[0470] Examples 1-14 [ka] Step 1: 2-Fluoro-4-hydroxy-5-nitrobenzaldehyde (1-14b)

[0471] 1-14a (1.50 g, 10.71 mmol) was dissolved in sulfuric acid (12.5 mL). A 65% nitric acid solution (690 μL, 10.71 mmol) was dissolved in sulfuric acid (2.5 mL) at -20°C and added to the reaction solution. The reaction mixture was stirred at -20°C for 2 hours, then quenched with ice water (100 mL), and extracted with ethyl acetate (100 mL x 2). The combined organic matter was dried over Na2SO4, concentrated, and purified by flash column chromatography to obtain 1-14b as a yellow solid (1.59 g, 80.2% yield). MS(ESI) m / z: 184.0 [MH] + ]

[0472] Step 2: (2S,3R,4S,5S,6S)-2-(5-fluoro-4-formyl-2-nitrophenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-14c)

[0473] 1-14b (1.59 g, 8.59 mmol) was added to a solution of acetobrom-α-D-glucuronide methyl ester (3.07 g, 7.73 mmol) and Ag2O (2.99 g, 12.88 mmol) in CH3CN (50 mL). The mixture was stirred in the dark at room temperature for 4 hours. The suspension was filtered through Celite, and the filtrate was partitioned between ethyl acetate (100 mL) and water (100 mL). The organic layer was washed three times with aqueous Na2CO3 (100 mL), dried over Na2SO4, and concentrated to obtain 1-14c as a yellow solid (3.64 g, 84.5% yield) without further purification. MS(ESI)m / z: 524.5[M+Na] +

[0474] Step 3: (2S,3R,4S,5S,6S)-2-(5-fluoro-4-(hydroxymethyl)-2-nitrophenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-14d)

[0475] NaBH4 (0.27 g, 7.24 mmol) was added at 0°C to a solution of 1-14c (3.63 g, 7.24 mmol) in IPA / CH2Cl2 (10 / 40 mL). The mixture was allowed to rise to room temperature, stirred for 3 hours, and then quenched with aqueous NH4Cl. The mixture was partitioned between water and dichloromethane. The organic layer was collected, concentrated, and purified by flash column chromatography (petroleum ether / siRNA = 50 / 50) to obtain 1-14d as a yellow solid (1.88 g, 51.6% yield). MS(ESI) m / z: 526.5[M+Na] +

[0476] Step 4: (2S,3R,4S,5S,6S)-2-(2-amino-5-fluoro-4-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-14e)

[0477] To a solution of 1-14d (980 mg, 1.95 mmol) in THF / MeOH / H2O (5 / 1 / 1 mL), Fe (1.09 g, 19.47 mmol) and NH4Cl (521 mg, 9.73 mmol) were added. The mixture was refluxed at 70°C for 4 hours. The mixture was partitioned between water and dichloromethane. The organic layer was collected and concentrated to obtain 1-14e as a colorless oil (880 mg, 95.5% yield). MS(ESI)m / z:474.5[M+H] + .

[0478] Step 5: (2S,3R,4S,5S,6S)-2-(2-(3-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)propanamide)-5-fluoro-4-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-14f)

[0479] EEDQ (919 mg, 3.72 mmol) was added to a solution of 1-14e (880 mg, 1.86 mmol) and Fmoc-β-alanine (694 mg, 2.23 mmol) in CH2Cl2 / DMF (10 / 1 mL). The mixture was stirred overnight at room temperature and purified by flash column chromatography (MeOH / CH2Cl2 = 5 / 95) to obtain 1-14f as a white solid (1.06 g, 74.4% yield). MS(ESI) m / z: 767.6 [M+H] + .

[0480] Step 6: (2S,3R,4S,5S,6S)-2-(2-(3-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)propanamide)-5-fluoro-4-((((4-nitrophenoxy)carbonyl)oxy)methyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate (1-14g)

[0481] A solution of 1-14f (190 mg, 0.25 mmol) and 4-nitrophenyl chloroformate (80 mg, 0.40 mmol) in anhydrous CH2Cl2 (4 mL) was mixed with DIPEA (90 μL, 0.50 mmol). The mixture was stirred at room temperature for 6 hours and purified by flash column chromatography (MeOH / CH2Cl2 = 5 / 95) to obtain 1-14 g as a white solid (190 mg, 82.3% yield). MS(ESI) m / z: 932.6 [M+H] + .

[0482] Step 7: (2S,3R,4S,5S,6S)-2-(2-(3-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)propanamide)-4-((5S,8S,11R,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amino)-1 -Methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-5-fluorophenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-14h) [ka]

[0483] To a solution of 1-14 g (190 mg, 0.20 mmol) in 2 mL of DMF, MMAE (146 mg, 0.20 mmol) and DIPEA (106 μL, 0.61 mmol) were added, followed by HOBt (6.9 mg, 0.069 mmol). The mixture was stirred at room temperature for 48 hours and then partitioned between water and ethyl acetate. The organic layer was purified by flash column chromatography (MeOH / CH2Cl2 = 5 / 95) to obtain the crude 1-14 h as a yellow solid (40 mg). MS(ESI) m / z: 1512.2[M+H] + .

[0484] Step 8: (2S,3S,4S,5R,6S)-6-(2-(3-aminopropanamide)-4-((5S,8S,11R,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-6-fluorophenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-14i) [ka]

[0485] A solution of crude 1-14h (40 mg, 0.026 mmol) in MeOH (1 mL) was mixed with 1N LiOH (400 μL) at 0°C. The mixture was stirred at room temperature for 20 minutes and quenched with HOAc (12 μL). The mixture was concentrated and then dissolved in DMF (1 mL). Et2NH (50 μL, 0.48 mmol) was added, and the mixture was stirred at room temperature for 30 minutes. The mixture was concentrated and purified by prep-HPLC (column: Sunfire Prep C18 OBD (trademark) 5 μm, 19*150 mm; mobile phase A: 0.1% FA in water, B: MeCN; gradient: 30%~60% B; flow rate: 20 mL / min), and lyophilized to obtain 1-14i (20 mg, 8.5% yield in 2 steps). MS(ESI) m / z: 1149.1[M+H] + .

[0486] Step 9: (2S,3S,4S,5R,6S)-6-(4-((5S,8S,11R,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl -3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-((R)-7-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)-2,2-dimethyl-4,11-dioxo-3,10-dioxa-5,12-diazapentadecane-15-amide)-5-fluorophenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-14j) [ka]

[0487] A solution of 1-14i (20 mg, 0.017 mmol) and Int-2 (11.74 mg, 0.026 mmol) in DMF (1 mL) was mixed with DIPEA (9.1 μL, 0.052 mmol). The mixture was stirred at room temperature for 3 hours and purified by prep-HPLC (column: Sunfire Prep C18 OBD (trademark) 5 μm, 19*150 mm; mobile phase A: 0.1% FA in water, B: MeCN; gradient: 30%~80% B; flow rate: 20 mL / min). The mixture was lyophilized to obtain 1-14j (13 mg, 51.2% yield). MS(ESI) m / z: 1459.3 [M+H] + .

[0488] Step 10: (2S,3S,4S,5R,6S)-6-(2-(3-((((R)-4-amino-3-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)butoxy)carbonyl)amino)propanamide)-4-((5S,8S,11R,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1 -phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-5-fluorophenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid(1-14) [ka]

[0489] 1-14j (13 mg, 8.92 μmol) was dissolved in CH2Cl2 / TFA (1 / 0.2 mL) and stirred at room temperature for 30 minutes. The mixture was purified by prep-HPLC (column: Xbridge Prep C18 OBD (trademark) 5 μm, 19*150 mm; mobile phase A: 0.1% TFA in water, B: MeCN; gradient: 35%~60% B; flow rate: 20 mL / min) and lyophilized to obtain 1-14 (4.3 mg, 35.5% yield). MS(ESI) m / z: 1359.1 [M+H] + .

[0490] Examples 1-15 [ka] Step 1 (2S,3R,4S,5S,6S)-2-(2-azido-4-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate (1-15a) [ka]

[0491] 1-4a (450 mg, 0.99 mmol) was dissolved in anhydrous THF (2 mL). The mixture was cooled to 0°C, then imidazole-1-sulfonyl azide (414.2 mg, 1.98 mmol) in 1 mL of MeOH was added, followed by K2CO3 in 1 mL of H2O and a catalytic amount of CuSO4. The mixture was stirred at room temperature under a nitrogen atmosphere for 2 hours. Depositphotos (5 mL) was added, the organic phase was collected, dried over Na2SO4, concentrated, and then purified by column chromatography (petroleum ether / Depositphotos = 50 / 50) to obtain 2-15a as a white solid (350 mg, 87.2%). MS(ESI) m / z: 504.2 [M + Na] + . 1 H NMR (400 MHz, CDCl3) δ 7.17 - 7.03 (m, 3H), 5.44 - 5.26 (m, 3H), 5.07 (d, J = 7.1 Hz, 1H), 4.64 (d, J = 2.2 Hz, 2H), 4.13 (dd, J = 6.8, 2.8 Hz, 1H), 3.83 - 3.71 (m, 3H), 2.17 - 2.00 (m, 9H).

[0492] Step 2 (2S,3R,4S,5S,6S)-2-(2-azido-4-((((4-nitrophenoxy)carbonyl)oxy)methyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-15b) [ka]

[0493] 1-15a (100 mg, 0.21 mmol) was dissolved in anhydrous DMF (0.5 mL). The mixture was cooled to 0°C, and 4-nitrophenyl chloroformate (50.24 mg, 0.25 mmol) was added, followed by DIPEA (79.58 μL, 0.42 mmol). The mixture was stirred at room temperature under a nitrogen atmosphere for 6 hours. The reaction solution was diluted with Depositphotos (5 mL) and washed with H₂O (5 mL). The organic phase was collected, dried over Na₂SO₄, concentrated, and then purified by column chromatography (petroleum ether / Depositphotos = 50 / 50) to obtain 1-15b as a pale yellow solid (70 mg, 52.2%). MS(ESI) m / z: 668.5 [M + Na] +

[0494] Step 3 (2S,3R,4S,5S,6S)-2-(2-azido-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-15c) [ka]

[0495] To a solution of 1-15b (70 mg, 0.11 mmol) in 1 mL of DMF, MMAE (85.51 mg, 0.12 mmol) and DIPEA (59.1 μL, 0.33 mmol) were added, followed by the addition of HOBt (4.4 mg, 0.032 mmol). The mixture was stirred at room temperature under a nitrogen atmosphere for 12 hours. The solvent was then removed by evaporation, and the crude product was purified by column chromatography (MeOH / CH2Cl2 = 5 / 95) to obtain 1-15c as a white solid (65 mg, 50%). MS(ESI) m / z: 1227.1 [M+H]+ .

[0496] Step 4 (2S,3S,4S,5R,6S)-6-(2-azido-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-15d) [ka]

[0497] A solution of 1-15c (65 mg, 0.05 mmol) in 2 mL of THF was mixed with 1 mL of 0.5 M LiOH solution at 0°C. The mixture was stirred under a nitrogen atmosphere at 0°C for 15 minutes. The reaction was quenched with AcOH, and then purified by prep-HPLC (method: column: XBridge Prep C18 OBD 5 μm 19*1250 mm; mobile phase: A-water (0.1% formic acid): B-acetonitrile; flow rate: 20 mL / min) to obtain 1-15d as a white solid after lyophilization (40 mg, 69.5% yield). MS(ESI) m / z: 1086.4[M+H] + .

[0498] Step 5: (9H-fluoren-9-yl)methyl(15-oxo-3,6,9,12-tetraoxa-16-azanonades-18-in-1-yl)carbamate(1-15e) [ka]

[0499] To a solution of 1-4b (200 mg, 0.41 mmol) in 2 mL of DMF, HATU (312 mg, 0.82 mmol) and DIPEA (224 μL, 1.23 mmol) were added at 0°C. The mixture was stirred at 0°C for 10 minutes. Propargylamine (25 μL, 0.45 mmol) was added. The mixture was stirred at room temperature under a nitrogen atmosphere for 1 hour. The solvent was then removed by evaporation, and the crude product was purified by column chromatography (MeOH / CH2Cl2 = 5 / 95) to obtain 1-15e as a brown oil (200 mg, 93% yield). MS(ESI) m / z: 525.8 [M+H] + .

[0500] Step 6: 1-amino-N-(prop-2-in-1-yl)-3,6,9,12-tetraoxapentadecane-15-amide(1-15f) [ka]

[0501] Diethylamine (0.98 mL, 9.5 mmol) was added to a solution of 1-15e (500 mg, 0.95 mmol) in 2 mL of DMF at 0°C. The mixture was stirred at 0°C for 30 minutes. The solvent was then removed by evaporation, and the crude product was purified by column chromatography (MeOH / CH2Cl2 = 5 / 95) to obtain 1-15f as a pale yellow solid (250 mg, 87% yield). MS(ESI) m / z: 303.3 [M+H] + .

[0502] Step 7: 1-(4-(5-(methylsulfonyl)-1,2,4-thiadiazole-3-yl)benzamide)-N-(prop-2-in-1-yl)-3,6,9,12-tetraoxapentadecane-15-amide (1-15g) [ka]

[0503] To a solution of 1-15f (10 mg, 0.033 mmol) in 0.5 mL of DMF, HATU (25 mg, 0.066 mmol) and DIPEA (18 μL, 0.099 mmol) were added at 0°C. The mixture was stirred at 0°C for 10 minutes. Int-4 (10.34 mg, 0.036 mmol) was added. The mixture was stirred at room temperature under a nitrogen atmosphere for 1 hour. The solvent was then removed by evaporation, and the crude product was purified by column chromatography (MeOH / CH2Cl2 = 5 / 95) to obtain 1-15 g as a white solid (15 mg, 80%). MS(ESI) m / z: 569.6 [M+H] + .

[0504] Step 8 (2S,3S,4S,5R,6S)-6-(4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxy So-2,13-dioxa-4,7,10-triazatetradecyl)-2-(4-(1-(4-(5-(methylsulfonyl)-1,2,4-thiadiazole-3-yl)phenyl)-1,17-dioxo-5,8,11,14-tetraoxa-2,18-diazanonadecane-19-yl)-1H-1,2,3-triazole-1-yl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-15) [ka]

[0505] To a solution of 1-15d (20 mg, 0.018 mmol) and 1-15g (12.6 mg, 0.022 mmol) in 0.5 mL of DMF, DIPEA (6.7 μL, 0.037 mmol) was added at 0°C, followed by the addition of a catalytic amount of CuI. The mixture was degassed with N2 for 10 minutes and then stirred under a nitrogen atmosphere. The reaction was quenched with AcOH and then purified by prep-HPLC (method: column: XBridge Prep C18 OBD 5 μm 19*1250 mm; mobile phase: A-water (0.1% formic acid): B-acetonitrile; flow rate: 20 mL / min) to obtain 1-15 as a white solid after lyophilization (7 mg, 23% yield). MS(ESI) m / z: 1654.9[M+H] + .

[0506] Examples 1-16 [ka] Step 1: (9H-Fluoren-9-yl)methyl(2-(5-formyl-2-hydroxybenzamide)ethyl)carbamate(1-16b)

[0507] To a solution of 1-16a (235 mg, 1.42 mmol) in anhydrous DMF (10.0 mL), HATU (652 mg, 1.70 mmol) and DIPEA (0.50 mL, 2.83 mmol) were added and the mixture was stirred at room temperature for 10 minutes. (9H-Fluoren-9-yl)methyl(2-aminoethyl)carbamate (501 mg, 1.56 mmol) was added to the mixture and the mixture was stirred at room temperature for 20 minutes. The resulting solution was purified by prep-HPLC (Method: Column XBridge Prep C18 OBD 5um 19*150 mm; Mobile phase: A-Water (0.1% FA): B-Acetonitrile; Flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-16b (220 mg, 36.1% yield) as a white solid. MS(ESI) m / z: 431.4[M+H] + .

[0508] Step 2: (2S,3R,4S,5S,6S)-2-(2-((2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)ethyl)carbamoyl)-4-formylphenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-16c)

[0509] To a solution of 1-16b (210 mg, 0.49 mmol) in anhydrous MeCN (4 mL), (2S,3R,4S,5S,6S)-2-bromo-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate (215 mg, 0.54 mmol), 4 Å MS (500 mg), and Ag2O (232 mg, 1.95 mmol) were added under an N2 atmosphere and stirred at room temperature for 24 hours. The resulting solution was filtered and washed with MeCN (30 mL). The filtrate was concentrated and purified by silica column gel chromatography (Â / petroleum ether = 60 / 40) to obtain 1-16c (210 mg, 57.6% yield) as a white solid. MS(ESI)m / z:747.6[M+H] + .

[0510] Step 3: (2S,3R,4S,5S,6S)-2-(2-((2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)ethyl)carbamoyl)-4-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-16d) [ka]

[0511] A solution of 1-16c (210 mg, 0.28 mmol) in CH2Cl2 (4 mL) and isopropyl alcohol (1 mL) was mixed with sodium borohydride (16 mg, 0.42 mmol) at 0°C and stirred at room temperature for 1 hour. The resulting solution was quenched with saturated NH4Cl and extracted with ELISA (30 mL x 3). The organic phase was concentrated and purified by silica gel column chromatography (CH2Cl2 / MeOH = 95 / 5) to obtain 1-16d (150 mg, 71.2% yield) as a colorless oil. MS(ESI) m / z: 749.4[M+H] + .

[0512] Step 4: (2S,3R,4S,5S,6S)-2-(2-((2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)ethyl)carbamoyl)-4-((((4-nitrophenoxy)carbonyl)oxy)methyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-16e) [ka]

[0513] To a solution of 1-16d (140 mg, 0.19 mmol) in anhydrous DMF (2 mL), bis(4-nitrophenyl)carbonate (69 mg, 0.22 mmol) and DIPEA (0.05 mL, 0.28 mmol) were added at 0°C, and the mixture was stirred at room temperature for 4 hours. The resulting solution was quenched with saturated NH4Cl and extracted with siRNA (30 mL x 3). The organic phase was concentrated and purified by silica gel column chromatography (siRNA / petroleum ether = 80 / 20) to obtain 1-16e (110 mg, 64.4% yield) as a white solid. MS(ESI) m / z: 914.5[M+H]

[0514] Step 5: (2S,3R,4S,5S,6S)-2-(2-((2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)ethyl)carbamoyl)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl) Amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-16f) [ka]

[0515] A solution of 1-16e (110 mg, 0.12 mmol) and MMAE (96 mg, 0.13 mmol) in anhydrous DMF (2 mL) was mixed with HOBt (4.9 mg, 0.036 mmol) and DIPEA (0.65 mL, 0.36 mmol) at 0°C and stirred at room temperature for 24 hours. The resulting solution was quenched with saturated NH4Cl and extracted with siRNA (30 mL x 3). The organic phase was concentrated and purified by silica gel column chromatography (CH2Cl2 / MeOH = 95 / 5) to obtain 1-16f (110 mg, 61.2% yield) as a white solid. MS(ESI) m / z: 1494.3 [M+H]

[0516] Step 6: (2S,3S,4S,5R,6S)-6-(2-((2-aminoethyl)carbamoyl)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-16g) trifluoroacetate [ka]

[0517] A solution of 1-16f (110 mg, 0.074 mmol) in water (2 mL) and THF (2 mL) was mixed with 1N LiOH (0.52 mL, 0.52 mmol) at 0°C and stirred at room temperature for 2 hours. The pH was adjusted to approximately 4 by gradually adding FA. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-16 g (trifluoroacetate) (65 mg, 70.9% yield) as a white solid. MS(ESI) m / z: 1131.1[M+H] + .

[0518] Step 7: (2S,3S,4S,5R,6S)-6-(4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl Tyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-(((R)-8-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)-13,13-dimethyl-4,11-dioxo-5,12-dioxa-3,10-diazatetradecyl)carbamoyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-16h) [ka]

[0519] A solution of 1-16 g (trifluoroacetate) (35 mg, 0.028 mmol) and Int-2 (15 mg, 0.034 mmol) in anhydrous DMF (2 mL) was mixed with DIPEA (0.01 mL, 0.056 mmol) at 0°C and stirred at room temperature for 2 hours. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5 μm 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-16 H (23 mg, 56.8% yield) as a white solid. MS(ESI) m / z: 1441.4 [M+H] + .

[0520] Step 8: (2S,3S,4S,5R,6S)-6-(2-((2-((((R)-4-amino-3-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)butoxy)carbonyl)amino)ethyl)carbamoyl)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1- Phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-16)trifluoroacetate [ka]

[0521] A solution of 1-16h (23 mg, 0.016 mmol) in CH2Cl2 (1.2 mL) was mixed with TFA (0.3 mL, 3.99 mmol) at 0°C and stirred at room temperature for 30 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-16 (trifluoroacetate) (12.5 mg, 54.2% yield) as a white solid. MS(ESI) m / z: 1341.2[M+H] + .

[0522] Examples 1-17 [ka] Step 1: Methyl 4-hydroxy-3-(2-methyl-1,3-dioxolan-2-yl)benzoate (1-17b)

[0523] To a solution of compound 1-17a (4.0 g, 20.6 mmol) in toluene (40 mL), PTSA (490 mg, 2.06 mmol) and ethylene glycol (8.5 mL, 206 mmol) were added. The mixture was then prepared using 110 ml. o The mixture was stirred under an N2 atmosphere for 10 hours in 1C. The mixture was concentrated and purified by flash column chromatography (SiO2 / petroleum ether = 0-60%). Compound 1-17b (2.0 g, 40.7% yield) was obtained as a white solid. MS(ESI) m / z: 239.1[M+H] + .

[0524] Step 2: 4-(hydroxymethyl)-2-(2-methyl-1,3-dioxolan-2-yl)phenol(1-17c)

[0525] To a solution of compound 1-17b (500 mg, 2.09 mmol) in THF (5.0 mL), add LiAlH4 (159 mg, 4.19 mmol) o The compound was added with C. The mixture was stirred at 50°C for 2 hours. The mixture was quenched with water (160 μL), NaOH (160 μL, 15% in water), and H2O (0.5 mL). The mixture was filtered and concentrated to obtain compound 1-17c (300 mg, 68.3% yield) as a colorless oil. MS(ESI)m / z:211.2[M+H] + .

[0526] Step 3: 4-Hydroxy-3-(2-methyl-1,3-dioxolan-2-yl)benzaldehyde (1-17d)

[0527] A solution of compound 1-17c (300 mg, 1.428 mmol) in toluene (5.0 mL) was mixed with MnO2 (1.24 g, 14.28 mmol) at 0°C. The mixture was stirred at room temperature for 5 hours. The mixture was filtered, concentrated, and purified by flash column chromatography (toluene / petroleum ether = 0-50%). Compound 1-17d (180 mg, 60.3% yield) was obtained as a white solid. MS(ESI) m / z: 209.1[M+H] + .

[0528] Step 4: (2S,3R,4S,5S,6S)-2-(4-formyl-2-(2-methyl-1,3-dioxolan-2-yl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-17e)

[0529] To a solution of compound 1-17d (180 mg, 0.86 mmol) in MeCN (3.0 mL), (2R,3R,4S,5S,6S)-2-bromo-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate (515 mg, 1.29 mmol), 4Ams (300 mg), and Ag2O (797 mg, 3.44 mmol) were added. The mixture was stirred at room temperature for 10 hours. The mixture was filtered, concentrated, and purified by flash column chromatography (Âxy / petroleum ether = 0-50%). Compound 1-17e (250 mg, 55.4% yield) was obtained as a white solid. MS(ESI) m / z: 525.7[M+H] + .

[0530] Step 5: (2S,3R,4S,5S,6S)-2-(4-(hydroxymethyl)-2-(2-methyl-1,3-dioxolan-2-yl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-17f)

[0531] NaBH4 (21 mg, 0.57 mmol) is added to a solution of compound 1-17e (250 mg, 0.477 mmol) in THF (5.0 mL). o The compound was added with C. The mixture was stirred at 0°C for 3 hours. The mixture was quenched with saturated NH4Cl, extracted with toluene (30 mL x 3), concentrated, and purified by flash column chromatography (toluene / petroleum ether = 0-70%). Compound 1-17f (180 mg, 60.3% yield) was obtained as a colorless oil. MS(ESI) m / z: 527.3[M+H] + .

[0532] Step 6: (2S,3R,4S,5S,6S)-2-(2-acetyl-4-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate (1-17g)

[0533] To a solution of compound 1-17f (180 mg, 0.342 mmol) in DCM (3.0 mL), TFA (51 μL, 0.684 mmol) was added at 0°C. The mixture was stirred at room temperature for 3 hours. The reaction was quenched with saturated NaHCO3, extracted with HCl (30 mL x 3), concentrated, and purified by flash column chromatography (HCl / petroleum ether = 0-60%). Compound 1-17 g (140 mg, 84.9% yield) was obtained as a colorless oil. MS(ESI) m / z: 483.5[M+H] + .

[0534] Step 7: (2S,3R,4S,5S,6S)-2-(2-acetyl-4-((((4-nitrophenoxy)carbonyl)oxy)methyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate (1-17h)

[0535] A solution of 1-17 g (140 mg, 0.290 mmol) of compound and bis(4-nitrophenyl)carbonate (132 mg, 0.435 mmol) in DCM (3.0 mL) was mixed with DIPEA (101 μL, 0.58 mmol) at 0°C. The mixture was stirred at room temperature for 8 hours. The reaction product was concentrated and purified by flash column chromatography (siRNA / petroleum ether = 0-50%). Compound 1-17h (140 mg, 74.5% yield) was obtained as a yellow solid. MS(ESI) m / z: 648.7[M+H] + .

[0536] Step 8: (2S,3R,4S,5S,6S)-2-(2-acetyl-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-17i)

[0537] To a solution of compound 1-17h (140 mg, 0.216 mmol) and MMAE (155 mg, 0.216 mmol) in DMF (3.0 mL), DIPEA (113 μL, 0.648 mmol) and HOBt (7 mg, 0.054 mmol) were added at 0°C. The mixture was stirred at room temperature for 24 hours. The reaction product was concentrated and purified by flash column chromatography (MeOH / DCM = 0-5%). Compound 1-17i (160 mg, 60.3% yield) was obtained as a white solid. MS(ESI) m / z: 1227.8[M+H] + .

[0538] Step 9: (2S,3R,4S,5S,6S)-2-(2-((E)-1-((2-aminoethoxy)imino)ethyl)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy -2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-17j) [ka]

[0539] To a solution of compound 1-17i (160 mg, 0.130 mmol) in EtOH (3.0 mL), add ethaneamine, 2-(aminooxy)-, dihydrochloride (97 mg, 0.65 mmol) o The mixture was added with C. The mixture was stirred at 50°C for 5 hours. The reaction was concentrated to obtain the crude product 1-17j. MS(ESI)m / z:1284.8[M+H] + .

[0540] Step 10: (2S,3S,4S,5R,6S)-6-(2-((E)-1-((2-aminoethoxy)imino)ethyl)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-17k) [ka]

[0541] A solution of crude compound 1-17j in EtOH (3.0 mL) was mixed with LiOH (780 μL, 0.78 mmol, 1 N in water) at 0°C. The mixture was stirred at room temperature for 0.5 hours. The reaction was quenched with AcOH and purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min). The fraction was lyophilized to obtain 1-17k (trifluoroacetate) (75 mg, 50.4% yield, 2 steps) as a white solid. MS(ESI) m / z: 1145.4 [M+H] + .

[0542] Step 11: (2S,3S,4S,5R,6S)-6-(4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3 ,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-((R,E)-12-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)-17,17-dimethyl-8,15-dioxo-4,9,16-trioxa-3,7,14-triazaoctades-2-en-2-yl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid 1-17l [ka]

[0543] A solution of crude compounds 1-17k (30 mg, 0.0262 mmol) and Int-2 (13 mg, 0.0288 mmol) in DMF (1.0 mL) was mixed with DIPEA (13 μL, 0.0786 mmol) at 0°C. The mixture was stirred at room temperature for 5 hours. The reaction product was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5 μm 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-17l (trifluoroacetate) (20 mg, 52.5% yield) as a white solid. MS(ESI) m / z: 1456.1 [M+H] + .

[0544] Step 12: (2S,3S,4S,5R,6S)-6-(2-((R,E)-13-amino-12-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)-8-oxo-4,9-dioxa-3,7-diazatrides-2-en-2-yl)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R )-1-Hydroxy-1-phenylpropane-2-yl)amino)-1-Methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid 1-17 [ka]

[0545] To a solution of 1-17 liters (20 mg, 0.0175 mmol) of crude compound in DCM (1.0 mL), add TFA (0.2 mL). o The mixture was added with C. The mixture was stirred at room temperature for 10 minutes. The reaction product was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-17 (trifluoroacetate) (10 mg, 42.2% yield) as a white solid. MS(ESI) m / z: 1355.3[M+H] + .

[0546] Examples 1-18 [ka]

[0547] Step 1: (7R,17S)-17-((3-((5-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-(((2S,3R,4S, 5S,6S)-6-carboxy-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)oxy)phenyl)amino)-3-oxopropyl)carbamoyl)-7-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)-2,2,55,55-tetramethyl-4,11,15,23,51-pentaoxo-3,10,26,29,32,35,38,41,44,47-decaoxa-5,12,16,22,50,55-hexaazaoctapentacontan-55-ium-58-sulfonate(1-18a) [ka]

[0548] To a solution of Int-3 (25 mg, 0.063 mmol) in anhydrous DMF (2 mL), HATU (24 mg, 0.063 mmol) and DIPEA (0.019 mL, 0.10 mmol) were added and the mixture was stirred at room temperature for 10 minutes. Then, 1-2 m (100 mg, 0.052 mmol) was added to the mixture and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5 μm 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-18a (55 mg, 45.9% yield) as a white solid. MS(ESI) m / z: 2298.9[M+H]

[0549] Step 2: (42S,52R)-53-amino-42-((3-((5-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-((( 2S,3R,4S,5S,6S)-6-carboxy-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)oxy)phenyl)amino)-3-oxopropyl)carbamoyl)-52-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)-4,4-dimethyl-8,36,44,48-tetraoxo-12,15,18,21,24,27,30,33,49-nonanoxa-4,9,37,43,47-pentazatripentacontan-4-ium-1-sulfonate(1-18)trifluoroacetate [ka]

[0550] A solution of 1-18a (55 mg, 0.024 mmol) in CH2Cl2 (1.2 mL) was mixed with TFA (0.3 mL, 5.98 mmol) at 0°C and stirred at room temperature for 30 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-18 (trifluoroacetate) (30 mg, 54.2% yield) as a white solid. MS(ESI) m / z: 2199.3[M+H] + .

[0551] Examples 1-19 [ka] Step 1: (2S,3S,4S,5R,6S)-6-(4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-di Isopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-((E)-1-((2-(4-(5-(methylsulfonyl)-1,2,4-thiadiazole-3-yl)benzamide)ethoxy)imino)ethyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-19)

[0552] To a solution of crude compound 1-17k (20 mg, 0.0175 mmol) and Int-4 (6 mg, 0.0192 mmol) in DMF (1.0 mL), HATU (7.3 mg, 0.0192 mmol) and DIPEA (9 μL, 0.0525 mmol) were added at 0°C. The mixture was stirred at room temperature for 1 hour. The reaction product was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5 μm 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-19 (trifluoroacetate) (10 mg, 40.5% yield) as a white solid. MS(ESI) m / z: 1411.4[M+H] + .

[0553] Examples 1-20 [ka] Step 1: (2R,3R,4R,5S)-1-((tert-butyldiphenylsilyl)oxy)-6-(methylamino)hexane-2,3,4,5-tetraol(1-20b)

[0554] TBDPSCl was added at 0°C to a solution of 1-20a (1.00 g, 5.1 mmol) and imidazole (0.37 g, 5.4 mmol) in DMF (10 mL). The mixture was stirred at room temperature for 3 hours and purified by flash column chromatography (MeOH / CH2Cl2 = 10 / 90) to obtain 1-20b as a white solid (2.2 g, 99.0% yield). MS(ESI) m / z: 434.6 [M+H] +

[0555] Step 2: (2S,3S,4R,5S)-6-(((2S,3R,4R,5R)-6-((tert-butyldiphenylsilyl)oxy)-2,3,4,5-tetrahydroxyhexyl)(methyl)amino)-2,3,4,5-tetrahydroxyhexanoic acid (1-20c)

[0556] NaCNBH3 was added to a solution of 1-20b (0.50 g, 1.2 mmol) and glucuronic acid (0.67 g, 3.5 mmol) in 10 mL of MeOH. The mixture was stirred at room temperature at 60°C for 4 hours and quenched with aqueous NH4Cl. The mixture was purified by prep-HPLC (column: Xbridge Prep C18 OBD (trademark) 5 μm, 19*150 mm; mobile phase A: 0.1% FA in water, B: MeCN; gradient: 20%~50% B; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-20c (200 mg, 28.4% yield) as a white solid. MS(ESI) m / z: 612.7[M+H] +

[0557] Step 3: (2S,3S,4R,5S)-2,3,4,5-tetraacetoxy-6-(methyl((2S,3R,4R,5R)-2,3,4,5-tetraacetoxy-6-((tert-butyldiphenylsilyl)oxy)hexyl)amino)hexanoic acid (1-20d)

[0558] A solution of 1-20c (200 mg, 0.33 mmol) in acetic anhydride (5 mL) was mixed with DMAP (40 mg, 0.33 mmol). The mixture was stirred at room temperature for 2 hours and purified by flash column chromatography (petroleum ether / Â=50 / 50) to obtain 1-20d as a white solid (200 mg, 64.5% yield). MS(ESI)m / z:948.9[M+H] +

[0559] Step 4: (9S,10S,11R,12S,16S,17R,18R,19R)-1-(9H-fluoren-9-yl)-14,23,23-trimethyl-3,8-dioxo-22,22-diphenyl-2,21-dioxa-4,7,14-triaza-22-silatetracosan-9,10,11,12,16,17,18,19-octileoctaacetate(1-20e)

[0560] A solution of 1-20d (200 mg, 0.21 mmol) and Fmoc-DEA (80 mg, 0.25 mmol) in 2 mL of anhydrous DMF was mixed with DIPEA (136 μL, 0.78 mmol) and HATU (160 mg, 0.42 mmol). The mixture was stirred at room temperature for 3 hours. The mixture was partitioned between water and ethyl acetate. The organic layer was concentrated and purified by flash column chromatography (petroleum ether / siRNA = 34 / 66) to obtain 1-20e as a white solid (250 mg, 97.8% yield). MS(ESI) m / z: 1213.6[M+H] +

[0561] Step 5: (6R,7R,8R,9S,13S,14R,15S,16S)-20-amino-2,2,11-trimethyl-17-oxo-3,3-diphenyl-4-oxa-11,18-diaza-3-silicosan-6,7,8,9,13,14,15,16-octileoctaacetate(1-20f)

[0562] To a solution of 1-20e (275 mg, 0.23 mmol) in 2 mL of anhydrous THF, 200 μL of piperidine was added. The mixture was stirred at room temperature for 3 hours and concentrated to obtain the crude product 1-20f without further purification. MS(ESI)m / z:991.1[M+H] +

[0563] Process 6: (5S,30S,31S,32R,33S,37S,38R,39R,40R)-5-(tert-butoxycarbonyl)-1-(9H-fluoren-9-yl)-35,44,44-trimethyl-3,8,24,29-tetraoxo-43,43-diphenyl-2,12,15,18,21,42-hexaoxa-4,9,25,28,35-pentaza-43-silapentatetracontane-30,31,32,33,37,38,39,40-octileoctaacetate (1-20h) [ka]

[0564] HATU (127 mg, 0.33 mmol) was added to a solution of 1-20f, 1-20g (150 mg, 0.22 mmol) and DIPEA (78 μL, 0.45 mmol) in anhydrous DMF (2 mL). The mixture was stirred at room temperature for 3 hours and purified by flash column chromatography (MeOH / CH2Cl2 = 10 / 90) to obtain 1-20h as a white solid (189 mg, 51.5% yield in 2 steps). MS(ESI) m / z: 1646.3 [M+H] +

[0565] Step 7: (6R,7R,8R,9S,13S,14R,15S,16S,41S)-41-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)-6,7,8,9,13,14,15,16-Octaacetoxy-2,2,11-Trimethyl-17,22,38-Trioxo-3,3-Diphenyl-4,25,28,31,34-Pentaoxa-11,18,21,37-Tetraaza-3-Siladtetracontan-42-Acid(1-20i) [ka]

[0566] To a solution of 1-20h (174 mg, 0.11 mmol) in 2 mL of anhydrous CH2Cl2, 400 μL of TFA was added. The mixture was stirred at room temperature for 3 hours and concentrated to obtain crude product 1-20i without further purification. MS(ESI)m / z: 1587.8[MH] + ]

[0567] Step 8: (2S,3S,4S,5R,6S)-6-(2-((6R,7R,8R,9S,13S,14R,15S,16S,41S)-41-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)-6,7,8,9,13,14,15,16-Octaacetoxy-2,2,11-Trimethyl-17,22,38,42-Tetraoxo-3,3-Diphenyl-4,25,28,31,34-Pentaoxa-11,18,21,37,43-Pentaza-3-Silahexatetracontane-46-amide)-4-((5S,8S, 11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-20j) [ka]

[0568] 1-2k (120 mg, 0.11 mmol) was added to a solution of 1-20i, HATU (40 mg, 0.11 mmol), and DIPEA (37 μL, 0.21 mmol) in anhydrous DMF (2 mL). The mixture was stirred at room temperature for 6 hours and purified by prep-HPLC (column: Xbridge Prep C18 OBD® 5 μm, 19*150 mm; mobile phase A: 0.1% FA in water, B: MeCN; gradient: 25%~60% B; flow rate: 20 mL / min). The fraction was lyophilized to obtain 1-20j as a white solid (44 mg, 15.4% yield in 2 steps). MS(ESI) m / z: 2699.3[M+H] +

[0569] Step 9: (2S,3S,4S,5R,6S)-6-(2-((6S,31S,32S,33R,34S,38S,39R,40R,41R)-6-amino-31,32,33,34,38,39,40,41,42-nonahydroxy-36-methyl-5,9,25,30-tetraoxo-13,16,19,22-tetraoxa-4,10,26,29,36-pentaazadotetracontanamid)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-1 2-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-20k) [ka]

[0570] TBAF was added to a solution of 1-20j (44 mg, 0.016 mmol) in 2 mL of anhydrous THF. The mixture was stirred at 40°C for 3 hours. Then, 1N LiOH was added, and the mixture was stirred at room temperature for a further 20 minutes, followed by quenching with 20 μL of AcOH. The mixture was purified by prep-HPLC (column: Xbridge Prep C18 OBD (trademark) 5 μm, 19*150 mm; mobile phase A: 0.1% FA in water, B: MeCN; gradient: 20%~55% B; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-20k as a white solid (20 mg, 64.5% yield). MS(ESI) m / z: 1905.5[M+H] +

[0571] Step 10: (2S,3S,4S,5R,6S)-6-(4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-((6S,31S,32S,33R,34S,38S,39 R,40R,41R)-6-((R)-7-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)-2,2-dimethyl-4,11-dioxo-3,10-dioxa-5,12-diazapentadecane-15-amide)-31,32,33,34,38,39,40,41,42-nonahydroxy-36-methyl-5,9,25,30-tetraoxo-13,16,19,22-tetraoxa-4,10,26,29,36-pentazadotetracontanaamide)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-20l) [ka]

[0572] 1-20k (20 mg, 0.011 mmol) was added to a solution of Int-3 (4.3 mg, 0.011 mmol), HATU (4.1 mg, 0.011 mmol), and DIPEA (2.8 μL, 0.016 mmol) in anhydrous DMF (2 mL). The mixture was stirred at room temperature for 1 hour and purified by prep-HPLC (column: Xbridge Prep C18 OBD® 5 μm, 19*150 mm; mobile phase A: 0.1% FA in water, B: MeCN; gradient: 25%~65% B; flow rate: 20 mL / min). The fraction was lyophilized to obtain 1-20 L as a white solid (8.4 mg, 34.1% yield). MS(ESI) m / z: 2286.5[M+H] +

[0573] Step 11: (2S,3S,4S,5R,6S)-6-(2-((6S,31S,32S,33R,34S,38S,39R,40R,41R)-6-(3-((((R)-4-amino-3-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)butoxy)carbonyl)amino)propanamide)-31,32,33,34,38,39,40,41,42-nonahydroxy-36-methyl-5,9,25,30-tetraoxo-13,16,19,22-tetraoxa-4,10,26,29,36-pentazadotetracontanamide)-4 -((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid(1-20) [ka]

[0574] TFA (200 μL) was added to a solution of 1-20 L (8.4 mg, 3.7 μmol) in anhydrous CH2Cl2 (1 mL). The mixture was stirred at room temperature for 30 minutes and purified by prep-HPLC (column: Xbridge Prep C18 OBD (trademark) 5 μm, 19*150 mm; mobile phase A: 0.1% TFA in water, B: MeCN; gradient: 25%~55% B; flow rate: 20 mL / min). The fractions were lyophilized to obtain 1-20 as a white solid (5.5 mg, 68.5% yield). MS(ESI) m / z: 2186.9 [M+H] +

[0575] Examples 1-21 [ka] Step 1: Synthesis of compound 1-21b

[0576] To a solution of 1-21a (80 mg, 0.082 mmol) and 1-4b (45 mg, 0.091 mmol) in anhydrous DMF (1 mL), HATU (38 mg, 0.099 mmol) and DIPEA (0.045 mL, 0.25 mmol) were added and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5 μm 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-21b (89 mg, 75% yield) as a white solid. MS(ESI) m / z: 1442.5 [M+H] +

[0577] Step 2: Synthesis of Compound 1-21c [ka]

[0578] To a solution of 1-21b (69 mg, 0.026 mmol) in anhydrous DMF (2 mL), Et2NH (0.19 mL, 1.85 mmol) was added and the mixture was stirred at room temperature for 30 minutes. The resulting solution was concentrated to obtain the crude product 1-21c (75 mg, 99.6% yield), which was used directly in the next step. MS(ESI)m / z:1220.2[M+H] +

[0579] Step 3: Synthesis of compound 1-21d [ka]

[0580] To a solution of 1-21c (75 mg, 0.062 mmol) and Fmoc-L-glutamic acid 1-tert-butyl ester (29 mg, 0.068 mmol) in anhydrous DMF (1 mL), HATU (28 mg, 0.074 mmol) and DIPEA (0.033 mL, 0.18 mmol) were added and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5 μm 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-21d (79 mg, 79% yield) as a white solid. MS(ESI) m / z: 1627.7[M+H] +

[0581] Step 4: Synthesis of compound 1-21e [ka]

[0582] To a solution of 1-21d (79 mg, 0.049 mmol) in DCM (2 mL), TFA (0.36 mL, 4.86 mmol) was added and the mixture was stirred at room temperature for 3 hours. The mixture was concentrated and the solvent was removed. The residue was dissolved in THF (1 mL) and MeOH (1 mL). 1N LiOH (0.73 mL, 0.33 mmol) and 30% H2O2 (0.31 mL, 1.33 mmol) were added to the above mixture and the mixture was stirred at 0°C for 1 hour. The resulting solution was quenched with AcOH and saturated Na2S2O3, concentrated, and the solvent was removed. The residue was purified by prep-HPLC (Method: Column XBridge Prep C18 OBD 5um 19*150mm; Mobile phase: A-Water (0.1% FA): B-Acetonitrile; Flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-21e (58 mg, 76% yield) as a white solid. MS(ESI) m / z: 1571.9[M+H]

[0583] Step 5: Synthesis of compound 1-21f [ka]

[0584] To a solution of 1-21e (58 mg, 0.037 mmol) in anhydrous DMF (1 mL), HATU (14 mg, 0.037 mmol) and DIPEA (0.02 mL, 0.11 mmol) were added and the mixture was stirred at room temperature for 10 minutes. 1-2k (46 mg, 0.037 mmol) was added to the mixture and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-21f (69 mg, 69.6% yield) as a white solid. MS(ESI) m / z: 2682.5[M+H] +

[0585] Step 6: Synthesis of 1-21 g of compound [ka]

[0586] Et2NH (0.081 mL, 0.77 mmol) was added to a solution of 1-21f (69 mg, 0.026 mmol) in anhydrous DMF (2 mL) and stirred at room temperature for 30 minutes. The resulting solution was concentrated to obtain 1-21 g (60 mg, 94.8% yield) of crude product, which was used directly in the next step. MS(ESI)m / z:2461.0[M+H] +

[0587] Step 7: Synthesis of compound 1-21h [ka]

[0588] To a solution of Int-3 (10 mg, 0.024 mmol) in anhydrous DMF (1 mL), HATU (10 mg, 0.024 mmol) and DIPEA (0.013 mL, 0.073 mmol) were added and the mixture was stirred at room temperature for 10 minutes. 1-21 g (60 mg, 0.024 mmol) was added to the mixture and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-21 h (40 mg, 57.7% yield) as a white solid. MS(ESI) m / z: 1422.3 [M / 2+H] +

[0589] Step 8: Synthesis of Compounds 1-21 [ka]

[0590] A solution of 1-21h (40 mg, 0.014 mmol) in DCM (2 mL) was mixed with TFA (0.38 mL, 4.93 mmol) and stirred at room temperature for 30 minutes. The resulting solution was concentrated and the solvent was removed. The residue was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5 μm 19*150 mm; mobile phase: A-water (0.05% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-21 (trifluoroacetate) (21 mg, 52.2% yield) as a white solid. MS(ESI) m / z: 1372.4 [M / 2+H] +

[0591] Examples 1-22 [ka] Step 1: Synthesis of compound 1-22a

[0592] To a solution of 1-6a (48 mg, 0.072 mmol) and 1-21a (70 mg, 0.072 mmol) in anhydrous DMF (2 mL), HATU (39 mg, 0.1 mmol) and DIPEA (0.033 mL, 0.18 mmol) were added, and the mixture was stirred at room temperature for 1 hour. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5 μm 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-22a (90 mg, 77.2% yield) as a colorless oil. MS(ESI) m / z: 1618.6[M+H] +

[0593] Step 2: Synthesis of compound 1-22b

[0594] To a solution of 1-22a (90 mg, 0.056 mmol) in anhydrous DMF (2 mL), Et2NH (0.18 mL, 1.67 mmol) was added and the mixture was stirred at room temperature for 30 minutes. The resulting solution was concentrated to obtain the crude product 1-22b (78 mg, 100.5% yield), which was used directly in the next step. MS(ESI)m / z:1396.0[M+H]+

[0595] Step 3: Synthesis of Compound 1-22c

[0596] To a solution of 1-22b (78 mg, 0.056 mmol) and Fmoc-L-glutamic acid 1-tert-butyl ester (26 mg, 0.061 mmol) in anhydrous DMF (2 mL), HATU (26 mg, 0.067 mmol) and DIPEA (0.025 mL, 0.14 mmol) were added and the mixture was stirred at room temperature for 1 hour. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5 μm 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-22c (63 mg, 62.5% yield) as a colorless oil. MS(ESI) m / z: 1804.0[M+H] +

[0597] Step 4: Synthesis of compound 1-22d [ka]

[0598] To a solution of 1-22c (40 mg, 0.022 mmol) in DCM (2 mL), TFA (0.17 mL, 2.22 mmol) was added and the mixture was stirred at room temperature for 3 hours. The mixture was concentrated and the solvent was removed. The residue was dissolved in THF (1 mL) and MeOH (1 mL). 1N LiOH (0.29 mL, 0.33 mmol) and H2O2 (0.14 mL, 1.33 mmol) were added to the above mixture at 0°C and the mixture was stirred at 0°C for 1 hour. The resulting solution was quenched with saturated Na2S2O3 and AcOH, concentrated, and the solvent was removed. The residue was purified by prep-HPLC (Method: Column XBridge Prep C18 OBD 5um 19*150mm; Mobile phase: A-Water (0.1% FA): B-Acetonitrile; Flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-22d (25 mg, 64.5% yield) as a colorless oil. MS(ESI) m / z: 1747.7[M+H]

[0599] Step 5: Synthesis of compound 1-22e [ka]

[0600] To a solution of 1-22d (25 mg, 0.014 mmol) in anhydrous DMF (1 mL), HATU (6 mg, 0.016 mmol) and DIPEA (0.005 mL, 0.029 mmol) were added and the mixture was stirred at room temperature for 10 minutes. 1-2k (20 mg, 0.016 mmol) was added to the mixture and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-22e (12 mg, 62.5% yield) as a white solid. MS(ESI) m / z: 1430.4 [M / 2+H] +

[0601] Step 6: Synthesis of compound 1-22f [ka]

[0602] To a solution of 1-22e (12 mg, 0.004 mmol) in anhydrous DMF (1 mL), Et2NH (0.013 mL, 0.13 mmol) was added and the mixture was stirred at room temperature for 30 minutes. The resulting solution was concentrated to obtain the crude product 1-22f (11 mg, 99.4% yield), which was used directly in the next step. MS(ESI)m / z:1319.4[M / 2+H] +

[0603] Step 7: Synthesis of 1-22g of compound [ka]

[0604] To a solution of Int-3 (2.8 mg, 0.007 mmol) in anhydrous DMF (0.5 mL), HATU (2.7 mg, 0.007 mmol) and DIPEA (0.002 mL, 0.013 mmol) were added and the mixture was stirred at room temperature for 10 minutes. 1-22f (11 mg, 0.006 mmol) was added to the mixture and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-22 g (10 mg, 52.6% yield) as a white solid. MS(ESI) m / z: 1510.4 [M / 2+H] +

[0605] Step 8: Synthesis of Compounds 1-22 [ka]

[0606] Compound 1-22 (6 mg, 62% yield) was synthesized according to the synthesis procedure of step 11 in Example 1-2. MS(ESI)m / z:1460.3[M / 2+H] +

[0607] Examples 1-23 [ka] Step 1: (2S,3S,4S,5R,6S)-6-(2-(((S)-42-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-39,43-dioxo-2,5,8,11,14,17,20,23,26,29,32,35-dodecaoxa-38,44-diazahexatetracontan-46-yl)carbamoyl)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-( (1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-23a) [ka]

[0608] To a solution of 1-10c (111 mg, 0.12 mmol) in anhydrous DMF (2 mL), HATU (46 mg, 0.12 mmol) and DIPEA (0.065 mL, 0.36 mmol) were added and the mixture was stirred at room temperature for 10 minutes. 1-16 g (150 mg, 0.12 mmol) was added to the mixture and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-23a (150 mg, 61.5% yield) as a white solid. MS(ESI) m / z: 2024.9[M+H]+

[0609] Process 2: (2S,3S,4S,5R,6S)-6-(2-(((S)-42-amino-39,43-dioxo-2,5,8,11,14,17,20,23,26,29,32,35-dodecaoxa-38,44-diazahexatetracontan-46-yl)carbamoyl)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S ,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-23b) [ka]

[0610] To a solution of 1-23a (150 mg, 0.074 mmol) in anhydrous DMF (2 mL), Et2NH (0.23 mL, 2.22 mmol) was added and the mixture was stirred at room temperature for 30 minutes. The resulting solution was concentrated to obtain the crude product 1-23b (125 mg, 93.6% yield), which was used directly in the next step. MS(ESI)m / z:1802.5[M+H]+

[0611] Step 3: (2S,3S,4S,5R,6S)-6-(4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2- (((S)-42-((R)-7-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)-2,2-dimethyl-4,11-dioxo-3,10-dioxa-5,12-diazapentadecane-15-amide)-39,43-dioxo-2,5,8,11,14,17,20,23,26,29,32,35-dodecaoxa-38,44-diazahexatetracontan-46-yl)carbamoyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-23c) [ka]

[0612] To a solution of Int-3 (28 mg, 0.069 mmol) in anhydrous DMF (2 mL), HATU (26.7 mg, 0.069 mmol) and DIPEA (0.037 mL, 0.21 mmol) were added and the mixture was stirred at room temperature for 10 minutes. 1-23b (125 mg, 0.069 mmol) was added to the mixture and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150 mm; mobile phase: A-water (0.1% FA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-23c (100 mg, 66% yield) as a white solid. MS(ESI) m / z: 2184.0[M+H]+

[0613] Step 4: (2S,3S,4S,5R,6S)-6-(2-(((S)-42-(3-((((R)-4-amino-3-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)butoxy)carbonyl)amino)propanamide)-39,43-dioxo-2,5,8,11,14,17,20,23,26,29,32,35-dodecaoxa-38,44-diazahexatetracontan-46-yl)carbamoyl)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl) -12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-23)trifluoroacetate [ka]

[0614] A solution of 1-23c (100 mg, 0.046 mmol) in DCM (2 mL) was mixed with TFA (0.38 mL, 4.93 mmol) and stirred at 0°C for 30 minutes. The resulting solution was concentrated and the solvent was removed. The residue was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5 μm 19*150 mm; mobile phase: A-water (0.05% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-23 (trifluoroacetate) (49 mg, 48.7% yield) as a white solid. MS(ESI) m / z: 2083.3[M+H]+

[0615] Examples 1-24 [ka] Step 1: (2S,3S,4S,5R,6S)-6-(4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl -3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-((R,E)-16-(2,5-dioxopyrrolidine-1-yl)-21,21-dimethyl-8,12,19-trioxo-4,13,20-trioxa-3,7,11,18-tetraazadocos-2-en-2-yl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid 1-24a [ka]

[0616] To a solution of Int-3 (10.5 mg, 0.026 mmol) in anhydrous DMF (1 mL), HATU (10 mg, 0.026 mmol) and DIPEA (9 μL, 0.053 mmol) were added and the mixture was stirred at room temperature for 10 minutes. Then, 1-17k (20 mg, 0.0175 mmol) was added to the mixture and the mixture was stirred at room temperature for 10 minutes. The resulting solution was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5 μm 19*150 mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-24a (14 mg, 52.4% yield) as a white solid. MS(ESI) m / z: 1526.1[M+H]

[0617] Step 2: (2S,3S,4S,5R,6S)-6-(2-((R,E)-17-amino-16-(2,5-dioxopyrrolidine-1-yl)-8,12-dioxo-4,13-dioxa-3,7,11-triazaheptades-2-en-2-yl)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R) -1-Hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid 1-24 [ka]

[0618] To a solution of crude compound 1-24a (14 mg, 0.0091 mmol) in DCM (1.0 mL), add TFA (0.2 mL) o The mixture was added at C. The mixture was stirred at room temperature for 10 minutes. The reaction product was purified by prep-HPLC (method: column XBridge Prep C18 OBD 5um 19*150mm; mobile phase: A-water (0.1% TFA): B-acetonitrile; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-24 (trifluoroacetate) (7 mg, 53.9% yield) as a white solid. MS(ESI) m / z: 1426.8[M+H] + .

[0619] Examples 1-25 [ka] Step 1: (2S,3R,4S,5S,6S)-2-(2-amino-4-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate (1-4a)

[0620] PtO2 (22.5 mg, 15% w / w) was added to a solution of Int-1a (2.00 g, 4.1 mmol) in 10 mL of THF. The mixture was stirred at room temperature under an H2 atmosphere for 2 hours. The suspension was filtered through Celite, and the filtrate was concentrated to obtain 1-4a as a colorless oil without further purification. MS(ESI) m / z: 456.6[M+H] +

[0621] Step 2: (2S,3R,4S,5S,6S)-2-(2-((2-((tert-butoxycarbonyl)amino)ethyl)amino)-4-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-25c)

[0622] NaCNBH3 (778 mg, 12.4 mmol) was added to a solution of 1-4a and tert-butyl(2-oxoethyl)carbamate (1.31 g, 8.3 mmol) in 10 mL of MeOH. The mixture was stirred at room temperature for 3 hours. The mixture was partitioned between water and ethyl acetate. The organic layer was concentrated and purified by flash column chromatography (petroleum ether / ethyl acetate = 50 / 50) to obtain 1-25c (1.68 g, 68.0% yield in 2 steps) as a white solid. MS(ESI) m / z: 599.6[M+H] +

[0623] Step 3: (2S,3R,4S,5S,6S)-2-(2-((2-((tert-butoxycarbonyl)amino)ethyl)amino)-4-(((tert-butyldimethylsilyl)oxy)methyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate(1-25d)

[0624] TBSCl (423 mg, 2.8 mmol) was added to a solution of 1-25c (1.68 g, 2.8 mmol), imidazole (191 mg, 2.8 mmol), and DMAP (86 mg, 0.7 mmol) in 4 mL of DMF. The mixture was stirred at room temperature for 3 hours. The mixture was partitioned between water and ethyl acetate. The organic layer was concentrated and purified by flash column chromatography (PE / EA = 60 / 40) to obtain 1-25d (1.51 g, 75.5% yield) as a white solid. MS(ESI) m / z: 713.8 [M+H] +

[0625] Step 4: 2-((2-((tert-butoxycarbonyl)amino)ethyl)(4-(((tert-butyldimethylsilyl)oxy)methyl)-2-(((2S,3R,4S,5S,6S)-3,4,5-triacetoxy-6-(methoxycarbonyl)tetrahydro-2H-pyran-2-yl)oxy)phenyl)amino)-N,N,N-trimethyl-2-oxoethane-1-aminium(1-25e)

[0626] A solution of 1-25d (1.00 mg, 1.4 mmol), betaine (237 mg, 1.5 mmol), and pyridine (2.3 mL, 28.0 mmol) in 10 mL of CH3CN was mixed with POCl3 (144 μL, 1.5 mmol) at 0°C. The mixture was stirred at room temperature for 1 hour. The mixture was quenched with 5 mL of H2O and concentrated to obtain crude 1-25e as a white solid. MS(ESI)m / z:814.0[M+H] +

[0627] Step 5: 2-((2-((tert-butoxycarbonyl)amino)ethyl)(4-(hydroxymethyl)-2-(((2S,3R,4S,5S,6S)-3,4,5-triacetoxy-6-(methoxycarbonyl)tetrahydro-2H-pyran-2-yl)oxy)phenyl)amino)-N,N,N-trimethyl-2-oxoethane-1-aminium(1-25f)

[0628] Crude 1-25e was dissolved in AcOH / THF / H2O (6 / 2 / 2 mL) and stirred at room temperature for 30 minutes. The mixture was purified by prep-HPLC (column: Xbridge Prep C18 OBD (trademark) 5 μm, 19*150 mm; mobile phase A: 0.1% FA in water, B: MeCN; gradient: 25%~60% B; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-25f (500 mg, 51.0% yield in 2 steps) as a white solid. MS(ESI) m / z: 698.8 [M+H] +

[0629] Step 6: 2-((2-((tert-butoxycarbonyl)amino)ethyl)(5-((((4-nitrophenoxy)carbonyl)oxy)methyl)-2-(((2S,3R,4S,5S,6S)-3,4,5-triacetoxy-6-(methoxycarbonyl)tetrahydro-2H-pyran-2-yl)oxy)phenyl)amino)-N,N,N-trimethyl-2-oxoethane-1-aminium (1-25g)

[0630] DIPEA (125 μL, 0.72 mmol) was added to a solution of 1-25f (250 mg, 0.36 mmol) and bis(4-nitrophenyl)carbonate (125 mg, 0.41 mmol) in anhydrous CH2Cl2 (3 mL). The mixture was stirred at room temperature for 3 hours and purified by prep-HPLC (column: Xbridge Prep C18 OBD® 5 μm, 19*150 mm; mobile phase A: 0.1% FA in water, B: MeCN; gradient: 40%~60% B; flow rate: 20 mL / min). The fraction was lyophilized to obtain 1-25 g (145 mg, 46.9% yield) as a white solid. MS(ESI) m / z: 863.9 [M+H] +

[0631] Step 7: 2-((2-((tert-butoxycarbonyl)amino)ethyl)(5-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl) -5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-(((2S,3R,4S,5S,6S)-3,4,5-triacetoxy-6-(methoxycarbonyl)tetrahydro-2H-pyran-2-yl)oxy)phenyl)amino)-N,N,N-trimethyl-2-oxoethane-1-aminium(1-25h) [ka]

[0632] To a solution of 1-25 g (145 mg, 0.17 mmol) in 4 mL of DMF, HOBT (5.7 mg, 0.042 mmol) and DIPEA (58 μL, 0.34 mmol) were added, followed by MMAE (121 mg, 0.17 mmol). The mixture was stirred at room temperature for 24 hours and purified by prep-HPLC (column: Xbridge Prep C18 OBD® 5 μm, 19*150 mm; mobile phase A: 0.1% FA in water, B: MeCN; gradient: 25%~65% B; flow rate: 20 mL / min). The fraction was lyophilized to obtain 1-25 h (180 mg, 74.3% yield) as a white solid. MS(ESI) m / z: 1442.7[M+H] +

[0633] Step 8: 2-((2-aminoethyl)(5-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl Ropyru-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-(((2S,3R,4S,5S,6S)-3,4,5-triacetoxy-6-(methoxycarbonyl)tetrahydro-2H-pyran-2-yl)oxy)phenyl)amino)-N,N,N-trimethyl-2-oxoethane-1-aminium(1-25i) [ka]

[0634] 1-25h (100 mg, 0.069 mmol) was dissolved in anhydrous CH2Cl2 (1 mL), and 300 μL of HCl (4 M in 1,4-dioxane) was added. The mixture was stirred at room temperature for 30 minutes, concentrated, and crude 1-25i was obtained as a white solid without further purification. MS(ESI) m / z: 1342.8 [M+H] +

[0635] Step 9: (S)-42-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-47-(5-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6, 9-Trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-(((2S,3R,4S,5S,6S)-3,4,5-triacetoxy-6-(methoxycarbonyl)tetrahydro-2H-pyran-2-yl)oxy)phenyl)-N,N,N-trimethyl-39,43,48-trioxo-2,5,8,11,14,17,20,23,26,29,32,35-dodecaoxa-38,44,47-triazanonatetracontan-49-aminium(1-25j) [ka]

[0636] Crude 1-25i was dissolved in anhydrous DMF (1 mL). 1-10c (65 mg, 0.070 mmol), HATU (32 mg, 0.084 mmol), and DIPEA (37 μL, 0.21 mmol) were added to the mixture. The mixture was stirred at room temperature for 3 hours and purified by prep-HPLC (column: Xbridge Prep C18 OBD® 5 μm, 19*150 mm; mobile phase A: 0.1% FA in water, B: MeCN; gradient: 25%~60% B; flow rate: 20 mL / min). The fraction was lyophilized to obtain 1-25j (78 mg, 49.7% yield in 2 steps) as a white solid. MS(ESI) m / z: 2235.8[M+H] +

[0637] Step 10: (S)-42-amino-47-(5-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,1 3-Dioxa-4,7,10-Triazatetradecyl)-2-(((2S,3R,4S,5S,6S)-6-Carboxy-3,4,5-Trihydroxytetrahydro-2H-pyran-2-yl)oxy)phenyl)-N,N,N-Trimethyl-39,43,48-Trioxo-2,5,8,11,14,17,20,23,26,29,32,35-Dodecaoxa-38,44,47-Triazanonatetracontan-49-Aminium(1-25k) [ka]

[0638] A solution of 1-25j in THF (2 mL) and MeOH (2 mL) was mixed with 1N LiOH (209 μL) at 0°C. The mixture was stirred at room temperature for 15 minutes and quenched with 200 μL of AcOH. The crude product was purified by prep-HPLC (column: Sunfire Prep C18 OBD (trademark) 5 μm, 19*150 mm; mobile phase A: 0.1% FA in water, B: MeCN; gradient: 25%~55% B; flow rate: 20 mL / min), and the fraction was lyophilized to obtain 1-25k (50 mg, 76.5% yield) as a white solid. MS(ESI) m / z: 1873.7[M+H] +

[0639] Step 11: (S)-47-(5-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-(((2S,3R,4S,5S,6S)-6-cal Boxy-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)oxy)phenyl)-42-((R)-7-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)-2,2-dimethyl-4,11-dioxo-3,10-dioxa-5,12-diazapentadecane-15-amide)-N,N,N-trimethyl-39,43,48-trioxo-2,5,8,11,14,17,20,23,26,29,32,35-dodecaoxa-38,44,47-triazanonatetracontan-49-aminium(1-25l) [ka]

[0640] Compound 1-25L (40 mg, 70.9% yield) was synthesized according to the synthesis procedure of step 10 in Examples 1-20. MS(ESI)m / z:2255.3[M+H] +

[0641] Step 12: (S)-42-(3-((((R)-4-amino-3-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)butoxy)carbonyl)amino)propanamide)-47-(5-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5, 8-Diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-(((2S,3R,4S,5S,6S)-6-carboxy-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)oxy)phenyl)-N,N,N-trimethyl-39,43,48-trioxo-2,5,8,11,14,17,20,23,26,29,32,35-dodecaoxa-38,44,47-triazanonatetracontan-49-aminium(1-25) [ka]

[0642] Compound 1-25 (19 mg, 49.7% yield) was synthesized according to the synthesis procedure of step 11 in Examples 1-20. MS(ESI)m / z:2155.0[M+H] +

[0643] Examples 1-26 [ka] Step 1: 20-(carboxymethyl)-1-(9H-fluoren-9-yl)-3,19-dioxo-2,7,10,13,16-pentaoxa-4,20-diazadocosan-22-acid(1-26b)

[0644] A solution of 1-4b (680 mg, 1.4 mmol), HATU (583 mg, 1.5 mmol), and DIPEA (486 μL, 2.8 mmol) in 10 mL of anhydrous DMF was mixed with 2,2'-azanejyldiacetic acid (371 mg, 2.8 mmol). The mixture was stirred overnight at room temperature. The mixture was partitioned between water and ethyl acetate. The organic layer was concentrated and purified by flash column chromatography (CH2Cl2 / MeOH = 96 / 4) to obtain 1-26b (250 mg, 19.7% yield) as a white solid. MS(ESI) m / z: 603.2[M+H] +

[0645] Step 2: (9H-Fluoren-9-yl)methyl((21S,22R,23R,24R)-21,22,23,24,25-pentahydroxy-19-methyl-16-(2-(methyl((2S,3R,4R,5R)-2,3,4,5,6-pentahydroxyhexyl)amino)-2-oxoethyl)-15,18-dioxo-3,6,9,12-tetraoxa-16,19-diazapentacosyl)carbamate(1-26c)

[0646] N-methyl-D-glucamine (162 mg, 0.83 mmol) was added to a solution of 1-26b (250 mg, 0.41 mmol), HATU (315 mg, 0.83 mmol), and DIPEA (217 μL, 1.2 mmol) in 10 mL of anhydrous DMF. The mixture was stirred at room temperature for 4 hours and purified by prep-HPLC (column: Xbridge Prep C18 OBD® 5 μm, 19*150 mm; mobile phase A: 0.1% FA in water, B: MeCN; gradient: 25%~60% B; flow rate: 20 mL / min). The fraction was lyophilized to obtain 1-26c (246 mg, 62.0% yield) as a white solid. MS(ESI) m / z: 957.7[M+H] +

[0647] Step 3: tert-butyl(2S,27S,28R,29R,30R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-27,28,29,30,31-pentahydroxy-25-methyl-22-(2-(methyl((2S,3R,4R,5R)-2,3,4,5,6-pentahydroxyhexyl)amino)-2-oxoethyl)-5,21,24-trioxo-9,12,15,18-tetraoxa-6,22,25-triazahenetricontanoate(1-26d)

[0648] Et2NH (266 μL, 2.6 mmol) was added to a solution of 1-26c (246 mg, 0.26 mmol) in 3 mL of DMF. The mixture was stirred at room temperature for 30 minutes and concentrated to obtain the crude product. Fmoc-E( t Crude products were added to a solution of Bu)-COOH (125 mg, 0.29 mmol), HATU (134 mg, 0.35 mmol), and DIPEA (102 μL, 0.59 mmol). The mixture was stirred at room temperature for 1 hour and purified by prep-HPLC (column: Xbridge Prep C18 OBD (trademark) 5 μm, 19*150 mm; mobile phase A: 0.1% FA in water, B: MeCN; gradient: 25%~60% B; flow rate: 20 mL / min). The fraction was lyophilized to obtain 1-26d (200 mg, 59.6% yield) as a white solid. MS(ESI) m / z: 1143.0 [M+H] +

[0649] Step 4: (2S,27S,28R,29R,30R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-27,28,29,30,31-pentahydroxy-25-methyl-22-(2-(methyl((2S,3R,4R,5R)-2,3,4,5,6-pentahydroxyhexyl)amino)-2-oxoethyl)-5,21,24-trioxo-9,12,15,18-tetraoxa-6,22,25-triazahentricontanoic acid (1-26e) [ka]

[0650] To a solution of 1-26d in CH2Cl2 (2 mL), TFA (600 μL) was added at 0°C. The mixture was stirred at room temperature for 3 hours and then purified by prep-HPLC (column: Xbridge Prep C18 OBD (trademark) 5 μm, 19*150 mm; mobile phase A: 0.1% FA in water, B: MeCN; gradient: 25%~55% B; flow rate: 20 mL / min). The fraction was lyophilized to obtain 1-26e (123 mg, 64.7% yield) as a white solid. MS(ESI) m / z: 1086.7 [M+H] +

[0651] Step 5: (2S,3S,4S,5R,6S)-6-(2-((6S,31S,32R,33R,34R)-6-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)-31,32,33,34,35-Pentahydroxy-29-methyl-26-(2-(methyl((2S,3R,4R,5R)-2,3,4,5,6-Pentahydroxyhexyl)amino)-2-oxoethyl)-5,9,25,28-Tetraoxo-13,16,19,22-Tetraoxa-4,10,26,29-Tetraazapentatriakotanamide)-4-((5 S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-26f) [ka]

[0652] Compound 1-26f (125 mg, 50.2% yield) was synthesized according to the synthesis procedure of step 8 in Example 1-2. MS(ESI)m / z:2199.5[M+H] +

[0653] Step 6: (2S,3S,4S,5R,6S)-6-(4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)-2-((6S,31S,32R,33R,34R)-6-((R)-7-(2,5-dioxo-2,5- Dihydro-1H-pyrrole-1-yl)-2,2-dimethyl-4,11-dioxo-3,10-dioxa-5,12-diazapentadecane-15-amide)-31,32,33,34,35-pentahydroxy-29-methyl-26-(2-(methyl((2S,3R,4R,5R)-2,3,4,5,6-pentahydroxyhexyl)amino)-2-oxoethyl)-5,9,25,28-tetraoxo-13,16,19,22-tetraoxa-4,10,26,29-tetraazapentatriakotanamide)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-26g) [ka]

[0654] 1-26 g (40 mg, 67.8% yield) of the compound was synthesized according to the synthesis procedure in steps 9-10 of Examples 1-2. MS(ESI)m / z:1179.6[M+2H + ] / 2

[0655] Step 7: (2S,3S,4S,5R,6S)-6-(2-((6S,31S,32R,33R,34R)-6-(3-((((R)-4-amino-3-(2,5-dioxo-2,5-dihydro-1H-pyrrole-1-yl)butoxy)carbonyl)amino)propanamide)-31,32,33,34,35-pentahydroxy-29-methyl-26-(2-(methyl((2S,3R,4R,5R)-2,3,4,5,6-pentahydroxyhexyl)amino)-2-oxoethyl)-5,9,25,28-tetraoxo-13,16,19,22-tetraoxa-4,10,26,29-tetra Azapentatriakotanamide)-4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropane-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidine-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (1-26) [ka]

[0656] Compound 1-26 (20.8 mg, 54.3% yield) was synthesized according to the synthesis procedure of step 11 in Example 1-2. MS(ESI)m / z:2258.5[M+H] + .

[0657] The conjugator-linker-payload compounds 1-1 to 1-26 are summarized in Table 1 below.

[0658] Example 2: Preparation and Characterization of Antibody-Drug Conjugates Preparation of antibody-drug conjugates for DAR3-5. Antibodies in conjugation buffer (PBS buffer pH 6.0-8.5 with a concentration of 0.5-25 mg / mL) were incubated at a reducing temperature (0-40°C) for 10 minutes. 2-10 eq. of TECP solution (5 mM stock solution in PBS buffer) were added to the reaction mixture, and the reduction reaction was carried out at a reducing temperature for 1-8 hours. After cooling the reducing mixture to 0-25°C, organic solvents (e.g., DMSO, DMF, DMA, PG, acetonitrile, 0-25% v / v) and the prepared conjugator-linker-payload stock solution (6-25 eq., 10 mM stock solution in organic solvent) (see Tables 2A-2D) were added stepwise. The conjugation solution was left at 0-25°C for 1-3 hours, and the reaction could be quenched with N-acetylcysteine ​​(1 mM stock solution). The solution was subjected to buffer exchange (spin desalting column, ultrafiltration, and dialysis) with a storage buffer (e.g., histidine acetate buffer with pH 5.5-6.5 containing any additives, e.g., sucrose, trehalose, tween® 20, 60, 80).

[0659] After the conjugation step, the ADC was subjected to buffer exchange with a ring-opening buffer (pH 6.5-9.0, PBS, borate, or Tris buffer), and the solution was left to stand at 22 or 37°C for 1-48 hours. The maleimide ring-opening process was monitored via reducing LC-MS. Once the hydrolysis of the conjugated maleimide was complete, the resulting ADC was buffer-exchanged via dialysis to a basic Tris pH 8.0-8.5 buffer or an acidic histidine acetate pH 5.0-6.5 buffer.

[0660] LC-MS method for monitoring and determining maleimide hydrolysis. LC-MS analysis was performed under the following measurement conditions: LC-MS system: Vanquish Flex UHPLC and Orbitrap Exploris 240 Mass Spectrometer Column: MAbPac(TM) RP, 2.1*50mm, 4μm, 1,500Å, Thermo Scientific(TM) Column temperature: 80℃ Mobile phase A: 0.1% formic acid (FA) aqueous solution Mobile phase B: Acetonitrile solution containing 0.1% formic acid (FA) Gradient Program 1: 25%B~25%B (0 min~2 min), 25%B~50%B (2 min~18 min), 50%B~90%B (18 min~18.1 min), 90%B~90%B (18.1 min~20 min), 90%B~25%B (20 min~20.1 min), 25%B~25%B (20.1 min~25 min) Gradient Program 2 Injected sample volume: 2 μg MS parameters: Intact and denatured data were acquired in HMR mode with R=15k and deconvolved using the ReSpect algorithm and Sliding Window integration in Thermo Scientific® BioPharma Finder® 4.0 software.

[0661] ADC Characterization. The ADCs were characterized using the following analytical methods. The drug-to-antibody ratio (DAR) of the ADCs was determined by LC-MS or HIC. The SEC purity of all prepared ADCs was >95%.

[0662] LC-MS method: LC-MS analysis was performed under the following measurement conditions: LC-MS system: Vanquish Flex UHPLC and Orbitrap Exploris 240 Mass Spectrometer Column: MAbPac(TM) RP, 2.1*50mm, 4μm, 1,500Å, Thermo Scientific(TM) Column temperature: 80℃ Mobile phase A: 0.1% formic acid (FA) aqueous solution Mobile phase B: Acetonitrile solution containing 0.1% formic acid (FA) Gradient Programs: 25%B~25%B (0 min~2 min), 25%B~50%B (2 min~18 min), 50%B~90%B (18 min~18.1 min), 90%B~90%B (18.1 min~20 min), 90%B~25%B (20 min~20.1 min), 25%B~25%B (20.1 min~25 min) Injected sample volume: 1 μg MS parameters: Intact and denatured data were acquired in HMR mode with R=15k and deconvolved using the ReSpect algorithm and Sliding Window integration in Thermo Scientific® BioPharma Finder® 4.0 software.

[0663] HIC method: HPLC analysis was performed under the following measurement conditions: HPLC System: Waters ACQUITY ARC HPLC System Detector: Measurement wavelength: 280nm Column: Tosoh Bioscience 4.6 μm ID × 3.5 cm, 2.5 μm butyl-nonporous resin column Column temperature: 25℃ Mobile phase A: 1.5M ammonium sulfate, 50mM phosphate buffer, pH 7.0 Mobile phase B: 50 mM phosphate buffer, 25% (V / V) isopropanol, pH 7.0 Gradient Programs: 0%B~0%B (0 min~2 min), 0%B~100%B (2 min~15 min), 100%B~100%B (15 min~16 min), 100%B~0%B (16 min~17 min), 0%B~0%B (17 min~20 min) Injected sample volume: 20 μg

[0664] SEC method for determining ADC purity: HPLC analysis was performed under the following measurement conditions: HPLC System: Waters H-Class HPLC System Detector: Measurement wavelength: 280nm Column: ACQUITY UPLC BEH200 SEC 1.7um 4.6x150mm, Waters Column temperature: Room temperature Mobile phase A: 200 mM phosphate buffer, 250 mM potassium chloride, 15% isopropyl alcohol, pH 7.0 Gradient program: Isocratic elution at a flow rate of 0.3 mL / min for 10 minutes. Injected sample volume: 20 μg

[0665] ADC Hydrophobicity Evaluation: ADCs with higher hydrophobic properties will appear with slower retention times ("RT") from HIC (Hydrophobic Interaction Column) chromatography. Results are provided in Tables 2A–2D, with the DAR8 peak used as reference. [Table 1-1] [Table 1-2] [Table 1-3] [Table 1-4] [Table 1-5] [Table 1-6] [Table 2A] [Table 2B-1] [Table 2B-2] [Table 2B-3] [Table 2B-4] [Table 2B-5] [Table 2B-6] [Table 2B-7] [Table 2C] [Table 2D]

[0666] Antibody information Ifinatamab (anti-B7H3 antibody)

[0667] Light chain sequence (SEQ ID NO: 1)

[0668] EIVLTQSPATLSLSPGERATLSCRASSRLIYMHWYQQKPGQAPRPLIYATSNLASGIPARFSGSGSGTDFTLTISSLEPEDFAVYYCQQWNSNPPTFGQGTKVEIK RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0669] Heavy chain sequence (SEQ ID NO: 2)

[0670] QVQLVQSGAEVKKPGSSVKVSCKASGYTFTNYVMHWVRQAPGQGLEWMGYINPYNDDVKYNEKFKGRVTITADESTSTAYMELSSLRSEDTAVYYCARWGYYGSPLYYFDYWG QGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCD KTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK TISKAKGQPREPQVYTLPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

[0671] 6E7 (anti-CLL1 antibody)

[0672] Light chain sequence (SEQ ID NO: 3)

[0673] DIQMTQSPSSLSASVGDRVTITCRASQSVSTSSYNYMHWYQQKPGKPPKLLIKYASNLESGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQHSWEIPLTFGQGTKVE IKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0674] Heavy chain sequence (SEQ ID NO: 4)

[0675] EVQLVQSGAEVKKPGASVKVSCKASGYSFTDYYMHWVRQAPGQGLEWIGRINPYAGAAFYSQNFKDRVTLTVDTSTSTAYLELSSLRSEDTAVYYCAIERGADLEGYAMDYWG QGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCD KTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK TISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

[0676] Cofetuzumab (anti-PTK7 antibody)

[0677] Light chain sequence (SEQ ID NO: 5)

[0678] DIEMTQSPDSLAVSLGERATINCRSSQSVLYSSSNRNYLAWYQQNPGQPPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSLQAEDVAVYYCQQYYSTPRTFGQGTKV EIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0679] Heavy chain sequence (SEQ ID NO: 6)

[0680] QVQLVQSGPEVKKPGASVKVSCKASGYTFTDYAVHWVRQAPGKRLEWIGVISTYNDYTYNNQDFKGRVTMTRDTSASTAYMELSRLRSEDTAVYYCARGNSYFYALDYWGQG TSVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDK THTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK TISKAKGQPREPQVYTLPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

[0681] Example 3: Direct killing of ADCs in cancer strains U937, HL60, TF-1, NCI-H358, NCI-H1048, and MDA-MB-453 cell line U937 (ATCC, CRL-1593.2). U-937 is a monocyte-like cell line induced in 1974 from malignant cells obtained from the pleural effusion of a 37-year-old Caucasian male with histiocytic lymphoma. U937 was purchased from ATCC. The basal medium for U937 was RPMI-1640 medium (ATCC30-2001), formulated by ATCC. To prepare a full-growth medium, fetal bovine serum (Gibco, 10099-141C) was added to the basal medium to a final concentration of 10%. The cell line was grown at 37°C in a humidified 5% CO2 atmosphere, and the presence of mycoplasma was periodically tested using the MycoAlert® PLUS mycoplasma detection kit (Lonza, LT07-710).

[0682] HL60 (ATCC, CCL-240). HL-60 cells are promyeloblasts isolated from peripheral blood by leukocyte apheresis from a 36-year-old Caucasian woman with acute promyelocytic leukemia. HL60 cells were purchased from ATCC. The basal medium for HL60 was Iskov-modified Dulbecco's medium, catalog number 30-2005, formulated by ATCC. To prepare a full growth medium, fetal bovine serum (Gibco, 10099-141C) was added to the basal medium to a final concentration of 20%. The cell line was grown at 37°C in a humidified 5% CO2 atmosphere, and the presence of mycoplasma was periodically tested using the MycoAlert® PLUS mycoplasma detection kit (Lonza, LT07-710).

[0683] TF1 (ATCC, CRL-2003). TF-1 erythroblasts were isolated in 1987 from the bone marrow of a 35-year-old Asian male with severe pancytopenia. TF-1 was purchased from ATCC. The basal medium for TF-1 was RPMI-1640 medium, catalog number 30-2001, formulated by ATCC. To prepare a full growth medium, fetal bovine serum (Gibco, 10099-141C) was added to the basal medium to a final concentration of 10%. The cell line was grown at 37°C in a humidified 5% CO2 atmosphere, and the presence of mycoplasma was periodically tested using the MycoAlert® PLUS mycoplasma detection kit (Lonza, LT07-710).

[0684] NCI-H358 (ATCC, CRL-5807). NCI-H358 cells are epithelial-like cells isolated from the bronchioles of a male patient with bronchoalveolar carcinoma. NCI-H358 was purchased from ATCC. The basal medium for NCI-H358 was RPMI-1640 medium, ATCC30-2001, formulated by ATCC. To prepare a full growth medium, fetal bovine serum (Gibco, 10099-141C) was added to the basal medium to a final concentration of 10%. The cell line was grown at 37°C in a humidified 5% CO2 atmosphere, and the presence of mycoplasma was periodically tested using the MycoAlert® PLUS mycoplasma detection kit (Lonza, LT07-710).

[0685] NCI-H1048 (ATCC, CRL-5853). NCI-H1048 is a cell line exhibiting epithelial morphology. NCI-H1048 was purchased from ATCC. The basal medium for NCI-H1048 was DMEM:F12 (catalog number 30-2006) formulated by ATCC. To prepare a complete growth medium, fetal bovine serum (Gibco, 10099-141C) was added to the basal medium to a final concentration of 10%. The cell line was grown at 37°C in a humidified 5% CO2 atmosphere, and the presence of mycoplasma was periodically tested using the MycoAlert® PLUS mycoplasma detection kit (Lonza, LT07-710).

[0686] MDA-MB-453 (SIBS). MDA-MB-453 was derived from the exudate of a 48-year-old female patient with metastatic breast cancer, including nodules, brain tissue, and both pleural and pericardial cavities. MDA-MB-453 was purchased from SIBS. The basal medium for MDA-MB-453 was RPMI 1640 medium, HEPES (Gibco, 22400105). To prepare a full growth medium, fetal bovine serum (Gibco, 10099-141C) was added to the basal medium to a final concentration of 10%. The cell line was grown at 37°C in a humidified 5% CO2 atmosphere and periodically tested for the presence of mycoplasma using the MycoAlert® PLUS mycoplasma detection kit (Lonza, LT07-710). [Table 3] [Table 4]

[0687] Direct ADC Killing: Direct ADC killing was evaluated in U937, HL60, and TF1 cancer strains. Cells were seeded in 2D 96-well plates (Greiner: 655090) (U937 or HL60 (3E3 / well) or TF1 (6E3 / well)) (100 μl / well) (containing 150 μg / ml of Fc blocker) and incubated at 37°C, 5% CO2 for 1 hour. New growth medium containing various concentrations of ADC was added (50 μl / well) and incubated at 37°C, 5% CO2 for 6 days. Cell viability was detected by Cell Titer-Glo (Promega, G7573) (70 μl / well). The 2D plates were incubated at room temperature for 10 minutes to stabilize the luminescence signal. The plates were analyzed with a microplate reader.

[0688] Direct ADC killing was evaluated in NCI-H358, NCI-H1048, and MDA-MB-453 cancer strains. Cells were seeded in 3D 96-well plates (Corning: 4520) (80 μl / well) (2E3 / well) and incubated overnight at 37°C and 5% CO2. A new growth medium containing various concentrations of ADC was added (40 μl / well) and incubated at 37°C and 5% CO2 for 6 days. Cell viability was detected using 3D reagent (Promega, G9683) (100 μl / well). The luminescence signal was stabilized by incubating the 3D plates at room temperature for 30 minutes. The plates were analyzed using a microplate reader.

[0689] Direct ADC cell-killing data for U937, HL60, and TF1 cells are presented in Figures 1A-3I and Tables 5-13. Direct ADC cell-killing data for NCI-H358, NCI-H1048, and MDA-MB-453 cells are presented in Figures 4A-6B and Tables 14 and 15. [Table 5] [Table 6] [Table 7] [Table 8] [Table 9] [Table 10] [Table 11] [Table 12] [Table 13] [Table 14] [Table 15]

[0690] Example 4: ADC PK study in mice Female BALB / c nude mice (n=3 per group) were treated with freshly prepared ADC (5 mg / kg) via intravenous administration. Mice were anesthetized with isoflurane at specified time points after administration. Whole blood samples were collected from the orbital venous sinus into coagulation tubes (Kangjian, #041-0121) at 0.5, 2, 6, 8, 24, 48, 72, and 168 hours after administration. Blood samples were left at room temperature for approximately 30 minutes and then treated with centrifugation (4°C, 3000 × g, 7 min) to separate the plasma. Serum samples were transferred to new 1.5 mL tubes and kept in a -80°C freezer before analysis.

[0691] Serum payload concentration. To determine the serum payload concentration, verapamil (IS, 10 ng / mL) was added to a 15 μL aliquot of the sample in 150 μL of ACN / MeOH (1:1). The mixture was vortexed for 1 minute and centrifuged at 4000 rpm for 10 minutes at 4°C. An 80 μL aliquot of the supernatant was diluted with 80 μL of water, and the mixed sample was injected into an LC-MS / MS.

[0692] LC-MS / MS method for determining payload biological analysis. LC-MS / MS analysis was performed under the following measurement conditions: Equipment: LC-MS / MS (Triple Quad 6500 Plus) Monitor: MRM Column: Advanced Materials Technology, HALO AQ-C18 2.7μm 90Å, 50*2.1mm Column temperature: 40℃ Mobile phase A: H2O-0.1%FA Mobile phase B: ACN-0.1%FA Gradient programs for MMAE: 15%B~15%B (0 min~0.4 min), 15%B~30%B (0.4 min~0.8 min), 30%B~30%B (0.8 min~1.8 min), 30%B~90%B (1.8 min~1.9 min), 90%B~90%B (1.9 min~2.4 min), 90%B~15%B (2.4 min~2.5 min), 15%B~15%B (2.5 min~3.0 min) Injected sample volume: 10 μL (MMAE)

[0693] Plasma ADC and total Ab (Tab) concentrations. Plasma ADC and Tab concentrations were determined using an ELISA assay where the capture reagent was CLL1 or PTK7 (where applicable) ECD (extracellular domain). The detection reagents were anti-payload Ab for ADC concentration and anti-human IgG polyclonal Ab for Tab concentration.

[0694] For ADC concentration, microplates were coated with CLL1 or PTK7, or ECD, if applicable, at a concentration of 1 μg / mL (100 μL per well), and incubated overnight at 4°C. The microplates were washed three times with 0.05% PBST, and then 200 μL of 2% BSA solution per well was added. The plates were incubated at 37°C for 1 hour, followed by three washes with 0.05% PBST. Then 100 μL (for ADC samples or standard curves) was added to each well. The plates were incubated at 37°C for 1 hour, followed by three washes with 0.05% PBST. 100 μL of 1 μg / mL biotin anti-MMAE antibody per well was added. The plates were incubated again at 37°C for 1 hour, and then washed three times with 0.05% PBST. Next, 100 μL of 1:10000 SA-HRP was added to each well, incubated at 37°C for 0.5 hours, and then washed three times with 0.05% PBST. Finally, 100 μL of TMB was added to each well, the plate was incubated at room temperature for 15 minutes, and then 100 μL of stop solution was added to each well. The plate was read using an ELISA microplate reader at OD450.

[0695] Regarding the Tab concentration of the anti-CLL1 antibody, the aforementioned ADC method was used, except for the following change: 100 μL of anti-human IgG Fc HRP (1:10000) was added per well.

[0696] Regarding the Tab concentration of the anti-PTK1 antibody, the aforementioned ADC method was used, with the following changes: 100 μL of 1 μg / ml anti-human IgG Fc HRP (1:10000) was added per well.

[0697] Changes in ADC DAR in mouse serum PK samples. ADC stability, as reflected by the time-dependent decrease in DAR, was studied using the plasma samples described above. The post-dose time points for the tested samples are shown in Tables 16 and 17 below.

[0698] Human CLL1 or PTK7, or ECD where applicable, was biotinylated and immobilized on Dynabeads M-280 streptavidin. ADCs were captured from plasma samples by incubation with the ECD-bead system at 37°C for 2 hours. The captured ADCs were then washed with HBS-EP buffer (10 mM Hepes [pH 7.4], 150 mM NaCl, 3.4 mM ethylenediaminetetraacetic acid [EDTA], 0.005% surfactant P20) and digested with IdeS enzyme at 37°C for 1 hour. After extensive washing of the beads with HBS-EP, water, and 10% acetonitrile, the ADC analyte was eluted using 30% acetonitrile with 1% formic acid. The samples were then reduced with 100 mM TCEP for 45 minutes. Liquid chromatography-tandem mass spectrometry (LC-MS / MS) was used for ADC DAR analysis.

[0699] The mouse PK parameters of several anti-CLL1 ADCs (ADC3-A, ADC3-11, ADC3-12, ADC3-14, and ADC3-18) are presented in Table 16 below. The results demonstrate that ADC3-11, ADC3-12, ADC3-14, and ADC3-18 exhibit lower payload exposure and less DAR change than ADC3-A. [Table 16]

[0700] The mouse PK parameters of several anti-PTK7 ADCs (ADC5-A, ADC5-1, and ADC5-2) are presented in Table 17 and Figure 7. The results demonstrate that ADC5-1 and ADC5-2 showed less payload exposure and less DAR change than ADC5-A. [Table 17]

[0701] Example 5: Tumor growth inhibition in an NCI-H1650 xenograft model NCI-H1650 (ATCC) is an epithelial cell line isolated in 1987 from the lung tissue of a 27-year-old male smoker with stage 3 bronchoalveolar carcinoma. NCI-H1650 cells were cultured in RPMI-1640 medium supplemented with 10% (v / v) fetal bovine serum, 100 U / ml penicillin, and 100 μg / ml streptomycin. On the day of implantation, cultured cells were collected and resuspended in a cold (4°C) mixture of PBS and Matrigel (1:1). Cell density was set to 2.5 × 10⁶. 7 The solution was adjusted to the desired cell / mL concentration, and the cells were placed on ice before inoculation.

[0702] 5 x 10 female BALB / c nude mice 6 NCI-H1650 cells (mixed with Matrigel in a 1:1 ratio) were subcutaneously injected into the right flank. After inoculation, tumor length (a) and width (b) were measured with calipers, and tumor volume was calculated using the formula V = 0.5(a × b). 2 Calculated using ). Tumor volume is approximately 200 mm 3 When the tumors reached their size, mice were randomized on day 0 into four groups of six animals each: vehicle, ADC5-A, ADC5-1, and ADC5-2. On day 1 of treatment, animals were intravenously administered vehicle, ADC5-A (3 mg / kg), ADC5-1 (3 mg / kg), or ADC5-2 (3 mg / kg). Whole blood was collected from three mice per group at one time point at 2, 6, 48, and 168 hours post-treatment (samples at 2 and 48 hours were collected from one group of three mice; samples at 6 and 168 hours were collected from another group of three mice). Blood was centrifuged at 5000 × g for 10 minutes at 4°C to obtain serum for PK assay. Animal body weight and tumor volume were measured twice weekly. Tumor growth inhibition (TGI) was calculated using the following formula: %TGI=[1-(Treatment T t - Treatment T0) / (Vehicle T t -Vehicle T0) × 100% Treatment T t = Mean tumor volume of the drug-treated group on day t Mean tumor volume of the drug-treated group on treatment day T0 = 0 Vehicle T t = Mean tumor volume of the vehicle group on day t Mean tumor volume of the vehicle group on day T0 = 0

[0703] The results of evaluating the antitumor effects of ADC5-A, ADC5-1, and ADC5-2 in a subcutaneous NCI-H1650 xenograft model are presented in Table 18 and Figure 8. Data are presented as tumor volume ± standard error (SEM). Single-dose treatment with 3 mg / kg of ADC5-A, ADC5-1, and ADC5-2 showed antitumor activity with TGI of 65% (adjusted p=0.0450), 83% (adjusted p=0.0004), and 84% (adjusted p=0.0011), respectively, on day 18 of treatment. ADC5-1 and ADC5-2 showed better efficacy than ADC5-A at 3 mg / kg. ADCs were well tolerated in all treatment groups, and no animals in any treatment group showed significant weight loss or abnormal clinical observations. [Table 18]

[0704] Although the foregoing disclosure is presented in some detail by description and examples for the purpose of clarity of understanding, it will be apparent to those skilled in the art that certain minor changes and modifications will be made. Therefore, the description and examples should not be construed as limiting.

[0705] Where any prior art publication is referenced herein, it should be understood that such reference does not constitute an endorsement that the publication forms part of the common general knowledge in the art in any country.

[0706] All non-patent publications, patents, patent applications, and disclosures of published patent applications referenced herein by distinctive reference are incorporated herein by reference in their entirety.

Claims

1. Compound of formula (I): 【Chemical 318】 or a pharmaceutically acceptable salt, tautomer, solvate, or stereoisomer thereof (in the formula, BA is a binder selected from humanized, monoclonal, chimeric, or human antibodies or their antigen-binding fragments; The conjugator is given by formula (II) or (III): 【Chemical 319】 (In the formula, U is a bond, heteroarylene, or arylene; V is a bond or -C≡C-(CH ２ ) ｎ - and; n is an integer between 0 and 10 (including both ends); W ２ is -C(=O)-, -NH-, or -O-; RG is, 【Chem.320】 , -(succinimido-3-yl-N)-, or 【Chemistry 321】 And; RS is -NR １ａ R １ｂ And; R １ａ and R １ｂ Each of these independently represents H or substituted or unsubstituted C. １－４ It is alkyl; RE is a bond, -O-, -OC(=O)-, -OC(=O)NR ６ -, -NHC(=O)NR ６ -, -OS(=O) ２ NR ６ -, -NHS(=O) ２ NR ６ -, or -OC(=O)NHS(=O) ２ NR ６ -; R ６ is H or substituted or unsubstituted C １－４ It is alkyl; W ３ is -C(=O)-, -NH-, or -O-; Each of s and t is independently 0, 1, or 2; 【Chemistry 322】 This indicates a covalent bond site within the compound, (* indicates a connection in which the conjugator connects to the BA.) Having; The spacer is coupled with, ＊＊ -NH-(CH ２ CH ２ O) ｍ -CH ２ CH ２ -C(=O)-, ＊＊ -(CH ２ ) ｍ -C(=O)-, ＊＊ -CH ２ -C(=O)-NH-(CH ２ ) ｍ -C(=O)-, ＊＊ -(CH ２ CH ２ O) ｍ -CH ２ CH ２ -C(=O)-, ＊＊ -CH[-(CH ２ ) ｍ -COOH]-C(=O)-, ＊＊ -CH ２ -C(=O)-NH-(CH ２ ) ｍ -C(=O)-NH-(CH ２ ) ｍ -C(=O)-,​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​ m is an integer from 1 to 12 (including both ends), ** indicates the connection in which the spacer connects to the conjugator. And; The severable bodies are given by formulas (IVa), (IVb), (IVc), (Va), (Vb), (VIa), (VIb), (VIIa), or (VIIb): 【Chemical 323】 (In the formula, Su is the sugar portion, Each R ２ These are, independently, hydrogen, halogen, substituted or unsubstituted C １－４ Alkyl, -CN, or -NO ２ And, *** indicates a connection in which the severable body is connected to the spacer. Having; The payload is a payload residue; x is between 1 and 15 (including both ends).

2. The conjugator is the compound according to claim 1, having formula (II).

3. The compound according to claim 2, wherein U is arylene.

4. The compound according to claim 3, wherein U is phenylene.

5. The aforementioned U is, 【Chemical 324】 The compound according to claim 4.

6. The compound according to claim 2, wherein U is a heteroarylene.

7. The compound according to claim 6, wherein U is a divalent pyrimidine ring.

8. The aforementioned U is, 【Chemical 325】 The compound according to claim 7.

9. The compound according to claim 2, wherein U is a bond.

10. The compound according to any one of claims 1 to 9, wherein V is a bond.

11. The above V is -C≡C-(CH ２ ) ｎ - The compound according to any one of claims 1 to 9.

12. The above V is -C≡C-(CH ２ ) ３ - The compound according to claim 11.

13. The aforementioned W ２ The compound according to any one of claims 1 to 12, wherein is -C(=O)-.

14. The conjugator is the compound according to claim 1, having formula (III).

15. The aforementioned RS is -NH ２ or -N(CH ３ ) ２ The compound according to claim 14.

16. The aforementioned RS is -NH ２ The compound according to claim 15.

17. The compound according to any one of claims 14 to 16, wherein RE is -OC(=O)NH-.

18. The aforementioned RG is, 【Chemistry 326】 The compound according to any one of claims 14 to 17.

19. The compound according to any one of claims 14 to 18, wherein each of s and t is 2.

20. The aforementioned W ３ The compound according to any one of claims 14 to 19, wherein is -C(=O)- or -NH-.

21. The spacer connects, ＊＊ -NH-(CH ２ CH ２ O) ｍ -CH ２ CH ２ -C (=O)-, or ＊＊ -NH-(CH ２ ) ｍ A compound according to any one of claims 1 to 20, wherein the compound is -O-.

22. The compound according to any one of claims 1 to 21, wherein m is 2, 4, 6, or 8.

23. The aforementioned conjugator is given by formula (IIa1), (IIa2), (IIa3), or (IIIa): 【Chemistry 327】 The compound according to claim 1, having the following characteristics.

24. The severable body is defined by formulas (IVa1), (IVa2), (IVa3), (Va1), (VIa1), or (VIIa1): 【Chemical 328】 A compound according to any one of claims 1 to 23, having the following characteristics.

25. The payload is given by the following formula: 【Chemistry 329-1】 【Chemistry 329-2】 A compound according to any one of claims 1 to 24, wherein one of the residues is one of the residues.

26. The aforementioned payload is 【Chemistry 330】 The compound according to any one of claims 1 to 25.

27. The compound according to any one of claims 1 to 26, wherein the BA binds to one or more receptors selected from B7H3, PTK7, or CLL1.

28. The compound according to any one of claims 1 to 27, wherein BA is ifinatamab, 6E7, or cofetuzumab, or an antigen-binding fragment of ifinatamab, 6E7, or cofetuzumab.

29. The aforementioned compound, 【Chemistry 331】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 332】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemical 333】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 334】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 335】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 336】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 337】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemical 338】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 339】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 340】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 341】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 342】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Transformation 343】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Transformation 344】 (wherein Ab is 6E7 or its antigen-binding fragment); or 【Chemistry 345】 (In the formula, Ab is ifinatamab or its antigen-binding fragment.) The compound according to claim 1, or any of the pharmaceutically acceptable salts, tautomers, solvates, or stereoisomers described above.

30. The compound according to any one of claims 1 to 29, wherein x is approximately 3.5 to approximately 4.

5.

31. The aforementioned compound, 【Transformation 346】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Transformation 347】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Transformation 348】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 349】 (In the formula, Ab is 6E7 or its antigen-binding fragment); [Chemical 350] (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 351】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 352】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 353】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 354】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemical 355】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Transformation 356】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 357】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemical 358】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 359】 (wherein Ab is 6E7 or its antigen-binding fragment); or 【Chemical 360】 (In the formula, Ab is ifinatamab or its antigen-binding fragment.) The compound according to claim 30, or any of the pharmaceutically acceptable salts, tautomers, solvates, or stereoisomers described above.

32. Compound of formula (XI): 【Chemical 361】 or a pharmaceutically acceptable salt, tautomer, solvate, or stereoisomer thereof (in the formula, BA is a binder selected from humanized, monoclonal, chimeric, or human antibodies or their antigen-binding fragments; The conjugator is given by formula (II) or (III): 【Chemical 362】 (In the formula, U is a bond, heteroarylene, or arylene; V is a bond or -C≡C-(CH ２ ) ｎ - and; n is an integer between 0 and 10 (including both ends); W ２ is -C(=O)-, -NH-, or -O-; RG is, 【Chemical 363】 , -(succinimido-3-yl-N)-, or 【Chemical 364】 And; RS is -NR １ａ R １ｂ And; R １ａ and R １ｂ each independently is H or substituted or unsubstituted C １－４ alkyl; RE is a bond, -O-, -OC(=O)-, -OC(=O)NR ６ -, -NHC(=O)NR ６ -, -OS(=O) ２ NR ６ -, -NHS(=O) ２ NR ６ -, or -OC(=O)NHS(=O) ２ NR ６ -; R ６ is H or substituted or unsubstituted C １－４ It is alkyl; W ３ is -C(=O)-, -NH-, or -O-; Each of s and t is independently 0, 1, or 2; 【Chemical 365】 This indicates a covalent bond site within the compound, (* indicates a connection in which the conjugator connects to the BA.) Having; The spacer is bonded, ＊＊ -NH-(CH ２ CH ２ O) ｍ -CH ２ CH ２ -C(=O)-, ＊＊ -C(=O)-(CH ２ CH ２ O) ｍ -CH ２ CH ２ -NH-, ＊＊ -(CH ２ ) ｍ -C(=O)-, ＊＊ -CH ２ -C(=O)-NH-(CH ２ ) ｍ -C(=O)-, ＊＊ -(CH ２ CH ２ O) ｍ -CH ２ CH ２ -C(=O)-, ＊＊ -C(=O)-(CH ２ CH ２ O) ｍ -CH ２ CH ２ -NH-, ＊＊ -CH[-(CH ２ ) ｍ -COOH]-C(=O)-, ＊＊ -CH ２ -C(=O)-NH-(CH ２ ) ｍ -C(=O)-NH-(CH ２ ) ｍ -C(=O)-, ＊＊ -C(=O)-(CH ２ ) ｍ -C(=O)-, ＊＊ -NH-(CH ２ ) ｍ -C(=O)-, or ＊＊ -NH-(CH<�000151>) ｍ -O-(wherein, m is an integer from 1 to 12 (including both ends), ** indicates the connection in which the spacer connects to the conjugator. And; The severable bodies are given by formulas (IVa'), (IVb'), (IVc'), (Va'), (Vb'), (VIa'), (VIb'), (VIia'), or (VIib'): 【Chemical 366】 (In the formula, Su is the sugar portion, Each R ２ These are, independently, hydrogen, halogen, substituted or unsubstituted C １－４ Alkyl, -CN, or -NO ２ And, (# indicates a connection where the severable body is connected to the branched body.) Having; The branched area is given by equation (XIIa), (XIIb), (XIIc), or (XIId): 【Chemical 367】 (In the formula, Each of p and q is independently 1, 2, 3, or 4; The hydrophilic group is -NH-(CH ２ CH ２ O) ａ -(CH ２ ) ｂ CH ３ , -N(CH ３ )-( (CH ２ ) ｃ C(=O)N(CH ３ )) ａ -(CH ２ ) ｃ C(=O)NH ２ , -C(=O)-(CH ２ CH ２ O) ａ -(CH ２ ) ｂ CH ３ , -C(=O)-(CH ２ CH ２ O) ａ -(CH ２ ) ｂ NHC(=O)(CH ２ ) ３ N(CH ３ ) ２ (CH ２ ) ３ S(=O)[[ID=u]] ２ OH, -NH-(CH ２ CH ２ O) ａ -(CH ２ ) ｂ NHC(=O)(CH ２ ) ３ N(CH ３ ) ２ (CH ２ ) ３ S(=O) ２ OH, 【Chemical 368】 And; a is an integer between 1 and 18 (including both ends); b is 0, 1, or 2. c is 1, 2, 3, or 4. Having; The payload is a payload residue; x is between 1 and 15 (including both ends).

33. The conjugator is the compound according to claim 32, having formula (II).

34. The compound according to claim 33, wherein U is arylene.

35. The compound according to claim 34, wherein U is phenylene.

36. The aforementioned U is, 【Chemical 369】 The compound according to claim 35.

37. The compound according to claim 32, wherein U is a heteroarylene.

38. The compound according to claim 37, wherein U is a divalent pyrimidine ring.

39. The aforementioned U is, 【Chemistry 370】 The compound according to claim 38.

40. The compound according to claim 32, wherein U is a bond.

41. The compound according to any one of claims 32 to 40, wherein V is a bond.

42. The above V is -C≡C-(CH ２ ) ｎ - The compound according to any one of claims 32 to 40.

43. V is -C≡C-(CH ２ ) ３ - The compound according to claim 42.

44. The aforementioned W ２ The compound according to any one of claims 32 to 43, wherein is -C(=O)-.

45. The conjugator is the compound according to claim 32, having formula (III).

46. The aforementioned RS is -NH ２ or -N(CH ３ ) ２ The compound according to claim 45.

47. The aforementioned RS is -NH ２ The compound according to claim 46.

48. The compound according to any one of claims 45 to 47, wherein RE is -OC(=O)NH-.

49. The aforementioned RG is, 【Chemistry 371】 The compound according to any one of claims 45 to 48.

50. The compound according to any one of claims 45 to 49, wherein each of s and t is independently 0 or 2.

51. The aforementioned W ３ The compound according to any one of claims 45 to 50, wherein is -C(=O)-.

52. The spacer is a bond, the compound according to any one of claims 32 to 51.

53. The aforementioned conjugator is given by formula (IIa1), (IIa2), (IIa3), (IIa4), (IIa5), or (IIIa): 【Chemistry 372】 The compound according to claim 32, having the following characteristics.

54. The severable body is defined by formula (IVa'1), (IVc'1), or (Va'1): 【Chemistry 373】 A compound according to any one of claims 32 to 53, having the following characteristics.

55. The hydrophilic group is -NH-(CH ２ CH ２ O) ａ -(CH ２ ) ｂ CH ３ 、-N(CH ３ )-(CH ２ ) ｃ C(=O)N(CH ３ ) ａ -(CH ２ ) ｃ C(=O)NH ２ 、-C(=O)-(CH ２ CH ２ O) ａ -(CH ２ ) ｂ CH ３ 、-C(=O)-(CH ２ CH ２ O) ａ -(CH ２ ) ｂ NHC(=O)(CH ２ ) ３ N(CH ３ ) ２ (CH ２ ) ３ S(=O) ２ OH、 【Chemistry 374】 The compound according to any one of claims 32 to 54.

56. The compound according to any one of claims 32 to 55, wherein a is 4, 8, 11, or 12.

57. The compound according to any one of claims 32 to 56, wherein b is 0 or 2.

58. The compound according to any one of claims 32 to 57, wherein c is 1.

59. The hydrophilic group is -NH-(CH ２ CH ２ O)- １２ -CH ３ , -N(CH ３ )-(CH ２ C(=O)N(CH ３ )- １１ -(CH ２ ) ｃ C(=O)NH ２ , -C(=O)-(CH ２ CH ２ O)- １２ -CH ３ , -C(=O)-(CH ２ CH ２ O)- ８ -(CH ２ ) ２ NHC(=O)(CH ２ ) ３ N(CH ３ [[ID=4;6]]) ２ (CH ２ ) ３ S(=O) ２ OH, 【Chemistry 375】 The compound according to claim 55.

60. The compound according to any one of claims 32 to 59, wherein each of the aforementioned p and q is independently 2 or 4.

61. The payload is given by the following formula: 【Transformation 376】 A compound according to any one of claims 32 to 60, which is one residue among the following.

62. The aforementioned payload is 【Chemical 377】 The compound according to any one of claims 32 to 61.

63. The compound according to any one of claims 32 to 62, wherein the BA binds to one or more receptors selected from B7H3, PTK7, or CLL1.

64. The compound according to any one of claims 32 to 63, wherein BA is ifinatamab, 6E7, or cofetuzumab, or an antigen-binding fragment of ifinatamab, 6E7, or cofetuzumab.

65. The aforementioned compound, 【Chemistry 378】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 379】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemical 380】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 381】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 382】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 383】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemical 384】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chem.385】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemical 386】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 387】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemical 388】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 389】 (In the formula, Ab is ifinatamab or its antigen-binding fragment.) 【Chemical 390】 (wherein Ab is cofetuzumab or its antigen-binding fragment); or 【Chemistry 391】 (In the formula, Ab is cofetuzumab or its antigen-binding fragment.) The compound according to claim 32, or any of the pharmaceutically acceptable salts, tautomers, solvates, or stereoisomers described above.

66. The compound according to any one of claims 32 to 65, wherein x is approximately 3.5 to approximately 4.

5.

67. The aforementioned compound, 【Chemistry 392】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 393】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chem. 394】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemical 395】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 396】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemistry 397】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chem.398】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chem.399】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemical 400】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemical 401】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemical 402】 (In the formula, Ab is 6E7 or its antigen-binding fragment); 【Chemical 403】 (In the formula, Ab is ifinatamab or its antigen-binding fragment.) 【Chemical 404】 (wherein Ab is cofetuzumab or its antigen-binding fragment); or 【Chemical 405】 (In the formula, Ab is cofetuzumab or its antigen-binding fragment.) The compound according to claim 66, or any of the pharmaceutically acceptable salts, tautomers, solvates, or stereoisomers described above.

68. Compound of formula (I-I): 【Chemical 406】 or a pharmaceutically acceptable salt, tautomer, solvate, or stereoisomer thereof (in the formula, The conjugator is given by formula (I-II) or (I-III): 【Chemical 407】 (In the formula, U is a bond, heteroarylene, or arylene; V is a bond or -C≡C-(CH ２ ) ｎ - and; n is an integer between 0 and 10 (including both ends); W ２ is -C(=O)-, -NH-, or -O-; RG is, 【Chemical 408】 And; RS is -NR １ａ R １ｂ And; R １ａ and R １ｂ Each of these independently represents H or substituted or unsubstituted C. １－４ It is alkyl; RE represents bond, -O-, -OC(=O)-, -OC(=O)NR ６ -, -NHC(=O)NR ６ -, -OS (=O) ２ NR ６ -, -NHS (=O) ２ NR ６ - or -OC(=O)NHS(=O) ２ NR ６ - and; R ６ is H or substituted or unsubstituted C １－４ It is alkyl; W ３ is -C(=O)-, -NH-, or -O-; Each of s and t is independently 0, 1, or 2. 【Chemical 409】 (This indicates a covalent bond site within the compound.) It has, The spacer is bonded, ＊＊ -NH-(CH ２ CH ２ O) ｍ -CH ２ CH ２ -C(=O)-, ＊＊ -(CH ２ ) ｍ -C(=O)-, ＊＊ -CH ２ -C(=O)-NH-(CH ２ ) ｍ -C(=O)-, ＊＊ -(CH ２ CH ２ O) ｍ -CH ２ CH ２ -C(=O)-, ＊＊ -CH[-(CH ２ ) ｍ -COOH]-C(=O)-, ＊＊ -CH ２ -C(=O)-NH-(CH ２ ) ｍ -C(=O)-NH-(CH ２ ) ｍ -C(=O)-, ＊＊ -C(=O)-(CH ２ ) ｍ -C(=O)-, ＊＊ -NH-(CH ２ ) ｍ -C(=O)-, or ＊＊ -NH-(CH ２ ) ｍ -O-(wherein, m is an integer from 1 to 12 (including both ends), ** indicates the connection in which the spacer connects to the conjugator. And; The severable bodies are given by formulas (IVa), (IVb), (IVc), (Va), (Vb), (VIa), (VIb), (VIIa), or (VIIb): 【Chemical 410】 (In the formula, Su is the sugar portion, Each R ２ These are, independently, hydrogen, halogen, substituted or unsubstituted C １－４ Alkyl, -CN, or -NO ２ And, *** indicates a connection in which the severable body is connected to the spacer. Having; The payload is the payload residue.

69. The conjugator is the compound according to claim 68, having formula (I-II).

70. The compound according to claim 69, wherein U is arylene.

71. The compound according to claim 70, wherein U is phenylene.

72. The aforementioned U is, 【Chemical 411】 The compound according to claim 71.

73. The compound according to claim 69, wherein U is a heteroarylene.

74. The compound according to claim 73, wherein U is a divalent pyrimidine ring.

75. The aforementioned U is, 【Chemical 412】 The compound according to claim 74.

76. The compound according to claim 69, wherein U is a bond.

77. The compound according to any one of claims 68 to 76, wherein V is a bond.

78. The above V is -C≡C-(CH ２ ) ｎ - The compound according to any one of claims 68 to 76.

79. The above V is -C≡C-(CH ２ ) ３ - The compound according to claim 78.

80. The aforementioned W ２ The compound according to any one of claims 68 to 79, wherein is -C(=O)-.

81. The conjugator is the compound according to claim 68, having formula (I-III).

82. The aforementioned RS is -NH ２ or -N(CH ３ ) ２ The compound according to claim 81.

83. The aforementioned RS is -NH ２ The compound according to claim 82.

84. The compound according to any one of claims 81 to 83, wherein RE is -OC(=O)NH-.

85. The aforementioned RG is, 【Chemical 413】 The compound according to any one of claims 81 to 84.

86. The compound according to any one of claims 81 to 85, wherein each of s and t is 2.

87. The aforementioned W ３ The compound according to any one of claims 81 to 86, wherein is -C(=O)- or -NH-.

88. The spacer connects, ＊＊ -NH-(CH ２ CH ２ O) ｍ -CH ２ CH ２ -C (=O)-, or ＊＊ -NH-(CH ２ ) ｍ The compound according to any one of claims 68 to 87, wherein it is -O-.

89. The compound according to any one of claims 68 to 88, wherein m is 2, 4, 6, or 8.

90. The aforementioned conjugator is given by formula (I-IIa1) or (I-IIIa1): 【Chemical 414】 The compound according to claim 68, having the following characteristics.

91. The severable body is defined by formulas (IVa1), (IVa2), (IVa3), (Va1), (VIa1), or (VIIa1): 【Chemical 415】 A compound according to any one of claims 68 to 90, having the following characteristics.

92. The payload is given by the following formula: 【Chemical Engineering 416-1】 【Chemical Engineering 416-2】 A compound according to any one of claims 68 to 91, wherein one of the residues is one of the residues.

93. The aforementioned payload is 【Chemical 417】 The compound according to any one of claims 68 to 92.

94. The aforementioned compound, 【Chemical Engineering 418-1】 【Chemical 418-2】 【Chemical 418-3】 The compound according to claim 68, or a pharmaceutically acceptable salt, tautomer, solvate, or stereoisomer thereof.

95. Compound of formula (XV): 【Chemical 419】 or a pharmaceutically acceptable salt, tautomer, solvate, or stereoisomer thereof (in the formula, The conjugator is given by formula (I-II) or (I-III): 【Chemical 420】 (In the formula, U is a bond, heteroarylene, or arylene; V is a bond or -C≡C-(CH ２ ) ｎ - and; n is an integer between 0 and 10 (including both ends); W ２ is -C(=O)-, -NH-, or -O-; RG is, 【Chemistry 421】 And; RS is -NR １ａ R １ｂ And; R １ａ and R １ｂ Each of these independently represents H or substituted or unsubstituted C. １－４ It is alkyl; RE represents bond, -O-, -OC(=O)-, -OC(=O)NR ６ -, -NHC(=O)NR ６ -, -OS (=O) ２ NR ６ -, -NHS (=O) ２ NR ６ - or -OC(=O)NHS(=O) ２ NR ６ - and; R ６ is H or substituted or unsubstituted C １－４ It is alkyl; W ３ is -C(=O)-, -NH-, or -O-; Each of s and t is independently 0, 1, or 2. 【Chemistry 422】 (This indicates a covalent bond site within the compound.) It has, The spacer is bonded, ＊＊ -NH-(CH ２ CH ２ O) ｍ -CH ２ CH ２ -C(=O)-, ＊＊ -(CH ２ ) ｍ -C(=O)-, ＊＊ -CH ２ -C(=O)-NH-(CH ２ ) ｍ -C(=O)-, ＊＊ -(CH ２ CH ２ O) ｍ -CH ２ CH ２ -C(=O)-, ＊＊ -CH[-(CH ２ ) ｍ -COOH]-C(=O)-, ＊＊ -CH ２ -C(=O)-NH-(CH ２ ) ｍ -C(=O)-NH-(CH ２ ) ｍ -C(=O)-, ＊＊ -C(=O)-(CH ２ ) ｍ -C(=O)-, ＊＊ -NH-(CH ２ ) ｍ -C(=O)-, or ＊＊ -NH-(CH ２ ) ｍ -O-(wherein, m is an integer from 1 to 12 (including both ends), ** indicates the connection in which the spacer connects to the conjugator. And; The severable bodies are given by formulas (IVa'), (IVb'), (IVc'), (Va'), (Vb'), (VIa'), (VIb'), (VIia'), or (VIib'): 【Chemistry 423】 (In the formula, Su is the sugar portion, Each R ２ These are, independently, hydrogen, halogen, substituted or unsubstituted C １－４ Alkyl, -CN, or -NO ２ And, (# indicates a connection where the severable body is connected to the branched body.) Having; The branched areas are given by equations (XIIa), (XIIb), (XIIc), and (XIId): 【Chemistry 424】 (In the formula, Each of p and q is independently 1, 2, 3, or 4; The hydrophilic group is -NH-(CH ２ CH ２ O) ａ -(CH ２ ) ｂ CH ３ , -N(CH ３ )-( (CH ２ ) ｃ C(=O)N(CH ３ )) ａ -(CH ２ ) ｃ C(=O)NH ２ , -C(=O)-(CH ２ CH ２ O)[[ID=;33]] ａ -(CH ２ ) ｂ CH ３ , -C(=O)-(CH ２ CH ２ O) ａ -(CH ２ ) ｂ NHC(=O)(CH ２ ) ３ N(CH ３ ) ２ (CH ２ ) ３ S(=O) ２ OH, -NH-(CH ２ CH ２ O) ａ -(CH ２ ) ｂ NHC(=O)(CH ２ ) ３ N(CH ３ ) ２ (CH ２ ) ３ S(=O) ２ OH, 【Chemical 425】 And; a is an integer between 1 and 18 (including both ends); b is 0, 1, or 2. c is 1, 2, 3, or 4. Selected from the group consisting of; The payload is the payload residue.

96. The conjugator is the compound according to claim 95, having formula (I-II).

97. The compound according to claim 96, wherein U is arylene.

98. The compound according to claim 97, wherein U is phenylene.

99. The aforementioned U is, 【Chemistry 426】 The compound according to claim 98.

100. The compound according to claim 96, wherein U is a heteroarylene.

101. The compound according to claim 100, wherein U is a divalent pyrimidine ring.

102. The aforementioned U is, 【Chemistry 427】 The compound according to claim 101.

103. The compound according to claim 96, wherein U is a bond.

104. The compound according to any one of claims 95 to 103, wherein V is a bond.

105. The above V is -C≡C-(CH ２ ) ｎ - The compound according to any one of claims 95 to 103.

106. The above V is -C≡C-(CH ２ ) ３ - The compound according to claim 105.

107. The aforementioned W ２ The compound according to any one of claims 95 to 106, wherein is -C(=O)-.

108. The conjugator is the compound according to claim 95, having formula (I-III).

109. The aforementioned RS is -NH ２ or -N(CH ３ ) ２ The compound according to claim 108.

110. The aforementioned RS is -NH ２ The compound according to claim 109.

111. The compound according to any one of claims 108 to 110, wherein RE is -OC(=O)NH-.

112. The aforementioned RG is, 【Chemistry 428】 The compound according to any one of claims 108 to 111.

113. The compound according to any one of claims 108 to 112, wherein each of s and t is 2.

114. The spacer connects, ＊＊ -NH-(CH ２ CH ２ O) ｍ -CH ２ CH ２ -C (=O)-, or ＊＊ -NH-(CH ２ ) ｍ A compound according to any one of claims 95 to 113, wherein the compound is -O-.

115. The compound according to any one of claims 95 and 97-114, wherein m is 2, 4, 6, or 8.

116. The aforementioned conjugator is given by formula (I-IIa1) or (I-IIIa1): 【Chemistry 429】 The compound according to claim 95, having the following characteristics.

117. The severable body is defined by formula (IVa'1), (IVc'1), or (Va'1): 【Chemistry 430】 A compound according to any one of claims 95 to 116, having the following characteristics.

118. The payload is given by the following formula: 【Chemistry 431】 A compound according to any one of claims 95 to 117, wherein one of the residues is one of the residues.

119. The aforementioned payload is 【Chemistry 432】 The compound according to any one of claims 95 to 118.

120. The aforementioned compound, 【Chemistry 433-1】 【Chemistry 433-2】 【Chemistry 433-3】 The compound according to claim 95, or a pharmaceutically acceptable salt, tautomer, solvate, or stereoisomer thereof.

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