Somatostatin subtype 2 receptor (SST2R) targeted therapeutic drug and application thereof

Abstract

Described herein are SST2R-targeted therapeutic drugs that target tumor cells expressing somatostatin subtype 2 receptor (SST2R) and their use in the treatment of cancer.

Description

[0001] Cross-reference to Related Applications

[0002] This application claims the benefit of U.S. Provisional Patent Application No. 63 / 387,235, filed on December 13, 2022, and U.S. Provisional Patent Application No. US 63 / 597,871, filed on November 10, 2023; each of which is incorporated herein by reference in its entirety. TECHNICAL FIELD

[0003] Disclosed herein are somatostatin subtype-2 receptor (SST2R) drug conjugates and methods of using such drug conjugates as cancer therapeutics, diagnostic agents, or both. BACKGROUND OF THE INVENTION

[0004] Neoplasms are abnormal growths of cells and impose a significant medical burden on humans, including morbidity and mortality. Neoplasms include: benign or non-cancerous neoplasms (e.g., adenomas) that do not exhibit malignant characteristics and are generally less likely to become dangerous; malignant neoplasms that exhibit characteristics such as gene mutations, loss of normal function, rapid division, and the ability to metastasize (invade) into other tissues; and neoplasms with uncertain or unknown behavior. Malignant neoplasms (i.e., cancerous solid tumors) are the leading cause of death in industrialized countries. Non-cancerous neoplasms, including benign adenomas, can also cause significant morbidity and mortality. Although standard treatments can achieve significant effects in tumor growth inhibition and even tumor elimination, the drugs applied exhibit only slight selectivity for malignant tissues compared to healthy tissues, and their side effects limit their efficacy and use. Specifically targeting neoplastic cells without affecting healthy tissues is desirable for effective solid tumor therapies. Non-peptide SST2R ligands conjugated to a suitable drug cargo or payload represent a novel class of small molecule drug conjugates (SMDCs) for selective cancer therapeutics or diagnostic agents. SUMMARY OF THE INVENTION

[0005] Disclosed herein are SST2R modulators that deliver a payload to tumors expressing SSTR2 and their use in tumor treatment. The present disclosure provides an alternative and improved method for treating tumors. In some embodiments, the SMDCs disclosed herein provide an improved method for targeting tumor cells expressing SST2R compared to traditional therapies with a narrow therapeutic index.

[0006] In one aspect, disclosed herein is a compound having the structure of formula (I) or a pharmaceutically acceptable salt thereof:

[0007]

[0008] Wherein:

[0009] A is -N(H)- or -O-;

[0010] R a is hydrogen or a C1-C6 alkyl group;

[0011] R 2 is hydrogen or a C1-C6 alkyl group;

[0012] R 6 is chlorine or -C(=O)NH2;

[0013] L is -L 1 -L 2 -;

[0014] L 1 is an optional spacer; and

[0015] L 2 is an optional linker;

[0016] Wherein at least one of L 1 or L 2 is present; and

[0017] R d is an effector molecule moiety containing a chemotherapeutic agent.

[0018] In some embodiments, L 2 is present and is -(L 2a ) w -L 2b - or -L 2c -;

[0019] Each L 2a is independently selected from natural or unnatural amino acids, wherein any free amine in the amino acid is optionally independently substituted by -CH3;

[0020] L 2b is absent or is -N(R 10 )(unsubstituted or substituted benzyl)-OC(=O)-; wherein the substituted benzyl is substituted by -C(=O)NHR 12 or a monosaccharide;

[0021] Each R 10 is independently selected from hydrogen and C1-C6 alkyl groups;

[0022] Each R 12 is independently selected from hydrogen, C4-C 20Polyethylene glycol and unsubstituted or substituted C1-C6 alkyl, wherein the substituted C1-C6 alkyl is substituted with -NHR 13 、-C(=O)NHR 13 or -NHC(=O)R 13 ;

[0023] Each R 13 is independently selected from hydrogen, C4-C 20 polyethylene glycol and C4-C 20 polyethylene glycol-NH2;

[0024] Or when R 13 is present and at least one free carboxyl group of the amino acid of L 2a is present, then R 13 and the free carboxyl group of the amino acid of L 2a form a ring together;

[0025] w is 1, 2, 3, 4, 5 or 6; and

[0026] Each L 2c is N-maleimidomethyl-cyclohexane-1-carbonyl (MCC) or -S-.

[0027] In some embodiments, L 1 is present and is -X 2 -L 3 -L 4 -;

[0028] X 2 is -C(=O)(CH2) p -, -(CH2) p -, -C(=O)CH(CH2SO3H)NHC(=O)- or -(X 2a ) p -;

[0029] Each X 2a is independently selected from natural or unnatural amino acids, wherein any free amine in the amino acid is optionally independently substituted with -CH3;

[0030] p is 0, 1, 2, 3, 4, 5 or 6;

[0031] L 3 is absent or is unsubstituted or substituted C1-C 10 alkylene, unsubstituted or substituted C1-C 10 heteroalkylene, C4-C 20 polyethylene glycol or -(X 3 CH2CH2) t -;

[0032] Each X3 independently selected from O and NR 10 ;

[0033] each t is independently 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12;

[0034] L 4 is absent or is -L 4a -(CH2) u -L 4b -(CH2) u -L 4c -;

[0035] L 4a is absent or is -O-, -NR 10 -, -NR 10 C(=O)-, -C(=O)NR 10 - or -C(=O)-;

[0036] L 4b is absent or is an unsubstituted or substituted N-containing 5- to 10-membered heterocycloalkylene; wherein any free amine of the N-containing 5- to 10-membered heterocycloalkylene is optionally independently substituted with -CH2CO2H;

[0037] L 4c is absent or is -O-, -NR 10 -, -NR 10 C(=O)-, -C(=O)NR 10 -, -C(=O)NR 10 (CH2) u O(CH2) u C(=O)-, CH(CH2SO3H)C(=O)NR 10 (CH2) u O(CH2) u C(=O)-, -C(=O)-, -CH(=N)-, -CH(=N-NH)-, -CCH3(=N)-, -CCH3(=N-NH)-, -C(=O)-(C1-C6 alkylene)-, -C(=O)NR 10 -(C1-C6 alkylene)-, -NR 10 C(=O)-(C1-C6 alkylene)-, -NR 10 -(C1-C6 alkylene)- or C1-C6 alkylene-;

[0038] each u is independently 0, 1, 2, 3, 4, 5 or 6; and

[0039] each R 10Independently selected from hydrogen and C1-C6 alkyl.

[0040] In some embodiments, R d is:

[0041]

[0042] In another aspect, the present invention describes a method for treating cancer, which comprises administering to a mammal suffering from cancer an effective amount of a compound of formula (I) or a pharmaceutically acceptable salt thereof.

[0043] In another aspect, the present invention describes a method for treating a tumor, which comprises administering to a mammal suffering from a tumor an effective amount of a compound of formula (I) or a pharmaceutically acceptable salt thereof.

[0044] In some embodiments, the mammal suffers from anal cancer, bladder cancer, bowel cancer, brain cancer, breast cancer, colon cancer, colorectal cancer, endometrial cancer, esophageal cancer, gallbladder cancer, gastric cancer, heart cancer, kidney cancer, lung cancer, liver cancer, melanoma, uterine cancer, lymphoma, ovarian cancer, pancreatic cancer, prostate cancer, thymic cancer, pheochromocytoma, medullary thyroid cancer, head and neck cancer or melanoma. In some embodiments, the mammal suffers from endocrine cancer. In some embodiments, the endocrine cancer includes adrenal tumors, neuroendocrine tumors, parathyroid tumors, pituitary tumors or thyroid tumors. In some embodiments, the mammal suffers from neuroendocrine tumors. In some embodiments, the mammal suffers from somatostatin receptor-positive gastroenteropancreatic neuroendocrine tumors (GEP-NET).

[0045] In another aspect, the present invention describes a method for targeting the delivery of a chemotherapeutic agent to a tumor in a mammal, which comprises administering to a mammal suffering from a tumor a compound of formula (I) or a pharmaceutically acceptable salt thereof.

[0046] In another aspect, the present invention describes a method for killing tumors overexpressing somatostatin subtype-2 receptor (SST2R) in a mammal, which comprises administering to the mammal a compound of formula (I) or a pharmaceutically acceptable salt thereof.

[0047] In any of the embodiments disclosed herein, the mammal is a human.

[0048] Other objects, features and advantages of the compounds, methods and compositions described herein will become apparent from the following detailed description. However, it should be understood that although specific embodiments are indicated, the detailed description and specific examples are given by way of illustration only, since various changes and modifications within the spirit and scope of the present disclosure will become apparent to those skilled in the art from this detailed description. BRIEF DESCRIPTION OF THE DRAWINGS

[0049] Figure 1 Time-activity curves of the selected organ activities of In-compound 1 in female Swiss nude mice bearing AR4J2 pancreatic tumors; showing the uptake of In-compound 1 in tumor and normal tissues in tumor-bearing animals (mean ± SD). 111 Time-activity curves of the selected organ activities of In-compound 1 in female Swiss nude mice bearing AR4J2 pancreatic tumors; showing the uptake of In-compound 1 in tumor and normal tissues in tumor-bearing animals (mean ± SD). 111 Uptake of In-compound 1 (mean ± SD).

[0050] Figure 2 Depicting the uptake of In-compound 1 alone and in the presence of excess In-compound 1 at 2 h post-dose in a xenograft mouse model with tumors of AR42J origin; showing the organ activities (% ID / g tissue) at 2 h post-dose and the selective blockade of the uptake of In-compound 1 in the presence of excess In-compound 1; this study demonstrated that specific SST2R-mediated uptake in tumors was blocked by excess In-compound 1 (mean ± SD). 111 In-compound 1 and in the presence of excess 115 Uptake of In-compound 1; showing the organ activities (% ID / g tissue) at 2 h post-dose and the selective blockade of the uptake of In-compound 1 in the presence of excess 115 In-compound 1 111 Selective blockade of the uptake of In-compound 1; this study demonstrated that specific SST2R-mediated uptake in tumors was blocked by excess 115 In-compound 1 (mean ± SD). Detailed description

[0051] Cancer is a disease in which some cells undergo genetic changes in controlling their growth and replication, resulting in uncontrolled growth and spread, and is one of the leading causes of death worldwide. General types of cancer include solid tumors (cancers that usually originate in organs), carcinomas (cancers that originate in the skin or tissue of organs), sarcomas (cancers of connective tissues such as bone), leukemias (cancers of the bone marrow), and lymphomas and myelomas (cancers of the immune system). A neoplasm is an abnormal growth of cells that gives rise to a solid tumor, which can be benign (i.e., not exhibiting malignant characteristics and generally less likely to become dangerous, such as an adenoma), malignant (i.e., exhibiting characteristics such as gene mutations, loss of normal function, rapid division, and the ability to metastasize (invade) other tissues), and of uncertain or unknown behavior. The treatment of neoplasms in the prior art is achieved through a combination of surgical procedures, chemotherapy, and radiotherapy. In some cases, surgical procedures can be curative, but usually require multiple interventions and are combined with radiotherapy and chemotherapy. In many cases, chemotherapy has proven to be a powerful weapon against cancer and requires further optimization. Chemotherapy is typically carried out by systemic administration of potent cytotoxic drugs, but these compounds lack tumor selectivity and thus also kill healthy cells in the body. This non-specific toxicity causes the severe side effects that are commonly associated with chemotherapy. Radiotherapy is the use of high-energy radiation to kill cells. The radiation source can be external-beam radiation (applied using an external source), internal radiation (placing radioactive material near the target cells), or radiotherapy from systemic administration of radioactive material. Similar to chemotherapy, many radiotherapy options also lack the tumor cell identification properties required to achieve the ultimate goal of targeted tumor therapy using drug molecules or radionuclides.

[0052] GPCRs are widespread and diverse integral membrane receptors and are thus expressed in every cell type in the body. GPCRs are generally poorly antigenic, making them difficult targets for antibody-based strategies. For many GPCRs, at any given time, the majority of the protein population resides in intracellular compartments, reducing the total number of cell surface binding sites accessible to antibodies or peptides.

[0053] Many human tumors overexpress different GPCRs, usually at significantly higher densities than other tissues. For example, gastroenteropancreatic (GEP) neuroendocrine tumors (NETs) overexpress somatostatin receptors, namely SSTR2, SSTR3, and SSTR5. Breast cancer also overexpresses SSTR2. Due to the complex GPCR overexpression profiles in neoplasms, targeting multiple receptors simultaneously may address problems such as heterogeneity, resistance, and phenotypic changes during disease progression, which have hampered many current treatment options.

[0054] The classes of somatostatin receptors (SSTRs) consist of five members (SSTR1, SSTR2, SSTR2, SSTR4, SSTR5), which are widely expressed in different tissues in the body, including the nervous system, pituitary gland, kidney, lung, and immune cells. Their natural ligand is the neuropeptide somatostatin (SST), which exists in two bioactive subtypes: SST-14 and SST-28. Binding to their receptors, both subtypes act as inhibitory hormones. An important physiological function of the SSTR / SST axis is, for example, the inhibition of growth hormone release. SSTRs, especially SSTR subtype 2, are highly expressed in many neoplastic cells and tumor vasculature. Overexpression of SSTRs (especially SSTR2) has been found in various neuroendocrine tumors as well as other tumors (such as breast cancer, ovarian cancer, and lung cancer). SSTR2 targeting for drug delivery can be achieved by using stable cyclic somatostatin analogs (such as octreotate, octreotide, and lanreotide). For example, covalently linking the DOTA chelator to octreotide (DOTA-TATE, also known as DOTA-(Tyr 3 )-octreotate) enables the targeted delivery of radionuclides to tumor cells expressing somatostatin receptors. 177 Lu DOTA-TATE therapy is a form of peptide receptor radionuclide therapy (PRRT) that targets somatostatin receptors and is a form of targeted drug delivery.

[0055] Most currently available GPCR-targeted drugs act on receptors whose natural ligands are small molecules, such as histamine, adrenaline, and neurotransmitters. Drugs targeting GPCRs whose natural ligands are peptides or proteins are usually also peptides or proteins.

[0056] Peptides are inherently sensitive to proteolytic enzymes and peptidases present in most tissues and are rapidly degraded into multiple fragments that no longer have significant affinity for the intended receptor. There are various ways to stabilize peptides (e.g., incorporating peptidomimetic structures or using more stable D-amino acids in the peptide backbone), but these modifications may lead to loss of affinity and / or selectivity and have a negative impact on physicochemical properties (e.g., poor solubility and tendency to aggregate). In addition, peptides may cause unwanted immunogenic responses, complicating late-stage development by masking the therapeutic effect and affecting safety assessment.

[0057] When a peptide ligand is linked to a cytotoxic effector molecule, the resulting conjugate is typically rapidly degraded in plasma and generates cytotoxic peptide fragments that can bind non-specifically to tumor and normal tissues. This premature breakdown of peptide-drug conjugates (PDCs) and antibody-drug conjugates (ADCs) reduces the amount of cytotoxic effector molecule distributed to the targeted tumor, decreases the therapeutic efficacy, and may increase toxicity. In addition, peptides are likely to be excreted only via the kidney, which may limit the application of PDCs. The significant renal uptake of some peptide-based therapeutic agents limits their conventional use.

[0058] High-affinity small molecule ligands that bind to peptide GPCRs and protein GPCRs (such as chemokine GPCRs) have been described, are cell permeable, and can act on receptor populations in the endoplasmic reticulum and endosomal compartments. Due to the low molecular weight of non-peptide small molecules, vascular permeability and tumor penetration should be improved compared to peptide- and antibody-based high molecular weight conjugates. In many cases, the binding affinity of small molecule non-peptide ligands exceeds that of FDA-approved antibodies by several orders of magnitude.

[0059] Provided herein are SMDCs, which are SST2R ligand-drug conjugates. The conjugated drug carrier or effector molecule moiety is linked to the ligand in a manner that does not affect the binding affinity of the small molecule SST2R ligand for SST2R. The conjugated drug carrier or effector molecule moiety includes chemotherapeutic agents and radionuclides, which are linked to the ligand using a spacer and / or linker moiety.

[0060] Solid tumor: Benign and / or malignant neoplasm (cancer)

[0061] In one aspect, the compounds of formula (I) are used for the treatment of benign and / or malignant neoplasms (solid tumors), wherein the neoplasm comprises cells that overexpress a cell surface GPCR.

[0062] As used herein, the term "neoplasm" refers to the abnormal growth of cells, which can proliferate in an uncontrolled manner and may have the ability to metastasize (spread).

[0063] Neoplasms include solid tumors, adenomas, carcinomas, sarcomas, leukemias, and lymphomas at any stage of the disease, with or without metastasis.

[0064] A solid tumor is an abnormal mass of tissue that usually does not contain cysts or areas of fluid. Solid tumors can be benign (non-cancerous) or malignant (cancerous). Different types of solid tumors are named for the type of cells that form them. Examples of tumors are sarcomas, carcinomas, and lymphomas. Leukemia (blood cancer) generally does not form solid tumors.

[0065] Solid tumors are cancers that typically originate in organs such as the bladder, intestine, brain, breast, endometrium, heart, kidney, lung, liver, uterus, ovary, pancreas, or other endocrine organs (thyroid) and the prostate.

[0066] An adenoma is a non-cancerous tumor. It originates from glandular cells of epithelial tissue (the thin layer of tissue that covers internal organs, glands, and other structures). Adenomas can grow from many glandular organs, including the adrenal glands, pituitary gland, thyroid gland, prostate, etc. Over time, adenomas may turn malignant, at which point they are called adenocarcinomas. Even when benign, they have the potential to cause serious health complications by compressing other structures (mass effect) and by producing large amounts of hormones in an unregulated, feedback-independent manner (causing paraneoplastic syndrome).

[0067] Adenomas are commonly found in the colon (e.g., adenomatous polyps, which have a tendency to become malignant and lead to colon cancer), kidney (e.g., renal adenomas may be precursor lesions to kidney cancer), adrenal glands (e.g., adrenal adenomas such as pheochromocytomas; some adrenal glands secrete hormones such as cortisol (causing Cushing's syndrome), aldosterone (leading to Conn's syndrome), or androgens (causing hyperandrogenism)), thyroid gland (e.g., thyroid adenomas), pituitary gland (e.g., pituitary adenomas such as prolactinomas), parathyroid glands (e.g., adenomas of the parathyroid glands may inappropriately secrete abnormally high amounts of parathyroid hormone and thereby cause primary hyperparathyroidism), liver (e.g., hepatocellular adenomas), breast (e.g., fibroadenomas), appendix (e.g., cystadenomas), bronchi (e.g., bronchial adenomas may cause carcinoid syndrome, a paraneoplastic syndrome), prostate (e.g., prostatic adenomas), sebaceous glands (e.g., sebaceous adenomas), and salivary glands.

[0068] Metastasis is the spread of malignant cells to new areas of the body, typically via the lymphatic system or bloodstream. Metastatic tumors are tumors that have spread from the primary site of origin or the site where they began to different areas of the body. Metastatic tumors contain malignant cells that express cell surface GPCRs.

[0069] The tumor formed by the spread cells is called a secondary tumor. The tumor may have spread to areas near the primary site, called regional metastasis, or to more distant parts of the body, called distant metastasis.

[0070] In some embodiments, the tumor to be treated comprises tumor cells expressing a GPCR, wherein the tumor is a primary or metastatic tumor. In some embodiments, the tumor to be treated comprises tumor cells expressing a GPCR, wherein the tumor is a primary or metastatic tumor of gastrointestinal origin, such as colorectal cancer, gastric cancer, small intestine cancer or esophageal cancer. In some embodiments, the tumor to be treated comprises tumor cells expressing a GPCR, wherein the tumor is a primary or metastatic tumor of the pancreas. In some embodiments, the tumor to be treated comprises tumor cells expressing a GPCR, wherein the tumor is a primary or metastatic tumor of the lung, such as squamous cell carcinoma, adenosquamous carcinoma or adenocarcinoma. In some embodiments, the tumor to be treated comprises tumor cells expressing a GPCR, wherein the tumor is a primary or metastatic neuroectodermal tumor, such as aphaechromotcytoma or paraganglioma. In some embodiments, the tumor to be treated comprises tumor cells expressing a GPCR, wherein the tumor is a primary or metastatic bronchopulmonary or gastrointestinal neuroendocrine tumor. In some embodiments, the tumor to be treated comprises tumor cells expressing a GPCR, wherein the tumor is a primary or metastatic tumor of the rectum or colon.

[0071] In some embodiments, the compound of formula (I) is used for treating sarcoma, such as leiomyosarcoma or rhabdomyosarcoma.

[0072] In some embodiments, the compound of formula (I) is used for treating adenoma.

[0073] In another aspect, provided herein is a method for treating cancer in a mammal, which comprises administering to the mammal in need thereof an SMDC disclosed herein. In some embodiments, the cancer comprises tumor cells expressing one or more peptide hormone GPCRs. In some embodiments, the cancer comprises tumor cells overexpressing one or more GPCRs. In some embodiments, the cancer includes solid tumors. In some embodiments, the cancer includes sarcoma, carcinoma or lymphoma. In some embodiments, the cancer includes neuroendocrine tumors. In some embodiments, the cancer includes insulinoma. In some embodiments, the cancer includes peptide hormone GPCR-positive (e.g., somatostatin receptor-positive) gastroenteropancreatic neuroendocrine tumors (GEP-NET).

[0074] In some embodiments, the compound of formula (I) is administered to a tumor patient. In some embodiments, the tumor patient is diagnosed with carcinoma, sarcoma, primary tumor, metastatic tumor, solid tumor, non-solid tumor, hematological tumor, leukemia or lymphoma.

[0075] Cancers include, but are not limited to, esophageal cancer, hepatocellular carcinoma, basal cell carcinoma (a form of skin cancer), squamous cell carcinoma (in various tissues), bladder cancer (including transitional cell carcinoma (malignant neoplasm of the bladder)), bronchial carcinoma, colon cancer, colorectal cancer, gastric cancer, lung cancer (including small cell lung cancer and non-small cell lung cancer), adrenocortical carcinoma, thyroid cancer, pancreatic cancer, breast cancer, ovarian cancer, prostate cancer, adenocarcinoma, sweat gland carcinoma, sebaceous gland carcinoma, papillary carcinoma, papillary adenocarcinoma, cystadenocarcinoma, medullary carcinoma, renal cell carcinoma, ductal carcinoma in situ or cholangiocarcinoma, choriocarcinoma, seminoma, embryonal carcinoma, Wilms' tumor, cervical cancer, uterine cancer, testicular cancer, osteogenic carcinoma, epithelial carcinoma, and nasopharyngeal carcinoma, etc.

[0076] Sarcomas include, but are not limited to, fibrosarcoma, myxosarcoma, liposarcoma, chondrosarcoma, chordoma, osteosarcoma, osteogenic sarcoma, angiosarcoma, endotheliosarcoma, lymphangiosarcoma, lymphangioendotheliosarcoma, synovioma, mesothelioma, Ewing's sarcoma, leiomyosarcoma, rhabdomyosarcoma, and other soft tissue sarcomas.

[0077] Solid tumors include, but are not limited to, glioma, astrocytoma, medulloblastoma, craniopharyngioma, ependymoma, pinealoma, hemangioblastoma, acoustic neuroma, oligodendroglioma, cerebral hemangioma, melanoma, neuroblastoma, and retinoblastoma. Benign solid tumors include adenoma.

[0078] Leukemias include, but are not limited to: a) chronic myeloproliferative syndromes (neoplastic disorders of pluripotent hematopoietic stem cells); b) acute myeloid leukemia (neoplastic transformation of pluripotent hematopoietic stem cells or hematopoietic cells with limited lineage potential); c) chronic lymphocytic leukemia (CLL; clonal proliferation of immunologically immature and functionally incompetent small lymphocytes), including B-cell CLL, T-cell CLL, prolymphocytic leukemia, and hairy cell leukemia; d) acute lymphoblastic leukemia (characterized by the accumulation of lymphoblasts). Lymphomas include, but are not limited to, B-cell lymphomas (e.g., Burkitt lymphoma); Hodgkin lymphoma; etc.

[0079] Primary and metastatic tumors include, for example, lung cancer (including but not limited to lung adenocarcinoma, squamous cell carcinoma, large cell carcinoma, bronchioloalveolar carcinoma, non-small cell carcinoma, small cell carcinoma, mesothelioma); breast cancer (including but not limited to ductal carcinoma, lobular carcinoma, inflammatory breast cancer, clear cell carcinoma, mucinous carcinoma); colorectal cancer (including but not limited to colon cancer, rectal cancer); anal cancer; pancreatic cancer (including but not limited to pancreatic adenocarcinoma, islet cell carcinoma, neuroendocrine tumor); prostate cancer; ovarian cancer (including but not limited to ovarian epithelial carcinoma or surface epithelial stromal tumors, including serous tumors, endometrioid tumors, and mucinous cystadenocarcinoma, sex cord-stromal tumors); liver cancer and cholangiocarcinoma (including but not limited to hepatocellular carcinoma, cholangiocarcinoma, hemangioma); esophageal cancer (including but not limited to esophageal adenocarcinoma and squamous cell carcinoma); non-Hodgkin lymphoma; bladder cancer; uterine cancer (including but not limited to endometrial adenocarcinoma, uterine papillary serous carcinoma, uterine clear cell carcinoma, uterine sarcoma and leiomyosarcoma, mixed Müllerian tumors); glioma, glioblastoma, medulloblastoma and other brain tumors; kidney cancer (including but not limited to renal cell carcinoma, clear cell carcinoma, Wilms tumor); head and neck cancer (including but not limited to squamous cell carcinoma); gastric cancer (including but not limited to gastric adenocarcinoma, gastrointestinal stromal tumor); thymic cancer; multiple myeloma; testicular cancer; germ cell tumor; neuroendocrine tumor; cervical cancer; carcinoid tumors of the gastrointestinal tract, breast and other organs; and signet ring cell carcinoma.

[0080] Small Molecule SST2R Ligands

[0081] In one aspect, a small molecule SST2R drug conjugate is described herein.

[0082] In some embodiments, the SMDC is a compound having the structure of formula (A) or a pharmaceutically acceptable salt thereof:

[0083]

[0084] Wherein:

[0085] A is -N(H)- or -O-;

[0086] X 1 is -N(R a )-, -O-, -C(=O)-, -C(=O)N(R a )-, -S(=O)-, -(CH2)C(R a )=N-O-(CH2)-;

[0087] R a is hydrogen or C1-C6 alkyl;

[0088] R b is hydrogen or C1-C6 alkyl;

[0089] Or when R a and R b both exist, then R a and R b together with the intervening atom(s) to which they are attached form piperidine or pyrrolidine;

[0090] q is 1, 2 or 3;

[0091] R 1 is hydrogen;

[0092] R 2 is hydrogen or C1-C6 alkyl;

[0093] R 3 is hydrogen, -OR 8 , -N(R 8 )2, -CN, halogen, C1-C6 alkyl or C1-C6 fluoroalkyl;

[0094] Or R 2 and R 3 together with the intervening atom to which they are attached form morpholine;

[0095] Each R 4 and R 5 is independently hydrogen, halogen, C1-C6 alkyl, C1-C6 fluoroalkyl, substituted or unsubstituted C1-C6 heteroalkyl, -CN, -N(R 8 )2 or -OR 8 ;

[0096] m is 1, 2 or 3;

[0097] R c is

[0098] Each R 6 and R 7 is independently hydrogen, halogen, C1-C4 alkyl, C1-C4 fluoroalkyl, C2-C4 alkenyl, C2-C4 alkynyl, substituted or unsubstituted C1-C6 heteroalkyl, substituted or unsubstituted phenyl, substituted or unsubstituted C3-C6 cycloalkyl, -CN, -OR 8 , -CO2R 8 , -C(=O)N(R 8 )2, -N(R 8 )2, -NR 8 C(=O)R 9 , -NR 8 C(=O)OR 9 , -SR 8 , -S(=O)R 9 , -SO2R9 or -SO2N(R 8 )2;

[0099] n is 1, 2, or 3;

[0100] Each R 8 is independently hydrogen, C1-C4 alkyl, C1-C4 fluoroalkyl, substituted or unsubstituted C1-C4 heteroalkyl;

[0101] Each R 9 is independently C1-C4 alkyl, C1-C4 fluoroalkyl, substituted or unsubstituted C1-C4 heteroalkyl;

[0102] L is -L 1 -L 2 -;

[0103] L 1 is an optional spacer;

[0104] L 2 is an optional linker;

[0105] wherein at least one of L 1 or L 2 is present;

[0106] R d is an effector moiety that comprises: (i) a chemotherapeutic agent; or (ii) a chelating moiety or its radionuclide complex.

[0107] In some embodiments, the SMDC is a compound having the structure of formula (A) or a pharmaceutically acceptable salt thereof:

[0108]

[0109] wherein:

[0110] A is -N(H)- or -O-;

[0111] X 1 is -N(R a )-, -O-, -C(=O)-, -C(=O)N(R a )-, -S(=O)-, -(CH2)C(R a )=N-O-(CH2)-;

[0112] R a is hydrogen or C1-C6 alkyl;

[0113] R b is hydrogen or C1-C6 alkyl;

[0114] or when Ra and R b When both are present, then R a and R b together with the intervening atom(s) to which they are attached form a piperidine or pyrrolidine;

[0115] q is 1, 2 or 3;

[0116] R 1 is hydrogen;

[0117] R 2 is hydrogen or C1-C6 alkyl;

[0118] R 3 is hydrogen, -OR 8 , -N(R 8 )2, -CN, halogen, C1-C6 alkyl or C1-C6 fluoroalkyl;

[0119] Or R 2 and R 3 together with the intervening atom(s) to which they are attached form a morpholine;

[0120] Each R 4 and R 5 is independently hydrogen, halogen, C1-C6 alkyl, C1-C6 fluoroalkyl, substituted or unsubstituted C1-C6 heteroalkyl, -CN, -N(R 8 )2 or -OR 8 ;

[0121] m is 1, 2 or 3;

[0122] R c is

[0123] Each R 6 and R 7 is independently hydrogen, halogen, C1-C4 alkyl, C1-C4 fluoroalkyl, C2-C4 alkenyl, C2-C4 alkynyl, substituted or unsubstituted C1-C6 heteroalkyl, substituted or unsubstituted phenyl, substituted or unsubstituted C3-C6 cycloalkyl, -CN, -OR 8 , -CO2R 8 , -C(=O)N(R 8 )2, -N(R 8 )2, -NR 8 C(=O)R 9 , -NR 8 C(=O)OR 9 , -SR 8 , -S(=O)R 9 , -SO2R 9 or -SO2N(R 8 )2;

[0124] n is 1, 2 or 3;

[0125] Each R 8 is independently hydrogen, C1-C4 alkyl, C1-C4 fluoroalkyl, substituted or unsubstituted C1-C4 heteroalkyl;

[0126] Each R 9 is independently C1-C4 alkyl, C1-C4 fluoroalkyl, substituted or unsubstituted C1-C4 heteroalkyl;

[0127] L is -L 1 -L 2 -;

[0128] L 1 is an optional spacer;

[0129] L 2 is an optional linker;

[0130] wherein at least one of L 1 or L 2 is present;

[0131] R d is an effector moiety comprising a chelating moiety or a radionuclide complex thereof.

[0132] In some embodiments, R 2 is hydrogen; R 3 is hydrogen, -OH or -OCH3; or R 2 and R 3 together with the intervening atom to which they are attached form morpholine.

[0133] In some embodiments, R 2 is hydrogen; and R 3 is hydrogen.

[0134] In some embodiments, each R 4 and R 5 is independently hydrogen, F, Cl, Br, C1-C4 alkyl, C1-C4 fluoroalkyl, -CN, -N(R 8 )2 or -OR 8 .

[0135] In some embodiments, each R 4 and R 5 is independently hydrogen, F, Cl, Br, -CH3, -CH2F, -CHF2, -CF3, -CN, -NH2, NHCH3, -N(CH3)2, -OH, -OCH3 or -OCF3.

[0136] In some embodiments, A is -N(H)-; R a is hydrogen, -CH3, or -CH2CH3; R b is hydrogen, -CH3, or -CH2CH3; or when R a and R b both exist, then R a and R b together with the intervening atom to which they are attached form a piperidine; and R 1 is hydrogen, -CH3, or -CH2CH3.

[0137] In some embodiments, the compound of formula (A) or a pharmaceutically acceptable salt thereof has the structure of formula (I):

[0138]

[0139] Wherein:

[0140] A is -N(H)- or -O-;

[0141] R a is hydrogen or a C1-C6 alkyl;

[0142] R 2 is hydrogen or a C1-C6 alkyl;

[0143] R 6 is chlorine or -C(=O)NH2;

[0144] L is -L 1 -L 2 -;

[0145] L 1 is an optional spacer; and

[0146] L 2 is an optional linker;

[0147] Wherein at least one of L 1 or L 2 exists; and

[0148] R d is an effector molecule moiety comprising a chemotherapeutic agent.

[0149] In some embodiments, A is -N(H)-.

[0150] In some embodiments, R 2 is hydrogen.

[0151] In some embodiments, R a is hydrogen or methyl. In some embodiments, R a is hydrogen.

[0152] In some embodiments, R 6 is chloro.

[0153] In some embodiments, the compound of formula (A) or a pharmaceutically acceptable salt thereof has the structure of formula (Ia):

[0154]

[0155] In some embodiments, the compound of formula (A) or a pharmaceutically acceptable salt thereof has the structure of formula (Ib):

[0156]

[0157] The compounds of the present disclosure also include tautomeric forms. Tautomeric forms are generated by the exchange of a single bond with an adjacent double bond and the accompanying proton migration. Tautomeric forms include prototropic tautomers, which are isomeric protonated states having the same empirical formula and total charge. Examples of prototropic tautomers include keto-enol pairs, amide-imino pairs, lactam-lactim pairs, amide-imino pairs, enamine-imine pairs, and cyclic forms in which a proton can occupy two or more positions of a heterocyclic system, such as 1H- and 3H-imidazole, 1H-, 2H-, and 4H-1,2,4-triazole, 2H- and 2H-isoindole, and 2H- and 2H-pyrazole. Tautomeric forms can be in equilibrium or locked in one form spatially by appropriate substitution.

[0158] Effector moiety (R) containing a cytotoxic effector molecule / drug d )

[0159] In some embodiments, R d comprises a chemotherapeutic agent.

[0160] In some embodiments, R d is:

[0161]

[0162] In some embodiments, R d is:

[0163]

[0164] Spacer and linker

[0165] In one embodiment, the SST2R targeting ligand is covalently linked to the effector molecule / drug via a spacer (L 1 ) and / or a linker (L 2 ).

[0166] In some embodiments, the spacer (L 1) has a defined length to link the ligand and the effector molecule or the linker / effector molecule while allowing an appropriate distance between the two. The spacer can also modulate the pharmacological activity of the SMDC.

[0167] In some embodiments, the spacer includes a dendritic spacer for covalently linking more than one drug moiety and / or ligand via a branched multifunctional moiety. The dendritic spacer can increase the molar ratio of the ligand to the effector molecule (i.e., the payload), which is related to the potency of the conjugate.

[0168] In some embodiments, L 1 is absent, or L 1 is a present spacer and is -X 2 -L 3 -L 4 -:

[0169] X 2 is -C(=O)(CH2) p -, -(CH2) p -, -C(=O)CH(CH2SO3H)NHC(=O)- or -(X 2a ) p -;

[0170] Each X 2a is independently selected from natural or unnatural amino acids, wherein any free amine in the amino acid is optionally independently substituted by -CH3;

[0171] p is 0, 1, 2, 3, 4, 5 or 6;

[0172] L 3 is absent or is unsubstituted or substituted C1-C 10 alkylene, unsubstituted or substituted C1-C 10 heteroalkylene, C4-C 20 polyethylene glycol or -(X 3 CH2CH2) t -;

[0173] Each X 3 is independently selected from O and NR 10 ;

[0174] Each t is independently 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12;

[0175] L 4 is absent or is -L 4a -(CH2) u -L 4b -(CH2) u -L 4c -;

[0176] L 4a is absent or is -O-, -NR 10 -, -NR 10 C(=O)-, -C(=O)NR 10 -, or -C(=O)-;

[0177] L 4b is absent or is an unsubstituted or substituted N-containing 5- to 10-membered heteroalkylene; wherein any free amine of the N-containing 5- to 10-membered heteroalkylene is optionally independently substituted by -CH2CO2H;

[0178] L 4c is absent or is -O-, -NR 10 -, -NR 10 C(=O)-, -C(=O)NR 10 -, -C(=O)NR 10 (CH2) u O(CH2) u C(=O)-, CH(CH2SO3H)C(=O)NR 10 (CH2) u O(CH2) u C(=O)-, -C(=O)-, -CH(=N)-, -CH(=N-NH)-, -CCH3(=N)-, -CCH3(=N-NH)-, -C(=O)-(C1-C6 alkylene)-, -C(=O)NR 10 -(C1-C6 alkylene)-, -NR 10 C(=O)-(C1-C6 alkylene)-, -NR 10 -(C1-C6 alkylene)- or C1-C6 alkylene-;

[0179] Each u is independently 0, 1, 2, 3, 4, 5 or 6; and

[0180] Each R 10 is independently selected from hydrogen and C1-C6 alkyl.

[0181] In some embodiments, L 4b is absent or is

[0182] In some embodiments, X 2 is -C(=O)(CH2) p -; p is 0, 1, 2, 3 or 4; L 3 is an unsubstituted or substituted C1-C 10 heteroalkylene, C4-C 20 polyethylene glycol or -(X 3(CH2CH2) t -; each X 3 is independently selected from O and NR 10 ; and each t is independently 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12.

[0183] In some embodiments, L 4 is absent or is -L 4a -; and L 4a is -NR 10 -, -NR 10 C(=O)-, -C(=O)NR 10 - or -C(=O)-.

[0184] In some embodiments, L 1 is absent, or L 1 is a spacer group that is present and is:

[0185]

[0186]

[0187] In some embodiments, L 1 is a spacer group that is present and is:

[0188]

[0189] In some embodiments, the linking group (L 2 ) is incorporated between the spacer group (L 1 ) and the effector molecule (R d ) and comprises a peptide linkage. The peptide linkage comprises L-amino acids and / or D-amino acids. In some embodiments, D-amino acids are preferred in order to minimize immunogenicity and non-specific cleavage by background peptidases or proteases. The cellular uptake of oligomeric D-arginine sequences is known to be as good as or better than that of oligomeric L-arginine.

[0190] The linking group unit can be "self-immolative" or "non-self-immolative". A "non-self-immolative" linking group unit is a linking group unit in which some or all of the spacer sequence units remain bound to the drug moiety upon enzymatic (e.g., proteolytic) cleavage of the conjugate. A "self-immolative" linking group unit allows release of the drug moiety without a separate hydrolysis step.

[0191] In certain embodiments, the linker comprises a p-aminobenzyl unit. In one such embodiment, p-aminobenzyl alcohol is linked to the amino acid unit via an amide bond, and a carbamate, methylcarbamate, or carbonate is formed between the benzyl alcohol and the cytotoxic agent. In one embodiment, the linker comprises p-aminobenzyloxycarbonyl (PAB). In certain embodiments, the phenylene moiety of the p-aminobenzyl unit is substituted with Q m where Q is -C1-C8 alkyl, -O-(C1-C8 alkyl), -halogen, -nitro, or -cyano; and m is an integer ranging from 0-4.

[0192] In some embodiments, the linker is cleavable. In some embodiments, the linker is designed to cleave in the presence of specific conditions or in a specific environment, such as near such targeted cells, tissues, or regions. The cleavable linker relies on the inherent properties of the cytoplasmic compartment of the cell to selectively release the cytotoxic drug. Such linkers mainly include chemically cleavable linkers that respond to low pH (acid-labile linkers) or a reducing environment (disulfide linkers), and enzymatically cleavable linkers that are sensitive to the action of certain lysosomal enzymes (peptide linkers or β-glucuronide linkers).

[0193] In some embodiments, the linker is cleavable under physiological conditions. In some embodiments, the linker is cleavable under intracellular conditions. In some embodiments, the linker is chemically cleavable. In some embodiments, the linker is enzymatically cleavable. In some embodiments, the linker is pH-sensitive, i.e., sensitive to hydrolysis at certain pH values. For example, the pH-sensitive linker can be hydrolyzable under acidic conditions. For example, the linker can be an acid-labile linker (e.g., hydrazone, semicarbazone, thiosemicarbazone, cis-aconitate, orthoester, acetal, ketal, etc.) that is hydrolyzable in lysosomes. Such linkers can be relatively stable under neutral pH conditions, such as in blood, but unstable at pH values below 7.0, such as the approximate pH of lysosomes and / or endosomes, e.g., pH 6.5 to 4.5.

[0194] In some embodiments, the linker comprises one or more disulfide bonds.

[0195] In some embodiments, the linker is cleaved in or near tissues that are hypoxic, such as cancer cells and cancerous tissues. In some embodiments, the linker comprises a disulfide bond. In some embodiments, the linker comprising a disulfide bond is preferentially cleaved in hypoxic regions. Hypoxia is thought to cause cancer cells to become more resistant to radiation and chemotherapy and also to trigger angiogenesis. In a hypoxic environment such as in the presence of leaky or necrotic cells, free thiols and other reducing agents become available extracellularly, while O2, which normally keeps the extracellular environment oxidized, is significantly depleted. In some embodiments, this shift in the redox balance promotes the reduction and cleavage of the disulfide bond within the linker.

[0196] In some embodiments, the linker is cleaved by intracellular peptidases or proteases, including but not limited to lysosomal or endosomal proteases. In some embodiments, the linker is cleaved by glycosidases (e.g., glucuronidase). Small peptide sequences such as Val-Cit and Phe-Lys have been developed as linkers for ADCs. These dipeptide linkers show good stability in serum but can be recognized and rapidly hydrolyzed by certain lysosomal proteases such as cathepsin B after internalization. β-Glucuronide linkers can be readily cleaved by the abundant lysosomal enzyme β-glucuronidase, thus facilitating the gentle and selective release of the active drug. In other embodiments, the linker is non-cleavable.

[0197] In some embodiments, the linker is cleaved by proteases, matrix metalloproteinases, serine proteases, or a combination thereof. In some embodiments, the linker is cleaved by reducing agents. In some embodiments, the linker is cleaved by oxidants or oxidative stress.

[0198] In some embodiments, the linker is cleaved by matrix metalloproteinases (MMPs). The hydrolytic activity of matrix metalloproteinases (MMPs) is associated with the invasive migration of metastatic tumor cells.

[0199] In some embodiments, the linker is cleaved by proteolytic enzymes or a reducing environment, such as may be present near cancerous cells. Such an environment or such enzymes are generally not present near normal cells.

[0200] In some embodiments, the linker is cleaved by serine proteases, including but not limited to thrombin and cathepsins. In some embodiments, the linker is cleaved by cathepsin K, cathepsin S, cathepsin D, cathepsin E, cathepsin W, cathepsin F, cathepsin A, cathepsin C, cathepsin H, cathepsin Z, or any combination thereof. In some embodiments, the linker is cleaved by cathepsin K and / or cathepsin S.

[0201] In some embodiments, the linking group is cleaved in a necrotic environment. Necrosis typically results in the release of enzymes or other cellular contents that can be used to trigger cleavage of the linking group. In some embodiments, cleavage of the linking group occurs by necrotic enzymes (e.g., calpains).

[0202] In some embodiments, L 2 is an optional non-cleavable linking group or a cleavable linking group.

[0203] In some embodiments, L 2 is an optional cleavable linking group that is an acid-sensitive linking group, a protease-sensitive linking group, or a glutathione-sensitive linking group.

[0204] In some embodiments, L 2 is absent, or L 2 is a present linking group and is -(L 2a ) w -L 2b - or -L 2c -;

[0205] Each L 2a is independently selected from natural or unnatural amino acids, wherein any free amine in the amino acid is optionally independently substituted with -CH3;

[0206] L 2b is absent or is -N(R 10 )(unsubstituted or substituted benzyl)-OC(=O)-; wherein the substituted benzyl is substituted with -C(=O)NHR 12 or a monosaccharide;

[0207] Each R 10 is independently selected from hydrogen and C1-C6 alkyl;

[0208] Each R 12 is independently selected from hydrogen, C4-C 20 polyethylene glycol, and unsubstituted or substituted C1-C6 alkyl, wherein the substituted C1-C6 alkyl is substituted with -NHR 13 , -C(=O)NHR 13 or -NHC(=O)R 13 ;

[0209] Each R 13 is independently selected from hydrogen, C4-C 20 polyethylene glycol, and C4-C 20 polyethylene glycol-NH2;

[0210] Or when R 13 is present and L 2aWhen at least one free carboxyl group of the amino acid is present, then R 13 forms a ring together with the free carboxyl group of the amino acid of L 2a ;

[0211] w is 1, 2, 3, 4, 5 or 6; and

[0212] each L 2c is N-maleimidomethyl-cyclohexane-1-carbonyl (MCC) or -S-.

[0213] In some embodiments, L 2b is selected from:

[0214]

[0215] In some embodiments, each L 2a is independently selected from natural or unnatural amino acids, any free amine of the amino acid being optionally independently substituted with -CH3, wherein the natural or unnatural amino acid is selected from alanine (Ala), Ala(SO3H), 3-(1-piperidyl)alanine, cyclohexylalanine, arginine (Arg), asparagine (Asn), aspartic acid (Asp), cysteine (Cys), glutamine (Gln), glutamic acid (Glu), glycine (Gly), leucine (Leu), lysine (Lys), methionine (Met), phenylalanine (Phe), homophenylalanine, proline (Pro), serine (Ser), 3-homoserine, tyrosine (Tyr), Tyr(SO3H), valine (Val), citrulline, β-alanine, β3-homoserine, β3-homolysine and β3-homoglutamic acid.

[0216] In some embodiments, each L 2a is independently selected from natural or unnatural amino acids, any free amine of the amino acid being optionally independently substituted with -CH3, wherein the natural or unnatural amino acid is selected from alanine (Ala), Ala(SO3H), arginine (Arg), asparagine (Asn), aspartic acid (Asp), cysteine (Cys), glutamine (Gln), glutamic acid (Glu), glycine (Gly), leucine (Leu), lysine (Lys), methionine (Met), phenylalanine (Phe), serine (Ser), valine (Val) and citrulline.

[0217] In some embodiments, L 2 is -(L 2a ) w -L 2b -; and -(L 2a ) w- are valine-citrulline, valine-alanine, methionine-valine-lysine, glycine-phenylalanine-glycine-glycine, tyrosine-arginine-valine, arginine-valine, and phenylalanine-lysine.

[0218] In some embodiments, L 2 is -L 2c -, and L 2c is N-maleimidomethyl-cyclohexane-1-carbonyl (MCC) or -S-.

[0219] In some embodiments, L 2 is absent, or L 2 is a linker that is present and is:

[0220]

[0221]

[0222]

[0223]

[0224]

[0225]

[0226]

[0227]

[0228] In some embodiments, L 2 is absent, or L 2 is a linker that is present and is:

[0229]

[0230] Representative Spacer-Linker Moieties

[0231] In some embodiments, L is:

[0232]

[0233]

[0234]

[0235]

[0236]

[0237]

[0238]

[0239]

[0240]

[0241]

[0242]

[0243]

[0244]

[0245]

[0246]

[0247]

[0248]

[0249]

[0250]

[0251]

[0252]

[0253]

[0254]

[0255]

[0256]

[0257]

[0258]

[0259]

[0260]

[0261] Representative Spacer / Linker and Effector Moieties

[0262] In some embodiments, -L-R d is:

[0263]

[0264]

[0265]

[0266]

[0267]

[0268]

[0269]

[0270]

[0271]

[0272]

[0273]

[0274]

[0275]

[0276]

[0277]

[0278]

[0279]

[0280]

[0281]

[0282]

[0283]

[0284]

[0285]

[0286]

[0287]

[0288]

[0289]

[0290]

[0291]

[0292]

[0293]

[0294]

[0295]

[0296]

[0297]

[0298] The present invention contemplates any combination of the groups described above for the various variables. Throughout the specification, one of ordinary skill in the art can select the groups and their substituents to provide stable moieties and compounds.

[0299] Representative SMDC Conjugates

[0300] The compound numbers listed below correspond to those recited in the Examples.

[0301] In some embodiments, the compound of formula (I) is Compound 6.

[0302] In some embodiments, the compound of formula (I) is Compound 7.

[0303] In some embodiments, the compound of formula (I) is Compound 8.

[0304] In some embodiments, the compound of formula (I) is Compound 9.

[0305] In some embodiments, the compound of formula (I) is Compound 10.

[0306] In some embodiments, the compound of formula (I) is Compound 11.

[0307] In some embodiments, the compound of formula (I) is Compound 13.

[0308] In some embodiments, the compound of formula (I) is Compound 14.

[0309] In some embodiments, the compound of formula (I) is Compound 19.

[0310] In some embodiments, the compound of formula (I) is Compound 20.

[0311] In some embodiments, the compound of formula (I) is Compound 21.

[0312] In some embodiments, the compound of formula (I) is Compound 22.

[0313] In some embodiments, the compound of formula (I) is Compound 23.

[0314] In some embodiments, the compound of formula (I) is Compound 24.

[0315] In some embodiments, the compound of formula (I) is Compound 25.

[0316] In some embodiments, the compound of formula (I) is Compound 26.

[0317] In some embodiments, the compound of formula (I) is Compound 27.

[0318] In some embodiments, the compound of formula (I) is Compound 28.

[0319] In some embodiments, the compound of formula (I) is Compound 29.

[0320] In some embodiments, the compound of formula (I) is Compound 30.

[0321] In some embodiments, the compound of formula (I) is Compound 31.

[0322] In some embodiments, the compound of formula (I) is Compound 32.

[0323] In some embodiments, the compound of formula (I) is Compound 33.

[0324] In some embodiments, the compound of formula (I) is Compound 34.

[0325] In some embodiments, the compound of formula (I) is Compound 35.

[0326] In some embodiments, the compound of formula (I) is Compound 36.

[0327] In some embodiments, the compound of formula (I) is Compound 37.

[0328] In some embodiments, the compound of formula (I) is Compound 38.

[0329] In some embodiments, the compound of formula (I) is Compound 39.

[0330] In some embodiments, the compound of formula (I) is Compound 40.

[0331] In some embodiments, the compound of formula (I) is Compound 41.

[0332] In some embodiments, the compound of formula (I) is Compound 42.

[0333] In some embodiments, the compound of formula (I) is Compound 43.

[0334] In some embodiments, the compound of formula (I) is Compound 44.

[0335] In some embodiments, the compound of formula (I) is Compound 45.

[0336] In some embodiments, the compound of formula (I) is Compound 46.

[0337] In some embodiments, the compound of formula (I) is Compound 47.

[0338] In some embodiments, the compound of formula (I) is Compound 48.

[0339] In some embodiments, the compound of formula (I) is Compound 49.

[0340] In some embodiments, the compound of formula (I) is Compound 50.

[0341] In some embodiments, the compound of formula (I) is Compound 51.

[0342] In some embodiments, the compound of formula (I) is Compound 52.

[0343] In some embodiments, the compound of formula (I) is Compound 53.

[0344] In some embodiments, the compound of formula (I) is Compound 54.

[0345] In some embodiments, the compound of formula (I) is Compound 55.

[0346] In some embodiments, the compound of formula (I) is Compound 56.

[0347] In some embodiments, the compound of formula (I) is Compound 57.

[0348] In some embodiments, the compound of formula (I) is Compound 58.

[0349] In some embodiments, the compound of formula (I) is Compound 59.

[0350] In some embodiments, the compound of formula (I) is Compound 60.

[0351] In some embodiments, the compound of formula (I) is Compound 61.

[0352] In some embodiments, the compound of formula (I) is Compound 68.

[0353] In some embodiments, the compound of formula (I) is Compound 69.

[0354] In some embodiments, the compound of formula (I) is Compound 70.

[0355] In some embodiments, the compound of formula (I) is Compound 71.

[0356] In some embodiments, the compound of formula (I) is Compound 72.

[0357] In some embodiments, the compound of formula (I) is Compound 73.

[0358] In some embodiments, the compound of formula (I) is Compound 74.

[0359] In some embodiments, the compound of formula (I) is Compound 75.

[0360] In some embodiments, the compound of formula (I) is Compound 77.

[0361] In some embodiments, the compound of formula (I) is Compound 78.

[0362] In some embodiments, the compound of formula (I) is Compound 81.

[0363] In some embodiments, the compound of formula (I) is Compound 82.

[0364] In some embodiments, the compound of formula (I) is Compound 83.

[0365] In some embodiments, the compound of formula (I) is Compound 84.

[0366] In some embodiments, the compound of formula (I) is Compound 85.

[0367] In some embodiments, the compound of formula (I) is Compound 86.

[0368] In some embodiments, the compound of formula (I) is Compound 87.

[0369] In some embodiments, the compound of formula (I) is Compound 88.

[0370] In some embodiments, the compound of formula (I) is compound 89.

[0371] In some embodiments, the compound of formula (I) is compound 90.

[0372] In some embodiments, the compound of formula (I) is compound 91.

[0373] In some embodiments, the compound of formula (I) is compound 92.

[0374] In some embodiments, the compound of formula (I) is compound 93.

[0375] In some embodiments, the compound of formula (I) is compound 94.

[0376] In some embodiments, the compound of formula (I) is compound 95.

[0377] In some embodiments, the compound of formula (I) is compound 96.

[0378] In some embodiments, the compound of formula (I) is compound 97.

[0379] In some embodiments, the compound of formula (I) is compound 98.

[0380] In some embodiments, the compound of formula (I) is compound 99.

[0381] In some embodiments, the compound of formula (I) is compound 100.

[0382] In some embodiments, the compound of formula (I) is compound 101.

[0383] In some embodiments, the compound of formula (I) is compound 102.

[0384] In some embodiments, the compound of formula (I) is compound 103.

[0385] In some embodiments, the compound of formula (I) is compound 104.

[0386] In some embodiments, the compound of formula (I) is compound 105.

[0387] In some embodiments, the compound of formula (I) is compound 106.

[0388] In some embodiments, the compound of formula (I) is compound 107.

[0389] In some embodiments, the compound of formula (I) is compound 108.

[0390] In some embodiments, the compound of formula (I) is Compound 109.

[0391] In some embodiments, the compound of formula (I) is Compound 110.

[0392] In some embodiments, the compound of formula (I) is Compound 111.

[0393] In some embodiments, the compound of formula (I) is Compound 112.

[0394] In some embodiments, the compound of formula (I) is Compound 113.

[0395] In some embodiments, the compound of formula (I) is Compound 114.

[0396] In some embodiments, the compound of formula (I) is Compound 115.

[0397] In some embodiments, the compound of formula (I) is Compound 116.

[0398] In some embodiments, the compound of formula (I) is Compound 117.

[0399] In some embodiments, the compound of formula (I) is Compound 118.

[0400] In some embodiments, the compound of formula (I) is Compound 119.

[0401] In some embodiments, the compound of formula (I) is Compound 120.

[0402] In some embodiments, the compound of formula (I) is Compound 121.

[0403] In some embodiments, the compound of formula (I) is Compound 122.

[0404] In some embodiments, the compound of formula (I) is Compound 123.

[0405] In some embodiments, the compound of formula (I) is Compound 124.

[0406] In some embodiments, the compound of formula (I) is Compound 125.

[0407] In some embodiments, the compound of formula (I) is Compound 126.

[0408] In some embodiments, the compound of formula (I) is Compound 127.

[0409] In some embodiments, the compound of formula (I) is Compound 128.

[0410] In some embodiments, the compound of formula (I) is Compound 129.

[0411] In some embodiments, the compound of formula (I) is Compound 130.

[0412] In some embodiments, the compound of formula (I) is Compound 131.

[0413] In some embodiments, the compound of formula (I) is Compound 132.

[0414] In some embodiments, the compound of formula (I) is Compound 133.

[0415] In some embodiments, the compound of formula (I) is Compound 134.

[0416] In some embodiments, the compound of formula (I) is Compound 135.

[0417] In some embodiments, the compound of formula (I) is Compound 136.

[0418] In some embodiments, the compound of formula (I) is Compound 137.

[0419] In some embodiments, the compound of formula (I) is Compound 138.

[0420] In some embodiments, the compound of formula (I) is Compound 139.

[0421] In some embodiments, the compound of formula (I) is Compound 140.

[0422] In some embodiments, the compound of formula (I) is Compound 141.

[0423] In some embodiments, the compound of formula (I) is Compound 142.

[0424] In some embodiments, the compound of formula (I) is Compound 143.

[0425] In some embodiments, the compound of formula (I) is Compound 144.

[0426] In some embodiments, the compound of formula (I) is Compound 145.

[0427] In some embodiments, the compound of formula (I) is Compound 146.

[0428] In some embodiments, the compound of formula (I) is Compound 147.

[0429] In some embodiments, the compound of formula (I) is Compound 148.

[0430] In some embodiments, the compound of formula (I) is Compound 149.

[0431] In some embodiments, the compound of formula (I) is Compound 150.

[0432] In some embodiments, the compound of formula (I) is Compound 151.

[0433] In some embodiments, the compound of formula (I) is Compound 152.

[0434] In some embodiments, the compound of formula (I) is Compound 153.

[0435] In some embodiments, the compound of formula (I) is Compound 154.

[0436] In some embodiments, the compound of formula (I) is Compound 155.

[0437] In some embodiments, the compound of formula (I) is Compound 156.

[0438] In some embodiments, the compound of formula (I) is Compound 157.

[0439] In some embodiments, the compound of formula (I) is Compound 158.

[0440] In some embodiments, the compound of formula (I) is Compound 159.

[0441] Effector molecule moiety (R d ) containing a chelated radionuclide

[0442] Radiopharmaceuticals have increasingly become very useful tools for physicians to diagnose, stage, treat, and monitor the progression of various diseases, especially cancer. The main difference between radiopharmaceuticals and other drugs is that radiopharmaceuticals contain radionuclides. The nuclear decay characteristics of radionuclides determine whether a radiopharmaceutical is used as a diagnostic agent or a therapeutic agent in clinical practice. Diagnostic radiopharmaceuticals require radionuclides that emit gamma rays or positrons (β+), which subsequently annihilate with nearby electrons, producing two 511 keV annihilation photons emitted approximately 180° apart from each other. Radionuclides that emit gamma rays (e.g., 99m Tc, 111 In, 201 Tl, etc.) can be used for single photon emission computed tomography (SPECT), while radionuclides that emit positrons (e.g., 18 F, 89 Zr, 68 Ga, etc.) can be used for positron emission tomography (PET).

[0443] In contrast, therapeutic radiopharmaceuticals require radionuclides that emit particulate radiation, such as alpha particles, beta (β-) particles, or Auger electrons. These particles interact strongly with the target tissue (e.g., a cancerous tumor) and cause extensive local ionization, which can break chemical bonds in DNA molecules and may induce cytotoxicity.

[0444] For most nuclear medicine applications, diagnostic radiopharmaceuticals need to be paired with therapeutic radiopharmaceuticals. This concept is commonly referred to as "theranostics." As a first step in the theranostics concept, positron emission tomography (PET) or single photon emission computed tomography (SPECT) is used to quantitatively image tumor imaging biomarkers with a target molecule labeled with a diagnostic radionuclide. When it is demonstrated that a tumoricidal radiation absorbed dose can be delivered to the tumor and metastases using this targeted molecule, as a second step, the same or a similar target molecule labeled with a therapeutic radionuclide is administered.

[0445] In some embodiments, the chemical and pharmacokinetic behaviors of both the diagnostic radiopharmaceutical and the therapeutic radiopharmaceutical are matched. In some embodiments, the diagnostic radionuclide and the therapeutic radionuclide are a pair of chemically identical radioisotopes (also referred to as a "matched pair"). An example of a pair used for theranostic radiopharmaceutical applications is 123 I / 131 I pair, where the 123 I-labeled compound is used for diagnosis, while the 131 I-labeled compound is used for treatment. Other theranostic matched pairs include 44 Sc / 47 Sc, 64 Cu / 67 Cu, 72 As / 77 As, 86 Y / 90 Y and 203 Pb / 212 Pb, etc. Alternatively, when the chemical actions of pairs of radionuclides from different elements are very similar (e.g., 99m Tc / 186 / 188 Re) and there are no significant differences in the pharmacokinetic behaviors between the diagnostic analog and the therapeutic analog, they can be used for theranostic radiopharmaceutical development. Another example is the 68 Ga / 177 Lu pair, where 68 Ga is used for diagnosis and 177 Lu is used for treatment. For example, gastroenteropancreatic endocrine tumors express a large amount of sst2 receptors, which can be imaged using 68 Ga sst2 ligand conjugates ( 68Ga]Ga-DOTA-TATE (NETSPOT TM ) or 68 Ga]Ga-DOTA-TOC (DOTA-(D-Phe1,Tyr3)-octreotide (SomaKit )) are used in somatostatin receptor scintigraphy for targeting for diagnostic purposes, and then 177 Lu sst2 ligand conjugate ( 177 Lu]Lu-DOTA-TATE) is used for treatment to effectuate internal radiotherapy.

[0446] Chelating agents for radionuclides

[0447] As used herein, "chelating agent" and "chelating moiety" are used interchangeably.

[0448] In some embodiments, the chelating agent is capable of binding a radioactive atom. In some embodiments, the binding is direct; for example, the chelating agent forms a hydrogen bond or an electrostatic interaction with the radioactive atom. In some embodiments, the binding is indirect; for example, the chelating agent binds to a molecule containing the radioactive atom. In some embodiments, the chelating agent is or comprises a macrocycle.

[0449] In some embodiments, the chelating agent comprises one or more amine groups. In some embodiments, the metal chelating agent comprises two or more amine groups. In some embodiments, the chelating agent comprises three or more amine groups. In some embodiments, the chelating agent comprises four or more amine groups. In some embodiments, the chelating agent comprises 4 or more N atoms, 4 or more carboxylic acid groups, or a combination thereof. In some embodiments, the chelating agent does not comprise S. In some embodiments, the chelating agent comprises a ring. In some embodiments, the ring comprises O and / or N atoms. In some embodiments, the chelating agent is a ring comprising 3 or more N atoms, 3 or more carboxylic acid groups, or a combination thereof. In some embodiments, the chelating agent is a multidentate ligand, a bidentate ligand, or a monodentate ligand. Multidentate ligands have different numbers of atoms for bonding to a metal atom or ion. EDTA is a hexadentate ligand and is an example of a multidentate ligand that has six donor atoms with electron pairs that can be used for bonding to a central metal atom or ion. Bidentate ligands have two donor atoms that enable them to bind to a central metal atom or ion at two points. Ethylenediamine (en) and oxalate ion (ox) are examples of bidentate ligands.

[0450] In some embodiments, the chelating agents described herein include cyclic chelating agents or acyclic chelating agents. In some embodiments, the chelating agents described herein include cyclic chelating agents. In some embodiments, the chelating agents described herein include acyclic chelating agents.

[0451] In some embodiments, the chelator is or comprises DOTA, HBED-CC, DOTAGA, DOTA(GA)2, NOTA, and DOTAM. In some embodiments, the chelator is or comprises NODAGA, NOTA, DOTAGA, DOTA(GA)2, TRAP, NOPO, NCTA, DFO, DTPA, and HYNIC.

[0452] In some embodiments, the chelator comprises a macrocycle, e.g., a macrocycle comprising O and / or N atoms, DOTA, HBED-CC, DOTAGA, DOTA(GA)2, NOTA, DOTAM, one or more amines, one or more ethers, one or more carboxylic acids, EDTA, DTPA, TETA, DO3A, PCTA, or desferrioxamine.

[0453] In some embodiments, R d is a chelating moiety selected from the group consisting of: 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA); 1,4,7,10-tetraazacyclododecane-1,4,7-triacetic acid (DO3A); 1,4,7,10-tetraazacyclododecane-1,7-diacetic acid (DO2A); α,α',α”,α”'-tetramethyl-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTMA); 1,4,7,10-tetra(carbamoylmethyl)-1,4,7,10-tetraazacyclododecane (DOTAM); 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrapropionic acid (DOTPA); 2,2',2”-(10-(2-amino-2-oxoethyl)-1,4,7,10-tetraazacyclododecane-1,4,7-triyl)triacetic acid; benzyl-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (Bn-DOTA); 6,6'-(((pyridine-2,6-diylbis(methylene))bis((carboxymethyl)azanediyl))-bis(methylene))pyridine dicarboxylic acid (H4pypa); H4pypa-benzyl; 6,6',6”,6”'-(((pyridine-2,6-diylbis(methylene))bis(azanetriyl))-tetra(methylene))-tetrapyridinecarboxylic acid (H4py4pa); H4py4pa-benzyl; H4octapa-benzyl; 3,6,9,12-tetra(carboxymethyl)-3,6,9,12-tetraazatetradecanedioic acid (TTHA); or a radionuclide complex thereof.

[0454] In some embodiments, R dis 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA); or 1,4,7,10-tetraazacyclododecane-1,4,7-triacetic acid (DO3A); or a radionuclide complex thereof.

[0455] In some embodiments, R d is

[0456]

[0457] or a radionuclide complex thereof.

[0458] In some embodiments, R d is or a radionuclide complex thereof.

[0459] Radionuclide

[0460] In some embodiments, the conjugate comprises a radionuclide that emits Auger electrons, an α-emitting radionuclide, a β-emitting radionuclide, or a γ-emitting radionuclide. In some embodiments, the conjugate comprises a radionuclide that emits Auger electrons, which is indium-111 ( 111 In), gallium-67 ( 67 Ga), gallium-68 ( 68 Ga), technetium-99m ( 99m Tc), or platinum-195m ( 195m Pt). In some embodiments, the conjugate comprises an α-emitting radionuclide, which is actinium-225 ( 225 Ac), bismuth-213 ( 213 Bi), radium-223 ( 223 Ra), or lead-212 ( 212 Pb). In some embodiments, the conjugate comprises a β-emitting radionuclide, which is yttrium-90 ( 90 Y), lutetium-177 ( 177 Lu), iodine-131 ( 131 I), rhenium-186 ( 186 Re), rhenium-188 ( 188 Re), copper-64 ( 64 Cu), copper-67 ( 67 Cu), samarium-153 ( 153 Sm), strontium-89 ( 89 Sr), gold-198 ( 198 Au), erbium-169 ( 169 Er), dysprosium-165 ( 165 Dy), technetium-99m ( 99m Tc), zirconium-8989 zirconium (Zr) or manganese-52 ( 52 Mn). In some embodiments, the conjugate comprises a gamma-emitting radionuclide which is cobalt-60 ( 60 Co), palladium-103 ( 103 Pd), cesium-137 ( 137 Cs), ytterbium-169 ( 169 Yb), iridium-192 ( 192 Ir) or radium-226 ( 226 Ra).

[0461] In some embodiments, the conjugate comprises a radionuclide and a chelator configured to bind the radionuclide, wherein the radionuclide is suitable for positron emission tomography (PET) analysis, single photon emission computed tomography (SPECT) or magnetic resonance imaging (MRI). In some embodiments, the radionuclide is copper-64 ( 64 Cu), gallium-68 ( 68 Ga), indium-111 ( 111 In) or technetium-99m ( 99m Tc).

[0462] Auger electrons (AE) are electrons with very low energy emitted by radionuclides that decay by electron capture (EC) (e.g., 111 In, 67 Ga, 99m Tc, 195m Pt, 125 I and 123 I). This energy deposition occurs over nano-micron distances, resulting in high linear energy transfer, which effectively causes lethal damage to cancer cells. Therefore, radiotherapeutic agents that emit AE have great potential for treating cancer.

[0463] β-particles are electrons emitted from the atomic nucleus. They typically have a longer range in tissue (about 1 - 5 mm) and are the most commonly used.

[0464] An α-particle is a helium nucleus (two protons and two neutrons) emitted from the nucleus of a radioactive atom. Depending on the emission energy, they can travel 50 - 100 μm in tissue. They are positively charged and are several orders of magnitude larger than electrons. The amount of energy deposited per path length traveled by an α-particle (referred to as ‘linear energy transfer’) is approximately 400 times that of an electron. This results in substantially more damage along its path than that caused by an electron. α-Particle tracks result in a large number of complex and mostly irreparable DNA double-strand breaks. The absorbed dose required to achieve cytotoxicity is related to the number of α-particles traversing the cell nucleus. Using this as a measure, cytotoxicity may be achieved when the number of α-particles traversing the cell nucleus ranges from 1 to 20. The resulting high potency, along with the short range of α-particles (which reduces normal organ toxicity), has generated significant interest in the development of agents that emit α-particles. Commonly used α-particle emitters include bismuth-212, lead-212, bismuth-213, actinium-225, radium-223, and thorium-227.

[0465] In some embodiments, the conjugate comprises a diagnostic or therapeutic radionuclide.

[0466] Representative radionuclides

[0467]

[0468]

[0469] Radionuclides have available emission characteristics that can be used in diagnostic imaging techniques such as single photon emission computed tomography (SPECT, e.g., 67 Ga, 99m Tc, 111 In, 177 Lu) and positron emission tomography (PET, e.g., 68 Ga, 64 Cu, 44 Sc, 86 Y, 89 Zr), as well as in therapeutic applications (e.g., 47 Sc, 114 mIn, 177 Lu, 90 Y, 212 / 213 Bi, 212 Pb, 225 Ac, 186 / 188Re). The basic component of a radiopharmaceutical based on a radioactive metal is a chelator, which is a ligand system that binds the radioactive metal ion in a tight and stable coordination complex, enabling it to be correctly directed to the desired molecular target in the body. The art provides guidance on selecting the optimal match between the chelator and the radioactive metal for a particular use (see, e.g., Price et al., “Matching chelators to radiometals for radiopharmaceuticals”, Chem. Soc. Rev., 2014, 43, 260 - 290).

[0470] In some embodiments, R d comprises a chelated radionuclide suitable for positron emission tomography (PET) analysis or single photon emission computed tomography (SPECT). In some embodiments, R d comprises a chelated radionuclide suitable for single photon emission computed tomography (SPECT). In some embodiments, R d comprises a chelated radionuclide suitable for positron emission tomography (PET) analysis. In some embodiments, R d comprises a chelated radionuclide suitable for positron emission tomography imaging, positron emission tomography with computed tomography imaging, or positron emission tomography with magnetic resonance imaging.

[0471] Response and toxicity prediction are crucial for the rational implementation of cancer therapy. The biological effects of radionuclide therapy are mediated by a well - defined physical quantity, the absorbed dose (D), which is defined as the energy absorbed per unit mass of tissue.

[0472] Radiation dosimetry is the measurement, calculation, and assessment of the ionizing radiation dose absorbed by a target, typically the human body, and can be considered the equivalent form of the ability to perform a pharmacodynamic study in real - time in the treated patient. This applies both internally due to the ingestion or inhalation of radioactive substances and externally due to irradiation by a radiation source. Dosimetry analysis can be performed as part of the patient's treatment to calculate the absorbed dose in the tumor compared to normal organs and thus calculate the likelihood of treatment success.

[0473] In some embodiments, R d is or its radionuclide complex.

[0474] In some embodiments, -L - R d is: -CH2CH2NH - R d 、-C(=O)CH2NH - R d 、-C(=O)CH2CH2NH - Rd , -CH2CH2(OCH2CH2)2NH-R d , -CH2CH2(OCH2CH2)3NH-R d , -CH2CH2(OCH2CH2)4NH-R d , -CH2CH2(OCH2CH2)5NH-R d , -CH2CH2(OCH2CH2)6NH-R d , -CH2CH2(OCH2CH2)7NH-R d , -CH2CH2(OCH2CH2)8NH-R d , -C(=O)CH2CH2(OCH2CH2)2NH-R d , -C(=O)CH2CH2(OCH2CH2)3NH-R d , -C(=O)CH2CH2(OCH2CH2)4NH-R d , -C(=O)CH2CH2(OCH2CH2)5NH-R d , -C(=O)CH2CH2(OCH2CH2)6NH-R d , -C(=O)CH2CH2(OCH2CH2)7NH-R d or -C(=O)CH2CH2(OCH2CH2)8NH-R d ;

[0475] R d is or its radionuclide complex.

[0476] In some embodiments, -L 1 -R d is: -(PEG2)NH-R d , -(PEG3)NH-R d , -(PEG4)NH-R d , -(PEG5)NH-R d , -(PEG6)NH-R d , -(PEG7)NH-R d , -(PEG8)NH-R d , -C(=O)(PEG2)NH-R d , -C(=O)(PEG3)NH-R d , -C(=O)(PEG4)NH-R d , -C(=O)(PEG5)NH-R d , -C(=O)(PEG6)NH-Rd 、 -C(=O)(PEG7)NH-R d or -C(=O)(PEG8)NH-R d ;

[0477] R d is or a radionuclide complex thereof.

[0478] In some embodiments, -L 1 -R d is:

[0479]

[0480] or a radionuclide complex thereof.

[0481] In some embodiments, the radionuclide of the radionuclide complex is a lanthanide element or an actinide element.

[0482] In some embodiments, the radionuclide of the radionuclide complex is actinium, bismuth, cesium, cobalt, copper, dysprosium, erbium, gold, indium, iridium, gallium, lead, lutetium, manganese, palladium, platinum, radium, rhenium, samarium, strontium, technetium, ytterbium, yttrium or zirconium.

[0483] In some embodiments, the radionuclide of the radionuclide complex is a diagnostic or therapeutic radionuclide.

[0484] In some embodiments, the radionuclide of the radionuclide complex is a radionuclide that emits Auger electrons, an alpha-emitting radionuclide, a beta-emitting radionuclide or a gamma-emitting radionuclide.

[0485] In some embodiments, the radionuclide of the radionuclide complex is copper-64 ( 64 Cu), 67-copper ( 67 Cu), 111-indium ( 111 In), 115-indium ( 115 In), 67-gallium ( 67 Ga), 68-gallium ( 68 Ga), 70-gallium ( 70 Ga), 225-actinium ( 225 Ac), 175-lutetium ( 175 Lu), 177-lutetium ( 177 Lu) or 212-lead ( 212 Pb).

[0486] In some embodiments, the radionuclide of the radionuclide complex is 111-indium ( 111 In), 115-indium ( 115 In), 67-gallium (67 Ga), 68-Gallium( 68 Ga), 70-Gallium( 70 Ga), 225-Actinium( 225 Ac), 175-Lutetium( 175 Lu) or 177-Lutetium( 177 Lu).

[0487] Synthesis of Compounds

[0488] The compounds described herein are synthesized using standard synthetic techniques or using a combination of methods known in the art and the methods described herein.

[0489] Unless otherwise indicated, conventional methods of mass spectrometry and method, NMR, HPLC are employed.

[0490] The compounds are prepared using standard organic chemistry techniques, such as those described in, for example, March's Advanced Organic Chemistry, 6th Edition, John Wiley and Sons, Inc. Alternative reaction conditions for the synthetic transformations described herein can be employed, such as variations in solvents, reaction temperatures, reaction times, and different chemical reagents and other reaction conditions.

[0491] In one aspect, the compounds described herein are in the form of pharmaceutically acceptable salts.

[0492] The term "pharmaceutically acceptable salts" refers to a form of a therapeutic active agent that consists of a combination of the therapeutic active agent in cationic form with a suitable anion, or in alternative embodiments, a combination of the therapeutic active agent in anionic form with a suitable cation. Handbook of Pharmaceutical Salts: Properties, Selection and Use. International Union of Pure and Applied Chemistry, Wiley-VCH 2002. S.M. Berge, L.D. Bighley, D.C. Monkhouse, J. Pharm. Sci. 1977, 66, 1-19.

[0493] In some embodiments, the pharmaceutically acceptable salts are obtained by reacting the compound of formula (I) with an acid. In some embodiments, the compound of formula (I) (i.e., the free base form) is basic and reacts with an organic or inorganic acid.

[0494] In some embodiments, a pharmaceutically acceptable salt is obtained by reacting a compound of formula (I) with a base. In some embodiments, the compound of formula (I) is acidic and reacts with a base. In such cases, the acidic proton of the compound of formula (I) is replaced by a metal ion.

[0495] In some embodiments, the compound of formula (I) has one or more stereocenters, and each stereocenter independently exists in the R or S configuration. In some embodiments, the compound of formula (I) exists in the R configuration. In some embodiments, the compound of formula (I) exists in the S configuration. The compounds presented herein include all diastereoisomeric, individual enantiomeric, atropisomeric, and epimeric forms and their appropriate mixtures. The compounds and methods provided herein include all cis, trans, syn, anti, entgegen (E), and zusammen (Z) isomers and their appropriate mixtures.

[0496] If desired, individual stereoisomers are obtained by methods such as stereoselective synthesis and / or separation of stereoisomers by chiral chromatography columns or separation of diastereoisomers by achiral or chiral chromatography columns or crystallization and recrystallization in suitable solvents or solvent mixtures. In certain embodiments, the compound of formula (I) is prepared as its respective stereoisomers by reacting a racemic mixture of the compound with an optically active resolving agent to form a pair of diastereoisomeric compounds / salts, separating the diastereoisomers, and recovering the optically pure individual enantiomers. In some embodiments, covalent diastereomeric derivatives of the compounds described herein are used for the resolution of individual enantiomers. In another embodiment, the diastereomers are separated by separation / resolution techniques based on solubility differences. In other embodiments, the separation of stereoisomers is carried out by chromatography or by forming diastereomeric salts and separating them by recrystallization or chromatography or any combination thereof. Jean Jacques, Andre Collet, Samuel H. Wilen, “Enantiomers, Racemates and Resolutions”, John Wiley And Sons, Inc., 1981. In some embodiments, stereoisomers are obtained by stereoselective synthesis.

[0497] In some embodiments, the cytotoxic SMDCs of the present invention described herein are prepared as described in Schemes A - F. In the following schemes, the carbonyl moieties on both sides of “X” and “X” represent the spacer moiety L 1 。

[0498] Scheme A:

[0499]

[0500] a) DIEA, ACN; b) NBS, DMF; c) (3-fluoro-5-methylphenyl)boronic acid, Pd, toluene / H2O, catalyst; d) DMF / H2O; e) NH2(CH2) n NH2, 2,6-dimethylpyridine; f) acid, FDPP, NMM, DMF; g) Val-Cit-PAB-MMAE, FDPP, NMM, DMF; h) deprotection.

[0501] Scheme B:

[0502]

[0503] a) HATU, DIEA, DMF; b) TFA, DCM; c) acetate buffer pH = 5.6, DMF. Scheme C:

[0504]

[0505] a) TrtSCH2CH2NH2, HSTU, DIEA, DMF; b) TFA, TIS; c) ACN.

[0506] Scheme D:

[0507]

[0508] a) AcOH, DMF, 0.2M NaOAc; b) HO(O)C(CH2CH2O)nCH2CH2SH, MeOH, DCM; c) Compound V, HATU, DIEA, DMF; d) ZnBr2, DCM.

[0509] Scheme E

[0510]

[0511] a) 3-hydroxy-4-nitrobenzaldehyde, Ag2O; b) H2, Pd / C; c) protected amino acid, EEDQ; d) deprotection; e) HATU, protected amino acid; f) (p-NO2C6H4O)2CO, THF; g) R d , HOBT, DIEA; h) deprotection; i) Compound VI, FDPP, NMM, DMF; j) LiOH, THF; k) deprotection.

[0512] Scheme F:

[0513]

[0514] a) NHR, MeOH; b) protected amino acid, EEDQ, DCM; c) PNP-CO3, DIEA, DMF; d) R d , HOBT, DIEA, DMF; e) deprotection; f) compound V, FDPP, NMM, DMF; g) deprotection.

[0515] In some embodiments, the radiopharmaceutical SMDCs of the invention described herein are prepared as described in Scheme G-H.

[0516] Scheme G:

[0517]

[0518] a) HO(O)C(CH2CH2O) n NH2; b) DOTA(OtBu)3, HATU, DIEA, DMF; c) TFA; d) metal chloride, NaHCO3, ACN / H2O.

[0519] Scheme H:

[0520]

[0521] a) NH2(CH2CH2O) n NHBoc; b) TFA, DCM; c) DOTA(OtBu)3, HATU, DIEA, DMF; d) TFA; e) metal chloride, NaHCO3, ACN / H2O.

[0522] Pharmaceutical Compositions

[0523] In some embodiments, the compounds described herein are formulated into pharmaceutical compositions.

[0524] In some embodiments, the compounds described herein are administered alone or in combination with a pharmaceutically acceptable carrier, excipient, or diluent in a pharmaceutical composition.

[0525] Methods of Treatment

[0526] In some embodiments, the method comprises administering to a subject a therapeutically effective amount of a compound of formula (I) or a pharmaceutically acceptable salt thereof. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt thereof is administered in the form of a pharmaceutical composition. In some embodiments, the subject has cancer. In some embodiments, the cancer is a solid tumor or a hematological cancer. In some embodiments, the subject has a non-cancerous tumor. In some embodiments, the subject has an adenoma.

[0527] In an embodiment, treatment is sufficient to reduce or inhibit the growth of a subject's tumor, reduce the number or size of metastatic lesions, reduce tumor burden, reduce primary tumor burden, reduce invasiveness, prolong survival time, or maintain or improve quality of life, or a combination thereof.

[0528] In some embodiments, provided herein are methods for killing tumor cells, which comprise contacting the tumor cells with a compound of formula (I) or a pharmaceutically acceptable salt thereof.

[0529] In one aspect, provided herein are methods and compositions for treating cancer. Cancers include carcinogenesis of tissues and organs, including metastasis, such as gastrointestinal cancers (e.g., gastric cancer, esophageal cancer, pancreatic cancer, colorectal cancer, bowel cancer, anal cancer, liver cancer, gallbladder cancer, or colon cancer); lung cancer; thyroid cancer; skin cancer (e.g., melanoma); oral cancer; urinary tract cancer (e.g., bladder cancer or kidney cancer); blood cancer (e.g., myeloma or leukemia); or prostate cancer. In some embodiments, the present disclosure provides methods and compositions for treating a subject's gastrointestinal cancer by administering to the subject in need thereof an effective amount of a non-peptide targeted therapeutic compound disclosed herein. Non-limiting examples of gastrointestinal cancers that can be treated according to the methods of the present disclosure include gastric cancer, esophageal cancer, pancreatic cancer, lung cancer (small cell lung cancer and / or non-small cell lung cancer), colorectal cancer, bowel cancer, anal cancer, liver cancer, gallbladder cancer, or colon cancer. In some embodiments, the cancer is Hodgkin lymphoma or B-cell lymphoma.

[0530] In one aspect, provided herein are methods and compositions for treating adenoma.

[0531] In one aspect, provided herein are methods and compositions for treating cancers that express a peptide hormone G protein-coupled receptor. In some embodiments, the cancer that expresses a peptide hormone G protein-coupled receptor to be treated is a primary or metastatic cancer of gastrointestinal origin, such as colorectal cancer, gastric cancer, small intestine cancer, or esophageal cancer. In some embodiments, the cancer that expresses a peptide hormone G protein-coupled receptor to be treated is a primary or metastatic pancreatic cancer. In some embodiments, the cancer that expresses a peptide hormone G protein-coupled receptor to be treated is a primary or metastatic lung cancer, such as squamous cell carcinoma, adenosquamous carcinoma, or adenocarcinoma. In some embodiments, the cancer that expresses a peptide hormone G protein-coupled receptor to be treated is a sarcoma, such as leiomyosarcoma or rhabdomyosarcoma. In some embodiments, the cancer that expresses a peptide hormone G protein-coupled receptor to be treated is a primary or metastatic neuroectodermal tumor, such as pheochromocytoma or paraganglioma. In some embodiments, the cancer that expresses a peptide hormone G protein-coupled receptor is a primary or metastatic bronchopulmonary or gastrointestinal neuroendocrine tumor. In some embodiments, the cancer is colorectal cancer.

[0532] In another aspect, provided herein is a method for treating cancer in a mammal, which comprises administering to a mammal in need thereof a non-peptide targeting therapeutic compound disclosed herein. In some embodiments, the cancer expresses one or more peptide hormone G protein-coupled receptors. In some embodiments, the cancer comprises a peptide hormone G protein-coupled receptor-positive cancer. In some embodiments, the cancer comprises a solid tumor. In some embodiments, the cancer comprises a sarcoma, carcinoma, or lymphoma. In some embodiments, the cancer comprises a neuroendocrine tumor. In some embodiments, the cancer comprises an insulinoma. In some embodiments, the cancer comprises a peptide hormone G protein-coupled receptor-positive (e.g., somatostatin receptor-positive) gastroenteropancreatic neuroendocrine tumor (GEP-NET).

[0533] Administration methods and treatment regimens

[0534] In one embodiment, a compound of formula (I) or a pharmaceutically acceptable salt thereof is used for preparing a medicament for treating tumors in a mammal. A method for treating any disease or condition described herein in a mammal in need of such treatment involves administering to the mammal a pharmaceutical composition in a therapeutically effective amount, the pharmaceutical composition comprising at least one compound of formula (I) or a pharmaceutically acceptable salt thereof.

[0535] In some embodiments, the conjugate or a pharmaceutically acceptable salt thereof is administered in combination with another therapeutic agent. In certain embodiments, the therapeutic agent is a chemotherapeutic agent. Chemotherapeutic agents include, but are not limited to, lutetium Lu 177 dotatate

[0536] Certain Terms

[0537] Unless otherwise indicated, the following terms used in this application have the definitions given below. The use of the term "including" and other forms such as "include", "includes", and "included" is non-limiting. The section headings used herein are for organizational purposes only and should not be construed as limiting the subject matter described.

[0538] As used herein, C1-C x including C1-C2, C1-C3... C1-C x . By way of example only, a group named "C1-C6" means that there are one to six carbon atoms in the moiety, i.e., a group containing 1 carbon atom, 2 carbon atoms, 3 carbon atoms, or 4 carbon atoms. Thus, by way of example only, "C1-C4 alkyl" means that there are one to four carbon atoms in the alkyl, i.e., the alkyl is selected from methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, sec-butyl, and tert-butyl.

[0539] "Alkyl" refers to an aliphatic hydrocarbon group. The alkyl group can be branched or straight-chain. In some embodiments, "alkyl" has 1 to 10 carbon atoms, i.e., C1-C 10 alkyl. Whenever it appears herein, a numerical range such as "1 to 10" refers to each integer within the given range; for example, "1 to 10 carbon atoms" means that the alkyl group consists of 1 carbon atom, 2 carbon atoms, 3 carbon atoms, etc., up to and including 10 carbon atoms, but this definition also covers the case where the term "alkyl" appears without a specified numerical range. In some embodiments, the alkyl is a C1-C6 alkyl. In one aspect, the alkyl is methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, sec-butyl or tert-butyl. Typical alkyl groups include, but are not limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl, neopentyl or hexyl.

[0540] "Alkylene" refers to a divalent alkyl group. Any of the above monovalent alkyl groups can become an alkylene group by removing a second hydrogen atom from the alkyl group. In some embodiments, the alkylene is a C1-C6 alkylene. In other embodiments, the alkylene is a C1-C4 alkylene. Typical alkylene groups include, but are not limited to, -CH2-, -CH2CH2-, -CH2CH2CH2-, -CH2CH2CH2CH2-, etc. In some embodiments, the alkylene is -CH2-.

[0541] "Alkoxy" refers to the (alkyl)O- group, where alkyl is as defined herein.

[0542] The term "alkenyl" refers to an alkyl type in which there is at least one carbon-carbon double bond. In one embodiment, the alkenyl has the formula -C(R)=CR2, where R refers to the remainder of the alkenyl, which may be the same or different. In some embodiments, R is H or alkyl. In some embodiments, the alkenyl is selected from ethenyl (i.e., vinyl), propenyl (i.e., allyl), butenyl, pentenyl, pentadienyl, etc. Non-limiting examples of alkenyl include -CH=CH2, -C(CH3)=CH2, -CH=CHCH3, -C(CH3)=CHCH3 and -CH2CH=CH2.

[0543] The term "alkynyl" refers to an alkyl type in which there is at least one carbon-carbon triple bond. In one embodiment, the alkynyl has the formula -C≡C-R, where R refers to the remainder of the alkynyl. In some embodiments, R is H or alkyl. In some embodiments, the alkynyl is selected from ethynyl, propynyl, butynyl, pentynyl, hexynyl, etc. Non-limiting examples of alkynyl include -C≡CH, -C≡CCH3, -C≡CCH2CH3, -CH2C≡CH.

[0544] The term "heteroalkyl" refers to an alkyl group in which one or more of the backbone atoms of the alkyl group are selected from atoms other than carbon, such as oxygen, nitrogen (e.g., -NH-, -N(alkyl)-), sulfur, or combinations thereof. The heteroalkyl is attached to the remainder of the molecule at a carbon atom of the heteroalkyl. In one aspect, the heteroalkyl is a C1-C6 heteroalkyl.

[0545] The term "carbocyclic" or "carbocycle" refers to a ring or ring system in which all of the atoms forming the ring backbone are carbon atoms. Thus, this term distinguishes carbocycles from "heterocyclic" or "heterocycles" in which the ring backbone contains at least one atom other than carbon. In some embodiments, at least one of the two rings of a bicyclic carbocycle is aromatic. In some embodiments, both of the two rings of a bicyclic carbocycle are aromatic. Carbocycles include aryl and cycloalkyl.

[0546] As used herein, the term "aryl" refers to an aromatic ring in which each atom forming the ring is a carbon atom. In one aspect, the aryl is phenyl or naphthyl. In some embodiments, the aryl is phenyl. In some embodiments, the aryl is phenyl, naphthyl, indanyl, indenyl, or tetrahydronaphthyl. In some embodiments, the aryl is a C6-C 10 aryl. Depending on the structure, the aryl can be a monoradical or a biradical (i.e., arylene).

[0547] The term "cycloalkyl" refers to a monocyclic or polycyclic aliphatic non-aromatic radical in which each atom forming the ring (i.e., the backbone atom) is a carbon atom. In some embodiments, the cycloalkyl is a spiro or bridged compound. In some embodiments, the cycloalkyl is optionally fused to an aromatic ring, and the point of attachment is at a carbon of a non-aromatic ring carbon atom. Cycloalkyls include groups having 3 to 10 ring atoms. In some embodiments, the cycloalkyl is selected from cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cycloheptyl, cyclooctyl, spiro[2.2]pentyl, norbornyl, and bicyclo[1.1.1]pentyl. In some embodiments, the cycloalkyl is a C3-C6 cycloalkyl. In some embodiments, the cycloalkyl is a C3-C4 cycloalkyl.

[0548] The term "halo" or alternatively "halogen" or "halide" means fluorine, chlorine, bromine, or iodine. In some embodiments, the halo is fluorine, chlorine, or bromine.

[0549] The term "fluoroalkyl" refers to an alkyl group in which one or more hydrogen atoms are replaced by fluorine atoms. In one aspect, the fluoroalkyl is a C1-C6 fluoroalkyl.

[0550] The terms "heterocycle" or "heterocyclic" refer to heteroaromatic rings (also known as heteroaryl) and heterocycloalkyl rings containing one to four heteroatoms in the ring, where each heteroatom in the ring is selected from O, S, and N, where each heterocyclic group has 3 to 10 atoms in its ring system, and provided that no ring contains two adjacent O or S atoms. Non-aromatic heterocyclic groups (also known as heterocycloalkyl) include rings having 3 to 10 atoms in their ring systems, and aromatic heterocyclic groups include rings having 5 to 10 atoms in their ring systems. Heterocyclic groups include benzo-fused ring systems. Examples of non-aromatic heterocyclic groups are pyrrolidinyl, tetrahydrofuranyl, dihydrofuranyl, tetrahydrothienyl, oxazolidinonyl, tetrahydropyranyl, dihydropyranyl, tetrahydrothiopyranyl, piperidinyl, morpholinyl, thiomorpholinyl, thiazolidinyl, piperazinyl, aziridinyl, azetidinyl, oxetidinyl, thietanyl, homopiperidinyl, oxepanyl, thiepanyl, oxazepinyl, diazepinyl, thiazepinyl yl yl thiazepinyl, 1,2,3,6-tetrahydropyridinyl, pyrrolin-2-yl, pyrrolin-3-yl, indolinyl, 2H-pyranyl, 4H-pyranyl, dioxolanyl, 1,3-dioxolanyl, pyrazolinyl, dithiolanyl, dithiolanyl, dihydropyranyl, dihydrothienyl, dihydrofuranyl, pyrazolidinyl, imidazolinyl, imidazolidinyl, 3-azabicyclo[3.1.0]hexyl, 3-azabicyclo[4.1.0]heptyl, 3H-indolyl, indolin-2-one, isoindolin-1-one, isoindoline-1,3-dione, 3,4-dihydroisoquinolin-1(2H)-one, 3,4-dihydroquinolin-2(1H)-one, isoindoline-1,3-dithionyl, benz[d]oxazol-2(3H)-one, 1H-benz[d]imidazol-2(3H)-one, benz[d]thiazol-2(3H)-one, and quinazolinyl. Examples of aromatic heterocyclic groups are pyridinyl, imidazolyl, pyrimidinyl, pyrazolyl, triazolyl, pyrazinyl, tetrazolyl, furanyl, thienyl, isoxazolyl, thiazolyl, oxazolyl, isothiazolyl, pyrrolyl, quinolinyl, isoquinolinyl, indolyl, benzimidazolyl, benzofuranyl, cinnolinyl, indazolyl, indazinyl, phthalazinyl, pyridazinyl, triazinyl, isoindolyl, pteridinyl, purinyl, oxadiazolyl, thiadiazolyl, furazanyl, benzofurazanyl, benzothienyl, benzothiazolyl, benzoxazolyl, quinazolinyl, quinoxalinyl, naphthyridinyl, and furanopyridinyl. Where possible, the foregoing groups are C-attached (or C-linked) or N-linked. For example, groups derived from pyrrole include pyrrol-1-yl (N-linked) or pyrrol-3-yl (C-linked). In addition, groups derived from imidazole include imidazol-1-yl or imidazol-3-yl (both N-linked) or imidazol-2-yl, imidazol-4-yl, or imidazol-5-yl (all C-linked). Heterocyclic groups include benzo-fused ring systems. Non-aromatic heterocycles are optionally substituted with one or two oxo groups (=O), such as pyrrolidin-2-one. In some embodiments, at least one of the two rings of the bicyclic heterocycle is aromatic. In some embodiments, both of the two rings of the bicyclic heterocycle are aromatic.

[0551] The term "heteroaryl" or alternatively "heteroaromatic" refers to an aryl group containing one or more ring heteroatoms selected from nitrogen, oxygen, and sulfur. Illustrative examples of heteroaryl groups include monocyclic heteroaryl and bicyclic heteroaryl. Monocyclic heteroaryl groups include pyridyl, imidazolyl, pyrimidinyl, pyrazolyl, triazolyl, pyrazinyl, tetrazolyl, furyl, thienyl, isoxazolyl, thiazolyl, oxazolyl, isothiazolyl, pyrrolyl, pyridazinyl, triazinyl, oxadiazolyl, thiadiazolyl, and furazanyl. Monocyclic heteroaryl groups include indene, indole, benzofuran, benzothiophene, indazole, benzimidazole, purine, quinazoline, quinoline, isoquinoline, cinnoline, phthalazine, quinazoline, quinoxaline, 1,8-naphthyridine, and pteridine. In some embodiments, the heteroaryl group contains 0-4 N atoms in the ring. In some embodiments, the heteroaryl group contains 1-4 N atoms in the ring. In some embodiments, the heteroaryl group contains 0-4 N atoms, 0-1 O atom, and 0-1 S atom in the ring. In some embodiments, the heteroaryl group contains 1-4 N atoms, 0-1 O atom, and 0-1 S atom in the ring. In some embodiments, the heteroaryl group is a C1-C9 heteroaryl group. In some embodiments, the monocyclic heteroaryl group is a C1-C5 heteroaryl group. In some embodiments, the monocyclic heteroaryl group is a 5- or 6-membered heteroaryl group. In some embodiments, the bicyclic heteroaryl group is a C6-C9 heteroaryl group.

[0552] "Heterocycloalkyl" refers to a cycloalkyl group containing at least one heteroatom selected from nitrogen, oxygen, and sulfur. In some embodiments, the heterocycloalkyl group is fused to an aryl or heteroaryl group. In some embodiments, the heterocycloalkyl group is an oxazolidinone group, pyrrolidinyl group, tetrahydrofuryl group, tetrahydrothienyl group, tetrahydropyranyl group, tetrahydrothiopyranyl group, piperidinyl group, morpholinyl group, thiomorpholinyl group, piperazinyl group, piperidin-2-one group, pyrrolidine-2,5-diylsulfinyl group, pyrrolidine-2,5-dione group, pyrrolidinone group, imidazolidinyl group, imidazolidin-2-one group, or thiazolidin-2-one group. In one aspect, the heterocycloalkyl group is a C2-C 10 heterocycloalkyl group. In another aspect, the heterocycloalkyl group is a C4-C 10 heterocycloalkyl group. In some embodiments, the heterocycloalkyl group is monocyclic or bicyclic. In some embodiments, the heterocycloalkyl group is monocyclic and is a 3-, 4-, 5-, 6-, 7-, or 8-membered ring. In some embodiments, the heterocycloalkyl group is monocyclic and is a 3-, 4-, 5-, or 6-membered ring. In some embodiments, the heterocycloalkyl group is monocyclic and is a 3- or 4-membered ring. In some embodiments, the heterocycloalkyl group contains 0-2 N atoms in the ring. In some embodiments, the heterocycloalkyl group contains 0-2 N atoms, 0-2 O atoms, and 0-1 S atom in the ring.

[0553] The term "bond" or "single bond" refers to a chemical bond between two atoms or two moieties (when the atoms connected by the bond are considered parts of a larger substructure). In one aspect, when a group described herein is a bond, the referenced group does not exist, thus allowing a bond to form between the remaining identified groups.

[0554] The term "moiety" refers to a specific segment or functional group of a molecule. Chemical moieties are often recognized chemical entities embedded in or attached to a molecule.

[0555] The term "optionally substituted" or "substituted" means that the group is optionally substituted by one or more additional groups, which are independently selected from halogen, -CN, -NH2, -NH(alkyl), -N(alkyl)2, -OH, -CO2H, -CO2alkyl, -C(=O)NH2, -C(=O)NH(alkyl), -C(=O)N(alkyl)2, -S(=O)2NH2, -S(=O)2NH(alkyl), -S(=O)2N(alkyl)2, alkyl, cycloalkyl, fluoroalkyl, heteroalkyl, alkoxy, fluoroalkoxy, heterocycloalkyl, aryl, heteroaryl, aryloxy, alkylthio, arylthio, alkyl sulfoxide, aryl sulfoxide, alkyl sulfone and aryl sulfone. In some other embodiments, the optional substituents are independently selected from halogen, -CN, -NH2, -NH(CH3), -N(CH3)2, -OH, -CO2H, -CO2(C1-C4alkyl), -C(=O)NH2, -C(=O)NH(C1-C4alkyl), -C(=O)N(C1-C4alkyl)2, -S(=O)2NH2, -S(=O)2NH(C1-C4alkyl), -S(=O)2N(C1-C4alkyl)2, C1-C4alkyl, C3-C6cycloalkyl, C1-C4fluoroalkyl, C1-C4heteroalkyl, C1-C4alkoxy, C1-C4fluoroalkoxy, -SC1-C4alkyl, -S(=O)C1-C4alkyl and -S(=O)2C1-C4alkyl. In some embodiments, the optional substituents are independently selected from halogen -CN, -NH2, -OH, -NH(CH3), -N(CH3)2, -CH3, -CH2CH3, -CHF2, -CF3, -OCH3, -OCHF2 and -OCF3. In some embodiments, the substituted group is substituted by one or two of the foregoing groups. In some embodiments, the optional substituent on an aliphatic carbon atom (acyclic or cyclic) includes an oxo group (=O).

[0556] As used herein, the term "modulate" means to interact directly or indirectly with a target so as to change the activity of the target, including, by way of example only, enhancing the activity of the target, inhibiting the activity of the target, restricting the activity of the target or prolonging the activity of the target.

[0557] As used herein, the term "modulator" refers to a molecule that interacts directly or indirectly with a target. Interactions include, but are not limited to, agonist, partial agonist, inverse agonist, antagonist, degrader, or combinations thereof. In some embodiments, the modulator is an agonist.

[0558] As used herein, the terms "administer", "administering", "administration", etc. refer to methods that can be used to enable a compound or composition to be delivered to a desired site of biological action. These methods include, but are not limited to, oral route, duodenal route, parenteral injection (including intravenous, subcutaneous, intraperitoneal, intramuscular, intraarterial or infusion). Those skilled in the art are familiar with the administration techniques that can be employed with the compounds and methods described herein.

[0559] As used herein, the term "co-administer" and the like are intended to cover the administration of a selected therapeutic agent to a single patient and are intended to include treatment regimens in which the agents are administered by the same or different routes of administration or at the same or different times.

[0560] As used herein, the term "effective amount" or "therapeutically effective amount" refers to an amount of an agent or compound administered that will be sufficient to alleviate to some extent one or more symptoms of the disease or condition being treated. The results include reduction and / or alleviation of the signs, symptoms or causes of the disease, or any other desired change in the biological system. For example, an "effective amount" for therapeutic use is the amount of a composition comprising a compound disclosed herein required to effect a clinically significant reduction in the symptoms of the disease. The appropriate "effective" amount in any individual case is optionally determined using techniques such as dose escalation studies.

[0561] As used herein, the term "enhance" or "enhancing" means to increase or prolong the potency or duration of a desired action. Thus, with respect to enhancing the action of a therapeutic agent, the term "enhance" refers to the ability to increase or prolong the action of another therapeutic agent on a system in terms of potency or duration. As used herein, an "enhancing effective amount" is an amount sufficient to enhance the action of another therapeutic agent in a desired system.

[0562] As used herein, the term "drug combination" means a product formed by mixing or combining more than one active ingredient, and includes both fixed and non-fixed combinations of active ingredients. The term "fixed combination" means that both the active ingredient (e.g., a compound of formula (I) or a pharmaceutically acceptable salt thereof) and a co-agent are administered to a patient simultaneously in the form of a single entity or dose. The term "non-fixed combination" means that the active ingredient (e.g., a compound of formula (I) or a pharmaceutically acceptable salt thereof) and the co-agent are administered to a patient simultaneously, concurrently, or sequentially as separate entities without a specific time limit in between, where such administration provides effective levels of both compounds in the patient. The latter also applies to cocktail therapies, e.g., the administration of three or more active ingredients.

[0563] The terms "article" and "kit" are used synonymously.

[0564] The term "subject" or "patient" encompasses mammals. Examples of mammals include, but are not limited to, any member of the mammalian class: humans, non-human primates such as chimpanzees and other ape and monkey species; farm animals such as cattle, horses, sheep, goats, and pigs; domestic animals such as rabbits, dogs, and cats; laboratory animals including rodents such as rats, mice, and guinea pigs, etc. In one aspect, the mammal is a human.

[0565] As used herein, the term "treat (treat, treating, or treatment)" includes alleviating, reducing, or ameliorating at least one symptom of a disease or disorder, preventing additional symptoms, inhibiting a disease or disorder, e.g., arresting the development of a disease or disorder, alleviating a disease or disorder, causing regression of a disease or disorder, alleviating the conditions caused by a disease or disorder, or prophylactically and / or therapeutically halting the symptoms of a disease or disorder.

[0566] Examples

[0567] The following examples are provided for illustrative purposes only and do not limit the scope of the claims provided herein.

[0568] Synthesis of Compounds

[0569] Example A. 2-{7-[({17-[(2-{[4-(4-Aminopiperidin-1-yl)-3-(5-chloro-1H-1,3-benzodiazol- 2-yl))-5-(3-fluoro-5-methylphenyl)pyridin-2-yl]amino}ethyl)carbamoyl]-3,6,9,12,15-pentaoxa heptadecane-1-yl}carbamoylmethyl]-4,10-bis(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl}acetic (Compound 1)

[0570]

[0571] Step A-1, Preparation of Benzyl (1-(3-(5-chloro-1H-benzo[d]imidazol-2-yl)-2-((2,2-dimethyl-4,23-dioxo -3,8,11,14,17,20-hexaoxa-5,24-diazahexacosane-26-yl)amino)-5-(3-fluoro-5-methylphenyl) pyridin-4-yl)piperidin-4-yl)carbamate:To a mixture of 2,2-dimethyl-4-oxo-3,8,11,14,17,20-hexaoxa-5-azatricosane-23-carboxylic acid (150 mg, 1.29 eq, 366 μmol), perfluorophenyl diphenylphosphinate (150 mg, 1.38 eq, 390 μmol), and N-methylmorpholine (NMM) (70 mg, 2.4 eq, 0.69 mmol) in dimethylformamide (DMF) (2 mL) was added (1-(2-((2-aminoethyl)amino)-3-(5-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-4-yl)piperidin-4-yl)carbamic acid benzyl ester (178 mg, 1 eq, 283 μmol). The resulting reaction mixture was stirred at 25 °C for 2 h. The crude product was purified by preparative HPLC (Preparative HPLC-013) with the following conditions: column, SunFire Prep C18 OBD column, 120 g; mobile phase, water (0.1% trifluoroacetic acid (TFA)) and acetonitrile (CAN) (from 30.0% ACN to 98% in 7 min); total flow rate, 70 mL / min; detector, UV 220 nm. The collected fractions were combined and concentrated in vacuo to afford (1-(3-(5-chloro-1H-benzo[d]imidazol-2-yl)-2-((2,2-dimethyl-4,23-dioxo-3,8,11,14,17,20-hexaoxa-5,24-diazahexacosane-26-yl)amino)-5-(3-fluoro-5-methylphenyl)pyridin-4-yl)piperidin-4-yl)carbamic acid benzyl ester (120 mg, 41.5%). MS (M+H) + = 1019.5.

[0572] Step A-2, Preparation of Benzyl (1-(2-((1-amino-18-oxo-3,6,9,12,15-pentaoxa-19-azahenicosan- 21-yl)amino)-3-(5-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-4-yl)piperidin-4-yl)carbamate: Step A-3, Preparation of Tri-tert-butyl 2,2',2''-(10-(24-((4-(4-(((benzyloxy)carbonyl)amino)piperidin-1-yl)-3-A mixture of benzyl (1-(3-(5-chloro-1H-benzo[d]imidazol-2-yl)-2-((2,2-dimethyl-4,23-dioxo-3,8,11,14,17,20-hexaoxa-5,24-diazahexacos-26-yl)amino)-5-(3-fluoro-5-methylphenyl)pyridin-4-yl)piperidin-4-yl)carbamate (118 mg, 1 equiv, 116 μmol) and TFA (1 mL) in dichloromethane (DCM) (3 mL) was stirred at 25 °C for 1 h. The reaction mixture was concentrated and the crude product was adjusted to pH 8 with ammonium bicarbonate (NH4HCO3) and extracted with ethyl acetate (3 x 30 mL). The collected fractions were combined and concentrated in vacuo to afford benzyl (1-(2-((1-amino-18-oxo-3,6,9,12,15-pentaoxa-19-azahenicos-21-yl)amino)-3-(5-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-4-yl)piperidin-4-yl)carbamate (95 mg, 89%). MS (M+H) + = 919.3.

[0573] (5-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)-2,21-dioxotriyl) (5-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl]amino}ethyl)carbamoyl]-3,6,9,12,15-penta -6,9,12,15,18-pentaoxa-3,22-diazatetracosanyl)-1,4,7,10-tetraazacyclododecane-1,4,7- oxaheptadecane-1-yl}acetate: Tri-tert-butyl Triacetate:A mixture of 2-(4,7,10-tris(2-(tert-butoxy)-2-oxoethyl)-1,4,7,10-tetraazacyclododecan-1-yl)acetic acid (70 mg, 1.2 eq, 0.12 mmol), N,N-diisopropylethylamine (DIEA) (40 mg, 54 μL, 3.2 eq, 0.31 mmol), and N,N,N',N'-tetramethyl-O-(N-succinimidyl)uronium hexafluorophosphate (HSTU) (45 mg, 1.3 eq, 0.13 mmol) in DMF (1 mL) was stirred at 25 °C for 10 min, then (1-(2-((1-amino-18-oxo-3,6,9,12,15-pentaoxa-19-azahenicosan-21-yl)amino)-3-(5-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-4-yl)piperidin-4-yl)carbamic acid benzyl ester (90 mg, 1 eq, 98 μmol) was added. The resulting reaction mixture was stirred at 25 °C for 1 h. The mixture was purified by preparative HPLC under the following conditions: column, SunFire Prep C18 OBD column, 19*150 mm, 5 μm; mobile phase, water (0.1% TFA) and ACN (30% ACN to 65% in 7 min); total flow rate, 20 mL / min; detector, UV 220 nm. The collected fractions were combined and concentrated in vacuo to give tris(tert-butyl) 2,2',2''-(10-(24-((4-(4-(((benzyloxy)carbonyl)amino)piperidin-1-yl)-3-(5-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)-2,21-dioxo-6,9,12,15,18-pentaoxa-3,22-diaza-tetracosyl)-1,4,7,10-tetraazacyclododecane-1,4,7-triyl)triacetate (62 mg, 43%). MS (M / 2+H) + = 737.9.

[0574] Step A-4, preparation of 2-{7-[({17-[(2-{[4-(4-aminopiperidin-1-yl)-3-(5-chloro-1H-1,3-benzodioxol- oxaheptadecane-1-yl}acetate: yl}carbamoyl)methyl]-4,10-bis(carboxymethyl)-1,4,7,10-tetraazacyclododecane -1- {7-[({17-[(2-{[4-(4-Aminopiperidin-1-yl)-3-(5-chloro-1H-1,3-benzodiazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl]amino}ethyl)carbamoyl]-3,6,9,12,15-pentaoxaheptadecane-1-yl}carbamoylmethyl]-4,10-bis(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl}acetic A mixture of tris(tert-butyl) 2,2',2''-(10-(24-((4-(4-(((benzyloxy)carbonyl)amino)piperidin-1-yl)-3-(5-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)-2,2'-dioxido-6,9,12,15,18-pentaoxa-3,22-diazatetracosan-1-yl)-1,4,7,10-tetraazacyclododecane-1,4,7-triyl)triacetate (60 mg, 1 equiv, 41 μmol) and TFA (1 mL) was stirred at 60 °C for 2 h. The reaction mixture was concentrated and the crude product was purified by preparative HPLC (Preparative HPLC - 013) under the following conditions: column, SunFire Prep C18 OBD column, 19*150 mm, 5 μm; mobile phase, water (0.05% TFA) and ACN (30.0% ACN to 50.0% in 7 min); total flow rate, 20 mL / min; detector, UV 220 nm. The collected fractions were combined and concentrated in vacuo. The collected fractions were combined and concentrated in vacuo to afford 2-{7-[({17-[(2-{[4-(4-aminopiperidin-1-yl)-3-(5-chloro-1H-1,3-benzodiazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl]amino}ethyl)carbamoyl]-3,6,9,12,15-pentaoxaheptadec-1-yl}carbamoyl)methyl]-4,10-bis(carboxymethyl)-1,4,7,10-tetraazacyclododecan-1-yl}acetic acid (30 mg, 53%). MS (M+H) + = 1171.7.

[0575] 111 Radiochemical synthesis of [In]-Compound 1

[0576] To 111 In]InCl3 (20.0 MBq, 40.0 μL, 0.1 M HCl) and Compound 1 (2.9 nmol, 2.9 μ, 1.0 mM in DI water) were added to an NH4OAc solution (4.0 μL, 1.0 M). The resulting mixture was heated at 85 °C for 30 min in a thermal mixer. At the end of the labeling, Ca-DTPA (4.0 μL, 4 mM) was added. The radiochemical purity was 97.5% as determined by RP-HPLC. The radioactive tracer solution for in vivo studies was prepared by dilution with 0.9% saline.

[0577] Similar to Example A, the following conjugates were prepared using appropriate substitution reagents and substrates in different steps and they may require additional functional group modification using appropriate reagents via well-known chemistries.

[0578]

[0579]

[0580]

[0581]

[0582]

[0583]

[0584] Example B. N-(2-{[4-(4-Aminopiperidin-1-yl)-3-(5-chloro-1H-1,3-benzodiazol-2-yl)-5- (3-fluoro-5-methylphenyl)pyridin-2-yl]amino}ethyl)-1-(2-{3,16,19-trioxo-2,17,18-trioxa-5, 8,11,14-tetraaza-1-indanotricyclo[9.6.3.2,14]docosane-8-yl}acetamido)-3,6,9,12,15-penta oxaoctadecane-18-carboxamide (Compound 15)

[0585]

[0586] Preparation of N-(2-{[4-(4-Aminopiperidin-1-yl)-3-(5-chloro-1H-1,3-benzodiazol-2-yl)-5-(3-fluoro- 5- Methylphenyl)pyridin-2-yl]amino}ethyl)-1-(2-{3,16,19-trioxo-2,17,18-trioxa -5,8,11, 5-methylphenyl)pyridin-2-yl]amino}ethyl)-1-(2-{3,16,19-trioxo-2,17,18-trioxa-5,8,11,14-tetraaza-1-indanotricyclo[9.6.3.25,14]docosane-8-yl}acetamido)-3,6,9,12,15-pentaoxaoctadecane-18-carboxamide: ​2,2',2''-(10-(24-((4-(4-aminopiperidin-1-yl)-3-(5-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)-2,21-dioxo-6,9,12,15,18-pentaoxa-3,22-diaza-tetracosyl)-1,4,7,10-tetraazacyclododecane-1,4,7-triyl)triacetic acid (16 mg, 1 equiv, 14 μmol), indium(III) chloride (8 mg, 2 μL, 3 equiv, 0.04 mmol), and sodium bicarbonate (5 mg, 2 μL, 4 equiv, 0.06 mmol) in a mixture of ACN (0.2 mL) and aqueous solution (0.1 mL) were stirred at 80 °C for 2 h. The mixture was diluted with 4 mL of dimethyl sulfoxide (DMSO), filtered, and the filtrate was purified by preparative HPLC (HPLC-007) under the following conditions: column, SunFire Prep C18 OBD column, 19*150 mm 5um 10nm; mobile phase, water (0.05% TFA) and ACN (30% ACN to 75% in 15 min); total flow rate, 20 mL / min; detector, UV220 nm. The collected fractions were combined and concentrated in vacuo to give N-(2-{[4-(4-aminopiperidin-1-yl)-3-(5-chloro-1H-1,3-benzodiazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl]amino}ethyl)-1-(2-{3,16,19-trioxo-2,17,18-trioxa-5,8,11,14-tetraaza-1-indacenetricyclo[9.6.3.2 5,14 docos-8-yl}acetamido)-3,6,9,12,15-pentaoxaoctadecane-18-carboxamide (8.9 mg, 43%). MS (M+H) + = 1283.6.

[0587] Similar to Example B, the following conjugates were prepared using appropriate substituted reagents and substrates in different steps and they may require additional functional group modification via well-known chemistry using appropriate reagents.

[0588]

[0589]

[0590]

[0591] Example C. (2S,5S,6S,16E,18E,20R,21S)-11-chloro-21-hydroxy-12,20-dimethoxy-2,5, 9,16 - tetramethyl - 8,23 - dioxo - 4,24 - dioxo - 9,22 - diazatetracyclo[19.3.1.1 1 ,14.0 3 ,5]hexacosane alk-10,12,14(26),16,18-pentaen-6-yl (2R)-2-{1-[(2-{[4-(4-aminopiperidin-1-yl)-3-(6-chloro- 1H-benzoimidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl]amino}ethyl)carbamoyl]-N-methyl- 3,6,9-trioxa-12,13-dithiahexadecane-16-carboxamido}propionate (Compound 19)

[0592]

[0593] Step C-1, Preparation of (14S,33S,2S,4S,10E,12E,14R)-86-chloro-14-hydroxy-85,14-dimethoxy -33,2,7,10-Tetramethyl-12,6-dioxo-7-aza-1(6,4)-oxazinan-3(2,3)-epoxyethane-8(1,3)-benzene and cyclotetradecane-10,12-dien-4-yl N-methyl-N-(3-(pyridin-2-yldithio)propanoyl)-D-alaninate: A mixture of (14S,33S,2S,4S,10E,12E,14R)-86-chloro-14-hydroxy-85,14-dimethoxy-33,2,7,10-tetramethyl-12,6-dioxo-7-aza-1(6,4)-oxazinan-3(2,3)-epoxyethane-8(1,3)-benzocyclotetradecane-10,12-dien-4-yl N-(3-mercaptopropionyl)-N-methyl-D-alaninate (50 mg, 1 equiv, 68 μmol) and 1,2-bis(pyridin-2-yl) disulfane (45 mg, 3.0 equiv, 0.20 mmol) in DMF (2 mL) and acetic acid (AcOH) (0.2 mL) was stirred at 25 °C for 20 min. Then sodium acetate (0.5 mL, 0.2 M) was added to the reaction mixture. The crude product was purified by preparative HPLC (Preparative HPLC-013) under the following conditions: column, Atlantis Prep T3 OBD column, 19*150 mm 5um; mobile phase, water (0.05% TFA) and ACN (from 28% B phase to 53% in 6 min); 20 mL / min. Detector, UV 220, 254 nm. The collected fractions were combined and concentrated in vacuo to afford (14S,33S,2S,4S,10E,12E,14R)-86-chloro-14-hydroxy-85,14-dimethoxy-33,2,7,10-tetramethyl-12,6-dioxo-7-aza-1(6,4)-oxazinan-3(2,3)-epoxyethane-8(1,3)-benzocyclotetradecane-10,12-dien-4-yl N-methyl-N-(3-(pyridin-2-yldisulfanyl)propionyl)-D-alaninate (52 mg, 91%). MS (M+H) + = 847.

[0594] Step C-2, Preparation of (2R)-1-(((14S,33S,2S,4S,10E,12E,14R)-86-chloro-14-hydroxy-85,14- dimethoxy-33,2,7,10-tetramethyl-12,6-dioxo-7-aza-1(6,4)-oxazinan-3(2,3)-epoxyethane-8 (1,3)-benzenacyclotetradecane-10,12-dien-4-yl)oxy)-2,3-dimethyl-1,4-dioxo-11,14,17-trioxaHetero-7,8-dithia-3-azaicos-20-oic acid: A mixture of (14S,33S,2S,4S,10E,12E,14R)-86-chloro-14-hydroxy-85,14-dimethoxy-33,2,7,10-tetramethyl-12,6-dioxo-7-aza-1(6,4)-oxazinan-3(2,3)-epoxyethane-8(1,3)-benzocyclotetradecane-10,12-dien-4-yl N-methyl-N-(3-(pyridin-2-yldithio)propanoyl)-alaninate (25 mg, 1 equiv, 30 μmol) and 3-(2-(2-(2-mercaptoethoxy)ethoxy)ethoxy)propanoic acid (14.0 mg, 2.0 equiv, 58.7 μmol) in methanol (MeOH) (2 mL) was stirred at 25 °C for 1 h. The crude product was purified by preparative HPLC (Preparative HPLC-013) under the following conditions: column, Atlantis Prep T3 OBD column, 19*150 mm 5um; mobile phase, water (0.05% TFA) and ACN (from 28% B phase to 53% in 6 min); 20 mL / min. Detector, UV 220, 254 nm. The collected fractions were combined and concentrated in vacuo to give (2R)-1-(14S,33S,2S,4S,10E,12E,14R)-86-chloro-14-hydroxy-85,14-dimethoxy-33,2,7,10-tetramethyl-12,6-dioxo-7-aza-1(6,4)-oxazinan-3(2,3)-epoxyethane-8(1,3)-benzocyclotetradecane-10,12-dien-4-yl)oxy)-2,3-dimethyl-1,4-dioxo-11,14,17-trioxa-7,8-dithia-3-azaicos-20-oic acid (31 mg, 97.8%). MS (M+H) + = 975.

[0595] Step C-3, Preparation of (14S,33S,2S,4S,10E,12E,14R)-86-chloro-14-hydroxy-85,14-dimethoxy - 33,2,7,10-tetramethyl-12,6-dioxo-7-aza-1(6,4)-oxazinan-3(2,3)-epoxyethane-8(1,3)-benz cyclotetradecane-10,12-dien-4-yl (2R)-23-((4-(4-((tert-butoxycarbonyl)amino)piperidin-1- yl)-3-(6-chloro- 1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)-2,3-dimethyl-4,20-dioxo 11,14,17-trioxa-7,8-dithia-3,21-diazatricosanoate:A mixture of (2R)-1-(((14S,33S,2S,4S,10E,12E,14R)-86-chloro-14-hydroxy-85,14-dimethoxy-33,2,7,10-tetramethyl-12,6-dioxo-7-aza-1(6,4)-oxazinan-3(2,3)-epoxy-8(1,3)-benzenacyclotetradeca-10,12-dien-4-yl)oxy)-2,3-dimethyl-1,4-dioxo-11,14,17-trioxa-7,8-dithia-3-azadocos-20-oic acid (31 mg, 1 eq., 32 μmol) and O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HATU) (24 mg, 2.0 eq., 63 μmol) in DMF (2 mL) was stirred at 25 °C for 10 min. Then (tert-butyl (1-(2-((2-aminoethyl)amino)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-4-yl)piperidin-4-yl)carbamate (21 mg, 1.1 eq., 35 μmol) was added to the reaction mixture. The crude product was purified by preparative HPLC (Preparative HPLC-013) with the following conditions: column, Atlantis Prep T3 OBD column, 19*150 mm 5um; mobile phase, water (0.05% TFA) and ACN (from 28% B phase to 53% in 6 min); 20 mL / min. Detector, UV 220, 254 nm. The collected fractions were combined and concentrated in vacuo to give (14S,33S,2S,4S,10E,12E,14R)-86-chloro-14-hydroxy-85,14-dimethoxy-33,2,7,10-tetramethyl-12,6-dioxo-7-aza-1(6,4)-oxazinan-3(2,3)-epoxy-8(1,3)-benzenacyclotetradeca-10,12-dien-4-yl (2R)-23-((4-(4-((tert-butoxycarbonyl)amino)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)-2,3-dimethyl-4,20-dioxo-11,14,17-trioxa-7,8-dithia-3,21-diazatricosanoate (25 mg, 51%). MS (M+H) + = 1550.7.

[0596] Step C-4, Preparation of (2S,5S,6S,16E,18E,20R,21S)-11-chloro-21-hydroxy-12,20-dimethoxy -2,5,9,16-tetramethyl-8,23-dioxo-4,24-dioxa-9,22-diazatetracyclo [19.3.1.1 1 ,14.0 3 ,5] twenty hexa-10,12,14(26),16,18-pentaen-6-yl (2R)-2-{1-[(2-{[4-(4-aminopiperidin-1-yl)-3-(6-chloro- 1H-1,3-benzodiazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl]{2-[(4-{4-[(tert-Butoxycarbonyl)amino]piperidin-1-yl}-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino]ethyl}carbamoyl]-N-methyl-3,6,9-trioxa-12,13-dithiahexadecane-16-carboxamido} propionate: A mixture of (14S,33S,2S,4S,10E,12E,14R)-86-chloro-14-hydroxy-85,14-dimethoxy-33,2,7,10-tetramethyl-12,6-dioxo-7-aza-1(6,4)-oxazinan-3(2,3)-epoxyethane-8(1,3)-benzenacyclotetradecane-10,12-dien-4-yl (2R)-23-((4-(4-((tert-butoxycarbonyl)amino)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)-2,3-dimethyl-4,20-dioxo-11,14,17-trioxa-7,8-dithia-3,21-diazatricosanoate (17 mg, 1 equiv, 11 μmol) and zinc chloride (II) (12 mg, 8.0 equiv, 88 μmol) in DCM (2 mL) was stirred at 25 °C for 3 h. The crude product was purified by preparative HPLC (Preparative HPLC-013) with the following conditions: column, Atlantis Prep T3 OBD column, 19*150 mm 5um; mobile phase, water (0.05% TFA) and ACN (from 28% B phase to 53% in 6 min); 20 mL / min. Detector, UV220, 254 nm. The collected fractions were combined and concentrated in vacuo to give (2S,5S,6S,16E,18E,20R,21S)-11-chloro-21-hydroxy-12,20-dimethoxy-2,5,9,16-tetramethyl-8,23-dioxo-4,24-dioxo-9,22-diazatetracyclo[19.3.1.1 1 ,14.0 3,5 hexacos-10,12,14(26),16,18-pentaen-6-yl (2R)-2-{1-[(2-{[4-(4-aminopiperidin-1-yl)-3-(6-chloro-1H-1,3-benzodiazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl]amino}ethyl)carbamoyl]-N-methyl-3,6,9-trioxa-12,13-dithiahexadecane-16-carboxamido}propionate (2.1 mg, 11%). MS (M+H) + = 1451.5.

[0597] Similar to Example C, the following conjugates were prepared using appropriate substituted reagents and substrates in different steps and they may require additional functional group modifications using appropriate reagents via well-known chemistries.

[0598]

[0599]

[0600]

[0601]

[0602] Example D. {4-[(2S)-2-[(2S)-2-[3-(2-{2-[(2-{[4-(4-aminopiperidin-1-yl)-3-(6- chloro-1H-1,3-benzodiazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyrrolidin-2-yl]amino}ethyl)carbamoyl] ethoxy}ethoxy)propanamido]-3-methylbutanamido]-5-(carbamoylamino)pentanamido]phenyl}methyl N-[(1S)-1-{[(1S)-1-{[(3R,4S,5S)-1-[(2S)-2-[(1R,2R)-2-{[(1S,2R)-1-hydroxy-1-phenyl propan-2-yl]carbamoyl}-1-methoxy-2-methylethyl]pyrrolidin-1-yl]-3-methoxy-5-methyl-1-oxo hept-4-yl](methyl)carbamoyl}-2-methylpropyl]carbamoyl}-2-methylpropyl]-N-methylcarbamate (Compound 24)

[0603]

[0604] Step D-1, prepare 4-((S)-2-((S)-2-((tert-butoxycarbonyl)amino)-3-methylbutanamido)-5-ureidopentanamido)benzyl ((2R)-1-((1-(((3S,4R,5R)-1-((R)-2-((1S,2S)-3-(((1R,2S)-1-hydroxy yl- 1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-3-methoxy-5- methyl-1-oxohept-4-yl)(methyl)amino)-3-methyl-1-oxobutan-2-yl)amino)-3-methyl-1-oxobut -2-yl)(meth yl)carbamate:To a solution of N-((3S,4R,5R)-1-((R)-2-((1S,2S)-3-(((1R,2S)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-3-methoxy-5-methyl-1-oxohept-4-yl)-N,3-dimethyl-2-((R)-3-methyl-2-(methylamino)butanamido)butanamide (100 mg, 1 equiv, 139 μmol), ((S)-3-methyl-1-(((S)-1-((4-((((4-nitrophenoxy)carbonyl)oxy)methyl)phenyl)amino)-1-oxo-5-ureidopent-2-yl)amino)-1-oxobutan-2-yl)carbamic acid tert-butyl ester (108 mg, 1.2 equiv, 167 μmol), and HOBt (25.6 mg, 1.2 equiv, 167 μmol) in DMF (1 mL) was added DIEA (54.0 mg, 72.8 μL, 3 equiv, 418 μmol). The reaction mixture was stirred at 20 °C for 6 h. Then the reaction mixture was diluted with ethyl acetate, washed with water and brine, and concentrated to give the crude product 4-((S)-2-((S)-2-((tert-butoxycarbonyl)amino)-3-methylbutanamido)-5-ureidopentanamido)benzyl ((2R)-1-((1-(((3S,4R,5R)-1-((R)-2-((1S,2S)-3-(((1R,2S)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-3-methoxy-5-methyl-1-oxohept-4-yl)(methyl)amino)-3-methyl-1-oxobutan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)(methyl)carbamate. MS(M+H) + = 1223.9. This material was used in the next step without further purification.

[0605] Step D-2, prepare 4-((S)-2-((S)-2-amino-3-methylbutanamido)-5-ureidopentanamido)benzyl ((2R)-1-((1-(((3S,4R,5R)-1-((R)-2-((1S,2S)-3-(((1R,2S)-1-hydroxy-1-phenylpropan-2-yl) amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-3-methoxy-5-methyl-1-oxohept-4-yl) (methyl)amino)-3-methyl-1-oxobutan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)(methyl)aminoFormate: 2,2,2-Trifluoroacetic acid (15.8 mg, 1 equiv, 139 μmol) was added to a solution of 4-((S)-2-((S)-2-((tert-butoxycarbonyl)amino)-3-methylbutanamido)-5-ureidopentanamido)benzyl ((2R)-1-((1-(((3S,4R,5R)-1-((R)-2-((1S,2S)-3-(((1R,2S)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-3-methoxy-5-methyl-1-oxohept-4-yl)(methyl)amino)-3-methyl-1-oxobutan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)(methyl)carbamate (170 mg, 1 equiv, 139 μmol) in DCM (0.6 mL). The resulting mixture was stirred at 0 °C for 30 min. The crude reaction mixture was concentrated and then purified by C18 reverse-phase chromatography, eluting with MeCN (0.1% TFA) / water (0.1% TFA). The collected fractions were combined and concentrated in vacuo to afford 4-((S)-2-((S)-2-amino-3-methylbutanamido)-5-ureidopentanamido)benzyl ((2R)-1-((1-(((3S,4R,5R)-1-((R)-2-((1S,2S)-3-(((1R,2S)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-3-methoxy-5-methyl-1-oxohept-4-yl)(methyl)amino)-3-methyl-1-oxobutan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)(methyl)carbamate (71.8 mg, 46.0%). MS (M+H) + = 1124.1.

[0606] Step D-3, Preparation of (6S,9S)-1-amino-6-((4-((5R,11R,12S)-11-((R)-sec-but yl)-12-(2- ((R)-2-((1S,2S)-3-(((1R,2S)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2- yl-3-oxopropyl)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-tri oxo-2,13-dioxo a-4,7,10-triazatetradecyl)phenyl)carbamoyl)-9-isopropyl-1,8,11-trioxo-14,17-dioxo a-2,7,10-triazacosan-20-yl tert-butyl ester:To a solution of 3-(2-(3-(tert-butoxy)-3-oxopropoxy)ethoxy)propanoic acid (6.0 mg, 1 equiv, 23 μmol) and 2-(3H-[1,2,3]triazolo[4,5-b]pyridin-3-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate (V) (13 mg, 1.5 equiv, 34 μmol) in DMF (0.5 mL) was added N-ethyl-N-isopropylpropan-2-amine (12 mg, 16 μL, 4 equiv, 91 μmol) and 4-((S)-2-((S)-2-amino-3-methylbutanamido)-5-ureidopentanamido)benzyl ((2R)-1-((1-(((3S,4R,5R)-1-((R)-sec-butyl)-12-(2-((R)-2-((1S,2S)-3-(((1R,2S)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenyl)carbamoyl)-9-isopropyl-1,8,11-trioxo-14,17-dioxa-2,7,10-triazacosan-20-oic acid tert-butyl ester (1 / 1) (28 mg, 1 equiv, 23 μmol). The reaction mixture was stirred at 20 °C for 2 h. The reaction mixture was diluted with ethyl acetate, washed with water and brine and concentrated to afford the crude product (6S,9S)-1-amino-6-((4-((5R,11R,12S)-11-((R)-sec-butyl)-12-(2-((R)-2-((1S,2S)-3-(((1R,2S)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenyl)carbamoyl)-9-isopropyl-1,8,11-trioxo-14,17-dioxa-2,7,10-triazacosan-20-oic acid tert-butyl ester. MS (M+H) + = 1369.4. This material was used in the next step without further purification.

[0607] Step D-4, prepare (6S,9S)-1-amino-6-((4-((5R,11R,12S)-11-((R)-sec-but yl)-12-(2-((R)-2-((1S,2S)-3-(((1R,2S)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2- ylmethyl-3-oxo propyl)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo a-4,7,10-triazatetradecyl)phenyl)carbamoyl)-9-isopropyl-1,8,11-Trioxo-14,17-dioxa-2,7,10-triazacos-20-oic acid: TFA (0.7 g, 0.5 mL, 3e+2 eq, 6 mmol) was added to a solution of tert-butyl (6S,9S)-1-amino-6-((4-((5R,11R,12S)-11-((R)-sec-butyl)-12-(2-((R)-2-((1S,2S)-3-(((1R,2S)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenyl)carbamoyl)-9-isopropyl-1,8,11-trioxo-14,17-dioxa-2,7,10-triazacos-20-oate (31 mg, 1 eq, 23 μmol) in DCM (0.6 mL). The resulting mixture was stirred at 20 °C for 30 min. The crude reaction was concentrated and purified by C18 reverse-phase chromatography, eluting with ACN (0.1% TFA) / water (0.1% TFA). The collected fractions were combined and concentrated in vacuo to afford (6S,9S)-1-amino-6-((4-((5R,11R,12S)-11-((R)-sec-butyl)-12-(2-((R)-2-((1S,2S)-3-(((1R,2S)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenyl)carbamoyl)-9-isopropyl-1,8,11-trioxo-14,17-dioxa-2,7,10-triazacos-20-oic acid (13.6 mg, 46%). MS (M+H) + = 1312.3.

[0608] Step D-5, prepare 4-((2S,5S)-19-((4-(4-((tert-butoxycarbonyl)amino)piperidin-1-yl)-3-(6- Chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)-5-isopropyl-4,7,16- tri oxo-2-(3-ureidopropyl)-10,13-dioxo-3,6,17-triazanonadecanamido)benzyl ((2R)-1-((1-(((3S,4R,5R)-1-((R)-2-((1S,2S)-3-(((1R,2S)-1-hydroxy-1-phenylpropan-2-yl)am yl)-1-methoxy yl-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-3-methoxy-5-methyl-1-oxohept-4- ino)(methyl)amino)-3-methyl-1-oxobutan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)(methyl)amino yl formate:To a solution of (6S,9S)-1-amino-6-((4-((5R,11R,12S)-11-((R)-sec-butyl)-12-(2-((R)-2-((1S,2S)-3-(((1R,2S)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)phenyl)carbamoyl)-9-isopropyl-1,8,11-trioxo-14,17-dioxo-2,7,10-triazacos-20-oic acid (13.6 mg, 1 eq, 9.94 μmol) and 2-(3H-[1,2,3]triazolo[4,5-b]pyridin-3-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate (V) (5.67 mg, 1.5 eq, 14.9 μmol) in DMF (0.2 mL) was added N-ethyl-N-isopropylpropan-2-amine (5.14 mg, 6.93 μL, 4 eq, 39.8 μmol), and then (1-(2-((2-aminoethyl)amino)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-4-yl)piperidin-4-yl)carbamic acid tert-butyl ester (6.50 mg, 1.1 eq, 10.9 μmol). The reaction mixture was stirred at 20 °C for 2 h. The reaction mixture was diluted with ethyl acetate, washed with water and brine and concentrated to give 4-((2S,5S)-19-((4-(4-((tert-butoxycarbonyl)amino)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)-5-isopropyl-4,7,16-trioxo-2-(3-ureidopropyl)-10,13-dioxo-3,6,17-triazanonadecanamido)benzyl ((2R)-1-((1-(((3S,4R,5R)-1-((R)-2-((1S,2S)-3-(((1R,2S)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-3-methoxy-5-methyl-1-oxohept-4-yl)(methyl)amino)-3-methyl-1-oxobutan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)(methyl)carbamate. MS(M+H) + = 1888.0. This material was used in the next step without further purification.

[0609] Step D-6, prepare {4-[(2S)-2-[(2S)-2-[3-(2-{2-[(2-{[4-(4-aminopiperidin-1-yl)-3- (6-Chloro-1H-1,3-benzodiazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl]amino}ethyl)carbamoyl yl]ethoxy}ethoxy)propanamido]-3-methylbutanamido]-5-(carbamoylamino)pentanamido]phenyl}methyl N-[(1S)-1-{[(1S)-1-{[(3R,4S,5S)-1-[(2S)-2-[(1R,2R)-2-{[(1S,2R)-1-hydroxy yl-1-benz ylpropan-2-yl]carbamoyl}-1-methoxy-2-methylethyl]pyrrolidin-1-yl]-3-methoxy-5- methyl-1-oxohept-4-yl](methyl)carbamoyl}-2-methylpropyl]carbamoyl}-2-methylprop yl]-N-methylcarbamic ester: To a solution of 4-((2S,5S)-19-((4-(4-((tert-butoxycarbonyl)amino)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)-5-isopropyl-4,7,16-trioxo-2-(3-ureidopropyl)-10,13-dioxa-3,6,17-triazanonadecanamido)benzyl ((2R)-1-((1-(((3S,4R,5R)-1-((R)-2-((1S,2S)-3-(((1R,2S)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-3-methoxy-5-methyl-1-oxohept-4-yl)(methyl)amino)-3-methyl-1-oxobutan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)(methyl)carbamate (18.8 mg, 1 equiv, 9.96 μmol) in DCM (0.6 mL) was added TFA (0.7 g, 0.5 mL, 7e+2 equiv, 6 mmol). The resulting mixture was stirred at 0 °C for 30 min. The crude reaction was concentrated and then purified by C18 reverse-phase chromatography, eluting with ACN (0.1% TFA) / water (0.1% TFA). The collected fractions were combined and concentrated in vacuo to afford {4-[(2S)-2-[(2S)-2-[3-(2-{2-[(2-{[4-(4-aminopiperidin-1-yl)-3-(6-chloro-1H-1,3-benzodiazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl]amino}ethyl)carbamoyl]ethoxy}ethoxy)propanamido]-3-methylbutanamido]-5-(carbamoylamino)pentanamido]phenyl}methyl N-[(1S)-1-{[(1S)-1-{[(3R,4S,5S)-1-[(2S)-2-[(1R,2R)-2-{[(1S,2R)-1-hydroxy-1-phenylpropan-2-yl]carbamoyl}-1-methoxy-2-methylethyl]pyrrolidin-1-yl]-3-methoxy-5-methyl-1-oxohept-4-yl](methyl)carbamoyl}-2-methylpropyl]carbamoyl}-2-methylpropyl]-N-methylcarbamate (10.5 mg, 59.0%). MS (M+H) + = 1788.3.

[0610] Similar to Example D, the following conjugates were prepared using appropriate substitution reagents and substrates in different steps, and they may require additional functional group modifications using appropriate reagents via well-known chemistry.

[0611]

[0612]

[0613]

[0614]

[0615]

[0616]

[0617] Example E. (4S)-4-(3-{2-[(2-{[4-(4-aminopiperidin-1-yl)-3-(6-chloro-1H-1,3-benzod iazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl]amino}ethyl)carbamoyl]ethoxy}propanamido)- 4-{[(1S)-1-{[(1S)-1-({4-[({[(1S)-1-{[(1S)-1-{[(3R,4S,5S)-1-[(2S)-2-[(1R,2R)- 2-{[(1S,2R)-1-hydroxy-1-phenylpropan-2-yl]carbamoyl}-1-methoxy-2-methylethyl]pyrrolidin-1- yl]-3-methoxy-5-methyl-1-oxohept-4-yl](methyl)carbamoyl}-2-methylpropyl]carbamoyl}-2- methylpropyl](methyl)carbamoyloxy)methyl]phenyl}carbamoyl)ethyl]carbamoyl}-2-methylprop yl]carbamoyl}butyric acid (Compound 32)

[0618]

[0619] Step E-1, prepare 4-((S)-2-((S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-methylbut (amide group) propanamido) benzyl ((S)-1-(((S)-1-(((3R,4S,5S)-1-((S)-2-((1R,2R)-3-(((1S, 2R)-1-hydroxy-1-phenylpropan-2-yl) (amino)-1-methoxy-2-methyl-3-oxopropyl) pyrrolidin-1-yl)-3-methoxy-5-methyl-1-oxohept-4- yl)(methyl)amino)-3-methyl-1-oxobutan-2-yl)amino)-3-methyl-1-oxo butan-2-yl)(methyl)aminoFormate: To a mixture of (S)-N-((3R,4S,5S)-1-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-3-methoxy-5-methyl-1-oxoheptan-4-yl)-N,3-dimethyl-2-((S)-3-methyl-2-(methylamino)butanamido)butanamide (150 mg, 1 equiv, 209 μmol) in DMF (1.5 mL) was added (9H-fluoren-9-yl)methyl ((S)-3-methyl-1-(((S)-1-((4-((((4-nitrophenoxy)carbonyl)oxy)methyl)phenyl)amino)-1-oxopropan-2-yl)amino)-1-oxobutan-2-yl)carbamate (160 mg, 1.13 equiv, 235 μmol), 1H-benzo[d][1,2,3]triazol-1-ol hydrate (50 mg, 1.6 equiv, 0.33 mmol), and N-ethyl-N-isopropylpropan-2-amine (100 mg, 3.70 equiv, 774 μmol). The reaction mixture was stirred at 20 °C for 5 h. The mixture was purified directly by MPLC under the following conditions: column, C18 120 g, Spherical 20-40 μm; mobile phase, water (0.05% FA) and ACN (5% ACN to 5% ACN in 1 min, 30% ACN to 90% in 10 min, 95% ACN to 95% in 2 min); total flow rate, 70 mL / min; detector, UV 220 nm. The collected fractions were combined and concentrated in vacuo to afford 4-((S)-2-((S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-methylbutanamido)propanamido)benzyl ((S)-1-(((S)-1-(((3R,4S,5S)-1-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-3-methoxy-5-methyl-1-oxoheptan-4-yl)(methyl)amino)-3-methyl-1-oxobutan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)(methyl)carbamate (130 mg, 49.4%). MS (M+H) + = 1259.8.

[0620] Step E-2, prepare 4-((S)-2-((S)-2-amino-3-methylbutanamido)propanamido)-benzyl ((S)-1-(((S)-1-(((3R,4S,5S)-1-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl) amine yl)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-3-methoxy-5-methyl-1-oxohept-4-((methyl)amino)-3-methyl-1-oxobutan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)(methyl)amino yl formate: To a mixture of 4-((S)-2-((S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)-amino)-3-methylbutanamido)propanamido)benzyl ((S)-1-(((S)-1-(((3R,4S,5S)-1-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-3-methoxy-5-methyl-1-oxohept-4-yl)(methyl)amino)-3-methyl-1-oxobutan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)(methyl)carbamate (125 mg, 1 equiv, 99.2 μmol) in ACN (1.5 mL) was added 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) (50 mg, 50 μL, 3.3 equiv, 0.33 mmol). The reaction mixture was stirred at 20 °C for 1 h. The mixture was purified directly by MPLC under the following conditions: column, WelFlashTM, C18 120 g, Spherical 20 - 40 μm; mobile phase, water (0.05% FA) and ACN (5% ACN to 5% ACN in 1 min, 30% ACN to 90% in 10 min, 95% ACN to 95% in 2 min); total flow rate, 70 mL / min; detector, UV 220 nm. The collected fractions were combined and concentrated in vacuo to afford 4-((S)-2-((S)-2-amino-3-methylbutanamido)-propanamido)benzyl ((S)-1-(((S)-1-(((3R,4S,5S)-1-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-3-methoxy-5-methyl-1-oxohept-4-yl)(methyl)amino)-3-methyl-1-oxobutan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)(methyl)carbamate (110 mg, 91%, 85% purity). MS (M+H) + = 1037.8.

[0621] Step E-3, prepare (S)-4-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-5-(((S)-1-(((S)-1- ((4-((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy yl-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-2-oxoethyl)-5, (8 - Diisopropyl - 4,10 - dimethyl - 3,6,9 - trioxo - 2,13 - dioxo - 4,7,10 - triazatetradecyl)phenyl)amine )(1 - Oxopropan - 2 - yl)amino - 3 - methyl - 1 - oxobutan - 2 - yl)amino - 5 - oxopentanoic acid tert - butyl ester:To a mixture of (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-5-(tert-butoxy)-5-oxopentanoic acid (35 mg, 1.1 eq, 82 μmol) in DMF (1 mL) was added HATU (33 mg, 1.1 eq, 87 μmol) and DIEA (30 mg, 40 μL, 3.0 eq, 0.23 mmol). The reaction mixture was stirred at 20 °C for 10 min, then 4-((S)-2-((S)-2-amino-3-methylbutanamido)propanamido)benzyl ((S)-1-(((S)-1-(((3R,4S,5S)-1-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-3-methoxy-5-methyl-1-oxohept-4-yl)(methyl)amino)-3-methyl-1-oxobutan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)(methyl)carbamate (80 mg, 1 eq, 77 μmol) was added, and the reaction mixture was stirred at 20 °C for an additional 1.5 h. The mixture was purified directly by MPLC under the following conditions: column, C18 120 g, Spherical 20 - 40 μm; mobile phase, water (0.05% TFA) and ACN (5% ACN to 5% ACN in 1 min, 30% ACN to 78% in 9 min, 90% ACN to 90% in 3 min); total flow rate, 70 mL / min; detector, UV 220 nm. The collected fractions were combined and concentrated in vacuo to afford tert-butyl (S)-4-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-5-(((S)-1-(((S)-1-((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)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)phenyl)amino)-1-oxopropan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)amino)-5-oxopentanoate (60 mg, 54%). MS (M+H) + = 1445.0.

[0622] (S) - 4 - Amino - 5 - (((S) - 1 - (((S) - 1 - ((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)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9- trioxo - 2,13 - dioxo - 4,7,10 - triazatetradecyl)phenyl)amino) - 1 - oxopropan - 2 - yl)amino) - (3 - methyl - 1 - oxobutan - 2 - yl)amino) - 5 - oxopentanoic acid tert - butyl ester: To a mixture of (S)-4-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-5-(((S)-1-(((S)-1-((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)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenyl)amino)-1-oxopropan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)amino)-5-oxopentanoic acid tert-butyl ester (55 mg, 1 equiv, 38 μmol) in ACN (1 mL) was added DBU (20 mg, 20 μL, 3.5 equiv, 0.13 mmol). The reaction mixture was stirred at 20 °C for 1 h. The mixture was purified directly by MPLC under the following conditions: column, WelFlashTM, C18 120 g, Spherical 20 - 40 μm; mobile phase, water (0.05% TFA) and ACN (5% ACN to 5% ACN in 1 min, 30% ACN to 80% in 9 min, 90% ACN to 90% in 3 min); total flow rate, 70 mL / min; detector, UV 220 nm. The collected fractions were dried by lyophilization. The collected fractions were combined and concentrated in vacuo to give (S)-4-amino-5-(((S)-1-(((S)-1-((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)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenyl)amino)-1-oxopropan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)amino)-5-oxopentanoic acid tert-butyl ester (40 mg, 86%). MS (M+H) + = 1223.2.

[0623] (S) - 4 - (3 - (3 - ((2 - ((4 - ((tert - butoxycarbonyl)amino)piperidin - 1 - yl) - 3 - (6 - chloro - 1H Benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)ethyl)amino)-3-oxopropoxyyl) propanoic acid: A mixture of 3,3'-oxydipropionic acid (150 mg, 1.10 eq, 925 μmol), HATU (384 mg, 1.20 eq, 1.01 mmol), and diisopropylethylamine (327 mg, 437 μL, 3.01 eq, 2.53 mmol) in DMF (5 mL) was stirred at 25 °C for 10 min, and then (1-(2-((2-aminoethyl)amino)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-4-yl)piperidin-4-yl)carbamic acid tert-butyl ester (500 mg, 1 eq, 842 μmol) was added. The resulting reaction mixture was stirred at 25 °C for 2 h. Then the reaction was quenched with water (200 mL). The resulting solution was extracted with ethyl acetate (3 x 200 mL). The organic layer was dried over anhydrous sodium sulfate and concentrated in vacuo. The residue was applied to a silica gel column with ethyl acetate / petroleum ether (1:1). The collected fractions were combined and concentrated in vacuo to afford 3-(3-((2-((4-((tert-butoxycarbonyl)amino)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)ethyl)amino)-3-oxopropoxy)propionic acid (290 mg, 46.7%). MS (M+H) + = 738.5

[0624] (S) - 4 - (3 - (3 - ((2 - ((4 - ((tert - butoxycarbonyl)amino)piperidin - 1 - yl) - 3 - (6- Chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)ethyl)amino)-3-oxo propanoylamino) - 5 - (((S) - 1 - (((S) - 1 - ((4 - ((5S,8S,11S,12R) - 11 - ((S) - sec - but yl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2- yl) - 12 - (2 - ((S) - 2 - ((1R,2R) - 3 - (((1S,2R) - 1 - hydroxy - 1 - phenylpropan - 2 - yl)amino) - 1 - methoxy - 2 - methyl - 1 - oxopropyl)pyrrolidin - 1 - yl) - 2 - oxoethyl) - 5,8 - diisopropyl - 4,10 - dimethyl - 3,6,9 - tri oxo-2,13-dioxa-4,7,10-triazatetradecyl)phenyl)amino)-1-oxopropan-2-yl)amino)-3-methyl oxo - 2,13 - dioxo - 4,7,10 - triazatetradecyl)phenyl)amino) - 1 - oxopropan - 2 - yl)amino) - (3 - methyl - 1 - oxobutan - 2 - yl)amino) - 5 - oxopentanoic acid tert - butyl ester:To a mixture of 3-(3-((2-((4-(4-((tert-Butoxycarbonyl)amino)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)ethyl)amino)-3-oxopropoxy)propanoic acid (25 mg, 1.0 equiv, 34 μmol) in DMF (1 mL) was added 4-methylmorpholine (12 mg, 3.6 equiv, 0.12 mmol) and perfluorophenyl diphenylphosphinate (16 mg, 1.3 equiv, 42 μmol). The reaction mixture was stirred at 20 °C for 10 min, then (S)-4-amino-5-(((S)-1-(((S)-1-((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)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)phenyl)amino)-1-oxopropan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)amino)-5-oxopentanoic acid tert-butyl ester (40 mg, 1 equiv, 33 μmol) was added, and the reaction mixture was stirred at 20 °C for another 2 h. The mixture was diluted with 20 mL of water, extracted with ethyl acetate (20 mL x 3), and the combined organic layers were washed with water (10 mL x 2) and brine (20 mL), dried over anhydrous Na2SO4 and concentrated under reduced pressure to give (S)-4-(3-(3-((2-((4-(4-((tert-Butoxycarbonyl)amino)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)ethyl)amino)-3-oxopropoxy)propanamido)-5-(((S)-1-(((S)-1-((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)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)phenyl)amino)-1-oxopropan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)amino)-5-oxopentanoic acid tert-butyl ester (50 mg, 79%). MS (M+H) + = 1942.2. This material was used in the next step without further purification.

[0625] (4S) - 4 - (3 - {2 - [(2 - {[4 - (4 - aminopiperidin - 1 - yl) - 3 - (6 - chloro - 1H - 1,3 - benz oxazol - 2 - yl) - 5 - (3 - fluoro - 5 - methylphenyl)pyridin - 2 - yl]amino}ethyl)carbamoyl {(1S)-1-{[(1S)-1-({4-[({[(1S)-1-{[(1S)-1-{[(3R,4S,5S)-1-[(2S)-2-[(1R,2R)-2 yl} - 1 - methoxy - 2 - methyl - 1 - oxoethyl]pyrrolidin - 1- yl]-3-methoxy-5-methyl-1-oxohept-4-yl](methyl)carbamoyl}-2-methylpropyl]carbamoyl yl} - 2 - methylpropyl](methyl)carbamoyloxy)methyl]phenyl}carbamoylethyl)aminoform{[(4S)-4-(3-{2-[(2-{[4-(4-aminopiperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl]amino}ethyl)carbamoyl]ethoxy}propanamido)-4-{[(1S)-1-{[(1S)-1-({4-[({[(1S)-1-{[(1S)-1-{[(3R,4S,5S)-1-[(2S)-2-[(1R,2R)-2-{[(1S,2R)-1-hydroxy-1-phenylpropan-2-yl]carbamoyl}-1-methoxy-2-methylethyl]pyrrolidin-1-yl]-3-methoxy-5-methyl-1-oxohept-4-yl](methyl)carbamoyl}-2-methylpropyl]carbamoyl}-2-methylpropyl](methyl)carbamoyl}oxy)methyl]phenyl}carbamoyl)ethyl]carbamoyl}-2-methylpropyl]carbamoyl}butyric acid: To a mixture of tert-butyl (S)-4-(3-(3-((2-((4-(4-((tert-butoxycarbonyl)amino)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)ethyl)amino)-3-oxopropoxy)propanamido)-5-(((S)-1-(((S)-1-((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)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)phenyl)amino)-1-oxopropan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)amino)-5-oxopentanoate (45 mg, 1 equiv, 23 μmol) in DCM (0.2 mL) was added TFA (0.2 mL), and the reaction mixture was stirred for 30 min. The pH of the reaction mixture was adjusted to 7.0 with DIEA at 0 °C and dried under nitrogen. The crude product was purified by preparative HPLC with the following conditions: column: SunFire prep OBD 19*150 mm 5um; mobile phase A: water (0.05% TFA); mobile phase B: ACN; gradient: 20% B to 53% B in 15 min; flow rate: 20 mL / min; wavelength: 220 nm. The collected fractions were dried by lyophilization to afford (4S)-4-(3-{2-[(2-{[4-(4-aminopiperidin-1-yl)-3-(6-chloro-1H-1,3-benzodiazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl]amino}ethyl)carbamoyl]ethoxy}propanamido)-4-{[(1S)-1-{[(1S)-1-({4-[({[(1S)-1-{[(1S)-1-{[(3R,4S,5S)-1-[(2S)-2-[(1R,2R)-2-{[(1S,2R)-1-hydroxy-1-phenylpropan-2-yl]carbamoyl}-1-methoxy-2-methylethyl]pyrrolidin-1-yl]-3-methoxy-5-methyl-1-oxohept-4-yl](methyl)carbamoyl}-2-methylpropyl]carbamoyl}-2-methylpropyl](methyl)carbamoyl}oxy)methyl]phenyl}carbamoyl)ethyl]carbamoyl}-2-methylpropyl]carbamoyl}butyric acid (8.4 mg, 17%). MS (M+H) + = 1790.

[0626] Similar to Example E, the following conjugates were prepared using appropriate substitution reagents and substrates in different steps, and they may require additional functional group modifications using appropriate reagents via well-known chemistry.

[0627]

[0628]

[0629]

[0630]

[0631]

[0632]

[0633]

[0634]

[0635]

[0636]

[0637]

[0638]

[0639]

[0640]

[0641]

[0642]

[0643]

[0644]

[0645]

[0646]

[0647]

[0648]

[0649]

[0650]

[0651]

[0652]

[0653]

[0654]

[0655]

[0656]

[0657]

[0658]

[0659]

[0660]

[0661]

[0662]

[0663]

[0664]

[0665]

[0666]

[0667]

[0668] Example F. {4 - [(2S) - 2 - [(2S) - 2 - [(2S) - 2 - (3 - {2 - [(2 - {[4 - (4 - aminopiperidin - 1 - yl) - 3 - (6 - chloro - 1H - 1,3 - benzoxazol - 2 - yl) - 5 - (3 - fluoro - 5 - methylphenyl)pyridin - 2 - yl]amino}ethyl)carbamoyl yl]ethoxy}propanamido) - 4 - carbamoylbutanamido] - 3 - methylbutanamido] - 5 - (carbamoylamino) pentanamido]phenyl}methyl N - [(1S) - 1 - {[(1S) - 1 - {[(3R,4S,5S) - 1 - [(2S) - 2 - [(1R,2R) - 2 - {[(1S, 2R) - 1 - hydroxy - 1 - phenylpropan - 2 - yl]aminoformyl} - 1 - methoxy - 2 - methyl - 1 - oxoethyl]pyrrolidin - 1 - yl] - 3 - methoxy {[(5-methyl-1-oxoheptan-4-yl)(methyl)carbamoyl]-2-methylpropyl}carbamoyl]-2-methylpropyl]- N-methylcarbamate (Compound 56)

[0669]

[0670] Step F-1, (4S)-4-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-5-(((2S)-1-(((2S)-1-((4-((5S,11S,14S,17S,18R)-17-((S)-sec-butyl yl)-18-(2-((S)-2-((1R,2R)-3-(((1S,2R)- 1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-(Methyl-3-oxopropyl)pyrrolidin-1-yl)-2-oxoethyl)-5,8,11,14-tetraisopropyl-4,10,16-trimethyl -3,6,9,12,15-pentaoxo-2,19-dioxa-4,7,10, 13,16-pentaazaeicosanyl)phenyl)amino)-1-oxotert-Butyl (4S)-4-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-5-(((2S)-1-(((2S)-1-((4-((5S,11S,14S,17S,18R)-17-((S)-sec-butyl)-18-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-2-oxoethyl)-5,8,11,14-tetraisopropyl-4,10,16-trimethyl-3,6,9,12,15-pentaoxo-2,19-dioxo-4,7,10,13,16-pentaazaeicosanyl)phenyl)amino)-1-oxo-5-ureidopentan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)amino)-5-oxopentanoate: A mixture of (S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-5-(tert-butoxy)-5-oxopentanoic acid (40 mg, 1.6 eq, 94 μmol), HATU (40 mg, 1.8 eq, 0.11 mmol), and DIEA (40 mg, 54 μL, 5.2 eq, 0.31 mmol) in DMF (1 mL) was stirred at 20 °C for 10 min, then 4-((S)-2-((S)-2-amino-3-methylbutanamido)-5-ureidopentanamido)benzyl ((3R,4S,7S,10S,16S)-4-((S)-sec-butyl)-3-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-2-oxoethyl)-7,10,13-triisopropyl-5,11,17-trimethyl-6,9,12,15-tetraoxo-2-oxa-5,8,11,14-tetraazaoctadec-16-yl)(methyl)carbamate (80 mg, 1 eq, 60 μmol) was added, and the reaction mixture was stirred at 20 °C for 1 h. The mixture was purified directly by MPLC with the following conditions: column, WelFlashTM, C18 120 g, Spherical 20 - 40 μm; mobile phase, water (0.05% FA) and ACN (5% ACN to 5% ACN in 1 min, 30% ACN to 80% in 7 min, 95% ACN to 95% in 3 min); total flow rate, 70 mL / min; detector, UV 220 nm. The collected fractions were combined and concentrated in vacuo to give tert-Butyl (4S)-4-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-5-(((2S)-1-(((2S)-1-((4-((5S,11S,14S,17S,18R)-17-((S)-sec-butyl)-18-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-2-oxoethyl)-5,8,11,14-tetraisopropyl-4,10,16-trimethyl-3,6,9,12,15-pentaoxo-2,19-dioxo-4,7,10,13,16-pentaazaeicosanyl)phenyl)amino)-1-oxo-5-ureidopentan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)amino)-5-oxopentanoate (90 mg, 86%). MS (M+H) + = 1531.6.

[0671] Step F-2, prepare (4S)-4-amino -5-(((2S)-1-(((2S)-1-((4-((5S,11S,14S,17S, 18R)-17-((S)-sec-butyl -18-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2- methyl-3-oxopropyl)pyrrolidin-1-yl)-2-oxoethyl)-5,8,11,14-tetraisopropyl- 4,10,16-trimethyl -3,6,9,12,15-pentaoxo-2,19-dioxa-4,7,10,13,16-pentaazaeicosyl)phenyl)amino)-1-oxo -5-ureidopentan-2-yl)amino)-3-methyl-1-oxobutan-2-yl)amino)-5-oxopentanoic acid tert-butyl ester:(4S)-4-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)-5-(((2S)-1-(((2S)-1-((4-((5S,11S,14S,17S,18R)-17-((S)-sec-Butyl)-18-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-Hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-2-oxoethyl)-5,8,11,14-tetraisopropyl-4,10,16-trimethyl-3,6,9,12,15-pentaoxo-2,19-dioxa-4,7,10,13,16-pentaazaeicosyl)phenyl)amino)-1-oxo-5-ureidopent-2-yl)amino)-3-methyl-1-oxobutan-2-yl)amino)-5-oxopentanoic acid tert-butyl ester (85 mg, 1 eq, 49 μmol) and 2,3,4,6,7,8,9,10-octahydropyrimido[1,2-a]azepine (25 mg, 3.4 eq, 0.16 mmol) in DMF (1 mL) were stirred at 20 °C for 1 h. The mixture was purified directly by MPLC with the following conditions: column, WelFlashTM, C18 120 g, Spherical 20 - 40 μm; mobile phase, water (0.05% FA) and ACN (5% ACN to 5% ACN in 1 min, 20% ACN to 70% in 7 min, 95% ACN to 95% in 3 min); total flow rate, 70 mL / min; detector, UV 220 nm. The collected fractions were combined and concentrated in vacuo to give (4S)-4-amino-5-(((2S)-1-(((2S)-1-((4-((5S,11S,14S,17S,18R)-17-((S)-sec-Butyl)-18-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-Hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-2-oxoethyl)-5,8,11,14-tetraisopropyl-4,10,16-trimethyl-3,6,9,12,15-pentaoxo-2,19-dioxa-4,7,10,13,16-pentaazaeicosyl)phenyl)amino)-1-oxo-5-ureidopent-2-yl)amino)-3-methyl-1-oxobutan-2-yl)amino)-5-oxopentanoic acid tert-butyl ester (70 mg, 94%). MS (M+H) + = 1531.6.

[0672] Step F-3, preparation of (S)-4-(3-(3-((2-((4-((tert-butoxycarbonyl)amino)piperidin-1-yl)-3- (6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)ethyl)amino)-3-oxopropyloxy)propanamido)-5-(((S)-1-(((S)-1-((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)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-tri oxo-2,13- dioxa-4,7,10-triaza-tetradecanyl)phenyl)amino)-1-oxo-5-ureidopentan-2-yl)amine(4S)-4-Amino-5-(((2S)-1-(((2S)-1-((4-((5S,11S,14S,17S,18R)-17-((S)-sec-Butyl)-18-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-Hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-2-oxoethyl)-5,8,11,14-tetraisopropyl-4,10,16-trimethyl-3,6,9,12,15-pentaoxo-2,19-dioxa-4,7,10,13,16-pentaazaeicosyl)phenyl)amino)-1-oxo-5-ureidopent-2-yl)amino)-3-methyl-1-oxobutan-2-yl)amino)-5-oxopentanoic acid tert-butyl ester: A mixture of 3-(3-((2-((4-(4-((tert-Butoxycarbonyl)amino)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)ethyl)amino)-3-oxopropoxy)propanoic acid (40 mg, 1.3 equiv, 54 μmol), 4-methylmorpholine (20 mg, 4.6 equiv, 0.20 mmol), and diphenyl perfluorobenzenephosphonate (25 mg, 1.5 equiv, 65 μmol) in DMF (1 mL) was stirred at 20 °C for 10 minutes, then (4S)-4-amino-5-(((2S)-1-(((2S)-1-((4-((5S,11S,14S,17S,18R)-17-((S)-sec-butyl)-18-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-2-oxoethyl)-5,8,11,14-tetraisopropyl-4,10,16-trimethyl-3,6,9,12,15-pentaoxo-2,19-dioxa-4,7,10,13,16-pentaazaeicosyl)phenyl)amino)-1-oxo-5-ureidopent-2-yl)amino)-3-methyl-1-oxobutan-2-yl)amino)-5-oxopentanoic acid tert-butyl ester (65 mg, 1 equiv, 43 μmol) was added, and the reaction mixture was stirred at 20 °C for 1 hour.The mixture was diluted with 20 mL of water and extracted with EtOAc (20 mL x 3). The combined organic layers were washed with water (10 mL x 2) and brine (20 mL), dried over anhydrous Na2SO4 and concentrated under reduced pressure to afford tert-butyl (S)-4-(3-(3-((2-((4-((tert-butoxycarbonyl)amino)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)ethyl)amino)-3-oxopropoxy)propanamido)-5-(((S)-1-(((S)-1-((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)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)phenyl)amino)-1-oxo-5-ureidopent-2-yl)amino)-3-methyl-1-oxobutan-2-yl)amino)-5-oxopentanoate (70 mg, 81%). MS (M / 2+H). + = 1015.2. This material was used in the next step without further purification.

[0673] Step F-4, preparation of 4-((S)-2-((S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-methylbut amido)propanamido)benzyl ((S)-1-(((S)-1-(((3R,4S,5S)-1-((S)-2-((1R,2R)-3-(((1S, 2R)-1-hydroxy-1-phenylpropan-2-yl) amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-3-methoxy-5-methyl-1-oxohept-4- yl)(methyl)amino)-3-methyl-1-oxobutan-2-yl)amino)-3-methyl-1-oxo butan-2-yl)(methyl)aminoFormate: To a mixture of (4S)-4-(3-{2-[(2-{[4-(4-aminopiperidin-1-yl)-3-(6-chloro-1H-1,3-benzodiazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl]amino}ethyl)carbamoyl]ethoxy}propanamido)-4-{[(1S)-1-{[(1S)-4-(carbamoylamino)-1-({4-[({[(1S)-1-{[(1S)-1-{[(3R,4S,5S)-1-[(2S)-2-[(1R,2R)-2-{[(1S,2R)-1-hydroxy-1-phenylpropan-2-yl]carbamoyl}-1-methoxy-2-methylethyl]pyrrolidin-1-yl]-3-methoxy-5-methyl-1-oxohept-4-yl](methyl)carbamoyl}-2-methylpropyl]amino}carbamoyl}-2-methylpropyl](methyl)carbamoyl}oxy)methyl]phenyl}carbamoyl)butyl]amino}carbamoyl}-2-methylpropyl]amino}butyric acid (35 mg, 80 wt%, 1 eq, 15 μmol) in DMF (1 mL) was added ammonium chloride (20 mg, 25 eq, 0.37 mmol), 1H-benzo[d][1,2,3]triazol-1-ol hydrate (3 mg, 1 eq, 0.02 mmol), 3-(((ethylimino)methylene)amino)-N,N-dimethylpropan-1-amine hydrochloride (5 mg, 2 eq, 0.03 mmol) and N,N-dimethylpyridin-4-amine (3 mg, 2 eq, 0.02 mmol). The reaction mixture was stirred at 20 °C for 6 h. The crude product was purified by preparative HPLC with the following conditions: column: SunFire prep OBD 19*150 mm 5um; mobile phase A: water (0.05% TFA); mobile phase B: ACN; gradient: 25% B to 55% B in 16 min; flow rate: 20 mL / min; wavelength: 220 nm.The collected fractions were dried by lyophilization to afford {4-[(2S)-2-[(2S)-2-[(2S)-2-(3-{2-[(2-{[4-(4-aminopiperidin-1-yl)-3-(6-chloro-1H-1,3-benzodiazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl]amino}ethyl)aminoformyl]ethoxy}propanamido)-4-carbamoylbutanamido]-3-methylbutanamido]-5-(carbamoylamino)pentanamido]phenyl}methyl N-[(1S)-1-{[(1S)-1-{[(3R,4S,5S)-1-[(2S)-2-[(1R,2R)-2-{[(1S,2R)-1-hydroxy-1-phenylpropan-2-yl]carbamoyl}-1-methoxy-2-methylethyl]pyrrolidin-1-yl]-3-methoxy-5-methyl-1-oxohept-4-yl](methyl)carbamoyl}-2-methylpropyl]carbamoyl}-2-methylpropyl]-N-methylcarbamate (10.3 mg, 31%). MS (M+H). + = 1872.1.

[0674] Similar to Example F, the following conjugates were prepared using appropriate substituted reagents and substrates in different steps and they may require additional functional group modifications via well-known chemistry using appropriate reagents.

[0675]

[0676]

[0677]

[0678] Example G. (2S,3S,4S,5R,6S)-6-{2-[(2S)-2-[(2S)-2-(3-{2-[(2-{[4-(4-aminopiperidin-1-yl)-3-(6-chloro-1H-1,3-benzodiazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl]amino}ethyl)carbamoyl]ethoxy}propanamido)-3-methylbutanamido]-5-(carbamoylamino)pentanamido]-5-[({[(1S)-1-{[(1S)-1-{[(3R,4S,5S)-1-[(2S)-2-[(1R,2R)-2-{[(1S,2R)-1-hydroxy-1-phenylpropan-2-yl]carbamoyl}-1-methoxy-2-methylethyl]pyrrolidin-1-yl]-3-methoxy-5-methyl-1-oxohept-4-yl](methyl)carbamoyl}-2-methylpropyl]carbamoyl}-2-methylpropyl](methyl)carbamoyl}oxy)methyl]phenoxy}-3,4,5-trihydroxyoxane-2-carboxylic acid (Compound 61)

[0679]

[0680] Step G-1, preparation of (2S,3R,4S,5S,6S)-2-(5-formyl-2-nitrophenoxy)-6-(methoxycarbonyl ) tetrahydro-2H-pyran-3,4,5-triyltriacetate ((2S,3R,4S,5S,6S)-2-(5-formyl-2-nitrophenoxy)- 6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate): A mixture of 3-hydroxy-4-nitrobenzaldehyde (2.334 g, 1 eq., 13.97 mmol), (2R,3R,4S,5S,6S)-2-bromo-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (16.64 g, 3.000 eq., 41.90 mmol) and silver(I) oxide (25.9 g, 8.00 eq., 112 mmol) in ACN (140 mL) was stirred at 25 °C in the dark for 24 h. The mixture was filtered and the filtrate was concentrated under reduced pressure. The crude product was purified by MPLC under the following conditions: silica gel column 120 g, petroleum ether / ethyl acetate (PE / EtOAc) system, ratio of EtOAc from 0% to 85% in 15 min, flow rate: 70 mL / min; wavelength: 254 nm. The collected fractions were combined and concentrated in vacuo to give (2S,3R,4S,5S,6S)-2-(5-formyl-2-nitrophenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (3.8 g, 56%). MS (M+H) + = 484.4.

[0681] Step G-2, Preparation of (2S,3R,4S,5S,6S)-2-(2-amino-5-(hydroxymethyl)phenoxy)-6-(methoxy carbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate:To a mixture of (2S,3R,4S,5S,6S)-2-(5-formyl-2-nitrophenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (3.8 g, 1 equiv, 7.9 mmol) and triethylamine (0.16 g, 0.22 mL, 0.20 equiv, 1.6 mmol) in ethyl acetate (210 mL) under a nitrogen inert atmosphere was added Pearlman's catalyst (85 mg, 0.10 equiv, 0.80 mmol). The reaction mixture was flushed with hydrogen 3 times, then flushed with hydrogen and stirred at 25 °C under H2 gas pressure for 24 h. The reaction mixture was filtered through a pad of diatomaceous earth, and the filtrate was concentrated under reduced pressure to afford (2S,3R,4S,5S,6S)-2-(2-amino-5-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (2.7 g, 75%). MS (M+H) + = 456.4

[0682] Step G-3, prepare (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-((tert-butoxycarbonyl)amino)- 3- methylbutanamido)-5-ureidopentanamido)-5-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H- Pyr an-3,4,5-triyltriacetate: To a mixture of (2S,3R,4S,5S,6S)-2-(2-amino-5-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (1.05 g, 1 equiv, 2.31 mmol), 2-ethoxy-1-ethoxycarbonyl-1,2-dihydroquinoline (EEDQ) (685 mg, 1.20 equiv, 2.77 mmol) in DCM (10 mL) at room temperature was added (S)-2-((S)-2-((tert-butoxycarbonyl)amino)-3-methylbutanamido)-5-ureidopentanoic acid (865 mg, 1.00 equiv, 2.31 mmol), and the reaction mixture was stirred at 25 °C in the dark for 24 h. The reaction mixture was concentrated under reduced pressure. The crude product was purified by MPLC under the following conditions: silica gel column 120 g, PE / EtOAc system, ratio of EtOAc from 0% to 85% in 15 min, flow rate: 70 mL / min; wavelength: 254 nm. The collected fractions were combined and concentrated in vacuo to afford (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-((tert-butoxycarbonyl)amino)-3-methylbutanamido)-5-ureidopentanamido)-5-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (1.36 g, 72.7%). MS (M+H) + = 812.2

[0683] Step G-4, prepare (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-((tert-butoxycarbonyl)amino)- 3- -5-((((4-nitrophenoxy)carbonyl)oxy)methyl)phenoxy yl)-6- (methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate: A mixture of (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-((tert-butoxycarbonyl)amino)-3-methylbutanamido)-5-ureidopentanamido)-5-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (460 mg, 1 equivalent, 567 μmol), bis(4-nitrophenyl) carbonate (260 mg, 1.51 equivalents, 855 μmol), and N-ethyl-N-isopropylpropan-2-amine (150 mg, 2.05 equivalents, 1.16 mmol) in tetrahydrofuran (THF) (4.6 mL) was stirred at room temperature for 24 h. The reaction mixture was concentrated under reduced pressure. The crude product was purified by MPLC under the following conditions: silica gel column 120 g, PE / EtOAc system, ratio of EtOAc from 0% to 85% in 15 min, flow rate: 70 mL / min; wavelength: 254 nm. The collected fractions were combined and concentrated in vacuo to give (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-((tert-butoxycarbonyl)amino)-3-methylbutanamido)-5-ureidopentanamido)-5-((((4-nitrophenoxy)carbonyl)oxy)methyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (400 mg, 72.3%). MS (M+H) + = 977.1.

[0684] Step G-5, preparation of (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-((tert-butoxycarbonyl)amino)-3- methylbutanamido)-5-ureidopentanamido)-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-oxo propyl)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-tri oxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyr oxa-3,4,5-triyltriacetic ester:(2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-((tert-Butoxycarbonyl)amino)-3-methylbutanamido)-5-ureidopentanamido)-5-(((4-nitrophenoxy)carbonyl)oxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (170 mg, 1 eq, 174 μmol), (S)-N-((3R,4S,5S)-1-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-3-methoxy-5-methyl-1-oxohept-4-yl)-N,3-dimethyl-2-((S)-3-methyl-2-(methylamino)butanamido)butanamide (100 mg, 0.800 eq, 139 μmol), 1H-benzo[d][1,2,3]triazol-1-ol (30 mg, 1.3 eq, 0.22 mmol) and DIEA (70 mg, 94 μL, 3.1 eq, 0.54 mmol) in a mixture of DMF (1.7 mL) was stirred at 25 °C for 2 h. The mixture was purified directly by MPLC under the following conditions: column, WelFlashTM, C18 120 g, Spherical 20 - 40 μm; mobile phase, water (0.05% NH₃.H₂O) and ACN (5% ACN to 5% ACN in 1 min, 30% ACN to 98% in 6 min, 98% ACN to 98% in 3 min); total flow rate, 70 mL / min; detector, UV 254 nm. The collected fractions were combined and concentrated in vacuo to give (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-((tert-Butoxycarbonyl)amino)-3-methylbutanamido)-5-ureidopentanamido)-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)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (80 mg, 30%). MS (M + H) + = 1556.5.

[0685] Step G-6, prepare (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-amino-3-methylbutanamide yl)-5-ureidopentanamido)-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)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trimethylol oxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate: yl)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-((tert-butoxycarbonyl)amino)-3-methylbutyramido)-5-ureidopentanamido)-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- A mixture of (3-oxopropyl)pyrrolidin-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-triyl triacetate (76 mg, 1 eq., 49 μmol) and TFA (0.5 mL) in DCM (0.5 mL) was stirred at 0° C. for 2 min. The mixture was adjusted to pH 5 with DIEA. 9.0 and dried with nitrogen to give (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-amino-3-methylbutyramido)-5-ureidopentanamido)-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)pyrrolidin-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-triyl triacetate (60 mg, 84%). MS (M+H) + = 1456.5. This material was used in the next step without further purification.

[0686] Step G-7, preparation of (2S,3R,4S,5S,6S)-2-(2-((2S,5S)-16-((4-(4-(tert-butoxycarbonyl)amino yl)-5- 1-(2-(3-ureidopropyl)-5-isopropyl-4,7,13-trioxo-2-(3-ureidopropyl)-10-oxa-3,6,14-triazahexadecanoic acid amide ((5S,8S,11S,12R)-11-((S)-sec-butyl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy yl)-5,8-di-((1-Phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-2-oxoethyl isopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy)-6- (methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyltriacetate: yl)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino(2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-Amino-3-methylbutanamido)-5-ureidopentanamido)-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)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (60 mg, 1 equiv, 41 μmol), diphenyl perfluorobenzenephosphonate (20 mg, 1.3 equiv, 52 μmol), 4-methylmorpholine (13 mg, 14 μL, 3.1 equiv, 0.13 mmol), and 3-(3-((2-((4-((tert-butoxycarbonyl)amino)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)ethyl)amino)-3-oxopropoxy)propanoic acid (30 mg, 0.99 equiv, 41 μmol) in DMF (0.6 mL) were stirred at 25 °C for 2 h. The mixture was directly purified by MPLC under the following conditions: column, WelFlashTM, C18 120 g, Spherical 20 - 40 μm; mobile phase, water (0.05% NH₃·H₂O) and ACN (5% ACN to 5% ACN in 2 min, 30% ACN to 98% in 12 min, 98% ACN to 98% in 1 min); total flow rate, 70 mL / min; detector, UV 220 nm.The collected fractions were combined and concentrated in vacuo to afford (2S,3R,4S,5S,6S)-2-(2-((2S,5S)-16-((4-(4-((tert-butoxycarbonyl)amino)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)-5-isopropyl-4,7,13-trioxo-2-(3-ureidopropyl)-10-oxa-3,6,14-triazahexadecanamido)-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)pyrrolidin-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-triyl triacetate (64 mg, 71%). MS (M / 2 + H). + = 1088.6.

[0687] Step G-8, preparation of (2S,3S,4S,5R,6S)-6-(2-((2S,5S)-16-((4-(4-((tert-butoxycarbonyl)amino yl)-5- yl)-5-isopropyl-4,7,13-trioxo-2-(3-ureidopropyl)-10-oxa-3,6,14-triazahexadecanamide ((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)pyrrolidin-1-yl)-2-oxoeth yl)phenyl)phenoxy)-3, yl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecane 4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid: ​(2S,3R,4S,5S,6S)-2-(2-((2S,5S)-16-((4-(4-((tert-Butoxycarbonyl)amino)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)-5-isopropyl-4,7,13-trioxo-2-(3-ureidopropyl)-10-oxa-3,6,14-triazahexadecanamido)-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)pyrrolidin-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-triyl triacetate (64 mg, 1 equiv, 29 μmol) and lithium hydroxide (4 mg, 6 equiv, 0.2 mmol) in a mixture of THF (0.6 mL) and H2O (0.6 mL) were stirred at 25 °C for 2 h. The residue was diluted with water (10 ml) and then adjusted to pH 6 - 7 with AcOH (1 M). The resulting solution was extracted with ethyl acetate (3 × 2 mL). The organic layer was dried over anhydrous sodium sulfate and concentrated in vacuo to afford (2S,3S,4S,5R,6S)-6-(2-((2S,5S)-16-((4-(4-((tert-Butoxycarbonyl)amino)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)-5-isopropyl-4,7,13-trioxo-2-(3-ureidopropyl)-10-oxa-3,6,14-triazahexadecanamido)-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)pyrrolidin-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 (50 mg, 15 μmol, 52%, 62% purity). MS (M / 2 + H) + = 1019.2. This material was used in the next step without further purification.

[0688] Step G-9, prepare (2S,3S,4S,5R,6S)-6-{2-[(2S)-2-[(2S)-2-(3-{2-[(2-{[4-(4-amino (piperidin-1-yl)-3-(6-chloro-1H-1,3-benzodiazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl]amino} ethyl)carbamoyl]ethoxy}propanamido)-3-methylbutanamido]-5-(carbamoylamino)pentanoyl amino]- 5-[({[(1S)-1-{[(1S)-1-{[(3R,4S,5S)-1-[(2S)-2-[(1R,2R)-2-{[(1S,2R)-1-hydroxy-1-phenyl propan-2-yl]carbamoyl}-1-methoxy-2-methylethyl]pyrrolidin-1-yl]-3-methoxy-5-meth yl-1-oxohept-4-yl](methyl)carbamoyl}-2-methylpropyl]carbamoyl}-2-methylpropyl](meth yl)carbamoyl} oxymethyl]phenoxy}-3,4,5-trihydroxyoxane-2-carboxylic acid:A mixture of (2S,3S,4S,5R,6S)-6-(2-((2S,5S)-16-((4-(4-((tert-butoxycarbonyl)amino)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)-5-isopropyl-4,7,13-trioxo-2-(3-ureidopropyl)-10-oxa-3,6,14-triazapentadecanamido)-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)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (50 mg, 52 wt%, 1 eq, 13 μmol) and TFA (0.2 mL) in DCM (1.0 mL) was stirred at 0 °C for 30 min. The crude product was purified by preparative HPLC with the following conditions: column: SunFire prep OBD 19*150 mm 5um; mobile phase A: water (0.05% NH₃.H₂O); mobile phase B: ACN; gradient: 25% B to 65% B in 8 min; flow rate: 20 mL / min.H₂O; wavelength: 220 nm. The collected fractions were dried by lyophilization to give (2S,3S,4S,5R,6S)-6-{2-[(2S)-2-[(2S)-2-(3-{2-[(2-{[4-(4-aminopiperidin-1-yl)-3-(6-chloro-1H-1,3-benzodiazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl]amino}ethyl)carbamoyl]ethoxy}propanamido)-3-methylbutanamido]-5-(carbamoylamino)pentanamido]-5-[({[(1S)-1-{[(1S)-1-{[(3R,4S,5S)-1-[(2S)-2-[(1R,2R)-2-{[(1S,2R)-1-hydroxy-1-phenylpropan-2-yl]carbamoyl}-1-methoxy-2-methylethyl]pyrrolidin-1-yl]-3-methoxy-5-methyl-1-oxohept-4-yl](methyl)carbamoyl}-2-methylpropyl]amino}carbamoyl}-2-methylpropyl](methyl)carbamoyloxy)methyl]phenoxy}-3,4,5-trihydroxyoxane-2-carboxylic acid (8.1 mg, 33%). MS (M+H) + = 1936.0.

[0689] Similar to Example G, the following conjugates were prepared using appropriate substitution reagents and substrates in different steps, and they may require additional functional group modification via well-known chemistry using appropriate reagents.

[0690]

[0691]

[0692]

[0693]

[0694]

[0695]

[0696]

[0697]

[0698]

[0699]

[0700]

[0701]

[0702]

[0703]

[0704]

[0705]

[0706]

[0707]

[0708] Example H. (2S,3S,4S,5R,6S)-6-{2-[(2S)-2-[(2S)-2-(3-{2-[(2-{[4-(4-Aminopiperidin-1-yl)-3-(6-chloro-1H-1,3-benzodiazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl]amino}ethyl)carbamoyl]ethoxy}propanamido)-3-methylbutanamido]propanamido]-5-[({[(1S)-1-{[(1S)-1-{[(3R,4S,5S)-1-[(2S)-2-[(1R,2R)-2-{[(1S,2R)-1-hydroxy-1-phenylpropan-2-yl]carbamoyl}-1-methoxy-2-methylethyl]pyrrolidin-1-yl]-3-methoxy-5-methyl-1-oxohept-4-yl](methyl)carbamoyl}-2-methylpropyl]carbamoyl}-2-methylpropyl](methyl)carbamoyl}oxy)methyl]phenoxy}-3,4,5-trihydroxyoxane-2-carboxylic acid (Compound 68)

[0709]

[0710] Step H-1, Preparation of (2S,3R,4S,5S,6S)-2-(2-((S)-2-((tert-Butoxycarbonyl)amino)propanamide yl)-5-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-tris ester: A mixture of (2S,3R,4S,5S,6S)-2-(2-amino-5-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (800 mg, 1 equiv, 1.76 mmol), (tert-Butoxycarbonyl)-L-alanine (400 mg, 1.20 equiv, 2.11 mmol), and EEDQ (520 mg, 1.20 equiv, 2.10 mmol) in DMF (10 mL) was stirred at 25 °C for 24 h. The reaction was then quenched with water (100 mL) and the resulting solution was extracted with ethyl acetate (3 × 100 mL). The organic layer was dried over anhydrous sodium sulfate and concentrated in vacuo. The residue was applied to a silica gel column with ethyl acetate / petroleum ether (3:1). The collected fractions were combined and concentrated in vacuo to give (2S,3R,4S,5S,6S)-2-(2-((S)-2-((tert-Butoxycarbonyl)amino)propanamido)-5-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (770 mg, 70.0%). MS (M+H) + = 627.5.

[0711] Step H-2, prepare (2S,3R,4S,5S,6S)-2-(2-((S)-2-aminopropanamido)-5-(hydroxymethyl)(2S,3R,4S,5S,6S)-2-(2-((S)-2-((tert-Butoxycarbonyl)amino)propanamido)-5-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (770 mg, 1 equiv, 1.23 mmol) and TFA (2 mL) in DCM (6 mL) were stirred at 25 °C for 30 min. The resulting solution was concentrated in vacuo. The resulting mixture was dissolved in 20 mL of DCM and washed with 2 mL of saturated NaHCO3. The organic layer was dried over anhydrous sodium sulfate and concentrated to afford (2S,3R,4S,5S,6S)-2-(2-((S)-2-aminopropanamido)-5-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (650 mg, 100%). MS (M+H) + = 527.2. This material was used in the next step without further purification.

[0712] Step H-3, Preparation of (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-((((9H-Fluoren-9-yl)methoxy) carbonyl)amino)-3-methylbutanamido)propanamido)-5-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetraHydrogen-2H-pyran-3,4,5-triyl triacetate: A mixture of (2S,3R,4S,5S,6S)-2-(2-((S)-2-aminopropanamido)-5-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (650 mg, 1 equivalent, 1.23 mmol), HATU (580 mg, 1.24 equivalents, 1.53 mmol), and diisopropylethylamine (480 mg, 642 μL, 3.01 equivalents, 3.71 mmol) in DMF (6.5 mL) was stirred at 25 °C for 10 min. Then, (((9H-fluoren-9-yl)methoxy)carbonyl)-L-valine (420 mg, 1.00 equivalent, 1.24 mmol) was added and the resulting reaction mixture was stirred at 25 °C for 1 h. Then, the reaction was quenched with water (100 mL) and the resulting solution was extracted with ethyl acetate (3 x 100 mL). The organic layer was dried over anhydrous sodium sulfate and concentrated in vacuo. The residue was applied to a silica gel column with ethyl acetate / petroleum ether (2:1). The collected fractions were combined and concentrated in vacuo to give (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-methylbutanamido)propanamido)-5-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (620 mg, 59.2%). MS (M+H) + = 848.3.

[0713] Step H-4, prepare (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-((((9H-fluoren-9-yl)methoxy yl)carbonyl)amino)-3-methylbutanamido)propanamido)-5-((((4-nitrophenoxy)carbonyl)oxy)methyl)phenoxy yl)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-tris acetate: A mixture of (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-methylbutanamido)propanamido)-5-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (520 mg, 1 equivalent, 613 μmol) and bis(4-nitrophenyl) carbonate (280 mg, 1.50 equivalents, 920 μmol) in THF (5 mL) was stirred at 25 °C under a nitrogen atmosphere for 24 h. 70% of the product was detected by LCMS. The reaction mixture was worked up together with the next batch. MS (M+H) + = 1013.

[0714] Step H-5, prepare (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-((((9H-fluoren-9-yl)methoxy yl) carbonyl)amino)-3-methylbutanamido)propanamido)-5-((5S,8S,11S,12R)-11-((S)-sec-butyl yl)-12-(2-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2- methyl 3-oxopropyl)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-tri oxo-2,13- dioxo-4,7,10-triaza-tetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyr furan-3,4,5-triyl triacetate: A mixture of (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-methylbutanamido)propanamido)-5-((((4-nitrophenoxy)carbonyl)oxy)methyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (320 mg, 1 equiv, 316 μmol), (S)-N-((3R,4S,5S)-1-((S)-2-((1R,2R)-3-(((1S,2R)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-3-methoxy-5-methyl-1-oxohept-4-yl)-N,3-dimethyl-2-((S)-3-methyl-2-(methylamino)butanamido)butanamide (205 mg, 0.904 equiv, 286 μmol), 1-hydroxy-1H-benzotriazole (52 mg, 53 μL, 1.2 equiv, 0.38 mmol) and N-ethyl-N-isopropylpropan-2-amine (123 mg, 3.01 equiv, 952 μmol) in DMF (3.5 mL) was stirred at 25 °C for 16 h. 68% product was detected by LCMS. The reaction mixture was worked up together with the next batch. MS(M+H) + = 1591.6.

[0715] Step H-6, prepare (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-amino-3-methylbutanamido) propanamido)-5-((5R,11R,12S)-11-((R)-sec-butyl)-12-(2-((R)-2-((1S,2S)-3-(((1R,2S)-1- hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-2-oxoeth yl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triaza-tetradecyl) phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-tris acetate:At 0 °C, piperidine (45.0 mg, 52.0 μL, 7.01 eq, 528 μmol) was added to a mixture of (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-(((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-methylbutanamido)propanamido)-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)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (120 mg, 1 eq, 75.4 μmol) in DMF (2.8 mL), and the resulting reaction mixture was stirred at 0 °C for 5 h. The mixture was purified directly by MPLC under the following conditions: column, WelFlashTM, C18 120 g, Spherical 20 - 40 μm; mobile phase, water (0.05% NH.H2O) and ACN (5% ACN to 5% ACN in 1 min, 20% ACN to 98% in 8 min, 98% ACN to 98% in 1 min); total flow rate, 70 mL / min; detector, UV 220 nm. The collected fractions were combined and concentrated in vacuo to give (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-amino-3-methylbutanamido)propanamido)-5-((5R,11R,12S)-11-((R)-sec-butyl)-12-(2-((R)-2-((1S,2S)-3-(((1R,2S)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (120 mg, 58%). MS (M+H) + = 1369.6.

[0716] Step H-7, prepare (2S,3R,4S,5S,6S)-2-(2-((2S,5S)-16-((4-(4-((tert-butoxycarbonyl) amino yl)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amine yl)-5-isopropyl-2-methyl-4,7,13-trioxo-10-oxa-3,6,14-triazahexadecanamide-yl)-5-((5R,11R,12S)-11-((R)-sec-butyl)-12-(2-((R)-2-((1S,2S)-3-(((1R,2S)-1-hydroxy-1- phenylpropan-2- ((2S,5S)-16-((4-(4-((tert-Butoxycarbonyl)amino)-5-isopropyl-2-methyl-4,7,13-trioxo-10-oxa-3,6,14-triazacyclohexadecanamido)-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)pyrrolidin-1-yl)allyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)oxy)-3,4,5-trihydroxyoxan-2-yl)methyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13 diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxa-4,7,10-triazatetradecyl)phenoxy ​ ​The mixture of (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-amino-3-methylbutanamido)propanamido)-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)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (100 mg, 1 equivalent, 73.0 μmol), 3-(3-((2-((4-((tert-butoxycarbonyl)amino)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)ethyl)amino)-3-oxopropoxy)propanoic acid (55 mg, 1.0 equivalent, 74 μmol), diphenyl perfluorophenyl phosphite (34 mg, 1.2 equivalents, 88 μmol) and 4-methylmorpholine (22 mg, 24 μL, 3.0 equivalents, 0.22 mmol) in DMF (1 mL) was stirred at 25 °C for 10 min. Then (2S,3R,4S,5S,6S)-2-(2-((S)-2-((S)-2-amino-3-methylbutanamido)propanamido)-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)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (100 mg, 1 equivalent, 73.0 μmol) was added thereto. The resulting reaction mixture was stirred at 25 °C for 1 h. The mixture was directly purified by MPLC under the following conditions: column, Flash TM, C18 120 g, Spherical 20 - 40 μm; mobile phase, water (0.05% NH₃·H₂O) and ACN (5% ACN to 5% ACN in 2 min, 30% ACN to 98% in 12 min, 98% ACN to 98% in 1 min); total flow rate, 70 mL / min; detector, UV 220 nm.The collected fractions were combined and concentrated in vacuo to afford (2S,3R,4S,5S,6S)-2-(2-((2S,5S)-16-((4-(4-((tert-butoxycarbonyl)amino)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)-5-isopropyl-2-methyl-4,7,13-trioxo-10-oxa-3,6,14-triazapentadecanamido)-5-((5R,11R,12S)-11-((R)-sec-butyl)-12-(2-((R)-2-((1S,2S)-3-(((1R,2S)-1-hydroxy-1-phenylpropan-2-yl)amino)-1-methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-2-oxoethyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triyl triacetate (20 mg, 13%). MS (M / 2+H). + = 1050.5; MS (M+H) + = 2099.5.

[0717] ​ ​ yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino ​ ​ ​ ​ ​(2S,3R,4S,5S,6S)-2-(2-((2S,5S)-16-((4-(4-((tert-Butoxycarbonyl)amino)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)-5-isopropyl-2-methyl-4,7,13-trioxo-10-oxa-3,6,14-triazahexadecanamido)-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)pyrrolidin-1-yl)allyl)-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-triyl triacetate (18 mg, 1 eq., 8.6 μmol) and lithium hydroxide (2 mg, 1e+1 eq., 0.08 mmol) in a mixture of THF (0.1 mL) and H2O (0.02 mL) were stirred at 25 °C for 30 min. The residue was diluted with water (10 ml) and then adjusted to pH 6 - 7 with AcOH (1 M). The resulting solution was extracted with ethyl acetate (3 x 2 mL). The organic layer was dried over anhydrous sodium sulfate and concentrated in vacuo to give (2S,3S,4S,5R,6S)-6-(2-((2S,5S)-16-((4-(4-((tert-Butoxycarbonyl)amino)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)-5-isopropyl-2-methyl-4,7,13-trioxo-10-oxa-3,6,14-triazahexadecanamido)-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)pyrrolidin-1-yl)allyl)-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 (20 mg, 84%). MS (M / 2 + H) + = 975; MS (M + H) + = 1949. This material was used in the next step without further purification.

[0718] ​ ​{ethyl)carbamoyl]ethoxy}propanamido)-3-methylbutanamido]propanamide ​ ​ ​ ​ ​A mixture of (2S,3S,4S,5R,6S)-6-(2-((2S,5S)-16-((4-(4-((tert-butoxycarbonyl)amino)piperidin-1-yl)-3-(6-chloro-1H-benzo[d]imidazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl)amino)-5-isopropyl-2-methyl-4,7,13-trioxo-10-oxa-3,6,14-triazapentadecanamido)-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)pyrrolidin-1-yl)allyl)-5,8-diisopropyl-4,10-dimethyl-3,6,9-trioxo-2,13-dioxo-4,7,10-triazatetradecyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (20 mg, 1 equiv, 10 μmol) and TFA (0.1 mL) in DCM (0.5 mL) was stirred at 25 °C for 30 min. The crude product was purified by preparative HPLC (Preparative HPLC-013) with the following conditions: column, Atlantis Prep T3 OBD column, 19*150 mm 5um; mobile phase, water (0.1% TFA) and ACN (from 28% B phase to 53% in 10 min); 20 mL / min. Detector, UV 220, 254 nm. The collected fractions were combined and concentrated in vacuo to give (2S,3S,4S,5R,6S)-6-{2-[(2S)-2-[(2S)-2-(3-{2-[(2-{[4-(4-aminopiperidin-1-yl)-3-(6-chloro-1H-1,3-benzodiazol-2-yl)-5-(3-fluoro-5-methylphenyl)pyridin-2-yl]amino}ethyl)carbamoyl]ethoxy}propanamido)-3-methylbutanamido]propanamido}-5-[({[(1S)-1-{[(1S)-1-{[(3R,4S,5S)-1-[(2S)-2-[(1R,2R)-2-{[(1S,2R)-1-hydroxy-1-phenylpropan-2-yl]carbamoyl}-1-methoxy-2-methylethyl]pyrrolidin-1-yl]-3-methoxy-5-methyl-1-oxohept-4-yl](methyl)carbamoyl}-2-methylpropyl]amino}carbamoyl}-2-methylpropyl](methyl)carbamoyl}oxy)methyl]phenoxy}-3,4,5-trihydroxyoxane-2-carboxylic acid (4.7 mg, 21%). MS (M+H) + = 1850.0.

[0719] Biological assay

[0720] ​

[0721] General overview: All five SSTR subtypes are Gi-coupled G protein-coupled receptors (GPCRs) that, when activated by an agonist, lead to a decrease in intracellular cyclic AMP (cAMP). Therefore, measurement of intracellular cAMP levels can be used to evaluate whether a compound of the present invention is an agonist of an SSTR subtype (John Kelly, Troy Stevens, W. Joseph Thompson, and Roland Seifert, Current Protocols in Pharmacology, 2005, 2.2.1-2.2). An example of an intracellular cAMP assay is described below.

[0722] Functional assay of SSTR2 agonist

[0723] Four days prior to the assay, 2,000 Chinese hamster ovary cells (CHO-K1, ATCC#CCL-61) stably expressing human somatostatin receptor subtype 2 were plated into each well of a tissue culture-treated 96-well plate in Ham's F12 growth medium (ThermoFisher#10-080-CM), which was supplemented with 10% donor bovine serum (Gemini Bio-Products#100-506), 100 U / mL penicillin, 100 μg / mL streptomycin, 2 mM L-glutamine (Gemini Bio-Products#400-110), and 0.2 mg / mL hygromycin B (GoldBio#31282-04-9). The cells were cultured at 37 °C, 5% CO2, and 95% humidity. cAMP was measured using the HTRF Dynamic cAMP assay (Cisbio, #62AM5PEJ) according to the manufacturer's instructions. On the day of the assay, the medium was aspirated, and the cells were treated with 50 μL of stimulation buffer supplemented with 10.2 mM 3-isobutyl-1-methylxanthine (IBMX, Millipore Sigma#I5879) and 1.6 μM NKH477 (Tocris#1603) and various dilutions of the SMDC of the present invention. The cells were incubated at 37 °C for 20 minutes (the final concentration of the compound of the present invention was generally 0 - 10,000 nM). The cells were treated with 50 μL of lysis buffer (HRTF cAMP kit, Cisbio) and incubated at room temperature for 30 minutes with rotation shaking at 600 rpm, then diluted with 150 μL of stimulation buffer and shaken again at 300 rpm for five minutes. The lysate was transferred to a 384-well plate and incubated at room temperature for 1 - 24 hours, and cAMP accumulation was detected by d2-labeled cAMP and anti-cAMP-Cryptate. The time-resolved fluorescence signal was read using a m1000 Pro (Tecan) or CLARIOStar (BMG Labtech) microplate reader, where the samples were excited with light at 340 nm, and the emitted light was measured at 620 nm and 665 nm. The data was represented as the calculated result of the fluorescence ratio (665 nm / 620 nm). The intracellular cAMP concentration was calculated by regression to a standard curve and plotted against the concentration of the compound of the present invention. The EC 50 of the compound was calculated using standard methods. All data manipulations were performed in GraphPad Prism v9 (GraphPad, San Diego, CA).

[0724] Example J: Internalization Assay Protocol

[0725] The SMDCs of the present invention were evaluated in vitro using the human PathHunter CHO-K1 SST2β-arrestin2 and PathHunter CHO-K1 SST2 internalization assays from DiscoverX. CHO-K cells were seeded at a density of 2500 cells / well in 20 μL of cell culture medium in a white-wall 384-well tissue culture compatible plate. After incubation for 48 hours, the cells were treated with 5 μL of 1:3 serial dilutions of SST2 SMDCs (prepared at 5X concentration, final top compound concentration of 10 μM) at 37 °C and in 5% CO2 for 90 min. Eleven dilutions as well as a vehicle control without compound were tested. After incubation, the medium containing the treatment was discarded, and 20 μL of Beta-Glo working solution from the Promega Beta-Glo assay system was added to each well. The plate was then incubated at room temperature in the dark for 60 min. Luminescence was measured using a Tecan plate reader. IC 50 curves were generated using GraphPad Prism 9 with four-parameter logistic regression analysis. Results are the mean of replicates and were corrected for background.

[0726] Example K: H524 assay protocol

[0727] The SMDCs of the present invention were evaluated in an in vitro assay assessing inhibition of cell proliferation. NCI-H524 (ATCC) human small cell lung cancer cells were seeded at a concentration of 5,000 cells / well in 96-well V-bottom plates (Costar). After 24 h, the cells were treated with SMDCs for 2 h. The starting dose of SMDC was 1 μM and two-fold serial dilutions were performed for a total of 11 points. After 2 h of treatment, the cells were centrifuged, the drug-containing medium was removed, and fresh complete medium was added and used to resuspend the cells, followed by centrifugation again. After removing the medium, the cells were resuspended in complete medium and incubated for an additional 70 h. Cell proliferation was evaluated using the CellTiter Glo assay according to the standard protocol (Promega). Luminescence was measured using a TECAN plate reader. The percent proliferation inhibition was calculated using the following formula: % Inhibition = (luminescence control - luminescence treatment) / luminescence control * 100. IC 50 curves were generated using GraphPad Prism 9 with four-parameter logistic regression analysis. Exemplary biological activities of the compounds are shown in the table below.

[0728] Table 1: Representative activities

[0729]

[0730]

[0731]

[0732]

[0733]

[0734] n.d. = Not determined

[0735] Comparative activity

[0736] The compounds (25 and 68) of the present invention use an ethylenediamine moiety to link the targeting ligand to the linker or spacer of the SMDC. The inventors unexpectedly found that, compared to conjugates using a piperazine moiety to link the targeting ligand to the linker or spacer, using an ethylenediamine moiety significantly improved the internalization and H524 activity of the conjugate. This is illustrated in the comparative examples below, where the internalization of the SMDC with an ethylenediamine moiety was increased 5 to 10-fold compared to the corresponding conjugate with a piperazine moiety, as shown in Table 2.

[0737] The structures of comparative compounds 1 (Comp.1) and 2 (Comp.2) with a piperazine moiety are shown below:

[0738]

[0739] Comp.1

[0740]

[0741] Comp.2

[0742] The structures of the corresponding SMDCs of the present invention with an ethylenediamine moiety are shown below:

[0743]

[0744] Compound 25

[0745]

[0746] Compound 68

[0747] Table 2: Comparative activity

[0748]

[0749] Example L: H524 tumor biodistribution model

[0750] In this example, 5 million H524 (small cell lung cancer) cells in 1:1 Matrigel: cold medium (RPMI1640) were injected subcutaneously (SQ) into the flank of female athymic nude mice. Tumor volume and body weight were measured twice a week. Tumors were allowed to grow until their size reached 250 - 500 mm 3, the animals were then randomly grouped (n = 3 / time point). The SMDC of the present invention was intravenously injected into the animals as a single dose of 500 nmol / kg. At 1, 4, 24, and 72 hours after administration of the compound, the whole body was perfused with saline to remove any vascular signals, and then plasma and tissues were collected, and the concentrations of the conjugate and MMAE were measured by LC / MS-MS. The tumor and plasma levels of SMDC and MMAE are shown in Table 4. BLQ: Below the limit of quantification.

[0751] Table 3: Tumor and plasma concentrations of compound 68 and MMAE in H524 tumor-bearing nude mice after intravenous administration of 500 nmol / kg of compound 68.

[0752]

[0753] Table 4: Tumor and plasma concentrations of compound 49 and MMAE in H524 tumor-bearing nude mice after intravenous administration of 500 nmol / kg of compound 49.

[0754]

[0755]

[0756] Table 5: Tumor and plasma concentrations of compound 61 and MMAE in H524 tumor-bearing nude mice after intravenous administration of 500 nmol / kg of compound 61.

[0757]

[0758] Table 6: Tumor and plasma concentrations of compound 107 and MMAE in H524 tumor-bearing nude mice after intravenous administration of 500 nmol / kg of compound 107.

[0759]

[0760] Example M: 111 Biodistribution of In-compound 1 in female swiss nude mice bearing tumors derived from the AR42J rat pancreatic cancer cell line.

[0761] Study outline: 24 hours before the start of the biodistribution study, In-compound 1 was radiolabeled as described herein. On the day of the study, the animals received a single IV injection of 200 uL of In-labeled compound 1 (1 nmol) into the tail vein via a catheter, as shown in Table 2. The blocking study (Group 6) combined 1 nmol of In-compound 1 with 100 nmol of In-compound 1. 111 In-compound 1 111 In-labeled compound 1 (1 nmol), as shown in Table 2. The blocking study (Group 6) combined 1 nmol of In-compound 1 with 100 nmol of In-compound 1. 111 In-compound 1 115 In-compound 1.

[0762] Table 7: Research details

[0763]

[0764]

[0765] After drug administration, the animals were sacrificed at specific time points (0.5 h, 1 h, 2 h, 6 h, 22 h), and organs (blood, tumor, heart, kidney, pituitary, brain, liver, spleen, lung, intestine, adrenal gland, pancreas, stomach, bone (femur) and tail) were collected, weighed and the radioactivity of each organ / tissue was evaluated. The activity was quantified and expressed as %ID / g (percentage of the initial dose per gram of tissue).

[0766] Biodistribution results:

[0767] In the AR42J xenograft tumor model, 111 In - Compound 1 showed high and persistent uptake in SST2R - positive tumors. It was demonstrated by competition studies that 111 the uptake of In - Compound 1 was SST2R - specific, where co - administration of 100 - fold molar excess of non - radioactive 115 In - Compound 1 significantly reduced the tumor uptake of 111 In - labeled Compound - 15 ( ​ ). Only the kidney showed non - specific uptake of 111 In - Compound 1, and this uptake was not blocked by co - administration of an excess of non - radioactive 115 In - Compound 1, indicating that urinary excretion was 111 the major route of elimination of In - Compound 1.

[0768] The examples and embodiments described herein are for illustrative purposes only, and various modifications or variations known to those skilled in the art will be included within the spirit and scope of this application and the scope of the appended claims.

Claims

1. A compound having the structure of formula (I) or a pharmaceutically acceptable salt thereof: Wherein: A is -N(H)- or -O-; R a is hydrogen or a C1-C6 alkyl group; R 2 is hydrogen or a C1-C6 alkyl group; R 6 is chlorine or -C(=O)NH2; L is -L 1 -L 2 -; L 1 is an optional spacer group; and L 2 is an optional linking group; wherein L 1 or L 2 exists at least one; and R d is an effector molecule moiety that includes a chemotherapeutic agent.

2. The compound or a pharmaceutically acceptable salt thereof according to claim 1, wherein A is N(H)-.

3. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 1-2, wherein R 2 is hydrogen.

4. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 1-3, wherein R a is hydrogen or methyl.

5. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 1-4, wherein R a is hydrogen.

6. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 1-5, wherein R 6 is chlorine.

7. The compound according to any one of claims 1-6 or a pharmaceutically acceptable salt thereof, wherein L 2 is present and is -(L 2a ) w -L 2b - or -L 2c -; Each L 2a is independently selected from natural or unnatural amino acids, wherein any free amine in the amino acid is optionally and independently substituted by -CH3; L 2b is absent or -N(R 10 )(unsubstituted or substituted benzyl)-OC(=O)-; wherein the substituted benzyl is substituted by -C(=O)NHR 12 or monosaccharide; Each R 10 is independently selected from hydrogen and C1-C6 alkyl; Each R 12 is independently selected from hydrogen, C4-C 20 polyethylene glycol, and unsubstituted or substituted C1-C6 alkyl, wherein the substituted C1-C6 alkyl is substituted by -NHR 13 , -C(=O)NHR 13 or -NHC(=O)R 13 ; Each R 13 is independently selected from hydrogen, C4-C 20 polyethylene glycol and C4-C 20 polyethylene glycol-NH2; Or when R 13 is present and at least one free carboxyl group of the amino acid of L 2a is present, then R 13 and the free carboxyl group of the amino acid of L 2a form a ring together; w is 1, 2, 3, 4, 5 or 6; and Each L 2c is N-maleimidomethyl-cyclohexane-1-carbonyl (MCC) or -S-.

8. The compound or a pharmaceutically acceptable salt thereof according to claim 7, wherein L 2b is selected from:

9. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 7-8, wherein the compound has the structure of formula (Ia):

10. The compound according to any one of claims 7-9 or a pharmaceutically acceptable salt thereof, wherein each L 2a is independently selected from natural or unnatural amino acids, any free amine of the amino acid being optionally independently substituted by -CH3, and the natural or unnatural amino acid being selected from alanine (Ala), Ala(SO3H), 3-(1-piperidinyl)alanine, cyclohexylalanine, arginine (Arg), asparagine (Asn), aspartic acid (Asp), cysteine (Cys), glutamine (Gln), glutamic acid (Glu), glycine (Gly), leucine (Leu), lysine (Lys), methionine (Met), phenylalanine (Phe), norleucine, proline (Pro), serine (Ser), 3-homoserine, tyrosine (Tyr), Tyr(SO3H), valine (Val), citrulline, β-alanine, β3-homoserine, β3-homolysine, and β3-homoglutamic acid.

11. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 7-10, wherein each L 2a is independently selected from natural or unnatural amino acids, any free amine of the amino acid is optionally independently substituted by -CH3, and the natural or unnatural amino acids are selected from alanine (Ala), Ala(SO3H), arginine (Arg), asparagine (Asn), aspartic acid (Asp), cysteine (Cys), glutamine (Gln), glutamic acid (Glu), glycine (Gly), leucine (Leu), lysine (Lys), methionine (Met), phenylalanine (Phe), serine (Ser), valine (Val) and citrulline.

12. The compound according to any one of claims 7-11 or a pharmaceutically acceptable salt thereof, wherein L 2 is -(L 2a ) w -L 2b -; and -(L 2a ) w - is valine-citrulline, valine-alanine, methionine-valine-lysine, glycine-phenylalanine-glycine-glycine, tyrosine-arginine-valine, arginine-valine, and phenylalanine-lysine.

13. The compound or a pharmaceutically acceptable salt thereof according to claim 7, wherein the compound has the structure of formula (Ib):

14. The compound or a pharmaceutically acceptable salt thereof according to claim 1, wherein L 2 is absent or is:

15. The compound or a pharmaceutically acceptable salt thereof according to claim 1, wherein L 2 is absent or is:

16. The compound according to any one of claims 1-10 or a pharmaceutically acceptable salt thereof, wherein L 1 is present and is -X 2 -L 3 -L 4 -; X 2 is -C(=O)(CH2) p -, -(CH2) p -, -C(=O)CH(CH2SO3H)NHC(=O)- or -(X 2a ) p -; Each X 2a is independently selected from natural or unnatural amino acids, wherein any free amine in the amino acid is optionally and independently substituted with -CH3; p is 0, 1, 2, 3, 4, 5 or 6; L 3 is absent or unsubstituted or substituted C1-C 10 alkylene, unsubstituted or substituted C1-C 10 heteroalkylene, C4-C 20 polyethylene glycol or -(X 3 CH2CH2) t -; Each X 3 is independently selected from O and NR 10 ; Each t is independently 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12; L 4 Absent or -L 4a -(CH2) u -L 4b -(CH2) u -L 4c -; L 4a is absent or is -O-, -NR 10 -, -NR 10 C(=O)-, -C(=O)NR 10 - or -C(=O)-; L 4b An N-containing 5- to 10-membered heterocycloalkylene which is absent or unsubstituted or substituted; any free amine of the N-containing 5- to 10-membered heterocycloalkylene is optionally and independently substituted by -CH2CO2H; L 4c is absent or is -O-, -NR 10 -, -NR 10 C(=O)-, -C(=O)NR 10 -, -C(=O)NR 10 (CH2) u O(CH2) u C(=O)-, CH(CH2SO3H)C(=O)NR 10 (CH2) u O(CH2) u C(=O)-, -C(=O)-, -CH(=N)-, -CH(=N-NH)-, -CCH3(=N)-, -CCH3(=N-NH)-, -C(=O)-(C1-C6 alkylene)-, -C(=O)NR 10 -(C1-C6 alkylene)-, -NR 10 C(=O)-(C1-C6 alkylene)-, -NR 10 -(C1-C6 alkylene)- or C1-C6 alkylene-; Each u is independently 0, 1, 2, 3, 4, 5 or 6; and Each R 10 is independently selected from hydrogen and C1-C6 alkyl.

17. The compound or a pharmaceutically acceptable salt thereof according to claim 11, wherein: X 2 is -C(=O)(CH2) p -; p is 0, 1, 2, 3 or 4; L 3 is unsubstituted or substituted C1-C 10 heteroalkylene, C4-C 20 polyethylene glycol or -(X 3 CH2CH2) t -; Each X 3 is independently selected from O and NR 10 ; and Each t is independently 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12.

18. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 10-11, wherein: L 4 Does not exist or is -L 4a -; and L 4a is -NR 10 -, -NR 10 C(=O)-, -C(=O)NR 10 - or -C(=O)-.

19. The compound or a pharmaceutically acceptable salt thereof according to claim 10, wherein L 1 is absent or is:

20. The compound or a pharmaceutically acceptable salt thereof according to claim 10, wherein L 1 is absent or is:

21. The compound or a pharmaceutically acceptable salt thereof according to claim 1, wherein L is:

22. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 1-16, wherein R d is:

23. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 1-17, wherein R d is:

24. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 1-15, wherein -L-R d is:

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

26. A compound having the structure of formula (A) or a pharmaceutically acceptable salt thereof: Wherein: A is -N(H)- or -O-; X 1 is -N(R a )-, -O-, -C(=O)-, -C(=O)N(R a )-, -S(=O)-, -(CH2)C(R a )=N-O-(CH2)-; R a is hydrogen or a C1-C6 alkyl group; R b is hydrogen or a C1-C6 alkyl group; Or when R a and R b both exist, then R a and R b together with the intervening atoms to which they are attached form piperidine or pyrrolidine; q is 1, 2 or 3; R 1 is hydrogen; R 2 is hydrogen or a C1-C6 alkyl group; R 3 is hydrogen, -OR 8 , -N(R 8 )2, -CN, halogen, C1-C6 alkyl or C1-C6 fluoroalkyl; or R 2 and R 3 form morpholine together with the intervening atoms to which they are attached; Each R 4 and R 5 are independently hydrogen, halogen, C1-C6 alkyl, C1-C6 fluoroalkyl, substituted or unsubstituted C1-C6 heteroalkyl, -CN, -N(R 8 )2 or -OR 8 ; m is 1, 2 or 3; R c is Each R 6 and R 7 independently is hydrogen, halogen, C1-C4 alkyl, C1-C4 fluoroalkyl, C2-C4 alkenyl, C2-C4 alkynyl, substituted or unsubstituted C1-C6 heteroalkyl, substituted or unsubstituted phenyl, substituted or unsubstituted C3-C6 cycloalkyl, -CN, -OR 8 , -CO2R 8 , -C(=O)N(R 8 )2, -N(R 8 )2, -NR 8 C(=O)R 9 , -NR 8 C(=O)OR 9 , -SR 8 , -S(=O)R 9 , -SO2R 9 or -SO2N(R 8 )2; n is 1, 2 or 3; Each R 8 is independently hydrogen, C1-C4 alkyl, C1-C4 fluoroalkyl, substituted or unsubstituted C1-C4 heteroalkyl; Each R 9 independently is C1-C4 alkyl, C1-C4 fluoroalkyl, substituted or unsubstituted C1-C4 heteroalkyl; L is -L 1 -L 2 -; L 1 is an optional spacer group; L 2 is an optional linking group; wherein at least one of L 1 or L 2 exists; R d is an effector molecule moiety that includes a chelating moiety or a radionuclide complex thereof.

27. A compound having the structure of formula (A) or a pharmaceutically acceptable salt thereof: Wherein: A is -N(H)- or -O-; X 1 is -N(R a )-, -O-, -C(=O)-, -C(=O)N(R a )-, -S(=O)-, -(CH2)C(R a )=N-O-(CH2)-; R a is hydrogen or a C1-C6 alkyl group; R b is hydrogen or a C1-C6 alkyl group; Or when R a and R b both exist, then R a and R b together with the intervening atoms to which they are attached form piperidine or pyrrolidine; q is 1, 2 or 3; R 1 is hydrogen; R 2 is hydrogen or a C1-C6 alkyl group; R 3 is hydrogen, -OR 8 , -N(R 8 )2, -CN, halogen, C1-C6 alkyl or C1-C6 fluoroalkyl; or R 2 and R 3 form morpholine together with the intervening atoms to which they are attached; Each R 4 and R 5 independently is hydrogen, halogen, C1-C6 alkyl, C1-C6 fluoroalkyl, substituted or unsubstituted C1-C6 heteroalkyl, -CN, -N(R 8 )2 or -OR 8 ; m is 1, 2 or 3; R c is Each R 6 and R 7 are independently hydrogen, halogen, C1-C4 alkyl, C1-C4 fluoroalkyl, C2-C4 alkenyl, C2-C4 alkynyl, substituted or unsubstituted C1-C6 heteroalkyl, substituted or unsubstituted phenyl, substituted or unsubstituted C3-C6 cycloalkyl, -CN, -OR 8 , -CO2R 8 , -C(=O)N(R 8 )2, -N(R 8 )2, -NR 8 C(=O)R 9 , -NR 8 C(=O)OR 9 , -SR 8 , -S(=O)R 9 , -SO2R 9 or -SO2N(R 8 )2; n is 1, 2 or 3; Each R 8 independently is hydrogen, C1-C4 alkyl, C1-C4 fluoroalkyl, substituted or unsubstituted C1-C4 heteroalkyl; Each R 9 is independently a C1-C4 alkyl group, a C1-C4 fluoroalkyl group, a substituted or unsubstituted C1-C4 heteroalkyl group; L is -L 1 -L 2 -; L 1 is an optional spacer group; L 2 is an optional linking group; wherein at least one of L 1 or L 2 exists; R d is an effector moiety that includes a chelating moiety or a radionuclide complex thereof.

28. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 26-27, wherein R 2 is hydrogen; R 3 is hydrogen, -OH or -OCH3; or R 2 and R 3 together with the intervening atoms to which they are attached form morpholine.

29. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 26 - 28, wherein each R 4 and R 5 is independently hydrogen, F, Cl, Br, C1 - C4 alkyl, C1 - C4 fluoroalkyl, -CN, -N(R 7 )2 or -OR 7 .

30. A compound or a pharmaceutically acceptable salt thereof according to any one of claims 26-29, wherein each R 4 and R 5 is independently hydrogen, F, Cl, Br, -CH3, -CH2F, -CHF2, -CF3, -CN, -NH2, NHCH3, -N(CH3)2, -OH, -OCH3 or -OCF3.

31. A compound or a pharmaceutically acceptable salt thereof according to any one of claims 26 - 30, wherein A is -N(H)-; R a is hydrogen, -CH3 or -CH2CH3; R b is hydrogen, -CH3 or -CH2CH3; or when R a and R b are both present, then R a and R b together with the intervening atoms to which they are attached form a piperidine; and R 1 is hydrogen, -CH3 or -CH2CH3.

32. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 26-31, wherein the compound has the structure of formula (II), 33. A compound or a pharmaceutically acceptable salt thereof according to any one of claims 26 - 32, wherein is 34. A compound or a pharmaceutically acceptable salt thereof according to any one of claims 26-33, wherein each R 6 and R 7 is independently hydrogen, F, Cl, Br, -CH3, -CH2CH3, -CO2H, -CO2CH3, -CO2CH2CH3, -CH2F, -CHF2, -CF3, -CH=CH2, -C≡CH, -CN, -OH, -OCH3, -OCH2CH3, -OCF3, -C(=O)NH2, -C(=O)NHCH3, -C(=O)N(CH3)2, -NH2, -NHCH3, -N(CH3)2, -NHC(=O)CH3, -NCH3C(=O)CH3, -SO2CH3, -SO2NH2, -SO2NHCH3, or -SO2N(CH3)2.

35. A compound or a pharmaceutically acceptable salt thereof according to any one of claims 26 - 34, wherein is 36. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 26-35, wherein R d is or a radionuclide complex thereof.

37. A compound according to any one of claims 26 - 35 or a pharmaceutically acceptable salt thereof, wherein -L-R d is: -CH2CH2NH-R d 、-C(=O)CH2NH-R d 、-C(=O)CH2CH2NH-R d 、-CH2CH2(OCH2CH2)2NH-R d 、-CH2CH2(OCH2CH2)3NH-R d 、-CH2CH2(OCH2CH2)4NH-R d 、-CH2CH2(OCH2CH2)5NH-R d 、-CH2CH2(OCH2CH2)6NH-R d 、-CH2CH2(OCH2CH2)7NH-R d 、-CH2CH2(OCH2CH2)8NH-R d 、-C(=O)CH2CH2(OCH2CH2)2NH-R d 、-C(=O)CH2CH2(OCH2CH2)3NH-R d 、-C(=O)CH2CH2(OCH2CH2)4NH-R d 、-C(=O)CH2CH2(OCH2CH2)5NH-R d 、-C(=O)CH2CH2(OCH2CH2)6NH-R d 、-C(=O)CH2CH2(OCH2CH2)7NH-R d or -C(=O)CH2CH2(OCH2CH2)8NH-R d ; R d is or its radionuclide complex.

38. A compound or a pharmaceutically acceptable salt thereof according to any one of claims 26-35, wherein -L 1 -R d is: -(PEG2)NH-R d 、-(PEG3)NH-R d 、-(PEG4)NH-R d 、-(PEG5)NH-R d 、-(PEG6)NH-R d 、-(PEG7)NH-R d 、-(PEG8)NH-R d 、-C(=O)(PEG2)NH-R d 、-C(=O)(PEG3)NH-R d 、-C(=O)(PEG4)NH-R d 、-C(=O)(PEG5)NH-R d 、-C(=O)(PEG6)NH-R d 、-C(=O)(PEG7)NH-R d or -C(=O)(PEG8)NH-R d ; R d is or a radionuclide complex thereof.

39. A compound or a pharmaceutically acceptable salt thereof according to any one of claims 26-35, wherein -L 1 -R d is: or a radionuclide complex thereof.

40. A compound or a pharmaceutically acceptable salt thereof according to any one of claims 26 - 39, wherein the radionuclide of the radionuclide complex is copper - 64( 64 Cu), copper - 67( 67 Cu), indium - 111( 111 In), indium - 115( 115 In), gallium - 67( 67 Ga), gallium - 68( 68 Ga), actinium - 225( 225 Ac), lutetium - 175( 175 Lu), lutetium - 177( 177 Lu) or lead - 212( 212 Pb).

41. A compound or a pharmaceutically acceptable salt thereof according to any one of claims 26 - 39, wherein the radionuclide of the radionuclide complex is indium-111 ( 111 In), indium-115 ( 115 In), gallium-67 ( 67 Ga), gallium-68 ( 68 Ga), actinium-225 ( 225 Ac), lutetium-175 ( 175 Lu) or lutetium-177 ( 177 Lu).

42. A compound or a pharmaceutically acceptable salt thereof, consisting of:

43. A pharmaceutical composition comprising the compound or a pharmaceutically acceptable salt thereof according to any one of claims 1-42 and at least one pharmaceutically acceptable excipient.

44. A method for treating cancer, comprising administering to a mammal suffering from cancer an effective amount of the compound or a pharmaceutically acceptable salt thereof according to any one of claims 1-42.

45. A method for treating a tumor, comprising administering to a mammal suffering from a tumor an effective amount of the compound or a pharmaceutically acceptable salt thereof according to any one of claims 1-42.

46. The method according to any one of claims 44-45, wherein the mammal suffers from anal cancer, bladder cancer, bowel cancer, brain cancer, breast cancer, colon cancer, colorectal cancer, endometrial cancer, esophageal cancer, gallbladder cancer, gastric cancer, heart cancer, kidney cancer, lung cancer, liver cancer, melanoma, uterine cancer, lymphoma, ovarian cancer, pancreatic cancer, prostate cancer, thymic cancer, pheochromocytoma, medullary thyroid cancer, or head and neck cancer.

47. The method according to any one of claims 44-45, wherein the mammal suffers from endocrine cancer.

48. The method according to claim 47, wherein the endocrine cancer includes adrenal tumor, neuroendocrine tumor, parathyroid tumor, pituitary tumor or thyroid tumor.

49. The method according to any one of claims 44-45, wherein the mammal suffers from a neuroendocrine tumor.

50. The method according to any one of claims 44-45, wherein the mammal suffers from a somatostatin receptor-positive gastroenteropancreatic neuroendocrine tumor (GEP-NET).

51. A method for targeted delivery of a chemotherapeutic agent to a tumor in a mammal, comprising administering to the mammal suffering from a tumor a compound according to any one of claims 1-42 or a pharmaceutically acceptable salt thereof.

52. A method for killing a tumor overexpressing somatostatin subtype-2 receptor (SST2R) in a mammal, comprising administering to the mammal a compound according to any one of claims 1-42 or a pharmaceutically acceptable salt thereof.

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