Method for treating IGF-1R-related cancers using insulin-like growth factor 1 receptor ligands conjugated with cytotoxic agents.

Abstract

The subject matter described herein relates to a method for treating insulin-like growth factor 1 receptor (IGF-1R) related cancer in a subject, comprising administering a conjugate comprising an IGF-1R ligand, or a part or variant thereof, and a cytotoxic agent to the subject.

Description

【Technical Field】 【0001】 (Cross - Reference to Related Applications) This application claims the benefit of priority to U.S. Provisional Patent Application No. 63 / 505,345, filed May 31, 2023, and U.S. Provisional Patent Application No. 63 / 550,459, filed Feb. 6, 2024, which are hereby incorporated by reference in their entirety. 【0002】 The subject matter of the present disclosure generally relates to methods of treating cancer by administering an IGF - 1R ligand conjugated to a cytotoxic agent, in particular. 【0003】 (Reference to Sequence Listing) The sequence listing described in the file named 614073SEQLIST.xml is 16.7 kilobytes, was created on May 23, 2024, and is hereby incorporated by reference in its entirety. 【Background Art】 【0004】 The insulin - like growth factor - 1 receptor (IGF - 1R) is widely involved in the regulation of both normal immunity and autoimmune diseases. IGF - 1 is a peptide of 70 amino acid residues that has 40% identity with pro - insulin. (Daughaday, W. H., et al., 1989, Endocrine Revs. 10:68). Insulin and IGF - 1 have some cross - reactivity with each other's receptors. (Soos, M. A., et al., 1993, Biochem. J. 290:419). IGF - 1 is secreted into the circulation by the liver and stimulates the growth of many cell types. IGF - 1 is also produced by many cell types throughout the body, including many cancers, due to autocrine and paracrine effects. IGF - 1 production is stimulated by growth hormone. (Stewart, C. H., et al., 1996, Physiol. Revs. 76:1005, Yakar, S., et al., 2002, Endocrine 19:239). 【0005】 IGF-1R is often found to be expressed at higher levels in adult cancer cells than in normal cells of the same tissue type. Elevated IGF-1R activity promotes cancer cell proliferation, migration, and invasion, and is associated with tumor metastasis, treatment resistance, poor prognosis, and shortened survival in patients with cancer. Furthermore, epidemiological studies have reported a positive association between circulating IGF-1 levels and various primary cancers, including breast cancer, colorectal cancer, and prostate cancer. A series of studies have shown that high levels of IGF-1 are associated with an increased risk of tumors, including prostate cancer, premenopausal and postmenopausal breast cancer, lung cancer, thyroid cancer, and colorectal cancer (Ma et al., 1999; Renehan et al., 2004; Shi et al., 2001). 【0006】 There is currently a need for treatments for IGF-1R-related cancers. The subject matter described herein addresses this need. [Overview of the project] 【0007】 In certain embodiments, the subject matter described herein relates to a method for treating insulin-like growth factor 1 receptor (IGF-1R) related cancer in a subject, comprising administering a conjugate comprising an IGF-1R ligand, or a part or variant thereof, and a cytotoxic agent to the subject. 【0008】 In certain embodiments, the subject matter described herein relates to a method for treating IGF-1R-related cancer in a subject, comprising administering to the subject a conjugate comprising an IGF-1R ligand, or a portion or variant thereof, and a cytotoxic agent, wherein the IGF-1R-related cancer is selected from the group consisting of head and neck cancer, triple-negative breast cancer, bladder cancer, gastrointestinal stromal tumor, adenoid cystic carcinoma, lung cancer, colorectal cancer, prostate cancer, pancreatic cancer, liver cancer, esophageal cancer, ovarian cancer, kidney cancer, and gastric cancer. 【0009】 In certain embodiments, the subject matter described herein relates to a method for treating IGF-1R-related cancer in a subject, comprising administering to the subject a conjugate comprising an IGF-1R ligand, or a portion or variant thereof, and a cytotoxic agent, wherein the IGF-1R-related cancer is selected from the group consisting of head and neck cancer, triple-negative breast cancer, bladder cancer, gastrointestinal stromal tumor, and adenoid cystic carcinoma. 【0010】 In certain embodiments, the subject matter described herein relates to a method for treating IGF-1R-related cancer in a subject, comprising administering to the subject a conjugate comprising an IGF-1R ligand, or a portion or variant thereof, and a cytotoxic agent, wherein the IGF-1R ligand, or a portion or variant thereof, comprises wild-type insulin-like growth factor 1 (IGF-1) (SEQ ID NO: 3), wild-type insulin (SEQ ID NO: 10 and SEQ ID NO: 11), wild-type insulin-like growth factor 2 (IGF-2) (SEQ ID NO: 12), a variant of wild-type IGF-1 (SEQ ID NO: 3), a variant of wild-type insulin (SEQ ID NO: 10 and SEQ ID NO: 11), or a variant of wild-type IGF-2 (SEQ ID NO: 12). 【0011】 In certain embodiments, the subject matter described herein relates to a method for treating IGF-1R-related cancer in a subject, comprising administering to the subject a conjugate comprising an IGF-1R ligand, or a portion or variant thereof, and a cytotoxic agent, wherein the IGF-1R ligand, or a portion or variant thereof, comprises a variant of wild-type insulin-like growth factor 1 (IGF-1) (SEQ ID NO: 3). 【0012】 In certain embodiments, the subject matter described herein relates to a method for treating IGF-1R-related cancer in a subject, comprising administering to the subject a conjugate comprising an IGF-1R ligand, or a portion or variant thereof, and a cytotoxic agent, wherein the IGF-1R ligand, or a portion or variant thereof, comprises SEQ ID NO: 2. 【0013】 In certain embodiments, the subject matter described herein relates to a method for treating IGF-1R-related cancer in a subject, comprising administering a conjugate comprising an IGF-1R ligand, or a portion or variant thereof, and a cytotoxic agent to the subject, wherein the cytotoxic agent is a chemotherapeutic agent. 【0014】 In certain embodiments, the subject matter described herein relates to a method for treating IGF-1R-related cancer in a subject, comprising administering to the subject a conjugate comprising an IGF-1R ligand, or a portion or variant thereof, and a cytotoxic agent, wherein the cytotoxic agent is methotrexate. 【0015】 In certain embodiments, the subject matter described herein relates to a method for treating IGF-1R-related cancer in a subject, comprising administering a conjugate comprising an IGF-1R ligand, or a portion or variant thereof, and a cytotoxic agent to the subject, wherein the cytotoxic agent comprises a toxin. 【0016】 In certain embodiments, the subject matter described herein relates to a method for treating IGF-1R-associated cancer in a subject, comprising administering to the subject a conjugate comprising an IGF-1R ligand, or a portion or variant thereof, and a cytotoxic agent, wherein the cytotoxic agent comprises Clostridium perfringens enterotoxin, diphtheria toxin, lysine chain A, Pseudomonas exotoxin, A-chain toxin, ribosome inactivating protein, α-sarcin, aspergiline, or ribonuclease. 【0017】 In certain embodiments, the subject matter described herein is a method for treating IGF-1R-related cancer in a subject, comprising administering to the subject a conjugate comprising an IGF-1R ligand, or a portion or variant thereof, and a cytotoxic agent, wherein the IGF-1R ligand, or a portion or variant thereof, comprises SEQ ID NO: 2, the cytotoxic agent is methotrexate, the methotrexate is covalently bound to the lysine of SEQ ID NO: 2, and the IGF-1R-related cancer is selected from the group consisting of head and neck cancer, triple-negative breast cancer, bladder cancer, gastrointestinal stromal tumor, adenoid cystic carcinoma, lung cancer, colorectal cancer, prostate cancer, pancreatic cancer, liver cancer, esophageal cancer, ovarian cancer, kidney cancer, and gastric cancer. 【0018】 These and other embodiments are described in full below. [Brief explanation of the drawing] 【0019】 [Figure 1] Figures 1A and 1C show the dose-response curves and IC50s of the conjugates described herein for head and neck cancer cell lines, FaDU(A) and SCC-25(C). Figures 1B and 1D show the IGF-1R expression levels in FaDU(B) and SCC-25(D). [Figure 2] Figures 2A and 2C show the dose-response curves and IC50 of the conjugates described herein for the triple-negative breast cancer cell lines BT-20(A) and HCC1143(C). Figures 2B and 2D show the IGF-1R expression levels in BT-20(B) and HCC1143(D). [Figure 3] Figures 3A and 3C show the dose-response curves and IC50 of the conjugates described herein for bladder cancer cell lines, 5637(A) and T-24(C). Figures 3B and 3D show the IGF-1R expression levels in 5637(B) and T-24(D). [Figure 4] Figures 4A to 4C show the dose-response curves and IC50s of the conjugates described herein for lung cancer cell lines A549 (A), NCI-H2122 (B), and NCI-H526 (C). [Figure 5] Figures 5A-5D show the dose response curves and IC50 of the conjugates described herein against colorectal cancer cell lines, COLO 205 (A) and HT-29 (B), and prostate cancer cell lines, VCaP (C) and DU 145 (D). [Figure 6] Figures 6A-6D show the dose response curves and IC50 of the conjugates described herein against pancreatic cancer cell lines, Capan-2 (A) and PANC-1 (B), and liver cancer cell lines, Hep G2 (C) and Huh-7 (D). [Figure 7] Figures 7A-7D show the dose response curves and IC50 of the conjugates described herein against esophageal cancer cell lines, TE-1 (A) and KYSE-70 (B), and ovarian cancer cell lines, OVCAR-8 (C) and Caov-3 (D). [Figure 8] Figures 8A-8D show the dose response curves and IC50 of the conjugates described herein against renal cancer cell lines, 786-O (A) and Caki-1 (B), and gastric cancer cell lines, MKN74 (C) and NUGC-4 (D). **DETAILED DESCRIPTION** 【0020】 The subject matter described herein relates to a method of treating insulin-like growth factor 1 receptor (IGF-1R)-related cancers using a targeted therapy against IGF-1R that includes IGF-1 or a variant thereof in combination with a cytotoxic payload. Attempts to inhibit IGF-1R with naked antibodies or small molecules without a payload have shown some clinical activity in the past, but to date, no resulting approved therapies have been available. 【0021】 A variety of invasive cancers with unaddressed needs have established associations with the IGF-1R pathway, and gene mutations and / or high expression of IGF-1R that activate the IGF-1R pathway are observed. Unfortunately, these are often correlated with poor prognosis. A specific conjugate known as LX-101 demonstrated good tolerability and efficacy as a monotherapy in a previous Phase 1 trial in adult cancer patients with advanced and previously treated cancer. However, further research, including additional cancer species, is needed to improve treatment for patients in need. This specification discloses research supporting the clinical development of the method described herein to address unaddressed needs in the treatment of cancers including head and neck cancer, triple-negative breast cancer, bladder cancer, gastrointestinal stromal tumors, and adenoid cystic carcinoma, in addition to demonstrating that the conjugate complex has potent antitumor activity against various cancer cell lines associated with IGF-1R. 【0022】 Herein, the subject matter of this disclosure will be described more fully. However, a person skilled in the art will, by benefiting from the teachings disclosed herein, come up with many variations and other embodiments of the disclosures herein. Therefore, it should be understood that the subject matter of this disclosure is not limited to the specific embodiments disclosed, and that modifications and other embodiments are intended to be included in the appended claims. In other words, the subject matter described herein encompasses all substitutes, modifications, and equivalents. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by a person skilled in the art. All publications, patent applications, patents, and other references referenced herein are incorporated in their entirety by reference. If one or more of the incorporated documents, patents, and similar materials differ from or contradict this application, including but not limited to defined terms, use of terms, and described techniques, this application shall control. 【0023】 I. Definition As used herein, patient, or subject, is any mammal having IGF-1R-related cancer. As used herein, the term “mammal” includes, but is not limited to, humans, mice, rats, guinea pigs, monkeys, dogs, cats, horses, cattle, pigs, and sheep. In certain embodiments, patient, or subject, is a human adult or child. In certain embodiments, the method involves treating a human subject (i.e., a pediatric subject) who is approximately 21 years of age or under 21 years of age at the time of diagnosis or treatment. The term “pediatric” can be further divided into various subgroups, including neonates (from birth to the first 28 days of life), infants (29 days to under 2 years of age), children (2 years to under 12 years of age), and adolescents (12 years to 21 years of age (excluding their 22nd birthday)). 【0024】 As used herein, the term “conjugate” refers to a molecule comprising an IGF-1R ligand, a part thereof, or a variant thereof, and a cytotoxic agent. 【0025】 As used herein, the term “insulin-like growth factor 1 receptor (IGF-1R)-related cancer” refers to cancers affecting subjects that have an etiology involving IGF-1R overexpression or in which genetic modification implies IGF-1R signaling pathways such as IGF-1R pathway activation, and which are primarily, but not always, found in adult human subjects. Such cancers primarily affect adult subjects and are known to those skilled in the art. 【0026】 As used herein, the term “cytotoxic agent” means any agent that, when administered according to the methods described herein, can prevent, delay, reduce, and / or reverse the activity, severity, and / or progression of a disease. Any suitable cytotoxic agent that results in cell death may be used in methods for treating conjugates and IGF-1R-associated cancers. 【0027】 As used herein, the terms “residue” or “residue of” a chemical moiety or compound means a chemical moiety or compound that is bonded to a molecule, thereby, through the bond, at least one covalent bond displaces at least one atom of the original chemical moiety or compound, resulting in a chemical moiety or compound residue within the molecule. 【0028】 As used herein, a subject is “refractory” to prior treatment if it has failed to achieve a response to therapy to which the treatment is determined to be ineffective, for example, if it has failed to reach a clinical endpoint including any of the following: response, extension of the duration of response, extension of disease-free survival, extension of relapse-free survival, and progression-free survival. 【0029】 As used herein, “and / or” means and encompasses one or more possible combinations of the related enumerated items, as well as the absence of any combination as interpreted as an alternative (or). 【0030】 As used herein, the term “approximately” means that when referring to a measurable value such as the amount of a compound or drug, such as the current subject, dose, time, or temperature, it includes a variation of ±20%, ±10%, ±5%, ±1%, ±0.5%, or ±0.1% of the specified amount. 【0031】 When used herein, the conditional language used herein, such as in particular "can," "could," "might," "may," and "for example," is generally intended to convey that a particular embodiment includes certain features, elements, and / or steps, while other embodiments do not, unless otherwise stated or understood in the context in which they are used. Therefore, such conditional language is generally not intended to imply that features, elements, and / or steps are required in any way to one or more embodiments, or that one or more embodiments necessarily include logic for determining, with or without author input or instruction, whether these features, elements, and / or steps are included in or implemented in any particular embodiment. The terms "comprising," "including," and "having" are synonymous and used in a comprehensive, open manner, without excluding additional elements, features, actions, behaviors, etc. Furthermore, the term "or" is used in its inclusive sense (rather than its exclusive sense), for example, when used to connect a list of elements, meaning one, some, or all of the elements in the list. 【0032】 Additional definitions of terms may be provided below. 【0033】 II. Treatment Methods In certain embodiments, the subject matter described herein relates to a method for treating insulin-like growth factor 1 receptor (IGF-1R) related cancer in a subject, comprising administering a conjugate comprising an IGF-1R ligand, or a portion thereof, or a variant thereof, and a cytotoxic agent to the subject. 【0034】 In certain embodiments, the patient is an adult patient approximately 21 years of age or older. 【0035】 In certain embodiments, the subjects are children under approximately 21 years of age. In certain embodiments, patients are those born within the first 28 days, from 29 days to under 2 years of age, from 2 years to under 12 years of age, or from 12 years to 21 years of age (excluding their 21st birthday). In certain embodiments, patients are those born up to 28 days of age, from 29 days to under 1 year of age, from 1 month to under 4 months of age, from 3 months to under 7 months of age, from 6 months to under 1 year of age, from 1 year to under 2 years of age, from 2 years to under 3 years of age, from 2 years to under 7 years of age, from 3 years to under 5 years of age, from 5 years to under 10 years of age, from 6 years to under 13 years of age, from 10 years to under 15 years of age, or from 15 years to under 21 years of age. 【0036】 In certain embodiments, the method may include treating the subject requiring treatment and performing a morphological diagnosis before administering the conjugate. The method may further include performing molecular testing before administering the conjugate. In certain embodiments, the method includes performing both morphological diagnosis and molecular testing before administering the conjugate. 【0037】 In certain embodiments, IGF-1R is overexpressed in tumor cells of IGF-1R-associated cancer. In certain embodiments, IGF-1R is overexpressed on tumor cells compared to non-tumor cells. In other embodiments, overexpression of IGF-1R in tumor cells of the IGF-1R-associated cancer results in a poor prognosis. In certain embodiments, overexpression of IGF-1R on tumor cells is measured by flow cytometry or immunohistochemistry. 【0038】 In certain embodiments, IGF-1R-associated cancers have one or more genetic modifications that activate the IGF-1R signaling pathway. In certain embodiments, the genetic modifications are mutations, gene fusions, gene amplifications, or translocations. 【0039】 In certain embodiments, IGF-1R-related cancers are selected from the group consisting of head and neck cancers, triple-negative breast cancers, bladder cancers, gastrointestinal stromal tumors, adenoid cystic carcinomas, lung cancers, colorectal cancers, prostate cancers, pancreatic cancers, liver cancers, esophageal cancers, ovarian cancers, kidney cancers, and gastric cancers. In certain embodiments, IGF-1R-related cancers are selected from the group consisting of head and neck cancers, triple-negative breast cancers, bladder cancers, gastrointestinal stromal tumors, and adenoid cystic carcinomas. In certain embodiments, IGF-1R-related cancers are selected from the group consisting of head and neck cancers, triple-negative breast cancers, and bladder cancers. 【0040】 In some embodiments, IGF-1R-related cancers are head and neck cancers, or squamous cell carcinomas (HNSCCs) of the head and neck region. HNSCCs are classified based on the presence or absence of HPV. Patients with HPV, or HPV(+), associated HNSCCs are generally younger, respond well to chemotherapy and radiation, and have a better overall prognosis than patients with HPV(-) disease. Of the approximately 50,000 patients diagnosed with HNSCC in the United States each year, an estimated 30% have HPV(-) disease. Treatment for HPV(-) HNSCCs includes chemotherapy, radiation therapy, and surgery. The most frequently used systemic therapies include pembrolizumab, 5-FU, and nivolumab. Historically, these treatments have resulted in a 45% 5-year overall survival rate for HPV(-) cases and approximately 80% for HPV(+) associated cases. Despite these efforts, clinical trials using new treatment methods are still considered the best option for treating HPV(-)HNSCC when surgery is not an option, according to the National Comprehensive Cancer Network guidelines. Other targeted therapies currently being investigated include tipifarnib, FTase inhibitors, the HGF inhibitor ficratuzumab, and the EGFR inhibitor SCT200. 【0041】 In some embodiments, IGF-1R-related cancer is myoinvasive bladder cancer. Approximately 80,000 new cases of bladder cancer are diagnosed annually in the United States. These cancers range from non-invasive, slowly progressing tumors to highly aggressive, rapidly growing malignancies. One key defining characteristic of bladder cancer is whether the tumor has invaded the muscles surrounding the bladder. Cancer confined to the muscular wall of the bladder is considered non-myoinvasive bladder cancer. These patients account for about 70% of cases and respond well to current treatment options, although recurrence is common (the literature indicates a range of 30–65%). The other 30% of cases are cancers where the tumor has invaded the muscles surrounding the bladder or metastasized beyond the muscles; this is called myoinvasive bladder cancer. Historically, patients with myoinvasive bladder cancer have a poor response to available therapies and a poor prognosis. The current standard treatment for myoinvasive bladder cancer is multi-agent chemotherapy followed by surgery. This treatment regimen has proven insufficient, as even with successful surgery, the 5-year overall survival rate remains low at 40%. However, due to the heterogeneity of the disease, there are many other treatment options for muscular bladder cancer that target various potential weaknesses of the malignant tumor, with varying success rates. Importantly, many of the recent regulatory approvals for muscular bladder cancer are based on single-arm, non-randomized studies using overall response rate endpoints. Four checkpoint inhibitors—Opdivo, Keytruda, Bavencio, and Imfinzi—were approved in a second-line setting after clinical trials of approximately 200 patients each and ORRs ranging from 13 to 29% (some are undergoing larger trials to obtain full approval and / or remove the restrictions on their labeling for second-line diseases). In addition, three targeted therapies—Varvasa, Troderbi, and Padsev—were approved in a third-line setting after clinical trials of approximately 100 patients each and ORRs ranging from 28 to 44%. 【0042】 In some embodiments, IGF-1R-associated cancer is triple-negative breast cancer. Triple-negative breast cancer is a type of breast cancer in which there is little to no expression of estrogen receptors, progesterone, and human epidermal growth factor receptor-2. Compared to other breast cancer subtypes, patients with triple-negative breast cancer have a poorer prognosis and reduced survival rate due to a more invasive disease, limited treatment options, and a higher frequency of recurrence and metastasis. Approximately 15–20% of all breast cancers are triple-negative, with approximately 45,000 new cases diagnosed annually in the United States. Advances have been made in treatment options for specific patients with triple-negative breast cancer over the past decade. The primary treatment option remains chemotherapy, including taxanes and anthracyclines. Patients with BRCA mutations account for approximately 10–15% of patients and have shown some success with targeted therapies, Lymphalza and Tarzena, with response rates of 52% and 50%, respectively, and median PFS of 7.0 months and 8.6 months. In 2020, Trodelvy received FDA accelerated approval in previously treated metastatic triple-negative breast cancer based on an ORR of 33.3% and a median response time of 7.7 months in a single-arm study of 108 patients. Other approvals in specific subsets of triple-negative breast cancer include Keytruda for patients expressing programmed death ligand 1, and Rozlietrek or Vitrakvy for patients with NTRK mutations. 【0043】 In some embodiments, IGF-1R-related cancers are gastrointestinal stromal tumors. Gastrointestinal stromal tumors are mesenchymal neoplasms originating from the digestive system. Approximately 6,000 patients are diagnosed with gastrointestinal stromal tumors annually, and 90% of these patients carry gain-of-function mutations in either the KIT or PDGFRA proto-oncogene. The remaining 10% of patients who do not have either of these mutations are considered to have wild-type disease and consist almost entirely of pediatric patients. Numerous therapies targeting common mutations in gastrointestinal stromal tumors are approved and available, including imatinib, sunitinib, and other chemotherapy. However, wild-type disease has historically been far more difficult to treat. Importantly, however, patients with disease without these mutations (i.e., wild-type disease) are typically pediatric patients and generally do not respond to approved therapies, including chemotherapy, thus constituting a major unmet medical need. 【0044】 In some embodiments, IGF-1R-related cancer is adenoid cystic carcinoma. Adenoid cystic carcinoma is a malignant tumor primarily of the secretory glands. It is driven by a gene translocation resulting in the MYB-NFIB fusion protein. In the United States, approximately 1,700 patients are diagnosed with adenoid cystic carcinoma each year, with 5,000 and 10,500 prevalence pools at 5 years and 16 years, respectively. Although the disease progresses slowly, it is constant, making tumor recurrence the biggest challenge with limited current treatment options. This is indicated by the low overall survival rate. The 5-year, 10-year, and 15-year overall survival rates for adenoid cystic carcinoma are 60%, 50%, and 29%, respectively, due to the high incidence of metastasis and recurrence. Currently, the standard treatment for adenoid cystic carcinoma is surgery, sometimes combined with radiotherapy. While this surgery helps extend survival, more than half of patients have metastatic or recurrent disease, including 30% of patients with distant metastases after recurrence, making surgery an unsuitable option and severely limiting treatment. There are no approved systemic therapies for adenoid cystic carcinoma. The most commonly used chemotherapy combinations, cisplatin, doxorubicin, and cyclophosphamide, as well as cisplatin and vinorelbine, yield overall response rates of 18%–31%. Many targeted therapies have been tested in adenoid cystic carcinoma with unfavorable results and have not been approved. 【0045】 In other embodiments, IGF-1R-related cancers include lung cancer, colorectal cancer, prostate cancer, ovarian cancer, gastric cancer, liver cancer, pancreatic cancer, kidney cancer, or esophageal cancer. 【0046】 In this embodiment or any of the embodiments of the present invention, the conjugate may include a conjugate in which an IGF-1R ligand and a cytotoxic agent are chemically linked together, either directly or via a chemical linker. In other embodiments, the conjugate is a recombinant in which the conjugate is expressed as a single polypeptide. If the conjugate is a recombinant conjugate, the translated conjugate preferably includes a toxin, or a portion thereof, or a variant linked to the IGF-1R ligand via a peptide bond. In certain embodiments, the conjugate is a fusion protein described in whole in U.S. Patent No. 9,675,671, which is incorporated herein by reference. 【0047】 Methods for preparing the conjugates described herein are known in the art. The nucleotide sequences encoding the IGF-1R ligand can be generated by standard recombinant DNA techniques or protein synthesis techniques, cloned into a suitable expression vector using standard molecular biology techniques, expressed in bacterial, insect, or mammalian cells, and purified by any method known in the art for protein purification. The conjugates of the present invention, comprising the IGF-1R ligand and chemotherapeutic agents, can be prepared by standard chemical and protein conjugate techniques, as described in U.S. Patents 7,811,982, 9,675,671, and 9,801,923, which are incorporated herein by reference. The conjugates described herein, comprising the IGF-1R ligand and toxins, can be prepared as fusion proteins by standard recombinant DNA techniques, as described in U.S. Patent 8,017,102, which is incorporated herein by reference. 【0048】 IGF-1R is a heterotetramer consisting of two extracellular ligand-binding subunits and two transmembrane subunits that possess kinase activity to mediate signal transduction. The native ligands for IGF-1R are IGF-1, IGF-2, and insulin. IGF-1R has the highest affinity for IGF-1, followed by affinity for IGF-2, and can bind to insulin with an affinity 50–100 times lower. IGF-1R can also form a hybrid receptor by dimerization with the insulin receptor. See Hakuno et al. J Mol Endocrinol. 61(1):T69-T86(2018). 【0049】 In certain embodiments, the IGF-1R ligand in the conjugate of the present invention comprises wild-type IGF-1 (SEQ ID NO: 3), wild-type insulin (SEQ ID NO: 10, and SEQ ID NO: 11), and mature insulin consists of two chains connected by a disulfide bond between chain A, corresponding to SEQ ID NO: 10, and chain B, corresponding to SEQ ID NO: 11, and thus comprises the enumeration of the two SEQ ID NOs, or wild-type IGF-2 (SEQ ID NO: 12). In other embodiments, the IGF-1R ligand in the conjugate comprises a variant of wild-type IGF-1 (SEQ ID NO: 3), a variant of wild-type insulin (SEQ ID NO: 10, and SEQ ID NO: 11), or a variant of wild-type IGF-2 (SEQ ID NO: 12). In certain embodiments, a variant of wild-type IGF-1 is at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identical to IGF-1 (SEQ ID NO: 3), a variant of wild-type insulin is at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identical to insulin (SEQ ID NO: 10 and SEQ ID NO: 11), or a variant of wild-type IGF-2 is at least 70%, 75%, 80%, 85%, 90%, 95%, or 99% identical to IGF-2 (SEQ ID NO: 12). 【0050】 In certain embodiments, the IGF-1R ligand in the conjugate includes a variant of IGF-1 with reduced binding affinity to IGFBP compared to wild-type IGF-1 (SEQ ID NO: 3), or a variant of IGF-2 with reduced binding affinity to IGFBP compared to wild-type IGF-2 (SEQ ID NO: 12). IGFBPs belong to a family of at least six proteins that bind to IGF-1 and IGF-2 with high affinity. IGFBPs bind to the majority of circulating IGF, increasing its half-life, regulating its bioavailability, and generally inhibiting its ability to bind to IGF receptors. See Am J Physiol Endocrinol Metab., 278(6):E967-76 (2000) and Allard et al. Front Endocrinol (Lausanne). 9;9:117(2018). Therefore, variants of IGF-1 or IGF-2 with reduced binding to IGFBP have higher bioactivity in vivo. 【0051】 IGF-1 variants with reduced binding affinity to IGFBP are known in the art and include IGF132 (US Patent No. 4,876,242) (where the first 17 amino acids of the insulin B chain (SEQ ID NO: 11) are replaced with the first 16 amino acids of human IGF-1 (SEQ ID NO: 3)), R3-IGF-1 (SEQ ID NO: 6) (where the glutamic acid at position 3 of natural human IGF-1 (SEQ ID NO: 3) is replaced with arginine), and des(1-3)IGF-1 (SEQ ID NO: 7) (where the first three amino acids of human IGF-1 (SEQ ID NO: 3) are missing). R3-IGF-1 and des(1-3)IGF-1 are described in Francis et al., J Mol Endocrinol. 8(3):213-23 (1992). In some embodiments, the conjugate includes IGF132 (SEQ ID NO: 4), R3-IGF-1 (SEQ ID NO: 6), or des(1-3)-IGF-1 (SEQ ID NO: 7). 【0052】 In certain embodiments, a variant of IGF-1 has a higher affinity for IGF-1R than wild-type IGF-1 (SEQ ID NO: 3), or a variant of IGF-2 has a higher affinity for IGF-1R than wild-type IGF-2 (SEQ ID NO: 12). 【0053】 In some embodiments, the IGF-1R ligand in the conjugate includes 765IGF (SEQ ID NO: 2), long-R3-IGF-1 (SEQ ID NO: 5), long-IGF-1 (SEQ ID NO: 8), or long-G3-IGF-1 (SEQ ID NO: 9). 765IGF, long-R3-IGF-1, long-IGF-1, and long-G3-IGF-1 have an N-terminal leader sequence that facilitates protein purification and provides a site for conjugation of cytotoxic agents, as described above. 765IGF (SEQ ID NO: 2) contains SEQ ID NO: 1, followed by R3-IGF-1 (SEQ ID NO: 6); long-R3-IGF-1 (SEQ ID NO: 5) contains the first 11 amino acids of methionyl porcine growth hormone, followed by Val-Asn dipeptide, followed by R3-IGF-1 (SEQ ID NO: 6); long-IGF-1 (SEQ ID NO: 8) contains the first 11 amino acids of methionyl porcine growth hormone, followed by Val-Asn dipeptide, followed by human IGF-1 (SEQ ID NO: 3); and long-G3-IGF-1 contains the first 11 amino acids of methionyl porcine growth hormone, followed by Val-Asn dipeptide, followed by a variant of human IGF-1 in which the glutamic acid at position 3 of natural human IGF-1 (SEQ ID NO: 3) is substituted with glycine. 【0054】 In certain embodiments, the IGF-1R ligand includes wild-type insulin-like growth factor 1, wild-type insulin, or wild-type insulin-like growth factor 2 (IGF-2). In certain embodiments, wild-type insulin-like growth factor 1 (IGF-1) includes SEQ ID NO: 3, wild-type insulin includes SEQ ID NO: 10 or SEQ ID NO: 11, and wild-type insulin-like growth factor 2 (IGF-2) includes SEQ ID NO: 12. 【0055】 In certain embodiments, the IGF-1R ligand includes a variant of wild-type IGF-1, a variant of wild-type insulin, or a variant of wild-type IGF-2. In certain embodiments, the wild-type IGF-1 variant is at least 90% identical to SEQ ID NO: 3, the wild-type insulin variant is at least 90% identical to SEQ ID NO: 10 or 11, or the wild-type IGF-2 variant is at least 90% identical to SEQ ID NO: 12.12. In certain embodiments, (i) the wild-type IGF-1 variant has a reduced binding affinity to insulin-like growth factor-binding protein (IGFBP) compared to wild-type IGF-1, or the wild-type IGF-2 variant has a reduced binding affinity to IGFBP compared to wild-type IGF-2, and / or (ii) the wild-type IGF-1 variant has an increased affinity to IGF-1R compared to wild-type IGF-1, or the wild-type IGF-2 variant has an increased affinity to IGF-1R compared to wild-type IGF-2. 【0056】 In certain embodiments, the IGF-1R ligand, or a portion thereof, or a variant thereof, includes a leader sequence. In certain embodiments, the leader sequence includes SEQ ID NO: 1. 【0057】 In certain embodiments, the IGF-1R ligand includes 765IGF (SEQ ID NO: 2), IGF-132 (SEQ ID NO: 4), long-R3-IGF-1 (SEQ ID NO: 5), R3-IGF-1 (SEQ ID NO: 6), des(1-3)-IGF-1 (SEQ ID NO: 7), long-IGF-1 (SEQ ID NO: 8), or long-G3-IGF-1 (SEQ ID NO: 9). 【0058】 In certain embodiments, the IGF-1R ligand includes 765IGF (SEQ ID NO: 2). 【0059】 In certain embodiments, the IGF-1R ligand, or a portion or variant thereof, is covalently bound to a cytotoxic agent. 【0060】 In certain embodiments, the cytotoxic agent includes a chemotherapeutic agent. In certain embodiments, the chemotherapeutic agent is amsacrin, azacitidine, bleomycin, busulfan, capecitabine, carboplatin, carmustine, chlorambucil, cisplatin, cladribine, cyclophosphamide, cytarabine, dactinomycin, daunorubicin, decarbazine, docetaxel, doxorubicin, epirubicin, estramustine, etoposide, fulocuridine, fludarabine, fluorouracil, gemcitabine, hexamethylmelamine, idarubicin, ifosfamide, These are irinotecan, lomustine, mechloretamine, melphalan, mercaptopurine, methotrexate, mitomycin C, mitotane, mitoxantrone, oxaliplatin, paclitaxel, pemetrexed, pentostatin, plicamycin, procarbazine, laritrexed, semustine, streptozocin, temozolamide, teniposide, thioguanine, thiotepa, topotecan, trimitrexate, barrubicin, vincristine, vinblastine, vindestine, or vinorelbine. In certain embodiments, the chemotherapeutic agent is methotrexate. In certain embodiments, the chemotherapeutic agent is covalently bonded to a lysine residue in the leader sequence. In certain embodiments, the chemotherapeutic agent is one or more methotrexate residues covalently bonded to any available lysine residue in the leader sequence. 【0061】 In certain embodiments, the conjugate comprises two or more cytotoxic agents bound to the IGF-1R ligand. In certain embodiments, the conjugate may comprise 1 to 12 cytotoxic agents, or 6 to 10 cytotoxic agents, or about 8 cytotoxic agents. In certain embodiments, the conjugate may comprise 1 to 12 covalently bound cytotoxic agents, or 6 to 10 covalently bound cytotoxic agents, or about 8 covalently bound cytotoxic agents. In certain embodiments, the chemotherapeutic agent is covalently bound to any available position on the IGF-1R ligand. In certain embodiments, the chemotherapeutic agent is covalently bound to any available lysine residue. In certain embodiments, the chemotherapeutic agent, if present, is covalently bound to any available lysine in the leader sequence. 【0062】 The leader sequence can incorporate tags such as polyhistidine tags, facilitate protein purification, and provide sites for the conjugate of cytotoxic agents. In certain embodiments, the leader sequence includes Sequence ID No. 1. 【0063】 In some embodiments, the cytotoxic agent includes a toxin. In certain embodiments, the toxin includes Clostridium perfringens enterotoxin, diphtheria toxin, lysine chain A, Pseudomonas exotoxin, A-chain toxin, ribosome inactivating protein, α-sarcin, aspergiline, or ribonuclease. In certain embodiments, the toxin includes Clostridium perfringens enterotoxin, or a part thereof, or a variant thereof. In certain embodiments, the toxin includes SEQ ID NO: 14 or SEQ ID NO: 15. In certain embodiments, the toxin includes diphtheria toxin, or a part thereof, or a variant thereof. In certain embodiments, the toxin includes SEQ ID NO: 13 or SEQ ID NO: 16. 【0064】 In certain embodiments, the subject matter described herein relates to a method for treating IGF-1R-related cancer in a subject, comprising administering to the subject a conjugate comprising an IGF-1R ligand, or a portion or variant thereof, and a cytotoxic agent, wherein the IGF-1R ligand, or a portion or variant thereof, comprises SEQ ID NO: 2, the cytotoxic agent is methotrexate, the methotrexate is covalently bound to the lysine of SEQ ID NO: 2, and the IGF-1R-related cancer is selected from the group consisting of head and neck cancer, triple-negative breast cancer, and bladder cancer. In certain embodiments of these embodiments, the conjugate is LX-101 as described above (the conjugate described above, wherein the IGF-1R ligand is SEQ ID NO: 2, the cytotoxic agent is methotrexate, and at least 6 to a maximum of 10, or at least 6 to a maximum of 9, or at least 7 to a maximum of 9, or at least 8 to a maximum of 9 methotrexate residues are each bound to the IGF-1R ligand. Methotrexate can be covalently bound to the IGF-1R ligand, particularly to the lysine residue of SEQ ID NO: 2. The average number of methotrexate residues per SEQ ID NO: 2 is 8). In certain embodiments, the subject matter described herein is a method for treating IGF-1R-related cancer in a subject, comprising administering LX-101 to the subject, wherein the IGF-1R-related cancer is a head and neck cancer. In certain embodiments, the subject matter described herein relates to a method for treating IGF-1R-related cancer in a subject, comprising administering LX-101 to the subject, wherein the IGF-1R-related cancer is triple-negative breast cancer. In certain embodiments, the subject matter described herein relates to a method for treating IGF-1R-related cancer in a subject, comprising administering LX-101 to the subject, wherein the IGF-1R-related cancer is bladder cancer. In certain embodiments, the number of methotrexate residues per conjugate is 6, 7, 8, 9, or 10. In certain embodiments, the average number of methotrexate residues per conjugate in the composition is 6, 7, 8, 9, or 10. 【0065】 In certain embodiments, the subjects are (i) not previously treated for IGF-1R-related cancer, (ii) previously treated for IGF-1R-related cancer, (iii) relapsed from previous treatment for IGF-1R-related cancer, (iv) refractory to previous treatment for IGF-1R-related cancer, or (v) susceptible to adverse reactions from other treatments for IGF-1R-related cancer. Therefore, in certain embodiments, IGF-1R-related cancer relapses if the patient has relapsed after previous treatment. In certain embodiments, IGF-1R-related cancer is stage II, III, or IV. In certain embodiments, IGF-1R-related cancer progresses to stage II, III, or IV during or after initial therapy. 【0066】 In certain embodiments, the treatment results in a reduction of tumor cell growth in the subject. In certain embodiments, the reduction is caused by the killing of IGF-1R expressing cells. 【0067】 Table 1 provides a list of specific sequences referenced herein. [Table 1-1] [Table 1-2] 【0068】 In certain embodiments, the methods described herein are part of a combination therapy. In particular, the methods described herein can be used alone or in combination with standard treatment options for each type of cancer. Generally, standard treatment options include surgery, systemic chemotherapy (either pre- or post-operative), and / or radiotherapy. 【0069】 In certain embodiments, subjects having IGF-1R-related cancer treated according to the method described herein have not previously received treatment for IGF-1R-related cancer. 【0070】 In certain embodiments, subjects having IGF-1R-related cancer treated according to the method described herein have previously received treatment for IGF-1R-related cancer. 【0071】 In certain embodiments, the subjects are those whose cancer has recurred after prior treatment for IGF-1R-related cancer. 【0072】 In certain embodiments, the subjects were refractory to previous treatments for IGF-1R-related cancer. 【0073】 In all embodiments, the conjugate is administered at a dose and frequency appropriate to the subject, and IGF-1R-related cancers are treated at the physician's discretion. 【0074】 In certain embodiments, the conjugate is administered in doses of approximately 0.05, 0.10, 0.20, 0.40, 0.80, 1.0, 1.5, 1.6, 2.0, 2.5, 3.0, 3.5, 4.0, 4.5, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5, 9.0, 9.5, or 10.0 μequivalents / kg body weight, or approximately 0.05-10.0, 0.1-8.0, 0.2-4.0, 0.3-3, 0.4-2.5 The drug is administered in dose ranges of 0.05-0.5, 0.5-1.0, 1.0-1.5, 1.5-2.0, 2.0-2.5, 2.5-3.0, 3.0-3.5, 3.5-4.0, 4.0-4.5, 4.5-5.0, 5.0-5.5, 5.5-6.0, 6.0-6.5, 6.5-7.0, 7.0-7.5, 7.5-8.0, 8.0-8.5, 8.5-9.0, 9.0-9.5, or 9.5-10.0 μEq / kg body weight. In certain embodiments, the conjugate is administered at approximately 0.05, 0.10, 0.20, 0.40, 0.80, 1.6, or 2.5 μEq / kg body weight per kg of body weight. A μequivalent is equivalent to the μmol of a chemotherapeutic agent group conjugated to an IGF-1R ligand. In certain embodiments, the conjugate is approximately 0.05, 0.10, 0.15, 0.20, 0.25, 0.30, 0.35, 0.40, 0.45, 0.50, 0.55, 0.60, 0.65, 0.70, 0.75, 0.80, 0.85, 0.90, 0.95, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4 4.5, 4.6, 4.7, 4.8, 4.9, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7. 6, 7.7, 7.8, 7.9, 8.0, 8.1, 8.2, 8.3, 8.4, 8.5, 8.6, 8.7, 8.8, 8.9, 9.0, 9.1, 9.2, 9.3, 9.4, 9.5, 9.6, 9.7, 9.8, 9.9, 10.0, 10.1, 10.2, 10.3, 10.4, 10.5, 10.6, 10.7, 10.8, 10.9, 11.0, 11.1, 11.2, 11.3, 11.4, 11.5, 11.6, 11.7, 11.8, 11.9, 12.0, 12.1, 12.2, 12.3, 12.4, 12.5, 12.6, 12.7, 12.8, 12.9, 13.0, 13.1, 13.2, 13.3, 13.4, 13.5, 13.6, 13.7, 13.8, 13. In doses of 9, 14.0, 14.1, 14.2, 14.3, 14.4, 14.5, 14.6, 14.7, 14.8, 14.9, 15.0, 15.1, 15.2, 15.3, 15.4, 15.5, 15.6, 15.7, 15.8, 15.9, 16.0, 16.1, 16.2, 16.3, 16.4, or 16.5 mg / kg body weight, or approximately 0.05-0.5, 0.5-1 0.0, 1.0~1.5, 1.5~2.0, 2.0~2.5, 2.5~3.0, 3.0~3.5, 3.5~4.0, 4.0~4.5, 4.5~5.0, 5.0~5.5, 5.5~6.0, 6.0~6.5, 6.5~7.0, 7.0~7.5, 7.5~8.0, 8.0~8.5, 8.5~9.0, 9.0~9.5, 9.5~10.0, 10.0~10.5, 10.5~11 It is administered in dose ranges of 0.0, 11.0-11.5, 11.5-12.0, 12.0-12.5, 12.5-13.0, 13.0-13.5, 13.5-14.0, 14.0-14.5, 14.5-15.0, 15.0-15.5, 15.5-16.0, or 16.0-16.5 mg / kg body weight, where mg refers to the amount of IGF-1R ligand present in the conjugate. 【0075】 In certain embodiments, the conjugate is administered daily, every other day, every three days, every four days, every five days, every six days, once a week, once every two weeks, once every three weeks, once every four weeks, once a month, once every two months, or once every three months. 【0076】 In certain embodiments, the conjugate may be administered at a lower dose and / or frequency than the cytotoxic agent would be effective when administered as a monotherapy. 【0077】 In some embodiments, the conjugate is administered at the maximum tolerated dose (MTD). As used herein, "MTD" refers to the maximum dose of the drug that an individual patient can tolerate, as determined by a physician. In other words, the side effects in a given patient can determine the MTD. 【0078】 In certain embodiments, the method does not cause significant or unacceptable hyperglycemia in the subject. Hyperglycemia is another term for hyperglycemia, which can occur when there is insufficient insulin in the body or when the body is unable to adequately utilize insulin. Unacceptable hyperglycemia refers to adverse effects of grade 3 or higher as determined by the attending physician, and / or adverse effects that cannot be controlled with diabetes medications and lead to discontinuation of treatment with the conjugate of the present invention. 【0079】 In certain embodiments, the method does not cause significant or unacceptable hyperglycemia in the subject. Hyperglycemia is another term for hyperglycemia, which can occur when there is insufficient insulin in the body or when the body is unable to adequately utilize insulin. Unacceptable hyperglycemia refers to adverse effects of grade 3 or higher as determined by the attending physician, and / or adverse effects that cannot be controlled with diabetes medications and lead to discontinuation of treatment with the conjugate of the present invention. 【0080】 The conjugates described herein can be formulated in pharmaceutical compositions for use in the methods described herein. In some embodiments, the pharmaceutical composition comprises an effective amount of the conjugate and a pharmaceutically acceptable carrier or vehicle. Such a pharmaceutical composition can be formulated to be suitable for administration to a subject and may be in any form that allows the composition to be administered to a subject. 【0081】 The materials used in the preparation of the pharmaceutical composition may be nontoxic in the amount used. It will be apparent to those skilled in the art that the optimal dose of the active ingredient in the pharmaceutical composition depends on various factors. These factors include, but are not limited to, the type of subject (e.g., human), the overall health status of the subject, the type of condition the subject has, the use of the composition as part of a multi-drug regimen, the specific form of the composition of the present invention, and the method of administration. The pharmaceutical composition contains an effective amount of the composition so as to yield a suitable dose. 【0082】 The term "carrier" refers to a diluent, adjuvant, or excipient on which the composition of the present invention is administered. Optional auxiliaries, stabilizers, thickeners, lubricants, and colorants may be used. In one embodiment, when administered to a subject, the composition of the present invention and the pharmaceutically acceptable carrier are sterilized. Water may be the carrier when the composition of the present invention is administered intravenously. Saline solution, as well as aqueous solutions of dextrose and glycerol, may also be used as liquid carriers, particularly in injectable solutions. The composition may also contain a small amount of pH buffering agent if desired. 【0083】 Any of the liquid pharmaceutical compositions, solutions, suspensions, or other similar forms of the present invention may also include one or more of the following: a sterile diluent such as water for injection; a fixative oil such as synthetic mono or diglycerides that can function as a solvent or suspension medium, such as polyethylene glycol, glycerin, cyclodextrin, propylene glycol, or other solvents; an antimicrobial agent such as benzyl alcohol or methylparaben; an antioxidant such as ascorbic acid or sodium sulfite; a chelating agent such as ethylenediaminetetraacetic acid; a buffer such as acetate, citrate, or phosphate; a pH adjuster such as hydrochloric acid; and a tonic adjuster such as sodium chloride or dextrose. Parenteral compositions may be enclosed in ampoules, disposable syringes, or multi-dose vials made of glass, plastic, or other materials. In some embodiments, physiological saline is an adjuvant. Injectable compositions may be sterile. 【0084】 This composition may take the form of a solution, suspension, tablet, pill, pellet, capsule, liquid-containing capsule, powder, sustained-release formulation, suppository, emulsion, aerosol, spray, suspension, or any other form suitable for use. Examples of suitable pharmaceutical carriers are described in Remington's Pharmaceutical Sciences by EW Martin. 【0085】 In some embodiments, the composition is formulated according to routine procedures as a pharmaceutical composition adapted for intravenous administration to human subjects. Typically, the carrier or vehicle for intravenous administration is a sterile isotonic aqueous buffer solution. Optionally, the composition may also include a solubilizer. The composition for intravenous administration may optionally include a local anesthetic, such as a lignocaine, to reduce pain at the injection site. Generally, the components are supplied in unit dosage forms, either separately or mixed together, as dry lyophilized powder or anhydrous concentrate, in sealed containers such as ampoules or sachets indicating the amount of the active agent. When the composition is administered by infusion, it can be dispensed using, for example, an infusion bottle containing sterile pharmaceutical-grade water or saline. When the composition is administered by injection, ampoules of sterile water or saline may be provided so that the components can be mixed before administration. 【0086】 Pharmaceutical compositions can be prepared using methodologies well known in the pharmaceutical art. For example, a composition intended to be administered by injection can be prepared by combining the composition with water to form a solution. Surfactants can be added to facilitate the formation of a homogeneous solution or suspension. Surfactants are conjugates that can interact non-covalently with the composition of the present invention to facilitate its dissolution or homogeneous suspension in an aqueous delivery system. 【0087】 The conjugates described herein may be administered by any convenient route, for example, by infusion or bolus injection, or by absorption through the epithelium or mucosal lining (e.g., oral mucosa, rectum, and intestinal mucosa). Administration may be systemic or topical. Various delivery systems, such as microparticles, microcapsules, and capsules, are known and may be useful for administering the compositions of the present invention. Methods of administration include, but are not limited to, oral and parenteral administration, intradermal, intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal, epidural, sublingual, intracerebral, intraventricular, subarachnoid, vaginal, transdermal, rectal, inhalation, or topical administration to the ear, nose, eye, or skin. The mode of administration is at the discretion of the physician and in part depends on the site of the condition. In certain embodiments, the conjugate is administered intravenously, subcutaneously, or intramuscularly. 【0088】 In certain embodiments, the conjugate is administered parenterally. In certain embodiments, the conjugate is administered intravenously. In certain embodiments, the conjugate is administered by continuous infusion. In certain embodiments, the conjugate is administered by infusions lasting 15 minutes, 20 minutes, 30 minutes, 45 minutes, 1 hour, or 2 hours. 【0089】 In certain embodiments, it may be desirable to administer the conjugate topically to the area requiring treatment. This can be achieved, for example, by injection, by catheter, by suppository, or by implant, for example, by local injection during surgery or by topical application in conjunction with postoperative wound dressings, where the implant is made of porous, non-porous, or gelatinous material, including a membrane such as a cialastic membrane or fiber. In certain embodiments, the conjugate may be injected intraperitoneally. 【0090】 In certain embodiments, the conjugate may be delivered via a controlled emission system. 【0091】 The following examples are provided for illustrative purposes only and not for limiting purposes. [Examples] 【0092】 Example 1 - IGF-1R Expression Assay [Table 2] 【0093】 IGF-1R expression was determined via FACS in the cell lines shown in Table 3. The data results are shown in Figures 1 to 3. The MCF7 cell line was used as a positive control. [Table 3] 【0094】 First, 1,000,000 cells were resuspended in 100 μL of PBS containing 2.5 μg of human Fc block (BD Biosciences) and incubated in the dark at room temperature for 10 minutes to block. For staining, PE CD221 (IGF-1R) antibody (clone 1H7, BD Biosciences) and eFluor780 bio / dead stain (eBiosciences) were added to each sample and incubated in the dark at 4°C for 30 minutes. Isotype controls for each cell line were established using PE mouse IgG1, κ isotype control (BD Biosciences). Next, 2 mL of PBS was added to each sample, the cells were gently resuspended, and the cells were centrifuged at 500 × g for 5 minutes, with the supernatant discarded. The cells were washed twice more in PBS and then resuspended in 200 μL of IC fixation buffer (eBioscience) and incubated in the dark at room temperature for 30 minutes. Cells were washed with 2 mL of PBS and resuspended in 250 μL of PBS for acquisition. Data were acquired using a CytoFLEX S flow cytometer (Beckman Coulter) and analyzed using a Kaluza 2.0 (Beckman Coulter). 【0095】 The obtained data are shown in Figures 1 to 3. The greater fluorescence intensity of cells stained with PE CD221 antibody compared to cells stained with isotype controls indicates that the cell lines FADU (Figure 1B), SCC-25 (Figure 1D), BT-20 (Figure 2B), HCC1143 (Figure 2D), 5637 (Figure 3B), and T-24 (Figure 3D) express IGF-1R on their cell surface. 【0096】 Example 2: Cytotoxicity assay in specific types of cancer cells 【0097】 LX-101-induced cytotoxicity was evaluated for specific types of cancer cells. Cell lines listed in Table 6 were treated with LX-101 at nine different concentrations using 2.5-fold dilutions, and cell viability was assessed. [Table 4] 【0098】 Approximately 7 days prior to treatment, the cryovials of frozen cells were thawed by transferring them to a water bath on dry ice. The contents of each cryovial were slowly transferred to a 15 mL tube containing 10 mL of the culture medium shown in Table 6. The cells were centrifuged at 125 × g for 5 minutes at room temperature. The cell pellet was resuspended in pre-warmed culture medium in a T-25 or T-75 flask and incubated at 37°C with 5% CO2. When the cell culture reached approximately 80% confluence, the cells were subcultured using TrypLE at a subculturing ratio of 1:3 to 1:6. [Table 5] 【0099】 One day prior to LX-101 treatment, cells were collected and counted during logarithmic growth using TrypLE. The number of cells shown in Table 7 were seeded in 100 μL per well of a 96-well plate according to the plate layout shown below. The plates were incubated overnight at 37°C and 5% CO2. [Table 6] [Table 7] [Table 8] 【0100】 A 1 / 10 working stock solution of LX-101 was prepared in 1 mM HCl, with a concentration of 0.4 μEq / ml of the drug in 1.9 mM HCl. Sterile Eppendorf tubes were prepared to contain three times the final well concentration in the culture medium corresponding to each cell line (Table 6). The first dilution was prepared using a 1 / 10 working stock diluted in culture medium. Subsequent 2.5-fold dilutions were prepared using culture medium. When the compound was diluted in culture medium, visual inspection was performed for signs of precipitation. 【0101】 For the vehicle control, a 10 mM HCl stock solution was prepared by diluting HCl in cell culture-grade water, and the pH was confirmed to be between 1 and 3. Next, a working stock solution was prepared with a final concentration of 1.9 mM HCl. In an Eppendorf filter, 75 μL of the working stock solution was added to 425 μL of culture medium to triple the final concentration of 285 μM HCl. 【0102】 As shown in Table 8, 50 μL of a prepared dilution of LX-101 or HCl was added to the appropriate wells to obtain a total volume of 150 μL per well. Unused wells were filled with 150 μL of PBS. [Table 9] 【0103】 After treatment, the plates were incubated at 37°C in 5% CO2. After 4 days, the plates were removed and allowed to equilibrate at room temperature for 30 minutes. A black sticker was placed at the bottom of the plate to block out light. Next, 75 μL of CellTiter-Glo 2.0 reagent (Promega) was added to each well, and the contents were mixed on an orbital shaker for 2 minutes. The plates were incubated at room temperature for a further 10 minutes to stabilize the luminescence signal. The luminescence was recorded using an Envision 2104 multi-label microplate reader (PerkinElmer) with an integration time of 0.25–1 second per well. 【0104】 The LX-101 concentration based on IGF-1 variant protein content was derived by dividing the drug concentration based on methotrexate content by 8, which is the average number of methotrexate groups per IGF-1 protein determined by MALDI-TOF. 50 The values ​​were calculated using GraphPad PRISM software. Survival rate (percentage of control) = (Lum 被験物質 -Lum ブランク対照 ) / (Lum ビヒクル -Lum ブランク対照 ) × 100%. 【0105】 result The cytotoxicity of lot LIR00223 of LX-101 was tested as a reference on MCF7 breast cancer cells. As shown in Table 9, the mean IC of three independent experiments using lot LIR0023. 50 It was found to be 31 nM. [Table 10] 【0106】 Following the same protocol, the cytotoxicity of LX-101 (lot LIR0023) was determined for the cell lines shown in Table 10. LX-101 had an absolute IC50 nM of less than 70 nM in pharyngeal cancer, triple-negative breast cancer, bladder cancer, lung cancer, colorectal cancer, prostate cancer, pancreatic cancer, liver cancer, esophageal cancer, ovarian cancer, kidney cancer, and gastric cancer cell lines.50 It exhibits substantial anticancer properties, based on the concentration of IGF-1R ligand containing eight covalently bound methotrexate molecules. In particular, LX-101 shows high cytotoxicity against pharyngeal cancer cell line FaDU, bladder cancer cell line 5637, prostate cancer cell line DU 145, liver cancer cell line Hep G2, esophageal cancer cell line KYSE-70, and kidney cancer cell line 786-O, with an extremely low absolute IC of less than 10 nM. 50 The value was shown. [Table 11] 【0107】 While efforts have been made to ensure accuracy regarding the numbers used (e.g., quantity, temperature, etc.), some experimental errors and deviations should be taken into consideration. 【0108】 Those skilled in the art will recognize many methods and materials similar to or equivalent to those described herein that may be used in carrying out the subject matter described herein. This disclosure is not limited to the methods and materials described herein. 【0109】 Unless otherwise defined, technical and scientific terms used herein have the same meanings as those generally understood by those skilled in the art to which this subject belongs. 【0110】 Throughout this specification and the claims, unless otherwise required by context, the terms “comprise,” “comprises,” and “comprising” are used in a non-exclusive sense. Embodiments described herein are understood to include embodiments consisting of and / or essentially consisting of embodiments. 【0111】 Where a range of values ​​is provided, unless the context explicitly indicates otherwise, it is understood that each intervening value up to one-tenth of the lower limit unit is included between the upper and lower limits of the range, and any other listed or intervening values ​​within that listed range. The upper and lower limits of any smaller ranges that may independently be included in these smaller ranges are also included and are subject to any specifically excluded limits within the specified range. If the listed range includes one or both limits, it also includes the range that excludes one or both of those limits. 【0112】 Many modifications and other embodiments of the invention described herein will be apparent to those skilled in the art who benefit from the teachings presented in the foregoing description and the accompanying drawings. It should be understood that the subject matter of this disclosure is not limited to the specific embodiments disclosed, and that modifications and other embodiments are intended to be included in the appended claims. Certain terms are used herein, but they are used only in a general and descriptive sense, and not for limiting purposes.

Claims

[Claim 1] A method for treating insulin-like growth factor 1 receptor (IGF-1R) related cancer in a subject, comprising administering to the subject a conjugate comprising an IGF-1R ligand, a part thereof, or a variant thereof, and a cytotoxic agent. [Claim 2] The method according to claim 1, wherein the IGF-1R-related cancer is selected from the group consisting of head and neck cancer, triple-negative breast cancer, bladder cancer, gastrointestinal stromal tumor, adenoid cystic carcinoma, lung cancer, colorectal cancer, prostate cancer, pancreatic cancer, liver cancer, esophageal cancer, ovarian cancer, kidney cancer, and gastric cancer. [Claim 3] The method according to claim 2, wherein the IGF-1R-related cancer is selected from the group consisting of head and neck cancer, triple-negative breast cancer, bladder cancer, gastrointestinal stromal tumor, and adenoid cystic carcinoma. [Claim 4] The method according to claim 3, wherein the IGF-1R-related cancer is selected from the group consisting of head and neck cancer, triple-negative breast cancer, and bladder cancer. [Claim 5] The method according to any one of claims 1 to 4, wherein IGF-1R is overexpressed in tumor cells of the IGF-1R-related cancer. [Claim 6] The method according to any one of claims 1 to 5, wherein the IGF-1R-related cancer has one or more gene changes that activate the IGF-1R signaling pathway. [Claim 7] The method according to claim 6, wherein the gene modification is a mutation, gene fusion, gene amplification, or translocation. [Claim 8] The method according to any one of claims 1 to 7, wherein the IGF-1R ligand comprises wild-type insulin-like growth factor 1, wild-type insulin, or wild-type insulin-like growth factor 2 (IGF-2). [Claim 9] The method according to claim 8, wherein the wild-type insulin-like growth factor 1 (IGF-1) comprises SEQ ID NO: 3, the wild-type insulin comprises SEQ ID NO: 10 or 11, and the wild-type insulin-like growth factor 2 (IGF-2) comprises SEQ ID NO:

12. [Claim 10] The method according to any one of claims 1 to 7, wherein the IGF-1R ligand comprises a variant of wild-type IGF-1, a variant of wild-type insulin, or a variant of wild-type IGF-2. [Claim 11] The method according to claim 10, wherein the wild-type IGF-1 variant is at least 90% identical to SEQ ID NO: 3, the wild-type insulin variant is at least 90% identical to SEQ ID NO: 10 or 11, and the wild-type IGF-2 variant is at least 90% identical to SEQ ID NO:

12. [Claim 12] The method according to claim 10 or 11, wherein (i) the wild-type IGF-1 variant has a reduced binding affinity to insulin-like growth factor-binding protein (IGFBP) compared to wild-type IGF-1, or the wild-type IGF-2 variant has a reduced binding affinity to IGFBP compared to wild-type IGF-2, and / or (ii) the wild-type IGF-1 variant has an increased affinity to IGF-1R compared to wild-type IGF-1, or the wild-type IGF-2 variant has an increased affinity to IGF-1R compared to wild-type IGF-2. [Claim 13] The method according to any one of claims 1 to 11, wherein the IGF-1R ligand, or a part thereof, or a variant thereof, comprises a leader sequence. [Claim 14] The method according to claim 13, wherein the leader array includes sequence number 1. [Claim 15] The method according to any one of claims 1 to 7 and 10 to 14, wherein the IGF-1R ligand includes 765IGF (SEQ ID NO: 2), IGF-132 (SEQ ID NO: 4), long-R3-IGF-1 (SEQ ID NO: 5), R3-IGF-1 (SEQ ID NO: 6)des(1-3)-IGF-1 (SEQ ID NO: 7), long-IGF-1 (SEQ ID NO: 8), or long-G3-IGF-1 (SEQ ID NO: 9). [Claim 16] The method according to claim 15, wherein the IGF-1R ligand comprises 765IGF (SEQ ID NO: 2). [Claim 17] The method according to any one of claims 1 to 16, wherein the IGF-1R ligand, or a part thereof, or a variant thereof, is covalently bound to the cytotoxic agent. [Claim 18] The method according to any one of claims 1 to 17, wherein the cytotoxic agent comprises a chemotherapeutic agent. [Claim 19] The aforementioned chemotherapeutic agents include amsacrin, azacitidine, bleomycin, busulfan, capecitabine, carboplatin, carmustine, chlorambucil, cisplatin, cladribine, cyclophosphamide, cytarabine, dactinomycin, daunorubicin, decarbazine, docetaxel, doxorubicin, epirubicin, estramustine, etoposide, phloxuridine, fludarabine, fluorouracil, gemcitabine, hexamethylmelamine, idarubicin, ifosfamide, irinotecan, and romus. The method according to claim 18, wherein the agent is tin, mechloretamine, melphalan, mercaptopurine, methotrexate, mitomycin C, mitotane, mitoxantrone, oxaliplatin, paclitaxel, pemetrexed, pentostatin, plicamycin, procarbazine, laritrexed, semustine, streptozocin, temozolamide, teniposide, thioguanine, thiotepa, topotecan, trimitrexate, barrubicin, vincristine, vinblastine, vindestine, or vinorelbine. [Claim 20] The method according to claim 19, wherein the chemotherapeutic agent is methotrexate. [Claim 21] The method according to any one of claims 1 to 17, wherein the cytotoxic agent comprises a toxin. [Claim 22] The method according to claim 21, wherein the toxin comprises Clostridium perfringens enterotoxin, diphtheria toxin, lysine chain A, Pseudomonas exotoxin, A-chain toxin, ribosome inactivating protein, α-sarcin, aspergiline, or ribonuclease. [Claim 23] The method according to claim 22, wherein the toxin comprises Clostridium perfringens enterotoxin, a part thereof, or a variant thereof. [Claim 24] The method according to claim 23, wherein the toxin comprises SEQ ID NO: 14 or SEQ ID NO:

15. [Claim 25] The method according to claim 22, wherein the toxin comprises diphtheria toxin, a part thereof, or a variant thereof. [Claim 26] The method according to claim 25, wherein the toxin comprises SEQ ID NO: 13 or SEQ ID NO:

16. [Claim 27] The method according to any one of claims 1 to 26, wherein the subject is (i) not previously treated for the IGF-1R-related cancer, (ii) previously treated for the IGF-1R-related cancer, (iii) relapsed from the previous treatment for the IGF-1R-related cancer, (iv) refractory to the previous treatment for the IGF-1R-related cancer, or (v) susceptible to side effects from other treatments for the IGF-1R-related cancer. [Claim 28] The method according to any one of claims 1 to 27, wherein the conjugate is administered in combination with one or more other therapies. [Claim 29] The method according to claim 28, wherein the one or more other therapies include one or more of surgery, systemic chemotherapy (either preoperative or postoperative), and radiotherapy. [Claim 30] The aforementioned conjugate is administered in doses of approximately 0.05, 0.10, 0.20, 0.40, 0.80, 1.0, 1.5, 1.6, 2.0, 2.5, 3.0, 3.5, 4.0, 4.5, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5, 9.0, 9.5, or 10.0 μequivalents / kg body weight, or approximately 0.05 to 10.0, 0.1 to 8.0, 0.2 to 4.0, 0.3 to 3, 0.4 to 2.5, 0.05 to 0.5, 0.5 to The method according to any one of claims 1 to 29, administered in a dose range of 1.0, 1.0–1.5, 1.5–2.0, 2.0–2.5, 2.5–3.0, 3.0–3.5, 3.5–4.0, 4.0–4.5, 4.5–5.0, 5.0–5.5, 5.5–6.0, 6.0–6.5, 6.5–7.0, 7.0–7.5, 7.5–8.0, 8.0–8.5, 8.5–9.0, 9.0–9.5, or 9.5–10.0 μequivalents / kg body weight. [Claim 31] The method according to claim 30, wherein the conjugate is administered at a dose of approximately 0.05, 0.10, 0.20, 0.40, 0.80, 1.6, or 2.5 μequivalents / kg body weight. [Claim 32] The method according to any one of claims 15 to 20 and 27 to 31, wherein the IGF-1R ligand is SEQ ID NO: 2, the cytotoxic agent is methotrexate, there are 6 to 10 methotrexate molecules for each IGF-1R ligand of SEQ ID NO: 2, and the IGF-1R-related cancer is head and neck cancer. [Claim 33] The method according to any one of claims 15 to 20 and 27 to 31, wherein the IGF-1R ligand is SEQ ID NO: 2, the cytotoxic agent is methotrexate, there are 6 to 10 methotrexate molecules for each IGF-1R ligand of SEQ ID NO: 2, and the IGF-1R-related cancer is triple-negative breast cancer. [Claim 34] The method according to any one of claims 15 to 20 and 27 to 31, wherein the IGF-1R ligand is SEQ ID NO: 2, the cytotoxic agent is methotrexate, there are 6 to 10 methotrexate molecules for each IGF-1R ligand of SEQ ID NO: 2, and the IGF-1R-related cancer is bladder cancer. [Claim 35] The method according to any one of claims 15 to 20 and 27 to 31, wherein the IGF-1R ligand is SEQ ID NO: 2, the cytotoxic agent is methotrexate, there are 6 to 10 methotrexate molecules for each IGF-1R ligand of SEQ ID NO: 2, and the IGF-1R-related cancer is a gastrointestinal stromal tumor. [Claim 36] The method according to any one of claims 15 to 20 and 27 to 31, wherein the IGF-1R ligand is SEQ ID NO: 2, the cytotoxic agent is methotrexate, there are 6 to 10 methotrexate molecules for each IGF-1R ligand of SEQ ID NO: 2, and the IGF-1R-related cancer is adenoid cystic carcinoma. [Claim 37] The method according to any one of claims 1 to 32, wherein the head and neck cancer is HPV-. [Claim 38] The method according to any one of claims 1 to 31 and 35, wherein the gastrointestinal stromal tumor is wild-type. [Claim 39] The method according to claim 1 or 2, wherein the IGF-1R-related cancer is a head and neck cancer. [Claim 40] The method according to claim 1 or 2, wherein the IGF-1R-related cancer is triple-negative breast cancer. [Claim 41] The method according to claim 1 or 2, wherein the IGF-1R-related cancer is bladder cancer. [Claim 42] The method according to claim 1 or 2, wherein the IGF-1R-related cancer is lung cancer. [Claim 43] The method according to claim 1 or 2, wherein the IGF-1R-related cancer is colorectal cancer. [Claim 44] The method according to claim 1 or 2, wherein the IGF-1R-related cancer is prostate cancer. [Claim 45] The method according to claim 1 or 2, wherein the IGF-1R-related cancer is pancreatic cancer. [Claim 46] The method according to claim 1 or 2, wherein the IGF-1R-related cancer is liver cancer. [Claim 47] The method according to claim 1 or 2, wherein the IGF-1R-related cancer is esophageal cancer. [Claim 48] The method according to claim 1 or 2, wherein the IGF-1R-related cancer is ovarian cancer. [Claim 49] The method according to claim 1 or 2, wherein the IGF-1R-related cancer is renal cancer. [Claim 50] The method according to claim 1 or 2, wherein the IGF-1R-related cancer is gastric cancer.

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