WO2024088275A1 - 一种萘酰胺化合物治疗耐药性肿瘤的用途 - Google Patents

一种萘酰胺化合物治疗耐药性肿瘤的用途 Download PDF

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WO2024088275A1
WO2024088275A1 PCT/CN2023/126296 CN2023126296W WO2024088275A1 WO 2024088275 A1 WO2024088275 A1 WO 2024088275A1 CN 2023126296 W CN2023126296 W CN 2023126296W WO 2024088275 A1 WO2024088275 A1 WO 2024088275A1
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drug
resistant
egfr
tumor
compound
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French (fr)
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谢华
段文虎
丁健
詹正生
耿美玉
杨汉煜
吕永聪
冯芳
赵倩
张阳
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中国科学院上海药物研究所
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
    • A61K31/415—1,2-Diazoles
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
    • A61K31/415—1,2-Diazoles
    • A61K31/416—1,2-Diazoles condensed with carbocyclic ring systems, e.g. indazole
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
    • A61K31/4164—1,3-Diazoles
    • A61K31/4188—1,3-Diazoles condensed with other heterocyclic ring systems, e.g. biotin, sorbinil
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents

Definitions

  • the invention belongs to the field of medicine, and specifically relates to the application of a naphthamide compound in preparing a medicine for treating drug-resistant tumors.
  • Gliomas are tumors that occur in the neuroectoderm, including astrocytomas, glioblastomas, medulloblastomas, ependymomas, and choroid papilloma, etc., and gliomas are divided into brain gliomas and spinal cord gliomas according to the site of onset. Glioblastoma is the most common malignant primary brain tumor, accounting for about 57% of all gliomas and 48% of all primary malignant central nervous system tumors.
  • Temozolomide is an imidazole tetrazine-derived alkylating agent. TMZ is rapidly hydrolyzed to 5-(3-methyltriazine-1-yl)imidazole-4-carboxamide (MTIC) at physiological pH, which then affects DNA replication through methylation at the O 6 and N 7 sites of guanine in deoxyribonucleic acid (DNA), thereby exerting a cytotoxic effect. Due to its oral administration and easy penetration of the blood-brain barrier, TMZ was approved by the FDA in 2005 and 2009 for the treatment of patients with astrocytoma and glioblastoma, respectively. It is also the only oral chemotherapy drug for the first-line treatment of glioma.
  • MTIC 5-(3-methyltriazine-1-yl)imidazole-4-carboxamide
  • TMZ-treated patients do not respond to TMZ, which is mainly caused by the overexpression of O 6 -methylguanine methyltransferase (MGMT) and the lack of DNA repair pathways in GBM cells.
  • MGMT can repair the methylation of guanine O 6 caused by TMZ, making tumor cells insensitive to TMZ.
  • the expression of MGMT is mainly affected by the methylation level of its promoter region. The higher the methylation level, the lower the MGMT expression, and the more sensitive it is to TMZ.
  • the MGMT methylation positivity rate in glioma patients is about 40%, and it is even lower in the most malignant glioblastoma. This means that without considering secondary resistance to TMZ treatment, at least half of glioma patients are resistant to TMZ.
  • the clinical recommendation is to use methylation-specific PCR (polymerase chain reaction) combined with immunohistochemistry to determine the expression of MGMT.
  • methylation-specific PCR polymerase chain reaction
  • immunohistochemistry to determine the expression of MGMT.
  • Non-small cell lung cancer (NSCLC), as the main histological type of the disease, accounts for more than 80% of lung cancer patients, with a 5-year survival rate of less than 15%.
  • High expression or abnormal activation of epidermal growth factor receptor (EGFR) is an important target that promotes the deterioration and development of NSCLC.
  • EGFR inhibitors In recent years, with the in-depth study of the pathogenesis of lung cancer, molecular targeted therapy represented by EGFR inhibitors has made breakthrough progress. So far, three generations of EGFR inhibitors have been approved for marketing, bringing huge clinical benefits to patients with EGFR mutant NSCLC, but EGFR inhibitors develop acquired resistance within about one year of use, which greatly limits the clinical application of the drug. Therefore, the development of strategies that can inhibit EGFR inhibitor resistance has become a hot topic in current NSCLC research.
  • the reported EGFR inhibitor resistance mechanisms can be divided into two categories: the first category is EGFR-dependent resistance mechanisms, which are mainly caused by changes in EGFR itself, such as EGFR T790M mutation, which is the main cause of resistance to first-generation EGFR inhibitors (accounting for about 60% of all resistant patients), or EGFR C797S/G , EGFR L798I , EGFR L792H and other point mutations and EGFR T790M loss mediating resistance to third-generation EGFR inhibitors. There is no treatment for tumors with EGFR C797S mutation.
  • the second category is EGFR-independent resistance mechanisms, including MET and HER2 amplification, FGFR1 amplification, PIK3CA mutation, small cell transformation, RAS-MAPK pathway activation and KRAS mutation and amplification.
  • EGFR-independent resistance mechanisms including MET and HER2 amplification, FGFR1 amplification, PIK3CA mutation, small cell transformation, RAS-MAPK pathway activation and KRAS mutation and amplification.
  • Compound (I) is a VEGFR/CSF1R dual-target inhibitor with excellent activity, which can inhibit tumor cell angiogenesis and promote tumor immunity to exert anti-tumor effects.
  • the purpose of the present invention is to provide a use of a naphthamide compound in treating drug-resistant tumors.
  • the first aspect of the present invention provides a use of a compound (I) or a pharmaceutically acceptable salt thereof for preparing a drug for treating drug-resistant tumors;
  • the compound (I) has the following structure:
  • the drug-resistant tumor is selected from the following group: glioma and lung cancer.
  • the drug-resistant tumor is selected from the following group: temozolomide-resistant tumor, EGFR inhibitor-resistant tumor.
  • the drug-resistant tumor is a tumor with high expression of O 6 -methylguanine methyltransferase.
  • the drug-resistant tumor is a glioma with high expression of O 6 -methylguanine methyltransferase.
  • the temozolomide-resistant tumor is a temozolomide-resistant glioma.
  • the temozolomide-resistant tumor is a glioma with high expression of O 6 -methylguanine methyltransferase.
  • the glioma is selected from the group consisting of glioblastoma, astrocytoma, and medulloblastoma.
  • the temozolomide-resistant tumor is temozolomide-resistant medulloblastoma.
  • the EGFR inhibitor-resistant tumor is EGFR inhibitor-resistant lung cancer.
  • the lung cancer is selected from the group consisting of small cell lung cancer, non-small cell lung cancer, and lung adenocarcinoma.
  • the EGFR inhibitor-resistant tumor is a non-small cell tumor resistant to EGFR inhibitors.
  • Cell lung cancer is a non-small cell tumor resistant to EGFR inhibitors.
  • the EGFR inhibitor-resistant tumor is an EGFR mutation tumor.
  • the EGFR mutation is selected from the following group: EGFR T790M mutation and EGFR C797S mutation.
  • the EGFR inhibitor-resistant lung cancer is lung cancer with EGFR T790M mutation or lung cancer with EGFR C797S mutation.
  • the EGFR inhibitor-resistant lung cancer is non-small cell lung cancer with EGFR T790M mutation or non-small cell lung cancer with EGFR C797S mutation.
  • the EGFR inhibitor-resistant tumor is an EGFR-independent EGFR inhibitor-resistant tumor.
  • the EGFR-independent EGFR inhibitor-resistant tumor is a tumor caused by a mechanism selected from the following group: MET and HER2 amplification, FGFR1 amplification, PIK3CA mutation, small cell transformation, RAS-MAPK pathway activation, KRAS mutation and amplification.
  • the EGFR-independent EGFR inhibitor-resistant tumor is EGFR-independent EGFR inhibitor-resistant lung cancer.
  • the EGFR-independent EGFR inhibitor-resistant lung cancer is EGFR-independent EGFR inhibitor-resistant non-small cell lung cancer.
  • the EGFR inhibitor is selected from the following group: ASK120067 (Limertinib), osimertinib (AZD9291) or a salt thereof, ametinib or a salt thereof, vometinib or a salt thereof,
  • the EGFR inhibitor is ASK120067 (Limertinib), osimertinib (AZD9291) and salts thereof.
  • the second aspect of the present invention provides a pharmaceutical composition for treating drug-resistant tumors, the pharmaceutical composition comprising a therapeutically effective amount of compound (I) or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier;
  • the compound (I) has the following structure:
  • the drug-resistant tumor is as described above.
  • the pharmaceutical composition is an oral preparation.
  • a pharmaceutical combination comprising a first pharmaceutically active ingredient and a second pharmaceutically active ingredient in therapeutically effective amounts
  • the first active pharmaceutical ingredient is compound (I) or a pharmaceutically acceptable salt thereof;
  • the compound (I) has the following structure:
  • the second active ingredient of the drug is an anti-tumor drug selected from the following group: temozolomide and EGFR inhibitor.
  • the tumor is selected from the following group: temozolomide-resistant tumors and EGFR inhibitor-resistant tumors.
  • the temozolomide-resistant tumor is as described above.
  • the EGFR inhibitor-resistant tumor is as described above.
  • the EGFR inhibitor is selected from the following group: ASK120067 (Limertinib), osimertinib (AZD9291) or a salt thereof, ametinib or a salt thereof, and vometinib or a salt thereof.
  • the EGFR inhibitor is ASK120067 (Limertinib).
  • the mass ratio of the first pharmaceutically active ingredient to the second pharmaceutically active ingredient is 1-4, preferably 1-3, and preferably 2.
  • the fourth aspect of the present invention provides a use of compound (I) or a pharmaceutically acceptable salt thereof for preparing a drug for enhancing the efficacy of an anti-tumor drug;
  • the compound (I) has the following structure:
  • the anti-tumor drug is selected from the following group: temozolomide and EGFR inhibitor.
  • the tumor is selected from the following group: temozolomide-resistant tumors and EGFR inhibitor-resistant tumors.
  • the temozolomide-resistant tumor is as described above.
  • the EGFR inhibitor-resistant tumor is as described above.
  • the EGFR inhibitor is as described above.
  • the fifth aspect of the present invention provides a pharmaceutical composition comprising a therapeutically effective amount of the drug combination described in the third aspect of the present invention and a pharmaceutically acceptable carrier.
  • the pharmaceutical composition is an oral preparation.
  • the pharmaceutical composition is used to treat drug-resistant tumors.
  • the drug-resistant tumor is as described above.
  • the sixth aspect of the present invention provides a use of the drug combination described in the third aspect of the present invention for preparing a drug for treating drug-resistant tumors.
  • the drug-resistant tumor is as described above.
  • the patient with the drug-resistant tumor is a human.
  • the seventh aspect of the present invention provides a method for treating drug-resistant tumors, comprising the steps of:
  • the compound (I) has the following structure:
  • the drug-resistant tumor is as described above.
  • the anti-tumor drug is selected from the following group: temozolomide and EGFR inhibitors.
  • the EGFR inhibitor is as described above.
  • the method further comprises the following steps before step 1):
  • a genetic testing method is used to determine the patient's MGMT gene expression and/or EGFR gene mutation.
  • the patient is a patient with overexpression of MGMT gene.
  • the patient is a patient with EGFR gene mutation.
  • the patient is a patient with EGFR T790M mutation or a patient with EGFR C797S mutation.
  • the eighth aspect of the present invention provides a method for treating drug-resistant tumors, comprising the steps of:
  • the drug-resistant tumor is as described above.
  • the anti-tumor drug is selected from the following group: temozolomide and EGFR inhibitors.
  • the EGFR inhibitor is as described above.
  • the method further comprises the following steps before step 1):
  • a genetic testing method is used to determine the patient's MGMT gene expression and/or EGFR gene mutation.
  • the patient is a patient with overexpression of MGMT gene.
  • the patient is a patient with EGFR gene mutation.
  • the patient is a patient with EGFR T790M mutation or a patient with EGFR C797S mutation.
  • compound (I) has excellent inhibitory activity against drug-resistant tumors, can significantly inhibit the growth of drug-resistant tumors, especially temozolomide-resistant tumors and EGFR inhibitor-resistant tumors, and is expected to be developed as a drug against drug-resistant tumors.
  • the inventor completed the present invention.
  • pharmaceutically acceptable salt refers to a salt formed between a compound of the present invention and an acid or base that is suitable for use as a drug.
  • Pharmaceutically acceptable salts include inorganic salts and organic salts.
  • “Pharmaceutically acceptable carrier” refers to: one or more compatible solid or liquid fillers or gel substances, which are suitable for human use and must have sufficient purity and sufficiently low toxicity. "Compatibility” here means that the components in the composition can be mixed with the compound of the present invention and with each other without significantly reducing the efficacy of the compound.
  • compositions of the present invention are no particular limitation on the administration of the pharmaceutical composition of the present invention.
  • Representative administration methods include, but are not limited to, oral, intratumoral, rectal, parenteral (intravenous, intramuscular or subcutaneous) and topical administration.
  • the drug of the present invention can be prepared into various clinically acceptable dosage forms, including oral dosage forms, injection dosage forms, local administration dosage forms or external dosage forms, etc.
  • Solid dosage forms such as tablets, dragees, capsules, pills and granules may be prepared with coatings and shells, such as enteric coatings and other materials known in the art. They may contain opacifying agents and the release of the active compound or compounds in such compositions may be delayed in a certain part of the digestive tract. Examples of embedding compositions used are polymeric substances and waxes. If desired, the active compound can also be in microencapsulated form with one or more of the above-mentioned excipients.
  • compositions for parenteral injection may comprise physiologically acceptable sterile aqueous or nonaqueous solutions, dispersions, suspensions or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions.
  • Dosage forms for topical administration of the compounds of the invention include ointments, powders, patches, sprays and inhalants.
  • the active ingredient is mixed under sterile conditions with a physiologically acceptable carrier and any preservatives, buffers, or propellants that may be required.
  • the compounds of the present invention can be administered alone or in combination with other pharmaceutically acceptable compounds (such as anti-tumor drugs).
  • the treatment method of the present invention can be used alone or in combination with other treatment methods or therapeutic drugs.
  • the therapeutically effective amount of the present invention refers to the effective dosage considered in medicine, that is, the amount of active compound is sufficient to significantly improve the condition without causing serious side effects.
  • the daily dosage is usually 0.01 to 2000 mg, preferably 1 to 500 mg. It can be administered as a single dose once a day, can be administered multiple times a day, or can be used at intervals.
  • the specific dosage and frequency of administration should take into account factors such as the route of administration and the patient's health status, which can be determined by a skilled physician based on routine skills.
  • the present invention also provides a method for using the pharmaceutical composition, that is, applying a therapeutically effective amount of the compound of the present invention (I) or a pharmaceutically acceptable salt thereof or the pharmaceutical composition of the present invention to a mammal (such as a human) in need of treatment.
  • the method for using the pharmaceutical composition before applying a therapeutically effective amount of the compound of the present invention (I) or a pharmaceutically acceptable salt thereof to a mammal (such as a human) in need of treatment, also includes a step of genetic testing to determine the MGMT methylation status and/or EGFR gene mutation of the mammal (such as a human) in need of treatment.
  • the MGMT methylation status of mammals can be detected using a mgmt gene methylation detection reagent (such as a manufacturer of gene technology, national medical device registration number 20193400101).
  • a mgmt gene methylation detection reagent such as a manufacturer of gene technology, national medical device registration number 20193400101.
  • the most commonly used methods for detecting EGFR gene mutations include direct sequencing and amplification retardation mutation system (ARMS-PCR).
  • Direct DNA sequencing is currently the most direct and basic method for detecting gene mutations, and known and unknown gene mutations in tumor tissue samples can be detected by reading DNA base sequences.
  • the ARMS-PCR method uses designed specific primers to accurately amplify the mutant sequence by PCR, and uses real-time fluorescence PCR quantitative analysis to highly sensitively detect the EGFR gene in small biopsy samples and blood specimens.
  • EGFR gene mutations in mammals can be detected using an EGFR gene mutation detection kit (such as: Human EGFR Gene Mutation Detection
  • the present invention has the following main advantages:
  • the compound (I) has excellent inhibitory activity against drug-resistant tumors and can significantly inhibit the growth of drug-resistant tumors, especially temozolomide-resistant tumors and EGFR inhibitor-resistant tumors, and is expected to be developed as a drug against drug-resistant tumors;
  • the compound (I) is effective against EGFR T790M mutation, EGFR C797S mutation and EGFR-independent third-generation inhibitor-resistant tumors.
  • control compounds, reagents and raw materials used in the experiments were purchased commercially or prepared by us.
  • test compound (I) preparation was prepared according to the following prescription as shown in Table A:
  • Blank preparation same formulation as Table A, except that it does not contain compound (I)
  • the latent solvent was evaporated and removed during the preparation process, and the obtained test substance and blank preparation did not contain the latent solvent.
  • the preparation containing compound (I) or the blank preparation was diluted with water to the desired concentration or volume.
  • TMZ Prepare the required dose with 0.5% sodium carboxymethylcellulose solution.
  • ASK120067 and AZD9291 Prepare the required dose with water for injection containing 1% Tween 80.
  • D283 and NCI-H1975 were purchased from ATCC (American Type Culture Collection).
  • PC-9-OR and 67R were constructed by the Shanghai Institute of Materia Medica, Chinese Academy of Sciences using conventional methods. For details, see the methods in the literature (Cancer Science 2022; 113: 709–720 and Molecular Cancer 2020; 19: 90) to obtain cells.
  • Example 1 Inhibitory effect of compound (I) on the growth of human medulloblastoma D283 transplanted tumors in nude mice 1.
  • mice Female, age: 4-6 weeks.
  • Human brain medulloblastoma D283 cell line (TMZ-resistant tumor cells, MGMT high expression) was inoculated subcutaneously in the right axilla of nude mice, with a cell inoculation amount of 5 ⁇ 10 6 /mouse. After the tumor grew to an average volume of about 120 mm 3 , the animals were randomly divided into 3 groups.
  • the compound (I) 10 mg/kg group was orally administered twice a day for 21 consecutive days, and the solvent control group was given an equal volume of blank preparation to the 10 mg/kg compound (I) administration group, orally administered twice a day for 21 consecutive days.
  • the control drug TMZ 3 mg/kg group was orally administered once a day for 21 consecutive days. During the entire experiment, the diameter of the transplanted tumor was measured 3 times a week, and the mouse body weight was weighed at the same time.
  • the relative tumor volume (RTV) was calculated based on the measurement results.
  • T/C (%) ( TRTV / CRTV ) x 100%, TRTV : RTV of the treatment group (compound (I) group or TMZ group); CRTV : RTV of the solvent control group.
  • the results are shown in Table 1.
  • the control drug TMZ 3 mg/kg group was orally administered once a day. After 21 consecutive days of administration, it had no inhibitory effect on the growth of human brain medulloblastoma D283 nude mouse subcutaneous transplanted tumors.
  • the T/C percentage obtained on the 21st day was 104.1%, proving that the human brain medulloblastoma D283 cell line is a TMZ-resistant cell line.
  • the test compound (I) was administered at a dose of 10 mg/kg, and was orally administered twice a day for three consecutive weeks. It can significantly inhibit the growth of human brain medulloblastoma D283 nude mouse subcutaneous transplanted tumors.
  • the T/C percentage obtained on the 21st day was 18.9%. In the experiment, all groups of mice were in good condition. The above results show that compound (I) can significantly inhibit the growth of human brain medulloblastoma D283 and show outstanding therapeutic effects on TMZ-resistant tumors.
  • Example 2 Inhibitory effect of compound (I) on the growth of human non-small cell lung cancer NCI-H1975 (EGFR T790M mutation) transplanted tumors in mice
  • Non-small cell lung cancer NCI-H1975 cell line (EGFR T790M mutation) was inoculated subcutaneously in the right axilla of NOD-SCID mice, with a cell inoculation amount of 5 ⁇ 10 6 /mouse. After the tumor grew to an average volume of about 135 mm 3 , the animals were randomly divided into groups. Compound (I) was divided into 10 mg/kg and 5 mg/kg groups, and was orally administered twice a day for 14 consecutive days. The solvent control group was given a blank preparation of the same volume as the 10 mg/kg compound (I) administration group, and was orally administered twice a day for 14 consecutive days.
  • V 0 is the tumor volume measured at the time of cage administration (i.e., d 0 )
  • V t is the tumor volume at each measurement.
  • T/C (%) ( TRTV / CRTV ) ⁇ 100%, TRTV : RTV of the treatment group (two groups of compound (I)); CRTV : RTV of the solvent control group.
  • the experimental results are shown in Table 2.
  • the test compound (I) 10 mg/kg and 5 mg/kg groups significantly delayed the growth of tumors, and the inhibitory effect increased with the increase in dose.
  • the T/C percentages obtained on the 14th day were 9.3% and 10.5%, respectively.
  • the growth of the tumors in the mice in the 10 mg/kg and 5 mg/kg groups of compound (I) was almost completely stagnant during the two-week experimental treatment.
  • the weight of the mice decreased slightly during the experiment, but the overall condition was good.
  • the above results show that compound (I) can significantly inhibit the growth of NCI-H1975 (EGFR T790M mutation) mouse transplanted tumors and has outstanding therapeutic effects on EGFR T790M mutation tumors.
  • Example 3 Inhibitory effect of compound (I) on the growth of human non-small cell lung cancer transplanted tumors in mice harboring EGFR C797S mutation
  • the human non-small cell lung cancer cell line PC-9-OR containing the EGFR C797S mutation (a cell line containing the EGFR C797S mutation constructed by the Shanghai Institute of Materia Medica, Chinese Academy of Sciences on the PC-9 cell line, the construction method is described in the literature Cancer Science 2022; 113: 709-720.) was inoculated subcutaneously in the right axilla of nude mice, with a cell inoculation amount of 5 ⁇ 10 6 /mouse, and the tumor was grown to an average volume. The animals were randomly divided into 4 groups after the diameter of the transplanted tumor was about 100 mm 3.
  • Compound (I) was divided into 10 mg/kg and 5 mg/kg groups, which were orally administered twice a day for 49 consecutive days.
  • the solvent control group was given a blank preparation of the same volume as the 10 mg/kg compound (I) administration group, which was orally administered twice a day for 49 consecutive days.
  • the control drug AZD9291 (third-generation EGFR inhibitor) 10 mg/kg group was orally administered once a day for 49 consecutive days.
  • the diameter of the transplanted tumor was measured 3 times a week, and the weight of the mice was weighed at the same time.
  • Example 4 Compound (I) enhances the anti-tumor efficacy of the third-generation EGFR inhibitor ASK120067
  • NOD-SCID mice female, age: 4-6 weeks.
  • the EGFR third-generation inhibitor ASK120067-resistant non-small cell lung cancer cell line 67R (a resistant cell line of the EGFR third-generation inhibitor ASK120067 constructed by the Shanghai Institute of Materia Medica, Chinese Academy of Sciences on the basis of NCI-H1975, whose EGFR itself has not mutated or deleted, is an EGFR-independent resistant cell, and the construction method is described in the literature Molecular Cancer 2020; 19:90) was injected subcutaneously into the right axilla of NOD-SCID mice.
  • the diameter of the subcutaneous transplanted tumor of BALB/c mice was measured with a vernier caliper, and the animals were randomly divided into groups after the tumor grew to an average volume of about 110 mm 3.
  • Compound (I) (5 mg/kg alone or in combination) was orally administered twice a day for 28 consecutive days; ASK120067 (5 mg/kg alone and in combination) was orally administered once a day for 28 consecutive days.
  • the solvent control group was given an equal volume of blank preparation without compound (I) as the 5 mg/kg compound (I) administration group, and the administration was orally administered twice a day for 28 consecutive days.
  • the diameter of the transplanted tumor was measured twice a week, and the mice were weighed at the same time.
  • T/C (%) ( TRTV / CRTV ) ⁇ 100%
  • TRTV RTV of treatment group (compound (I) alone group, ASK120067 alone group, compound (I) and ASK120067 combination group);
  • ASK120067 was combined with compound (I) at a dose of 5 mg/kg twice a day, tumor growth was significantly inhibited. The tumor volume did not increase substantially.
  • the T/C of the combination group was 8.5%.
  • the combination index was 1.69, indicating that the combination of compound (I) can enhance the tumor inhibition effect of ASK120067 in the 67R resistant tumor model.
  • the above results show that in the EGFR-independent EGFR inhibitor resistance model, the combination of compound (I) and EGFR inhibitor is effective, can significantly inhibit tumor growth, and reverse EGFR inhibitor resistance.
  • the compound (I) of the present invention can have significant in vivo inhibitory activity on the growth of human brain medulloblastoma D283 nude mouse transplanted tumor (TMZ-resistant tumor model).
  • the test compound (I) was administered at a dose of 10 mg/kg, orally twice a day, and for three consecutive weeks, it was able to significantly inhibit the growth of human brain medulloblastoma D283 nude mouse subcutaneous transplanted tumors, and the T/C obtained on the 21st day was 18.9%.
  • the above results show that compound (I) can significantly inhibit the growth of TMZ-resistant tumors and has a significant effect on TMZ-resistant tumors.
  • Compound (I) has significant in vivo inhibitory activity against PC-9-OR (containing EGFR C797S mutant cells) human non-small cell lung cancer mouse transplant tumor model.
  • the 10 mg/kg and 5 mg/kg groups of compound (I) can inhibit the growth of mouse PC-9-OR tumors in a dose-dependent manner, and the T/C percentages obtained on day 49 are 13.0% and 21.6%, respectively.
  • the above results show that compound (I) can significantly inhibit the growth of EGFR C797S mutant tumors.
  • Compound (I) enhances the anti-tumor efficacy of the third-generation EGFR inhibitor ASK120067.
  • compound (I) was combined with a dose of 5 mg/kg twice a day, and tumor growth was significantly inhibited, and the tumor volume basically did not increase.
  • the T/C of the combination group was 8.5%.
  • the combination drug index was 1.69, indicating that the combination of compound (I) can enhance the tumor inhibition effect of ASK120067 in the 67R resistant tumor model. This indicates that in EGFR-independent EGFR third-generation inhibitor-resistant tumors, the combination of compound (I) and EGFR third-generation inhibitors is effective and can significantly inhibit tumor growth and overcome EGFR third-generation inhibitor resistance.
  • the above experimental results show that the compound (I) has excellent inhibitory activity against drug-resistant tumors and can significantly inhibit the growth of drug-resistant tumors, especially temozolomide-resistant tumors and EGFR inhibitor-resistant tumors, and is expected to be developed as a drug against drug-resistant tumors.

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Abstract

本发明涉及一种萘酰胺化合物治疗耐药性肿瘤的用途。具体地，本发明公开了化合物(I)或其药学上可接受的盐的用于制备用于治疗耐药性肿瘤的药物的用途，尤其是替莫唑胺耐药性肿瘤、EGFR抑制剂耐药性肿瘤。本发明还公开了化合物(I)或其药学上可接受的盐与EGFR三代抑制剂ASK120067联用可逆转EGFR抑制剂耐药。

Description

一种萘酰胺化合物治疗耐药性肿瘤的用途 技术领域
本发明属于医药领域，具体涉及一种萘酰胺化合物在制备治疗耐药性肿瘤的药物中的应用。
背景技术
恶性肿瘤是威胁人类健康的重大疾病，尽管肿瘤治疗在手术、放疗、化疗等方面取得了长足的进展，也切实提高了癌症的生存期，然而肿瘤对药物产生耐药性仍然是肿瘤治疗学领域的重大难题。
胶质瘤(又称神经胶质瘤)是发生于神经外胚层的肿瘤，包括星形细胞瘤、胶质母细胞瘤、髓母细胞瘤、室管膜瘤和脉络从***状瘤等，而且胶质瘤根据发病部位又分为大脑神经胶质瘤和脊髓神经胶质瘤。胶质母细胞瘤是最常见的恶性原发性脑肿瘤，约占所有胶质瘤的57％，占所有原发性恶性中枢神经***肿瘤的48％。尽管近年来胶质母细胞瘤的多种治疗方法取得了进展，包括手术、放疗、化疗和支持治疗，但患者中位生存期也只有10～12个月，总体预后仍然较差，长期生存率较低，复发率接近100％。
替莫唑胺(TMZ)是一种咪唑四嗪衍生的烷化剂，TMZ在生理pH值时迅速水解为5-(3-甲基三氮-1-基)咪唑-4-甲酰胺(MTIC)，后者再通过脱氧核糖核酸(DNA)鸟嘌呤的O6和N7位点上的甲基化影响DNA复制从而发挥细胞毒作用。由于其可口服和易于透过血脑屏障，TMZ于2005年和2009年被FDA分别批准用于星型胶质瘤和胶质母细胞瘤患者的治疗，也是目前胶质瘤一线治疗的唯一口服化疗药。但是临床用药中发现至少50％的TMZ治疗患者对TMZ无反应，这主要是O6-甲基鸟嘌呤甲基转移酶(MGMT)过度表达和缺乏GBM细胞中的DNA修复途径所造成的。MGMT可修复TMZ引起的鸟嘌呤O6位的甲基化，使肿瘤细胞对TMZ不敏感。而MGMT的表达主要受到其启动子区的甲基化水平影响，甲基化水平越高MGMT表达量越低，对TMZ也就越敏感。在胶质瘤病人中MGMT甲基化阳性率约为40％，在恶性程度最高的胶质母细胞瘤中更低。这样就意味着在不考虑TMZ治疗继发耐药的情况下，至少一半以上的胶质瘤患者对TMZ耐药。
目前，临床推荐使用甲基化特异性PCR(聚合酶链式反应)结合免疫组化的方法来确定MGMT表达量。根据脑胶质瘤诊疗规范(2018版)，对于MGMT启动子区非甲基化和甲基化情况不明确者，只能选择放疗同步并辅助TMZ化疗等获益很有限的治疗手段。此外，几种靶向MGMT的药物正处在临床研究中，例如开发MGMT的抑制剂(O6-BG，6-BTG)，然而这些药物的研究进展缓慢且由于非特异性靶向正常细胞引起的全身性毒性不容忽视。综上，胶质瘤患者尤其是MGMT高表达患者急需有效的治疗药物或治疗方案。
肺癌是目前世界范围内癌症死亡的主要原因，其中非小细胞肺癌(Non-small cell lung cancer,NSCLC)作为该疾病的主要组织学类型，占肺癌患者的80％以上，5年生存率低于15％。表皮生长因子受体(Epidermal growth factor receptor,EGFR)的高表达或异常激活是促进NSCLC恶化发展的重要靶点。近年来，随着对肺癌发病机制的深入研究，以EGFR抑制剂为代表的分子靶向治疗取得了突破性进展。迄今为止，已有三代EGFR抑制剂陆续获批上市，为EGFR突变型NSCLC患者带来了巨大的临床获益，但EGFR抑制剂在用药一年左右的时间内出现获得性耐药，极大限制了药物的临床应用。因此开发能够抑制EGFR抑制剂耐药的策略成为当前NSCLC研究的热点。
已报道的EGFR抑制剂耐药机制主要分为两大类：第一类是EGFR依赖的耐药机制，主要是EGFR自身发生变化，例如在EGFR一代抑制剂耐药中占据主要原因的EGFRT790M突变(占所有耐药病人的60％左右)，或EGFRC797S/G、EGFRL798I、EGFRL792H等多种点突变和EGFRT790M丢失介导EGFR三代抑制剂的耐药，其中发生EGFRC797S突变的肿瘤尚无治疗药物。第二类是EGFR非依赖的耐药机制，包括MET和HER2扩增、FGFR1扩增、PIK3CA突变、小细胞转化、RAS-MAPK途径激活与KRAS突变和扩增等。目前，靶向EGFRC797S的四代EGFR抑制剂均处于早期临床研究阶段，尚无药物上市；此外，针对EGFR非依赖的耐药途径，由于肿瘤的高度异质性导致耐药机制复杂，且缺乏***性的耐药机制研究，因此克服耐药研究进展缓慢，远不能满足临床需求。综上，克服EGFR抑制剂耐药问题仍然面临严峻挑战，亟待寻求新的治疗策略。
化合物(I)首次公开于CN104860885A中，结构式如下式(I)所示，
化合物(I)是一种具有优异活性的VEGFR/CSF1R双靶抑制剂，能够抑制肿瘤细胞血管增生和促进肿瘤免疫发挥抗肿瘤作用。
发明内容
本发明的目的在于提供一种萘酰胺化合物治疗耐药性肿瘤的用途。
本发明的第一方面，提供了一种化合物(I)或其药学上可接受的盐的用途，用于制备药物，所述药物用于治疗耐药性肿瘤；
所述化合物(I)具有如下结构：
在另一优选例中，所述耐药性肿瘤选自下组：胶质瘤、肺癌。
在另一优选例中，所述耐药性肿瘤选自下组：替莫唑胺耐药性肿瘤、EGFR抑制剂耐药性肿瘤。
在另一优选例中，所述耐药性肿瘤为O6-甲基鸟嘌呤甲基转移酶高表达的肿瘤。
在另一优选例中，所述耐药性肿瘤为O6-甲基鸟嘌呤甲基转移酶高表达的胶质瘤。
在另一优选例中，所述替莫唑胺耐药性肿瘤为替莫唑胺耐药性胶质瘤。
在另一优选例中，所述替莫唑胺耐药性肿瘤为O6-甲基鸟嘌呤甲基转移酶高表达的胶质瘤。
在另一优选例中，所述胶质瘤选自下组：胶质母细胞瘤、星型胶质瘤、髓母细胞瘤。
在另一优选例中，所述替莫唑胺耐药性肿瘤为替莫唑胺耐药性髓母细胞瘤。
在另一优选例中，所述EGFR抑制剂耐药性肿瘤为EGFR抑制剂耐药性肺癌。
在另一优选例中，所述肺癌选自下组：小细胞肺癌、非小细胞肺癌、肺腺癌。
在另一优选例中，所述EGFR抑制剂耐药性肿瘤为EGFR抑制剂耐药性非小 细胞肺癌。
在另一优选例中，所述EGFR抑制剂耐药性肿瘤为EGFR突变肿瘤。
在另一优选例中，所述EGFR突变选自下组：EGFRT790M突变、EGFRC797S突变。
在另一优选例中，所述EGFR抑制剂耐药性肺癌为EGFRT790M突变的肺癌或者EGFRC797S突变的肺癌。
在另一优选例中，所述EGFR抑制剂耐药性肺癌为EGFRT790M突变的非小细胞肺癌或者EGFRC797S突变的非小细胞肺癌。
在另一优选例中，所述EGFR抑制剂耐药性肿瘤为EGFR不依赖的EGFR抑制剂耐药性肿瘤。
在另一优选例中，所述EGFR不依赖的EGFR抑制剂耐药性肿瘤为选自下组机制导致的肿瘤：MET和HER2扩增、FGFR1扩增、PIK3CA突变、小细胞转化、RAS-MAPK途径激活、KRAS突变和扩增。
在另一优选例中，所述EGFR不依赖的EGFR抑制剂耐药性肿瘤为EGFR不依赖的EGFR抑制剂耐药性肺癌。
在另一优选例中，所述EGFR不依赖的EGFR抑制剂耐药性肺癌为EGFR不依赖的EGFR抑制剂耐药性非小细胞肺癌。
在另一优选例中，所述EGFR抑制剂选自下组：ASK120067(Limertinib)、奥希替尼(AZD9291)或其盐、阿美替尼或其盐、伏美替尼或其盐，
在另一优选例中，所述EGFR抑制剂为ASK120067(Limertinib)、奥希替尼(AZD9291)及其盐。
本发明的第二方面，提供了一种用于治疗耐药性肿瘤的药物组合物，所述药物组合物包含治疗有效量的化合物(I)或其药学上可接受的盐和药学上可接受的载体；
所述化合物(I)具有如下结构：
在另一优选例中，所述耐药性肿瘤如上文所述。
在另一优选例中，所述药物组合物为口服制剂。
本发明的第三方面，提供了一种药物组合，所述药物组合包含治疗有效量的第一药物活性成分和第二药物活性成分；
所述第一药物活性成分为化合物(I)或其药学上可接受的盐；
所述化合物(I)具有如下结构：
所述第二药物活性成分为选自下组的抗肿瘤药物：替莫唑胺、EGFR抑制剂。
在另一优选例中，所述肿瘤选自下组：替莫唑胺耐药性肿瘤、EGFR抑制剂耐药性肿瘤。
在另一优选例中，所述替莫唑胺耐药性肿瘤如上文所述。
在另一优选例中，所述EGFR抑制剂耐药性肿瘤如上文所述。
在另一优选例中，所述EGFR抑制剂选自下组：ASK120067(Limertinib)、奥希替尼(AZD9291)或其盐、阿美替尼或其盐、伏美替尼或其盐。
在另一优选例中，所述EGFR抑制剂为ASK120067(Limertinib)。
在另一优选例中，所述第一药物活性成分和所述第二药物活性成分的质量比为1-4，较佳地1-3，优选为2。
本发明的第四方面，提供了一种化合物(I)或其药学上可接受的盐的用途，用于制备药物，所述药物用于增强抗肿瘤药物的药效；
所述化合物(I)具有如下结构：
所述抗肿瘤药物选自下组：替莫唑胺、EGFR抑制剂。
在另一优选例中，所述肿瘤选自下组：替莫唑胺耐药性肿瘤、EGFR抑制剂耐药性肿瘤。
在另一优选例中，所述替莫唑胺耐药性肿瘤如上文所述。
在另一优选例中，所述EGFR抑制剂耐药性肿瘤如上文所述。
在另一优选例中，所述EGFR抑制剂如上文所述。
本发明的第五方面，提供了一种药物组合物，包含治疗有效量的本发明第三方面所述药物组合和药学上可接受的载体。
在另一优选例中，所述药物组合物为口服制剂。
在另一优选例中，所述药物组合物用于治疗耐药性肿瘤。
在另一优选例中，所述耐药性肿瘤如上文所述。
本发明的第六方面，提供了一种本发明第三方面所述药物组合的用途，用于制备药物，所述药物用于治疗耐药性肿瘤。
在另一优选例中，所述耐药性肿瘤如上文所述。
在另一优选例中，所述耐药性肿瘤的患者为人。
本发明的第七方面，提供了一种治疗耐药性肿瘤的方法，包括步骤：
1)将治疗有效量的化合物(I)或其药学上可接受的盐或含化合物(I)或其药学上可接受的盐的药物组合物施用于患有耐药性肿瘤的患者或者对抗肿瘤药物不敏感的患者；
所述化合物(I)具有如下结构：
在另一优选例中，所述耐药性肿瘤如上文所述。
在另一优选例中，所述抗肿瘤药物选自下组：替莫唑胺、EGFR抑制剂。
在另一优选例中，所述EGFR抑制剂如上文所述。
在另一优选例中，所述方法在步骤1)之前还包括如下步骤：
使用基因检测方法确定所述患者的MGMT基因表达情况和/或EGFR基因突变情况。
在另一优选例中，所述患者为MGMT基因过度表达的患者。
在另一优选例中，所述患者为EGFR基因突变的患者。
在另一优选例中，所述患者为EGFRT790M突变的患者或EGFRC797S突变的患者。
本发明的第八方面，提供了一种治疗耐药性肿瘤的方法，包括步骤：
1)将治疗有效量的本发明第三方面所述药物组合施用于患有耐药性肿瘤的患者或者对抗肿瘤药物不敏感的患者。
在另一优选例中，所述耐药性肿瘤如上文所述。
在另一优选例中，所述抗肿瘤药物选自下组：替莫唑胺、EGFR抑制剂。
在另一优选例中，所述EGFR抑制剂如上文所述。
在另一优选例中，所述方法在步骤1)之前还包括如下步骤：
使用基因检测方法确定所述患者的MGMT基因表达情况和/或EGFR基因突变情况。
在另一优选例中，所述患者为MGMT基因过度表达的患者。
在另一优选例中，所述患者为EGFR基因突变的患者。
在另一优选例中，所述患者为EGFRT790M突变的患者或EGFRC797S突变的患者。
应理解，在本发明范围内中，本发明的上述各技术特征和在下文(如实施例)中具体描述的各技术特征之间都可以互相组合，从而构成新的或优选的技术方案。限于篇幅，在此不再一一累述。
本发明人经过长期而深入的研究，意外地发现化合物(I)对于耐药性肿瘤具有优异的抑制活性，可显著抑制耐药性肿瘤的生长，尤其是替莫唑胺耐药性肿瘤、EGFR抑制剂耐药性肿瘤，有望开发为抗耐药性肿瘤的药物。在此基础上，发明人完成了本发明。
如本文所用，术语“药学上可接受的盐”指本发明化合物与酸或碱所形成的适合用作药物的盐。药学上可接受的盐包括无机盐和有机盐。
“药学上可以接受的载体”指的是：一种或多种相容性固体或液体填料或凝胶物质，它们适合于人使用，而且必须有足够的纯度和足够低的毒性。“相容性”在此指的是组合物中各组份能和本发明的化合物以及它们之间相互掺和，而不明显降低化合物的药效。
本发明药物组合物的施用方式没有特别限制，代表性的施用方式包括但并不限于：口服、瘤内、直肠、肠胃外(静脉内、肌肉内或皮下)和局部给药。
相应地，本发明药物可制成临床上可接受的各种剂型，包括口服剂型、注射剂型、局部给药剂型或外用剂型等。
固体剂型如片剂、糖丸、胶囊剂、丸剂和颗粒剂可采用包衣和壳材制备，如肠衣和其它本领域公知的材料。它们可包含不透明剂，并且，这种组合物中活性化合物或化合物的释放可以延迟的方式在消化道内的某一部分中释放。可 采用的包埋组分的实例是聚合物质和蜡类物质。必要时，活性化合物也可与上述赋形剂中的一种或多种形成微胶囊形式。
用于肠胃外注射的组合物可包含生理上可接受的无菌含水或无水溶液、分散液、悬浮液或乳液，和用于重新溶解成无菌的可注射溶液或分散液的无菌粉末。
用于局部给药的本发明化合物的剂型包括软膏剂、散剂、贴剂、喷射剂和吸入剂。活性成分在无菌条件下与生理上可接受的载体及任何防腐剂、缓冲剂，或必要时可能需要的推进剂一起混合。
本发明化合物可以单独给药，或者与其他药学上可接受的其他化合物(如抗肿瘤药物)联合给药。
本发明治疗方法可以单独施用，或者与其它治疗手段或者治疗药物联用。
本发明所述的治疗有效量是指药学上认为的有效给药剂量，即活性化合物的量足以明显改善病情，而不至于产生严重的副作用。对于60kg体重的人而言，日给药剂量通常为0.01～2000mg，优选1～500mg。可以每日一次单剂量施用，可以每天分多次施用，也可以间隔使用。具体剂量和给药频率应考虑给药途径、病人健康状况等因素，这些都是熟练医师根据常规技能可以确定的。
本发明还提供所述药物组合物的使用方法，即将治疗有效量的本发明化合物(I)或其药学上可接受的盐或本发明药物组合物施用于需要治疗的哺乳动物(如人)。在一些实施方式中，所述药物组合物的使用方法，在将治疗有效量的本发明化合物(I)或其药学上可接受的盐施用于需要治疗的哺乳动物(如人)之前，还包括基因检测的步骤，以确定需要治疗的哺乳动物(如人)的MGMT甲基化状态和/或EGFR基因突变。哺乳动物(如人)的MGMT甲基化状态可采用mgmt基因甲基化检测试剂(如厂商为基因技术，国械注准20193400101)进行检测。目前检测EGFR基因突变最常用的方法包括直接测序法和扩增阻滞突变***(ARMS-PCR)。DNA直接测序是目前检测基因突变最直接、最基础的一种方法，可通过读出DNA碱基序列检测肿瘤组织样本中已知和未知的基因突变。ARMS-PCR法利用设计的特异性引物对突变序列进行PCR精准扩增，同时采用实时荧光PCR定量分析，可高灵敏地检测活检小样本和血液标本中的EGFR基因 突变。哺乳动物(如人)的EGFR基因突变可采用EGFR基因突变检测试剂盒(如：人EGFR基因突变检测试剂盒，厂商为艾德生物，国械注准20143402001)进行检测。
与现有技术相比，本发明具有以下主要优点：
(1)所述化合物(I)对于耐药性肿瘤具有优异抑制活性，可显著抑制耐药性肿瘤的生长，尤其是替莫唑胺耐药性肿瘤、EGFR抑制剂耐药性肿瘤，有望开发为抗耐药性肿瘤的药物；
(2)在EGFR不依赖的EGFR三代抑制剂耐药肿瘤中，化合物(I)与EGFR三代抑制剂联合用药有效，可显著抑制肿瘤的生长，克服EGFR三代抑制剂耐药；
(3)所述化合物(I)制法简单；
(4)所述化合物(Ⅰ)对EGFRT790M突变和EGFRC797S突变以及EGFR不依赖的三代抑制剂耐药肿瘤均有效。
具体实施方式
下面结合具体实施例，进一步阐述本发明。应理解，这些实施例仅用于说明本发明而不用于限制本发明的范围。下列实施例中未注明具体条件的实验方法，通常按照常规条件或按照制造厂商所建议的条件。除非另外说明，否则百分比和份数按重量计算。
除非另行定义，文中所使用的所有专业与科学用语与本领域熟练人员所熟悉的意义相同。此外，任何与所记载内容相似或均等的方法及材料皆可应用于本发明方法中。文中所述的较佳实施方法与材料仅作示范之用。
实验材料来源或配制：
化合物(I)：参考CN104860885A实施例21制备得到。
对照化合物、实验中所用试剂、原材料均为商业购买或自行配制。
体内实验受试物化合物(I)制剂按如下处方进行制备见表A：
表A

空白制剂：配方同表A，区别为不含化合物(I)
所述潜溶剂在制备过程中蒸发去除，所得受试物和空白制剂中不含所述潜溶剂。
临用前将含化合物(I)的制剂或空白制剂以水稀释至所需浓度或所需体积。
TMZ：用0.5％羧甲基纤维素钠溶液配制到所需剂量后使用。
ASK120067和AZD9291：用含1％Tween 80的注射用水配到所需剂量后使用。
细胞株来源及获得：D283和NCI-H1975购于ATCC(美国模式培养物集存库)。PC-9-OR和67R由中国科学院上海药物研究所采用常规方法构建，具体见文献(Cancer Science 2022；113:709–720和Molecular Cancer 2020；19:90)上的方法进行构建以获得细胞。
实施例1：化合物(I)对人脑髓母细胞瘤D283裸小鼠移植瘤生长抑制作用1.实验方法
BALB/c裸小鼠，雌性，鼠龄：4-6周龄。用人脑髓母细胞瘤D283细胞株(TMZ耐药性肿瘤细胞、MGMT高表达)接种于裸小鼠右侧腋窝皮下，细胞接种量为5×106/只，待肿瘤生长至平均体积为120mm3左右后将动物随机分为3组。化合物(I)10mg/kg组，每天口服给药两次，连续给药21天，溶剂对照组给以10mg/kg化合物(Ⅰ)给药组等体积的空白制剂，每天口服给药两次，连续给药21天。对照药物TMZ 3mg/kg组每天口服给药一次，连续给药21天。整个实验过程中，每周3次测量移植瘤直径，同时称量小鼠体重。肿瘤体积(tumor volume,TV)的计算公式为：TV＝1/2×a×b2，其中a、b分别表示长、宽。根据测量的结果计算出相对肿瘤体积(relative tumor volume，RTV)，计算公式为：RTV＝Vt/V0。其中V0为分笼给药时(即d0)测量所得肿瘤体积，Vt为每一次测量时的肿瘤体积。抗肿瘤活性的评价指标为：相对肿瘤增殖率T/C(％)，计算公式如下：T/C(％)＝ (TRTV/CRTV)×100％，TRTV：治疗组(化合物(I)组或TMZ组)RTV；CRTV：溶剂对照组RTV。
2.实验结果
结果如表1所示。对照药物TMZ 3mg/kg组每天口服给药一次，连续给药21天后对人脑髓母细胞瘤D283裸小鼠皮下移植瘤的生长没有抑制作用，第21天所得T/C百分数为104.1％，证明人脑髓母细胞瘤D283细胞株为TMZ耐药细胞株。受试物化合物(I)给药剂量为10mg/kg，每天口服给药两次，连续三周则能显著抑制人脑髓母细胞瘤D283裸小鼠皮下移植瘤的生长，第21天所得T/C百分数为18.9％。实验中，各组小鼠均状态良好。以上结果表明化合物(I)能显著抑制人脑髓母细胞瘤D283生长，对TMZ耐药肿瘤表现出突出的疗效。
表1.化合物(I)对人脑髓母细胞瘤D283裸小鼠移植瘤的治疗作用
注：实验数据采用t检验分析统计，*p<0.05，**p<0.01，***p<0.001。
实施例2：化合物(I)对人非小细胞肺癌NCI-H1975(EGFRT790M突变)小鼠移植瘤生长抑制作用
1、实验方法
NOD-SCID小鼠，雌性，鼠龄：4-6周龄。用非小细胞肺癌NCI-H1975细胞株(EGFRT790M突变)接种于NOD-SCID小鼠右侧腋窝皮下，细胞接种量为5×106/只，待肿瘤生长至平均体积约为135mm3左右后将动物随机分组。化合物(I)分为10mg/kg和5mg/kg组，每天口服给药两次，连续给药14天，溶剂对照组给予与10mg/kg化合物(Ⅰ)给药组等体积的的空白制剂，每天口服给药两次，连续给药14天。整个实验过程中，每周3次测量移植瘤直径，同时称量小鼠体重。 肿瘤体积(tumor volume,TV)的计算公式为：TV＝1/2×a×b2，其中a、b分别表示长、宽。根据测量的结果计算出相对肿瘤体积(relative tumor volume，RTV)，计算公式为：RTV＝Vt/V0。其中V0为分笼给药时(即d0)测量所得肿瘤体积，Vt为每一次测量时的肿瘤体积。抗肿瘤活性的评价指标为：相对肿瘤增殖率T/C(％)，计算公式如下：T/C(％)＝(TRTV/CRTV)×100％，TRTV：治疗组(化合物(I)两个组)RTV；CRTV：溶剂对照组RTV。
2、实验结果
实验结果如表2所示。受试物化合物(I)10mg/kg和5mg/kg组，明显延缓了肿瘤的生长，且抑制效果随剂量增加而增加，第14天所得T/C百分数分别为9.3％和10.5％。化合物(I)的10mg/kg和5mg/kg组小鼠所荷肿瘤在实验治疗两周期间生长几乎完全停滞。实验期间小鼠体重略有下降但是整体状态良好。以上结果说明化合物(I)能够显著抑制NCI-H1975(EGFRT790M突变)小鼠移植瘤生长，对EGFRT790M突变的肿瘤有突出疗效。
表2.化合物(I)对非小细胞肺癌NCI-H1975小鼠移植瘤的治疗作用
备注：实验数据采用t检验分析统计，*p<0.05，**p<0.01，***p<0.001。
实施例3：化合物(I)对含EGFRC797S突变的人非小细胞肺癌小鼠移植瘤生长抑制作用
1、实验方法
BALB/c裸小鼠，雌性，鼠龄：4-6周龄。将含EGFRC797S突变的人非小细胞肺癌细胞株PC-9-OR(由中国科学研究院上海药物研究所在PC-9细胞株上构建的含EGFRC797S突变的细胞株，构建方法见文献Cancer Science 2022；113:709-720.)接种于裸小鼠右侧腋窝皮下，细胞接种量为5×106/只，待肿瘤生长至平均体积 为100mm3左右后将动物随机分为4组。化合物(I)分为10mg/kg和5mg/kg组，每天口服给药两次，连续给药49天，溶剂对照组给予与10mg/kg化合物(Ⅰ)给药组等体积的空白制剂，每天口服给药两次，连续给药49天。对照药物AZD9291(三代EGFR抑制剂)10mg/kg组每天口服给药一次，连续给药49天。整个实验过程中，每周3次测量移植瘤直径，同时称量小鼠体重。肿瘤体积(TV)的计算公式为：TV＝1/2×a×b2，其中a、b分别表示长、宽。根据测量的结果计算出相对肿瘤体积(relative tumor volume，RTV)，计算公式为：RTV＝Vt/V0。其中V0为分笼给药时(即d0)测量所得肿瘤体积，Vt为每一次测量时的肿瘤体积。抗肿瘤活性的评价指标为：相对肿瘤增殖率T/C(％)，计算公式如下：T/C(％)＝(TRTV/CRTV)×100％，TRTV：治疗组(化合物(I)组或AZD9291组)RTV；CRTV：溶剂对照组RTV。
2、实验结果
实验结果如表3所示。AZD9291几乎不抑制PC-9-OR肿瘤的生长，T/C在90％以上。化合物(I)分别在10mg/kg和5mg/kg剂量下显著延缓了肿瘤的生长，且抑制效果随剂量增加而增加，第49天所得T/C百分数分别为13.0％和21.6％。实验期间小鼠状态良好。以上结果说明化合物(I)能够显著抑制人非小细胞肺癌PC-9-OR(含EGFRC797S突变)的小鼠移植瘤生长，对EGFRC797S突变的肿瘤有显著疗效。
表3.化合物(I)对含EGFRC797S突变的人非小细胞肺癌细胞株PC-9-OR小鼠移植瘤的实验治疗作用
备注：实验数据采用t检验分析统计，*p<0.05，**p<0.01，***p<0.001。
实施例4：化合物(I)增敏EGFR三代抑制剂ASK120067抗肿瘤药效
1、实验方法
NOD-SCID小鼠，雌性，鼠龄：4-6周龄。在无菌条件下，将EGFR三代抑制剂ASK120067耐药非小细胞肺癌细胞株67R(由中国科学院上海药物研究所在NCI-H1975的基础上构建的EGFR三代抑制剂ASK120067的耐药细胞株，其EGFR自身并未发生突变或者缺失，为EGFR不依赖的耐药细胞，构建方法见文献Molecular Cancer 2020；19:90)注射于NOD-SCID小鼠右侧腋窝皮下。BALB/c小鼠皮下移植瘤用游标卡尺测量移植瘤直径，待肿瘤生长至平均体积约为110mm3左右后将动物随机分组。化合物(I)(5mg/kg单用或联用)每天口服给药两次，连续给药28天；ASK120067(5mg/kg单用和联用组)每天口服给药一次，连续给药28天。溶剂对照组给以5mg/kg化合物(Ⅰ)给药组等体积不含化合物(I)的空白制剂，每天口服给药两次，连续给药28天。整个实验过程中，每周2次测量移植瘤直径，同时称量小鼠体重。肿瘤体积的计算公式为：TV＝1/2×a×b2，其中a、b分别表示长、宽。根据测量的结果计算出相对肿瘤体积(relative tumor volume，RTV)，计算公式为：RTV＝Vt/V0。其中V0为分笼给药时(即d0)测量所得肿瘤体积，Vt为每一次测量时的肿瘤体积。抗肿瘤活性的评价指标为：相对肿瘤增殖率T/C(％)，计算公式如下：T/C(％)＝(TRTV/CRTV)×100％，TRTV：治疗组(化合物(I)单用组、ASK120067单用组、化合物(I)和ASK120067联用组)RTV；CRTV：溶剂对照组RTV。联合用药指数(Combination Ratio＝Expected-FTV/Observed-FTV)，Expected-FTV＝(药物A的FTV)×(药物B的FTV)，FTV(fractional tumor volume，肿瘤体积分数)＝实验组的最终肿瘤体积/对照组的最终肿瘤体积，Observed-FTV＝联合给药组的最终肿瘤体积/对照组的最终肿瘤体积。当联合用药指数大于1时，认为两药有协同作用，小于1时，则没有协同作用。
2、实验结果
化合物(I)在5mg/kg单用剂量下对67R肿瘤的体内生长具有部分抑制活性(T/C＝43.8％)；ASK120067(5mg/kg)单用组对肿瘤生长也有一定的抑制作用(T/C＝32.3％)，但肿瘤仍然持续缓慢增长。在使用ASK120067的基础上联合化合物(I)以每天两次5mg/kg的剂量给药，肿瘤生长显著被抑制，肿 瘤体积基本没有增长。实验终点时联用组T/C为8.5％。联合用药指数为1.69，显示联用化合物(I)能够增强ASK120067在67R耐药肿瘤模型中的抑瘤作用。以上结果表明在EGFR不依赖的EGFR抑制剂耐药模型中，化合物(I)与EGFR抑制剂联合用药有效，可显著抑制肿瘤的生长，逆转EGFR抑制剂耐药。
表4.化合物(I)联合ASK120067对ASK120067耐药非小细胞肺癌67R小鼠移植瘤的实验治疗作用
备注：实验数据采用t检验分析统计，*p<0.05，**p<0.01，***p<0.001。
表5.联合用药指数计算
体内研究显示：
(1)本发明化合物(I)能够对人脑髓母细胞瘤D283裸小鼠移植瘤(TMZ耐药性肿瘤模型)生长具有显著的体内抑制活性。受试物化合物(I)给药剂量为10mg/kg，每天口服给药两次，连续三周则能显著抑制人脑髓母细胞瘤D283裸小鼠皮下移植瘤的生长，第21天所得T/C为18.9％。以上结果表明化合物(I)能显著抑制TMZ耐药性肿瘤生长，对TMZ耐药肿瘤效果显著。
(2)本发明化合物(I)对人非小细胞肺癌NCI-H1975小鼠移植瘤(EGFRT790M 突变)生长具有显著的体内抑制活性。化合物(I)的10mg/kg和5mg/kg组小鼠所荷NCI-H1975肿瘤在实验治疗两周期间生长几乎完全停滞。实验期间小鼠体重略有下降但是整体状态良好。以上结果说明化合物(I)能够显著抑制EGFRT790M突变肿瘤生长。
(3)化合物(I)对于PC-9-OR(含EGFRC797S突变细胞)人非小细胞肺癌小鼠移植瘤模型具有显著体内抑制活性。化合物(I)的10mg/kg和5mg/kg组能够剂量依赖的抑制小鼠PC-9-OR肿瘤生长，第49天所得T/C百分数分别为13.0％和21.6％。以上结果表明化合物(I)能够显著抑制EGFRC797S突变肿瘤生长。
(4)化合物(I)增敏EGFR三代抑制剂ASK120067抗肿瘤药效。化合物(I)在5mg/kg单用剂量下对67R肿瘤(ASK120067耐药模型)的体内生长具有部分抑制活性(T/C＝43.8％)；ASK120067(5mg/kg)单用组对肿瘤生长也有一定的抑制作用(T/C＝32.3％)，但肿瘤仍然持续缓慢增长。在使用ASK120067的基础上联合化合物(I)以每天两次5mg/kg的剂量给药，肿瘤生长显著被抑制，肿瘤体积基本没有增长。实验终点时联用组T/C为8.5％。联合用药指数为1.69，显示联用化合物(I)能够增强ASK120067在67R耐药肿瘤模型中的抑瘤作用。这表明在EGFR不依赖的EGFR三代抑制剂耐药肿瘤中，化合物(I)与EGFR三代抑制剂联合用药有效，可显著抑制肿瘤的生长，克服EGFR三代抑制剂耐药。
以上实验结果表明，所述化合物(I)对于耐药性肿瘤具有优异抑制活性，可显著抑制耐药性肿瘤的生长，尤其是替莫唑胺耐药性肿瘤、EGFR抑制剂耐药性肿瘤，有望开发为抗耐药性肿瘤的药物。
在本发明提及的所有文献都在本申请中引用作为参考，就如同每一篇文献被单独引用作为参考那样。此外应理解，在阅读了本发明的上述讲授内容之后，本领域技术人员可以对本发明作各种改动或修改，这些等价形式同样落于本申请所附权利要求书所限定的范围。

Claims (10)

  1. 一种化合物(I)或其药学上可接受的盐的用途，其特征在于，用于制备药物，所述药物用于治疗耐药性肿瘤；
    所述化合物(I)具有如下结构：
  2. 如权利要求1所述用途，其特征在于，所述耐药性肿瘤选自下组：胶质瘤、肺癌；
    优选地，所述胶质瘤选自下组：星型胶质瘤、胶质母细胞瘤、髓母细胞瘤；
    优选地：所述肺癌选自下组：小细胞肺癌、非小细胞肺癌、肺腺癌。
  3. 如权利要求2所述用途，其特征在于，所述耐药性肿瘤选自下组：替莫唑胺耐药性肿瘤或EGFR抑制剂耐药性肿瘤，优选地，所述EGFR抑制剂选自下组：ASK120067或其盐、奥希替尼或其盐、阿美替尼或其盐、伏美替尼或其盐。
  4. 如权利要求3所述用途，其特征在于，所述替莫唑胺耐药性肿瘤为替莫唑胺耐药性胶质瘤或O6-甲基鸟嘌呤甲基转移酶高表达的胶质瘤；
    优选地，所述替莫唑胺耐药性肿瘤为替莫唑胺耐药性髓母细胞瘤。
  5. 如权利要求3所述用途，其特征在于，所述EGFR抑制剂耐药性肿瘤为EGFR抑制剂耐药性肺癌，或所述EGFR抑制剂耐药性肿瘤为EGFR不依赖的EGFR抑制剂耐药性肿瘤；
    优选地，所述EGFR抑制剂耐药性肺癌为EGFR抑制剂耐药性非小细胞肺癌。
  6. 如权利要求5所述用途，其特征在于，所述EGFR抑制剂耐药性肺癌为EGFRT790M突变的肺癌或者EGFRC797S突变的肺癌；
    优选地，所述EGFR抑制剂耐药性肺癌为EGFRT790M突变的非小细胞肺癌或者EGFRC797S突变的非小细胞肺癌。
  7. 一种药物组合，其特征在于，所述药物组合包含治疗有效量的第一药物活性成分和第二药物活性成分；
    所述第一药物活性成分为化合物(I)或其药学上可接受的盐；
    所述化合物(I)具有如下结构：
    所述第二药物活性成分为选自下组的抗肿瘤药物：替莫唑胺、EGFR抑制剂。
  8. 一种化合物(I)或其药学上可接受的盐的用途，其特征在于，用于制备药物，所述药物用于增强抗肿瘤药物的药效；
    所述化合物(I)具有如下结构：
    所述抗肿瘤药物选自下组：替莫唑胺、EGFR抑制剂。
  9. 一种药物组合物，其特征在于，包含治疗有效量的权利要求7所述药物组合和药学上可接受的载体。
  10. 一种权利要求7所述药物组合的用途，其特征在于，用于制备药物，所述药物用于治疗耐药性肿瘤。
PCT/CN2023/126296 2022-10-25 2023-10-24 一种萘酰胺化合物治疗耐药性肿瘤的用途 WO2024088275A1 (zh)

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US20180036304A1 (en) * 2015-03-06 2018-02-08 Beyondspring Pharmaceuticals, Inc. Method of treating cancer associated with a ras mutation
US20200246346A1 (en) * 2017-05-02 2020-08-06 Novartis Ag Combination therapy
CN115215847A (zh) * 2021-04-16 2022-10-21 中国科学院上海药物研究所 一类kras-sos1抑制剂、其制备方法及其应用

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20100087499A1 (en) * 2007-01-30 2010-04-08 Schering Corporation Pharmaceutical compositions and methods of using temozolomide and multi-targeted kinase inhibitors
CN104860885A (zh) * 2014-02-24 2015-08-26 中国科学院上海药物研究所 萘酰胺类化合物、其制备方法和用途
US20180036304A1 (en) * 2015-03-06 2018-02-08 Beyondspring Pharmaceuticals, Inc. Method of treating cancer associated with a ras mutation
US20200246346A1 (en) * 2017-05-02 2020-08-06 Novartis Ag Combination therapy
CN115215847A (zh) * 2021-04-16 2022-10-21 中国科学院上海药物研究所 一类kras-sos1抑制剂、其制备方法及其应用

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