A type of tetravalent platinum complex containing APR-246 and its preparation method and application

Abstract

The present invention discloses a tetravalent platinum complex containing APR-246, having a structure as shown in Formula I: wherein n is selected from 0, 1, 2, 3, or 4; R is selected from a branched or linear C1-C20 alkyl group; and R2 is selected from a branched or linear C1-C20 alkyl group. The tetravalent platinum compounds containing the p53 mutation activator APR-246 provided by the present invention can enhance the anti-tumor activity of APR-246 and leverage the anti-tumor advantages of APR-246 and divalent platinum drugs. By utilizing the unique anti-tumor mechanism of APR-246, they can work together with platinum drugs to achieve excellent synergistic anti-tumor efficacy, while also significantly helping to improve tumor drug resistance.

Description

Technical Field

[0001] The present invention belongs to the field of medical technology, and in particular relates to a tetravalent platinum complex containing a p53 mutation activator APR-246, a preparation method and an application thereof in the preparation of anti-tumor drugs. Background Art

[0002] Platinum-based anti-cancer drugs are widely used clinically to treat malignant tumors. However, these drugs can only be administered intravenously and have significant side effects, particularly ototoxicity and nephrotoxicity. Furthermore, long-term use of these drugs can easily lead to the development of drug resistance in tumors. Therefore, the design of a new generation of platinum-based drugs with low toxicity, oral compatibility, and no cross-resistance with existing platinum-based drugs has been a hot topic in recent years. Divalent platinum drugs can be oxidized with hydrogen peroxide to introduce two hydroxyl groups along the axial direction to produce tetravalent platinum compounds. Tetravalent platinum compounds are easily reduced in vivo, releasing the divalent platinum drug. Furthermore, additional antitumor pharmacophores can be introduced along the axial direction to exert synergistic antitumor effects. Tetravalent platinum compounds exhibit oral antitumor activity and have the potential to be developed into oral antitumor drugs. Compared to divalent platinum, tetravalent platinum is more stable in the bloodstream, can more completely reach cancer cells, and exhibits low toxicity. Currently, several tetravalent platinum compounds have entered clinical trials. For example, satraplatin is undergoing multiple trials in Phase I, II, and III clinical trials. In summary, quadrivalent platinum compounds are expected to solve the defects of existing platinum drugs, such as severe drug resistance, high toxicity, and only being able to be administered by injection, and are a hot topic in the research and development of a new generation of platinum drugs.

[0003] p53 is a key regulator of various stress signals. Upon activation by factors such as DNA damage, oncogene activation, hypoxia, and reactive oxygen species (ROS), p53 induces multiple cellular responses, including cell cycle arrest and apoptosis, to restore genomic integrity. p53 is considered the "guardian of the genome," preventing the accumulation of oncogenic mutations that lead to malignancy. In tumors, the anti-tumor function of p53 is suppressed. Half of tumors overexpress the MDM2 protein, which degrades p53 and inhibits its anti-tumor function. Meanwhile, half of tumors harbor p53 mutations, inhibiting its normal anti-tumor function. Most p53 mutations are missense mutations, resulting in the expression of the full-length mutant p53 protein. Mutant p53 not only loses the tumor suppressor function of wild-type p53 but also promotes tumor initiation or progression. Mutated p53 participates in cellular processes such as regulating glucose, lipids, and nucleotide metabolism in tumor cells. Furthermore, p53 mutations are closely associated with tumor resistance to radiotherapy and chemotherapy. Therefore, restoring the normal anti-tumor function of mutant p53 through small molecule drugs is a hot spot in the research and development of new anti-tumor drugs and is of great significance for solving tumor resistance.

[0004] p53 mutation activators can restore the anti-tumor function of mutant p53, induce apoptosis in tumor cells, and exert synergistic anti-tumor effects with existing anti-tumor drugs such as paclitaxel, cisplatin, azacitidine, and doxorubicin. They have been a hot topic in the research and development of anti-tumor drugs in recent years. Among them, APR-246 is the most representative p53 mutation activator and has progressed to Phase III clinical trials. Studies have found that APR-246 exhibits tumor inhibitory effects on tumor cells expressing mutant p53 from different sources. Clinical trials have confirmed that APR-246 has good safety and pharmacokinetic characteristics, and can induce p53-dependent biological effects in cancer patients with p53 mutations. Currently, multiple clinical trials of APR-246 are underway, including APR-246 combined with liposome doxorubicin for the treatment of platinum-resistant high-grade serous ovarian cancer (Phase II trial, NCT03268382), carboplatin combined with APR-246 for the treatment of recurrent high-grade serous ovarian cancer with p53 mutations (NCT02098343), APR-246 combined with azacitidine for the treatment of p53-mutant myelodysplastic syndrome (Phase III trial, NCT03745716), and APR-246 combined with azacitidine for the treatment of p53-mutant acute myeloid leukemia or myelodysplastic syndrome (Phase II trial, NCT03931291). Studies have shown that APR-246 can increase tumor sensitivity to chemotherapy drugs by depleting glutathione, which is of great significance for addressing tumor resistance to chemotherapy drugs.

[0005] The clinical development of p53 mutation activators is mainly combined with other anti-tumor drugs, but there are defects such as the inconvenience of administering multiple drugs simultaneously and the easy occurrence of drug-drug interactions. Summary of the Invention

[0006] The object of the present invention is to provide a tetravalent platinum complex containing APR-246.

[0007] The second object of the present invention is to provide a method for preparing the tetravalent platinum complex containing APR-246.

[0008] The third object of the present invention is to provide a use of the tetravalent platinum complex containing APR-246 in the preparation of anti-tumor drugs.

[0009] In order to achieve the above object, the technical solution adopted by the present invention is as follows:

[0010] The first aspect of the present invention provides a tetravalent platinum complex containing APR-246, the structure of which is shown in Formula I:

[0011]

[0012] wherein n is selected from 0, 1, 2, 3, and 4;

[0013] R is selected from

[0014] R1 is selected from a branched or linear C1 to C20 alkyl group;

[0015] R2 is selected from branched or linear C1-C20 alkyl groups.

[0016] Preferably, in the general formula I, R1 is selected from -CH3, -CH2CH3, -(CH2)2CH3, -(CH2)3CH3, -(CH2)4CH3, -(CH2)5CH3, -(CH2)7CH3, -(CH2) 11 CH3, -(CH2) 13 CH3, -(CH2) 15 CH3;

[0017] R2 is selected from -CH3, -CH2CH3, -(CH2)2CH3, -(CH2)3CH3, -(CH2)4CH3, -(CH2)5CH3, -(CH2)7CH3, -(CH2) 11 CH3, -(CH2) 13 CH3, -(CH2) 15 CH3.

[0018] Most preferably, the tetravalent platinum complex containing APR-246 is selected from one of the following structures:

[0019]

[0020] The second aspect of the present invention provides a method for preparing the tetravalent platinum complex containing APR-246, comprising the following steps:

[0021] In the first step, an excess of 30% hydrogen peroxide is added to cisplatin, i.e., compound II, and the mixture is reacted at 20-60°C for 1-4 hours. The mixture is then filtered and the filtrate is allowed to stand to obtain a yellow needle-like solid, i.e., compound III.

[0022]

[0023] In the second step, compound III and dianhydride IV at a molar ratio of 1:1 were dissolved in DMSO, reacted at room temperature, the solvent was evaporated, and the residue was recrystallized from acetone to obtain compound V;

[0024]

[0025] The dianhydride IV is selected from:

[0026] Step 3: Compound V and anhydride VI or isocyanate VI in a molar ratio of 1:1 are sequentially added to DMF, stirred at room temperature, and the solvent is evaporated to obtain compound VII.

[0027]

[0028] The acid anhydride VI is selected from n-hexanoic anhydride.

[0029] The isocyanate VI is selected from hexyl isocyanate, n-octyl isocyanate, dodecyl isocyanate, tetradecyl isocyanate, and hexadecyl isocyanate.

[0030] Step 4: Compound VII, APR-246, and EDCI at a molar ratio of 1:1:(1-2) were sequentially added to DMF, and a catalytic amount of DMAP was added. The mixture was stirred at room temperature. After the reaction, the solvent was evaporated and the mixture was purified by column chromatography to obtain Compound I.

[0031]

[0032] The third aspect of the present invention provides a use of the tetravalent platinum complex containing APR-246 in the preparation of anti-tumor drugs.

[0033] The tumor is selected from malignant tumors such as lung cancer, intestinal cancer, osteosarcoma, breast cancer, liver cancer, pancreatic cancer, and cisplatin-resistant lung adenocarcinoma.

[0034] The tumor cell line is selected from osteosarcoma SJSA-1, lung cancer A549, liver cancer HepG2, intestinal cancer HCT116, breast cancer MDA-MB-231, pancreatic cancer Panc-1, pancreatic cancer CFPAC-1, and cisplatin-resistant lung cancer A549.

[0035] Due to the adoption of the above technical solution, the present invention has the following advantages and beneficial effects:

[0036] The present invention provides a tetravalent platinum compound containing the p53 mutation activator APR-246. This compound can enhance the anti-tumor activity of APR-246 and can exert the anti-tumor advantages of APR-246 and divalent platinum drugs. By utilizing the unique anti-tumor mechanism of APR-246, it can work together with platinum drugs to exert excellent synergistic anti-tumor efficacy, and at the same time, it is also very helpful in improving the drug resistance of tumors.

[0037] The present invention provides a class of tetravalent platinum complexes containing the p53 mutation activator APR-246. This class of compounds can fully utilize the advantages of both, using the unique anti-tumor mechanism of the p53 mutation activator to solve the drug resistance of platinum drugs and exert excellent synergistic anti-tumor effects. It has a strong inhibitory effect on the proliferation process of osteosarcoma cells SJSA-1, pancreatic cancer cells Panc-1, pancreatic cancer cells CFPAC-1, liver cancer HepG2, lung cancer A549 and its cisplatin-resistant strains, intestinal cancer HCT116, and breast cancer MDA-MB-231, showing excellent broad-spectrum anti-tumor activity. Some compounds have shown excellent anti-tumor activity against cisplatin-resistant strains of lung cancer A549, such as compounds 4, 5, 6, and 7, whose anti-tumor activity is more than 21 times that of cisplatin.

[0038] The present invention provides a class of tetravalent platinum complexes containing the p53 mutation activator APR-246, which have a relatively significant proliferation inhibitory effect on liver cancer HepG2, lung cancer A549, intestinal cancer HCT116, breast cancer MDA-MB-231, and cisplatin-resistant lung adenocarcinoma. The anti-tumor activity of some compounds is significantly better than cisplatin, APR-246, and the combination of the two, and can be used as candidate anti-tumor drugs for further research. For example, compounds 4, 5, 6, and 7 generally show strong anti-tumor activity, with a half-maximal inhibitory concentration IC of 100% against osteosarcoma cells SJSA-1, pancreatic cancer cells Panc-1, pancreatic cancer cells CFPAC-1, liver cancer HepG2, lung cancer A549 and its cisplatin-resistant strain, intestinal cancer HCT116, and breast cancer MDA-MB-231. 50 were all lower than 3 μM, and showed excellent antitumor activity against cisplatin-resistant lung cancer A549 (IC 50 ≤1.80μM), and its effect is much better than cisplatin (IC 50 =39.52 μM). Other compounds, such as compound 3, showed significantly better antitumor activity against pancreatic cancer cells Panc-1, pancreatic cancer cells CFPAC-1, liver cancer cells HepG2, lung cancer cells A549 cisplatin-resistant strains, colorectal cancer cells HCT116, and breast cancer cells MDA-MB-231 than the combination of cisplatin and APR-246. Compounds 1 and 2 also showed stronger antitumor activity against colorectal cancer cells HCT116 and breast cancer cells MDA-MB-231 than cisplatin, APR-246, and their combination. The compounds provided by this invention possess novel skeletal structures and excellent antitumor activity, making them promising candidates for the development of antitumor drugs.

[0039] The tetravalent platinum complex containing the p53 mutation activator APR-246 provided by the present invention has a simple synthesis route, readily available synthetic raw materials, and an easy-to-implement synthesis method. DETAILED DESCRIPTION

[0040] In order to explain the present invention more clearly, the present invention is further described below in conjunction with preferred embodiments. Those skilled in the art should understand that the following specific description is illustrative rather than restrictive and should not be used to limit the scope of protection of the present invention.

[0041] The experimental methods in the following examples where specific conditions are not specified are generally carried out under conventional conditions or conditions recommended by the manufacturers.

[0042] The chemical structural formulas of the compounds prepared in the following examples are: 1 H-NMR - The HRMS data are shown in Table 1, where numbers 1 to 7 correspond to compounds 1 to 7 prepared in Examples 1 to 7, respectively.

[0043] Table 1. Chemical structures of target compounds 1 to 7 1 H-NMR and HRMS data

[0044]

[0045]

[0046] Example 1

[0047] Synthesis of compound 1:

[0048]

[0049] 1.0 g of cisplatin, compound II, was added to 25 mL of 30% hydrogen peroxide, reacted at 60° C. for 2 h, filtered, and the filtrate was allowed to stand to obtain 0.95 g of yellow needle-like solid cisplatin, compound III, with a yield of 85%.

[0050]

[0051] Compound III (0.20 g, 0.598 mmol) and succinic anhydride IV-1 (60 mg, 0.598 mmol) were dissolved in 3 mL of DMSO and reacted at room temperature for 48 h. After the reaction was complete, the solvent was evaporated and recrystallized from acetone to obtain a white solid, 0.25 g of compound V-1, with a yield of 96%.

[0052]

[0053] Compound V-1 (50 mg, 0.115 mmol) and n-hexanoic anhydride compound VI-1 (25 mg, 0.115 mmol) were added to 2 mL of DMF in sequence and stirred at room temperature for 24 h. After the reaction, the solvent was evaporated to obtain 59 mg of crude product VII-1, which was directly used in the next reaction.

[0054]

[0055] 59 mg of the crude compound VII-1 and APR-246 (30 mg, 0.10 mmol) from the previous step were added sequentially to 2 mL of DMF, and EDC hydrochloride (30 mg, 0.156 mmol) and DMAP (3 mg) were added. The mixture was stirred at room temperature for 12 h. After the reaction, the solvent was evaporated and the product was purified by column chromatography to obtain the target compound 1 as a white solid (15 mg, 21% yield).

[0056] Example 2

[0057] Synthesis of compound 2:

[0058]

[0059] Compound V-1 (50 mg, 0.115 mmol) and isocyanate compound VI-2 (15 mg, 0.115 mmol) were added to 2 mL of DMF in sequence and stirred at room temperature for 24 h. After the reaction, the solvent was evaporated to obtain 60 mg of crude product VII-2, which was directly used in the next reaction.

[0060]

[0061] 60 mg of the crude compound VII-2 and APR-246 (30 mg, 0.10 mmol) from the previous step were added sequentially to 2 mL of DMF, and EDC hydrochloride (30 mg, 0.156 mmol) and DMAP (3 mg) were added. The mixture was stirred at room temperature for 12 h. After the reaction, the solvent was evaporated and the target compound 2 was purified by column chromatography as a white solid (18 mg, 24% yield).

[0062] Example 3

[0063] Synthesis of compound 3:

[0064]

[0065] Refer to the synthesis method of compound 2. Replace the raw material hexyl isocyanate in Example 2 with n-octyl isocyanate, and perform other operations in the same manner. Purify by column chromatography to obtain the target compound 3 as a white solid (16 mg) with a yield of 21%.

[0066] Example 4

[0067] Synthesis of compound 4:

[0068]

[0069] Refer to the synthesis method of compound 2. Replace the raw material hexyl isocyanate in Example 2 with dodecyl isocyanate, and the other operations are the same. Purification by column chromatography afforded the target compound 4 as a white solid (20 mg) with a yield of 24%.

[0070] Example 5

[0071] Synthesis of compound 5:

[0072]

[0073] Refer to the synthesis method of compound 2. Replace the raw material hexyl isocyanate in Example 2 with tetradecyl isocyanate, and the other operations are the same. Purification by column chromatography afforded the target compound 5 as a white solid (16 mg) with a yield of 18%.

[0074] Example 6

[0075] Synthesis of compound 6:

[0076]

[0077]

[0078] Refer to the synthesis method of compound 2. Replace the raw material hexyl isocyanate in Example 2 with hexadecyl isocyanate, and the other operations are the same. Purification by column chromatography afforded the target compound 6 as a white solid (20 mg) with a yield of 18%.

[0079] Example 7

[0080] Synthesis of compound 7:

[0081]

[0082] Refer to the synthesis method of compound 2. Replace the raw materials succinic anhydride with glutaric anhydride and hexyl isocyanate with dodecyl isocyanate in Example 2. Other operations are the same. Purification by column chromatography afforded the target compound 7 as a white solid (18 mg) in a yield of 21%.

[0083] Example 8

[0084] Antitumor activity test of the compounds of the present invention

[0085] The compounds prepared in Examples 1 to 7 of the present invention were subjected to tumor cell proliferation inhibition tests using the conventional CKK-8 method.

[0086] The cell lines used were osteosarcoma SJSA-1, lung cancer A549, liver cancer HepG2, intestinal cancer HCT116, breast cancer MDA-MB-231, pancreatic cancer Panc-1, pancreatic cancer CFPAC-1 and cisplatin-resistant lung cancer A549 (A549 / CDDP), all purchased from the cell bank of Shanghai Institutes for Biological Sciences or Xiamen Yimo Biotechnology Co., Ltd.; the culture medium was DMEM + 10% NBS + double antibodies.

[0087] Sample solution preparation: Dissolve the test compound in DMSO to prepare a 10 mM stock solution. Dilute the stock solution serially with culture medium to prepare final drug concentrations of 50 μM, 25 μM, 10 μM, 5 μM, 1 μM, 0.5 μM, 0.25 μM, and 0.125 μM.

[0088] The anti-tumor compounds cisplatin, APR-246, and a 1:1 mixture of cisplatin and APR-246 were prepared into reference solutions under the same conditions.

[0089] The concentration of 8 × 10 4 100 μL of cell suspension (8000 cells / mL) was added to each well and placed in a 37°C, 5% CO2 incubator. After 24 hours, the upper culture medium was aspirated and the culture medium containing the sample and the control solution were added at 100 μL / well. The cells were incubated at 37°C for 72 hours. 10 μL of CKK-8 was added to each well and placed in an incubator. After 1 hour, the OD value at 450 nm was measured using an MK-2 fully automatic microplate reader to calculate the half-maximal inhibitory concentration (IC). 50 .

[0090] The antitumor activities of the compounds are detailed in Table 2, where samples 1 to 7 refer to the tetravalent platinum complexes containing APR-246 prepared in the corresponding examples, such as compound 1 represents the compound obtained in Example 1, and so on.

[0091] Table 2. In vitro antitumor activity of compounds 1 to 7

[0092]

[0093]

[0094] ND: Not tested.

[0095] The results in Table 2 show that the novel structural type of tetravalent platinum complexes containing the p53 mutation activator APR-246 of the present invention, the anti-tumor activity screening found that the anti-tumor activity of this type of compound is significantly better than cisplatin, APR-246 and the combination of cisplatin and APR-246. For example, compounds 4, 5, 6, and 7 generally exhibited strong anti-tumor activity, with a half-maximal inhibitory concentration IC of 0.05 for osteosarcoma cells SJSA-1, pancreatic cancer cells Panc-1, pancreatic cancer cells CFPAC-1, liver cancer HepG2, lung cancer A549 and its cisplatin-resistant strain, colorectal cancer HCT116, and breast cancer MDA-MB-231. 50 were all lower than 3 μM, and showed excellent antitumor activity against cisplatin-resistant lung cancer A549 (IC 50 ≤1.80μM), and its effect is much better than cisplatin (IC 50 =39.52 μM). Other compounds, such as compound 3, showed significantly better antitumor activity against pancreatic cancer cells Panc-1, pancreatic cancer cells CFPAC-1, liver cancer cells HepG2, lung cancer cells A549 cisplatin-resistant strains, intestinal cancer cells HCT116, and breast cancer cells MDA-MB-231 than the combined use of cisplatin and APR246. Compounds 1 and 2 also showed stronger antitumor activity against intestinal cancer cells HCT116 and breast cancer cells MDA-MB-231 than cisplatin, APR-246, and their combination. Therefore, the compounds provided by the present invention possess novel skeletal structures, excellent antitumor activity, and are promising for the development of antitumor drugs.

[0096] The above description is merely a preferred embodiment of the present invention and does not constitute any form of limitation to the present invention. Although the present invention has been disclosed as above in terms of a preferred embodiment, it is not intended to limit the present invention. Any technician familiar with this patent can make slight changes or modifications to equivalent embodiments using the above technical content without departing from the scope of the technical solution of the present invention. However, any simple modifications, equivalent changes, and modifications made to the above embodiments based on the technical essence of the present invention without departing from the content of the technical solution of the present invention are still within the scope of the solution of the present invention.

Claims

1. A tetravalent platinum complex containing APR-246, characterized in that: The structure is shown in general formula I: wherein n is selected from 0, 1, 2, 3, and 4; R is selected from R1 is selected from a branched or linear C1 to C20 alkyl group; R2 is selected from branched or linear C1-C20 alkyl groups.

2. The tetravalent platinum complex containing APR-246 according to claim 1, characterized in that In the general formula I, R1 is selected from -CH3, -CH2CH3, -(CH2)2CH3, -(CH2)3CH3, -(CH2)4CH3, -(CH2)5CH3, -(CH2)7CH3, -(CH2) 11 CH3, -(CH2) 13 CH3, -(CH2) 15 CH3; R2 is selected from -CH3, -CH2CH3, -(CH2)2CH3, -(CH2)3CH3, -(CH2)4CH3, -(CH2)5CH3, -(CH2)7CH3, -(CH2) 11 CH3, -(CH2) 13 CH3, -(CH2) 15 CH3.

3. The tetravalent platinum complex containing APR-246 according to claim 2, characterized in that The tetravalent platinum complex containing APR-246 is selected from one of the following structures:

4. A method for preparing a tetravalent platinum complex containing APR-246 according to any one of claims 1 to 3, characterized in that: The following steps are involved: In the first step, an excess of 30% hydrogen peroxide is added to cisplatin, i.e., compound II, and the mixture is reacted at 20-60°C for 1-4 hours. The mixture is then filtered and the filtrate is allowed to stand to obtain a yellow needle-like solid, i.e., compound III. In the second step, compound III and dianhydride IV at a molar ratio of 1:1 were dissolved in DMSO, reacted at room temperature, the solvent was evaporated, and the residue was recrystallized from acetone to obtain compound V; The dianhydride IV is selected from: Step 3: Compound V and anhydride VI or isocyanate VI in a molar ratio of 1:1 are sequentially added to DMF, stirred at room temperature, and the solvent is evaporated to obtain compound VII. The acid anhydride VI is selected from n-hexanoic anhydride; The isocyanate VI is selected from hexyl isocyanate, n-octyl isocyanate, dodecyl isocyanate, tetradecyl isocyanate, and hexadecyl isocyanate; Step 4: Compound VII, APR-246, and EDCI at a molar ratio of 1:1:(1-2) were sequentially added to DMF, and a catalytic amount of DMAP was added. The mixture was stirred at room temperature. After the reaction, the solvent was evaporated and the mixture was purified by column chromatography to obtain Compound I.

5. Use of a tetravalent platinum complex containing APR-246 according to any one of claims 1 to 3 in the preparation of an anti-tumor drug, characterized in that: The tumor is selected from lung cancer, intestinal cancer, osteosarcoma, breast cancer, liver cancer, and pancreatic cancer.

6. The use according to claim 5, characterized in that The lung cancer is selected from cisplatin-resistant lung cancer.

7. The use according to claim 6, characterized in that The cisplatin-resistant lung cancer cell line is selected from cisplatin-resistant lung cancer A549.

8. The use according to claim 5, characterized in that The tumor cell line is selected from osteosarcoma SJSA-1, lung cancer A549, liver cancer HepG2, intestinal cancer HCT116, breast cancer MDA-MB-231, pancreatic cancer Panc-1, and pancreatic cancer CFPAC-1.

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