A cisplatin (IV) prodrug with Chuanxiong-chalcone as ligand, and preparation method and application thereof

Abstract

The present invention belongs to the technical field of pharmaceutical chemistry, and discloses a cisplatin (IV) prodrug with Chuanxiong-chalcone as a ligand, and a preparation method and application thereof. The present invention introduces Chuanxiong-chalcone into the axis of a platinum (IV) complex, prepares a multifunctional platinum (IV) prodrug, and optimizes its structure. Studies have shown that this type of platinum (IV) complex has a significant external anti-tumor value-added effect, especially better than cisplatin in activity against pancreatic cancer cells, but has low toxicity to normal human cells (such as human umbilical vein cell line HUEVC and normal liver cell line L02). At equal doses, the representative product has better anti-tumor effect than cisplatin in a cisplatin-resistant pancreatic cancer transplantation model, but does not show obvious toxic side effects. Therefore, this type of cisplatin (IV) prodrug with Chuanxiong-chalcone as a ligand has a potential use in targeted treatment of pancreatic cancer.

Description

Technical Field

[0001] The invention belongs to the technical field of pharmaceutical chemistry and relates to the preparation of a chemical prodrug, in particular to a cisplatin (IV) prodrug with chuanxiong-chalcone as a ligand and a preparation method and application thereof. Background Art

[0002] Pancreatic cancer (PC) is one of the most common malignant tumors. Despite significant progress in diagnostic strategies and new targeted technologies, the 5-year survival rate is only about 10%. At present, traditional anti-tumor drugs are still one of the effective methods for treating primary tumors in clinical practice. Platinum anticancer drugs, such as cisplatin and its analogs, are important therapeutic drugs used in the clinical treatment of various malignant tumors including PC. However, CDDP generally has moderate to severe toxic side effects and drug resistance in various organs in clinical application. The combination of drugs can be used to intervene in CDDP resistance and improve efficacy, but due to the difficulty in predicting pharmacokinetic parameters and drug-drug interactions, the expected clinical effect may not be achieved. Therefore, the development of new platinum anticancer drugs that can delay or alleviate drug resistance while reducing tumor toxicity is of great significance for the treatment of cancer.

[0003] In order to overcome or alleviate the problems of reduced accumulation of platinum in tumor cells, enhanced DNA repair and drug inactivation, people pay more and more attention to the development of platinum (IV) prodrugs. On the one hand, these platinum (IV) prodrugs are more inert than platinum (II) drugs in kinetics, which can reduce off-target reactions with biological substances. On the other hand, the axial molecules modified by platinum (IV) prodrugs can obtain new compounds with ideal pharmacological characteristics, with advantages such as controllable pharmacokinetic parameters, enhanced cellular uptake, tumor target properties or enhanced biological activity. A large number of studies have shown that multifunctional platinum (IV) complexes show better anticancer activity than CDDP in CDDP-sensitive or resistant tumor xenograft models and have lower toxicity to major organs such as liver and kidney. Therefore, the development of multifunctional platinum (IV) prodrugs has become a promising strategy for exploring potential anti-PC drugs. Summary of the invention

[0004] In view of the shortcomings of the prior art, the purpose of the present invention is to provide a cisplatin (IV) prodrug with Chuanxiong-chalcone as a ligand. When the cisplatin (IV) prodrug of the present invention is used to treat pancreatic cancer, it overcomes the defects of existing cisplatin anti-tumor drugs in the treatment of pancreatic cancer, such as easy drug resistance and large toxic side effects. The present invention also provides a method for preparing the cisplatin (IV) prodrug.

[0005] The present invention is achieved through the following technical solutions:

[0006] A cisplatin (IV) prodrug with Chuanxiong-chalcone as a ligand, the general structural formula of which is shown below:

[0007]

[0008] Where n=1–5.

[0009] Further, in the formula, n=1, 3, 5, the specific structural formula is shown in Table 1:

[0010] Table 1 Platinum (IV) prodrug structures

[0011]

[0012] A further improvement of the present invention is:

[0013] A method for preparing a cisplatin (IV) prodrug using Chuanxiong-chalcone as a ligand comprises the following steps:

[0014] (1) reacting compound 5 with compound 6 or 7 under the catalysis of an inorganic base to synthesize compound 8 or 9;

[0015] (2) reacting compound 8 or 9 with compound 10 in the presence of an inorganic base and a catalyst to synthesize compound 11 or 12;

[0016] (3) hydrolyzing compound 11 or 12 under the action of an inorganic base to obtain compound 13 or 14;

[0017] (4) Compound 13 or 14 is reacted with a platinum (IV) intermediate in the presence of a condensing agent and an organic base to obtain compound 15 or 16; the synthetic route is as follows:

[0018]

[0019] Furthermore, the inorganic base in step (1) is a 50 wt% potassium hydroxide aqueous solution; the inorganic base in step (2) is K2CO3, and the catalyst is KI; the inorganic base in step (3) is lithium hydroxide monohydrate; the condensing agent in step (4) is O-benzotriazole-N,N,N,N-tetramethyluronium tetrafluoroborate, and the organic base is triethylamine.

[0020] Furthermore, in terms of molar amount, in step (1), compound 5: compound 6 or 7: inorganic base = 1: 1-3: 10-20, in step (2), compound 8 or 9: compound 10: inorganic base: catalyst = 1: 1-3: 2-4: 0.08-0.2, in step (3), compound 11 or 12: inorganic base = 1: 3-6, in step (4), compound 13 or 14: platinum (IV) intermediate: condensation agent: organic base = 1: 0.8-1.2: 1-2: 1-2.

[0021] Preferably, on a molar basis, in step (1), compound 5: compound 6 or 7: inorganic base = 1:1:10, in step (2), compound 8 or 9: compound 10: inorganic base: catalyst = 1:2:3:0.1, in step (3), compound 11 or 12: inorganic base = 1:4, in step (4), compound 13 or 14: platinum (IV) intermediate: condensation agent: organic base = 1:1:1.5:1.5.

[0022] Furthermore, the reaction temperature of step (1) is 20-30°C and the time is 8-16h; the reaction temperature of step (2) is 50-60°C and the time is 8-16h; the reaction temperature of step (3) is 20-30°C and the time is 3-5h; the reaction temperature of step (4) is 35-50°C and the time is 1-3h.

[0023] Furthermore, steps (1) to (4) also include a purification process.

[0024] A further improvement of the present invention is:

[0025] Application of a cisplatin (IV) prodrug with Chuanxiong-chalcone as ligand in the treatment of pancreatic cancer

[0026] Compared with the prior art, the present invention has the following beneficial effects:

[0027] The present invention introduces a chalcone derivative derived from Chuanxiong into the axial direction of a platinum (IV) intermediate to obtain a novel cisplatin (IV) prodrug with Chuanxiong-chalcone as a ligand. Studies have shown that this type of platinum (IV) complex has significant anti-tumor activity, especially against pancreatic cancer cells, but has low toxicity to normal human cells (HUEVC and L02 cell lines). In addition, the cisplatin (IV) prodrug with Chuanxiong-chalcone as a ligand of the present invention can better overcome the cisplatin resistance of pancreatic cancer cells and has a good anti-pancreatic cancer effect in vivo. It shows that this type of cisplatin (IV) prodrug has a potential use for targeted treatment of pancreatic cancer. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Figure 1 Schematic diagram of the in vivo anti-tumor effect of compound 16a in the embodiment of the present invention;

[0029] Among them, (A) images of resected tumors in each group, (B) tumor weights in each group, (C) changes in tumor volume in each group, and (D) changes in the weight of mice in each group over 15 days;

[0030] The dose of CDDP was selected as 5.0 mg / kg, and the other test compounds were tested at an equimolar amount of CDDP (16.7 mmol / kg). DETAILED DESCRIPTION

[0031] The present invention is described in detail below in conjunction with specific embodiments.

[0032] Experimental abbreviations: CDDP, cisplatin; ROS, reactive oxygen species; DCM, dichloromethane; MeOH, methanol; EA, ethyl acetate; PE, petroleum ether; GSH, glutathione; PANC-1, pancreatic cancer cells; PBS, phosphate buffered saline; A549, lung cancer alveolar basal epithelial cells; DMF, N,N-dimethylformamide; TLC, thin layer chromatography; EtOH, ethanol; P-gp, P-glycoprotein; Et3N, triethylamine ; TBTU, O-benzotriazole-N,N,N,N-tetramethyluronium tetrafluoroborate; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; MDA-MB-231, triple-negative breast cancer; MCF-7, breast cancer; DMSO, dimethyl sulfoxide; HCT-116, colon cancer cells; SGC-7901, gastric cancer cells; L02, hepatocytes; HUEVC, human umbilical vein endothelial cells.

[0033] Experimental materials: All chemicals used in the experiment were from reagent suppliers (Sinopharm, Aladdin, Anage, and McLean); solvents such as dichloromethane, petroleum ether, methanol, ethyl acetate, and N,N-dimethylformamide were of analytical grade and used directly without additional treatment (unless otherwise specified); cells used in the experiment, such as pancreatic cancer cells (PANC-1), lung cancer alveolar basal epithelial cells (A549), culture medium (DMEM), and serum (Gibco) were all from Keygene Biotechnology.

[0034] Experimental equipment: The heating device (DF-101), stirring device (KEMS-2SS), cooling device (DHJF-2005), and rotary evaporator (EXRE-207B) used in the experiment were all purchased from Nanjing Kerr Instrument Equipment Co., Ltd.; high-resolution mass spectrometer (ESI / TOF MS 6224, Agilent), nuclear magnetic resonance instrument (Bruker, 600MHz), high-performance liquid chromatography (waters, e2695system; 2489system), GmbH spectrometer (CDCl3 or DMSO-d6), reversed-phase high-performance liquid chromatography (RP-HPLC).

[0035] The synthesis of compound 5 was carried out with reference to the synthesis method reported in the literature (J.Med.Chem.2016,59,1747-1760). The raw materials used in the present invention were purchased from MacLean reagent unless otherwise stated. The structure of the compound prepared according to the method of the present invention was confirmed by hydrogen nuclear magnetic spectrum, carbon spectrum and high-resolution mass spectrum.

[0036] Example 1: Preparation of Compound 5

[0037] The preparation route is:

[0038]

[0039] Preparation of compound 2

[0040] 2,3,5,6-Tetramethyl-3,4-dihydropyridine 1-oxide (2). Compound 1 (5.0 g, 36.7 mmol) was dissolved in acetic acid (50 mL) and stirred at room temperature. Then 30% H2O2 (8.3 g, 73.4 mmol) was added dropwise to the suspension and heated at 70°C for 8 h. The reaction process was monitored by thin-layer chromatography (TLC). After the reaction was completed, the solvent was removed under reduced pressure. The obtained mixture was extracted with dichloromethane (DCM), and the combined organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure. Finally, the crude product was further purified by silica gel chromatography with a dichloromethane-methanol system (DCM:MeOH=80:1-30:1) to obtain compound 2. Yellow oily compound (4.2 g, 27.8 mmol) with a yield of 75.5%. 1 H NMR (600MHz, CDCl3) δ2.53 (s, 6H), 2.45 (s, 6H).

[0041] Preparation of compound 3

[0042] Methyl (3,5,6-trimethylpyrazine-2-yl)acetate (3). A solution of compound 2 (15.0 g, 98.5 mmol) was heated to reflux in acetic anhydride (100 mL) for 2 h, and the reaction process was monitored by thin layer chromatography. After the reaction was completed, the solvent was removed in vacuo. The mixture was extracted with ethyl acetate EA (200 mL × 3), and the combined organic phase was concentrated under reduced pressure. The crude product was further purified by ethyl acetate-petroleum ether system silica gel chromatography column (EA: PE = 1: 1) to obtain compound 3. Yellow powder (15.4 g, 79.1 mmol), yield 80.3%. 1 H NMR (600MHz, CDCl3) δ5.17(s,2H),2.52(s,3H),2.50(s,3H),2.49(s,3H),2.09(s,3H).

[0043] Preparation of compound 4

[0044] (3,5,6-Trimethylpyrazine-2-yl)methanol (4). Compound 3 (10.0 g, 51.5 mmol) was added to a mixture of MeOH (100 mL) and H2O (30 mL), and LiOH·H2O (4.3 g, 103.0 mmol) was added at room temperature and stirred for 2 h. The reaction process was monitored by TLC. After the reaction was completed, the organic solvent was removed, extracted with EA (200 mL×3), and the combined organic phase was concentrated in vacuo to obtain compound 4. Pale yellow solid powder (7.1 g, 46.4 mmol), yield 90.2%. 1 HNMR (600MHz, CDCl3) δ4.66 (s, 2H), 4.42 (s, 1H), 2.50 (s, 6H), 2.40 (s, 3H).

[0045] Preparation of compound 5

[0046] 3,5,6-Trimethylpyrazine-2-carboxaldehyde (5). Slowly add MnO2 (43.5 g, 500.0 mmol) to a DCM (250 mL) solution of compound 4, stir at room temperature for 4 h, and monitor the reaction process by TLC. After the reaction is completed, filter and remove the solvent under reduced pressure. The crude product is further purified by silica gel chromatography with a PE and EA system (1:1) to obtain compound 5. Pale yellow solid powder, yield 83.3%. 1 HNMR (600MHz, CDCl3) δ10.13(s,1H),2.78(s,3H),2.58(s,6H).

[0047] Example 2: Preparation of Compounds 8 and 9

[0048] Compound 5 (4.0 g, 26.6 mmol) and compound 6 or 7 (4.4 g, 26.6 mmol) in EtOH were added dropwise with 50% KOH (30.0 mL) at 0°C. After that, the solution was stirred at room temperature overnight. The reaction process was monitored by thin layer chromatography. After the reaction was completed, the solvent was removed under reduced pressure, the pH was adjusted to 5.5 with concentrated HCl, filtered and the organic solvent was removed by rotary evaporation. The crude product was further purified by silica gel chromatography with a dichloromethane and methanol system (80:1-30:1) to obtain compound 8 or 9.

[0049] (E)-1-(4-Hydroxy-3-methoxyphenyl)-3-(3,5,6-trimethylpyrazin-2-yl)prop-2-en-1-one (8). Yellow solid powder (4.2 g, 14.2 mmol), yield 53.3%. 1H NMR (600MHz, DMSO-d6) δ9.56(s,1H),8.05(d,J=14.9Hz,1H),7.80(d,J=14.9Hz,1H),7.59(dd,J=8.4,2.2 Hz,1H),7.48(d,J=2.2Hz,1H),7.08(d,J=8.5Hz,1H),3.87(s,3H),2.58(s,3H),2.52(s,3H),2.48(s,3H).

[0050] (E)-1-(3-Hydroxy-4-methoxyphenyl)-3-(3,5,6-trimethylpyrazin-2-yl)prop-2-en-1-one (9). Yellow solid powder (6.0 g, 20.0 mmol), yield 75.0%. 1 HNMR (600MHz, DMSO-d6) δ10.15(s,1H),8.07(d,J=14.9Hz,1H),7.79(d,J=14.9Hz,1H),7.65(dd,J=8.3,2. 0Hz, 1H), 7.54 (d, J = 2.0Hz, 1H), 6.94 (d, J = 8.2Hz, 1H), 3.86 (s, 3H), 2.58 (s, 3H), 2.51 (s, 3H), 2.48 (s, 3H).

[0051] Example 3: Preparation of Compounds 11a-12c

[0052] At the same temperature, K2CO3 (1.4 g, 10.1 mmol), catalyst KI (56 mg, 0.335 mmol) and compounds 10a-10c (6.70 mmol) were added to a DMF (25 mL) solution of compound 8 or 9 (1.0 g, 3.35 mmol), and the solution was stirred overnight at 55 ° C. The reaction process was monitored by thin layer chromatography. After the reaction was completed, the solvent was removed under reduced pressure and extracted with EA (200 mL×3). Drying with anhydrous sodium sulfate, the organic solvent was removed by rotation, and the crude product was further purified by silica gel chromatography with a DCM and MeOH system (80:1-30:1) to obtain compounds 11a-12c.

[0053] Ethyl (E)-2-(2-methoxy-4-(3-(3,5,6-trimethylpyrazin-2-yl)acryloyl)phenoxy)acetate (11a). Pale yellow solid (1.04 g, 2.72 mmol), yield 81.2%. 1H NMR (600MHz, CDCl3) δ8.17(d,J=14.9Hz,1H),7.94(d,J=14.9Hz,1H),7.83(dd,J=8.5,1.0Hz,1H),7.60(d,J=2.0Hz,1H),6.97( d,J=8.5Hz,1H),4.77(s,2H),4.28(q,J=7.1Hz,2H),3.97(s,3H),2.66(s,3H),2.57(s,3H),2.56(s,3H),1.30(t,J=7.1Hz,3H).

[0054] Ethyl (E)-4-(2-methoxy-4-(3-(3,5,6-trimethylpyrazin-2-yl)acryloyl)phenoxy)butanoic acid methyl ester (11b). Pale yellow solid (1.12 g, 2.72 mmol), yield 80.9%. 1 HNMR (600MHz, CDCl3) δ8.21(d,J=14.9Hz,1H),7.97(d,J=14.9Hz,1H),7.80(dd,J=8.4,2.0Hz,1H),7.67(d,J=2.0Hz,1H),6.96(d,J=8. 4Hz,1H),4.19-4.15(m,4H),3.96(s,3H),2.69(s,3H),2.59-2.58(m,3H),2.57-2.56(m,5H),2.25-2.17(m,2H),1.27(t,J=7.1Hz,3H).

[0055] Ethyl (E)-6-(2-methoxy-4-(3-(3,5,6-trimethylpyrazin-2-yl)acryloyl)phenoxy)hexanoic acid methyl ester (11c). Pale yellow solid (1.19 g, 2.70 mmol), yield 80.5%. 1 H NMR (600MHz, CDCl3) δ8.19(d,J=14.9Hz,1H),7.95(d,J=14.9Hz,1H),7.77(dd, J=8.4,2.0Hz,1H),7.64(d,J=2.0Hz,1H),6.93(d,J=8.4Hz,1H),4.14–4.10(m,4 H),3.95(s,3H),2.67(s,3H),2.57(s,3H),2.56(s,3H),2.34(t,J=7.5Hz,2H), 1.92-1.87(m,2H),1.73-1.69(m,2H),1.55-1.53(m,2H),1.25(t,J=7.1Hz,3H).

[0056] Ethyl (E)-2-(2-methoxy-5-(3-(3,5,6-trimethylpyrazin-2-yl)acryloyl)phenoxy)acetate (12a). Pale yellow solid (1.06 g, 2.76 mmol), yield 82.5%. 1 H NMR (600MHz, CDCl3) δ8.18(d,J=14.9Hz,1H),7.95(d,J=14.9Hz,1H),7.73(dd,J=8.4,2.0Hz,1H),7.68(d,J=2.0Hz,1H),6.85( d,J=8.4Hz,1H),4.77(s,2H),4.27(q,J=7.1Hz,2H),3.98(s,3H),2.67(s,3H),2.56(s,3H),2.55(s,3H),1.28(t,J=7.1Hz,3H).

[0057] Ethyl (E)-4-(2-methoxy-5-(3-(3,5,6-trimethylpyrazin-2-yl)acryloyl)phenoxy)butanoic acid methyl ester (12b). Pale yellow solid (1.15 g, 2.80 mmol), yield 83.4%. 1 H NMR (600MHz, CDCl3) δ8.20(d,J=14.9Hz,1H),7.96(d,J=14.9Hz,1H),7.77(dd,J=8.4,2.0Hz,1H),7.66(d,J=2.0Hz,1H),6.9 5(d,J=8.4Hz,1H),4.18-4.13(m,4H),3.95(s,3H),2.68(s,3H),2.58-2.55(m,8H),2.22-2.18(m,2H),1.26(t,J=7.1Hz,3H).

[0058] Ethyl (E)-6-(2-methoxy-5-(3-(3,5,6-trimethylpyrazin-2-yl)acryloyl)phenoxy)hexanoic acid methyl ester (12c). Pale yellow solid (1.23 g, 2.80 mmol), yield 83.6%. 1HNMR(600MHz, CDCl3)δ8.20(d,J=14.9Hz,1H),7.96(d,J=14.9Hz,1H),7.76(dd ,J=8.4,2.0Hz,1H),7.65(d,J=2.0Hz,1H),6.92(d,J=8.4Hz,1H),4.16-4.09(m, 4H),3.95(s,3H),2.67(s,3H),2.57(s,3H),2.56(s,3H),2.34(t,J=7.5Hz,2H), 1.93-1.88(m,2H),1.77-1.66(m,2H),1.56-1.51(m,2H),1.25(t,J=7.1Hz,3H).

[0059] Example 4: Preparation of Compounds 13a-14c

[0060] LiOH·H2O (588.0 mg, 14.0 mmol) was added to a solution of compound 11 or compound 12 (3.5 mmol) in MeOH (25 mL) and H2O (10 mL), and the mixture was stirred at room temperature for 4 h. The reaction process was monitored by thin layer chromatography. After the reaction was completed, the solvent was removed under reduced pressure, the pH was adjusted to 5.5, and the mixture was extracted with DCM (200 mL×3). The organic layer was collected, dried over anhydrous sodium sulfate, and the organic solvent was removed by rotation. The crude product was further purified by silica gel chromatography using a DCM and MeOH system (50:1-25:1) to obtain compounds 13a-14c.

[0061] (E)-2-(2-methoxy-4-(3-(3,5,6-trimethylpyrazin-2-yl)acryloyl)phenoxy)acetic acid (13a). Pale yellow solid (1.07 g, 3.0 mmol), yield 85.6%. 1 HNMR(600MHz,DMSO-d6)δ13.07(s,1H),8.07(d,J=14.9Hz,1H),7.80(dd,J=15.6,8.5Hz,2H),7.46(d,J=2.0H z,1H),7.16(d,J=8.5Hz,1H),4.79(s,2H),3.90(s,3H),2.59(s,3H),2.53(s,3H),2.50(d,J=3.6Hz,3H).[MH] - Calculate for C 19 H 20 N2O5355.1267, found 355.1265.

[0062] (E)-4-(2-methoxy-4-(3-(3,5,6-trimethylpyrazin-2-yl)acryloyl)phenoxy)butanoic acid (13b). Pale yellow solid (1.14 g, 2.96 mmol), yield 84.5%. 1 HNMR (600MHz, DMSO-d6) δ12.17(s,1H),8.08(d,J=14.9Hz,1H),7.81(d,J=14.9Hz,1H),7.73(dd,J=8.5,2.0Hz,1H),7.53(d,J=2.0Hz,1H),7. [MH] - Calculate for C 21 H 24 N2O5383.1572, found 383.1587.

[0063] (E)-6-(2-methoxy-4-(3-(3,5,6-trimethylpyrazin-2-yl)acryloyl)phenoxy)hexanoic acid (13c). Pale yellow solid (1.24 g, 3.02 mmol), yield 86.3%. 1 HNMR(600MHz,DMSO-d6)δ12.00(s,1H),8.10(d,J=14.9Hz,1H),7.83(d,J=14 .9Hz,1H),7.75(d,J=8.3Hz,1H),7.55(s,1H),7.13(d,J=8.4Hz,1H),4.08(t ,J=6.3Hz,2H),3.86(s,3H),2.60(s,3H),2.53(s,3H),2.51(s,3H),2.25(t, J=7.2Hz,2H),1.78-1.76(m,2H),1.61-1.56(m,2H),1.47-1.43(m,2H).[MH] - Calculate for C 23 H 28 N2O5411.1925, found 411.1900.

[0064] (E)-2-(2-Methoxy-5-(3-(3,5,6-trimethylpyrazin-2-yl)acryloyl)phenoxy)acetic acid (14a). Pale yellow solid (1.07 g, 3.01 mmol), yield 85.9%. 1HNMR(600MHz,DMSO-d6)δ13.06(s,1H),8.06(d,J=14.9Hz,1H),7.82-7.78(m,2H),7.45(d,J=2.0Hz ,1H),7.16(d,J=8.5Hz,1H),4.78(s,2H),3.90(s,3H),2.58(s,3H),2.52(s,3H),2.49(s,3H).[MH] - Calculate for C 19 H 20 N2O5355.1264, found355.1277.

[0065] (E)-4-(2-methoxy-5-(3-(3,5,6-trimethylpyrazin-2-yl)acryloyl)phenoxy)butanoic acid (14b). Pale yellow solid (1.17 g, 3.06 mmol), yield 87.4%. 1 H NMR (600MHz, DMSO-d6) 12.15(s,1H), δ8.07(d,J=14.9Hz,1H),7.84(s,1H),7.76(dd,J=8.5,2.0Hz,1H),7.53(d,J=2.0Hz,1H),7.13(d,J= 8.5Hz,1H),4.06(t,J=6.4Hz,2H),3.88(s,3H),2.58(s,3H),2.52(s,3H),2.48(s,3H),2.42(t,J=7.4Hz,2H),1.97(p,J=6.8Hz,2H).[MH] - Calculate for C 21 H 24 N2O5383.1577, found383.1584.

[0066] (E)-6-(2-methoxy-5-(3-(3,5,6-trimethylpyrazin-2-yl)acryloyl)phenoxy)hexanoic acid (14c). Pale yellow solid (1.22 g, 2.96 mmol), yield 84.8%. 1H NMR (600MHz, DMSO-d6) δ12.02(s,1H),8.08(d,J=14.9Hz,1H),7.83(d,J=14 .9Hz,1H),7.75(d,J=8.3Hz,1H),7.54(s,1H),7.12(d,J=8.4Hz,1H),4.07(t ,J=6.3Hz,2H),3.85(s,3H),2.59(s,3H),2.53(s,3H),2.50(s,3H),2.24(t, J=7.2Hz,2H),1.80-1.70(m,2H),1.63-1.53(m,2H),1.48-1.40(m,2H).[MH] - Calculate for C 23 H 28 N2O5411.1898, found 411.1896.

[0067] Example 5: Preparation of Compounds 15a-16c

[0068] In a DMF (5 mL) solution of compound 13 or 14 (0.40 mmol), O-benzotriazole-N,N,N,N-tetramethyluronium tetrafluoroborate (TBTU) (189.4 mg, 0.60 mmol), Et3N (59.6 mg, 0.59 mmol) and platinum (IV) (151.2 mg, 0.44 mmol) were added. Then, under nitrogen protection, the mixture was stirred at 40 ° C for 2 h. The reaction process was monitored by thin layer chromatography. After the reaction was completed, the solvent was removed under reduced pressure and extracted with DCM (100 mL×3). The organic layer was dried over anhydrous sodium sulfate and the solvent was removed by rotation. The crude product was further purified by silica gel chromatography using a DCM and MeOH system (40:1-15:1) to obtain compounds 15a-16c.

[0069] (OC-6-33)-Trichloroamine diamine ((E) 2-(2-methoxy-4-(3-(3,5,6-trimethylpyrazin-2-yl) acryloyl) phenoxy) acetic acid) platinum (IV) (15a). Pale yellow solid (97.6 mg, 0.14 mmol), yield 35.3%. 1 H NMR (600MHz, DMSO-d6) δ8.08(d,J=14.9Hz,1H),7.81(d,J=14.9Hz,1H),7.65(dd,J=8.5,1.9Hz,1H),7.54(d,J=1.9 Hz,1H),7.11(d,J=8.6Hz,1H),6.28-6.15(m,6H),4.75(s,2H),3.86(s,3H),2.58(s,3H),2.51(s,3H),2.48(s,3H).13 C NMR(150MHz,DMSO-d6)δ187.79,175.15,153.25,152.73,150.21,149.75,149.36,142.7 4,137.81,130.94,126.14,123.47,113.34,111.24,66.19,56.14,22.16,21.95,20.91.

[0070] (OC-6-33)-Trichloroamine diamine ((E)-4-(2-methoxy-4-(3-(3,5,6-trimethylpyrazin-2-yl)acryloyl)phenoxy)butyric acid methyl ester) platinum (IV) (15b). Pale yellow solid (105.8 mg, 0.15 mmol), yield 36.8%. 1 H NMR (600MHz, DMSO-d6) δ8.08(d,J=14.9Hz,1H),7.81(d,J=14.9Hz,1H),7.74(dd,J=8.4,1.7Hz,1H),7.53(d,J=1.8Hz,1H),7.14(d,J=8.4Hz, 1H),6.26-6.12(m,6H),4.14(t,J=6.5Hz,2H),3.86(s,3H),2.58(s,3H ),2.52(s,3H),2.48(s,3H),2.43(t,J=7.2Hz,2H),1.98-1.94(m,2H). 13 CNMR(150MHz,DMSO-d6)δ187.80,180.09,153.28,153.22,150.19,149.74,149.48,142.77,137 .77,130.53,126.18,123.90,112.31,110.99,68.17,56.04,32.84,25.52,22.15,21.95,20.92.

[0071] (OC-6-33)-Trichloroamine diamine ((E)-6-(2-methoxy-4-(3-(3,5,6-trimethylpyrazin-2-yl)acryloyl)phenoxy)hexanoic acid methyl ester) platinum (IV) (15c). Pale yellow solid (110.8 mg, 0.15 mmol), yield 37.1%. 1H NMR(600MHz,DMSO-d6)δ8.08(d,J=14.9Hz,1H),7.81(d,J=14.9Hz,1H),7.74- 7.73(m,1H),7.53(d,J=1.6Hz,1H),7.11(d,J=8.5Hz,1H),6.26-6.12(m,6H), 4.06(t,J=6.5Hz,2H),3.85(s,3H),2.58(s,3H),2.52(s,3H),2.48(s,3H),2. 27(t,J=7.3Hz,2H),1.78-1.73(m,2H),1.58-1.53(m,2H),1.47-1.42(m,2H). 13 C NMR(150MHz,DMSO-d6)δ187.78,181.08,153.38,153.22,150.19,149.74,149.52,142.77,137.77,130 .59,126.17,123.90,112.27,111.11,68.84,56.09,36.69,28.85,25.64,25.54,22.15,21.94,20.91.

[0072] (OC-6-33)-Trichloroamine diamine ((E)-2-(2-methoxy-5-(3-(3,5,6-trimethylpyrazin-2-yl)acryloyl)phenoxy)acetate) platinum (IV) (16a). Pale yellow solid (108.9 mg, 0.16 mmol), yield 39.4%. 1 H NMR(600MHz,DMSO-d6)δ8.06(d,J=15.0Hz,1H),7.81(d,J=15.0Hz,1H),7.76-7.75(m,1H),7.56(d,J=2.0Hz,1 H),7.14(d,J=8.5Hz,1H),6.28-6.16(m,6H),4.71(s,2H),3.88(s,3H),2.59(s,3H),2.53(s,3H),2.49(s,3H). 13 C NMR(150MHz,DMSO-d6)δ187.88,175.86,154.14,153.19,150.23,149.75,148.12,142.8 2,137.90,130.75,126.34,124.24,113.65,111.91,66.65,56.38,22.16,21.96,20.96.

[0073] (OC-6-33)-Trichloroamine diamine ((E) methyl 4-(2-methoxy-5-(3-(3,5,6-trimethylpyrazin-2-yl)acryloyl)phenoxy)butyrate) platinum (IV) (16b). Pale yellow solid (103.2 mg, 0.14 mmol), yield 35.9%. 1 HNMR (600MHz, DMSO-d6) δ8.07(d,J=14.9Hz,1H),7.81(d,J=14.9Hz,1H),7.80-7.74(m,1H),7.52(d,J=1.8Hz,1H),7.12(d,J=8.5Hz,1H) ,6.26-6.12(m,6H),4.08(t,J=6.4Hz,2H),3.88(s,3H),2.58(s,3H),2.52(s,3H),2.48(s,3H),2.44(t,J=7.3Hz,2H),1.98-1.94(m,2H). 13 C NMR (150MHz, DMSO-d6) δ187.85,180.23,154.06,153.22,150.20,149.74,148.78,142.77,137. 80,130.73,126.19,123.82,111.82,111.54,68.03,56.33,32.93,25.74,22.16,21.96,20.92.

[0074] (OC-6-33)-trichloroamine diamine ((E)-6-(2-methoxy-5-(3-(3,5,6-trimethylpyrazin-2-yl)acryloyl)phenoxy)hexanoic acid methyl ester)-platinum(IV) (16c). 1 HNMR(600MHz,DMSO-d6)δ8.07(d,J=14.8Hz,1H),7.81(d,J=14.8Hz,1H),7.75(d,J=7.1Hz,1H),7.52(s,1H),7.12(d,J=7.7Hz,1H), 6.20(s,6H),4.03(s,2H),3.88(s,3H),3.33(s,3H),2.58(s,3H),2.52(s,3H),2.27(s,2H),1.75(s,2H),1.56(s,2H),1.46(s,2H). 13C NMR(150MHz,DMSO-d6)δ187.84,181.07,154.14,153.23,150.20,149.75,148.81,142.77,137.80,130 .73,126.18,123.85,112.01,111.63,68.72,56.35,36.69,29.00,25.68,25.62,22.16,21.96,20.92.

[0075] Example 6: In vitro antiproliferative activity assay:

[0076] The antiproliferative activity of the compounds in human cancer cell lines (PANC-1, pancreatic cancer; MDA-MB-231, triple-negative breast cancer; MCF-7, breast cancer; A549, lung cancer alveolar basal epithelial cells; HCT-116, colon cancer cells; and HUEVC, human umbilical vein endothelial cells) was evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay.

[0077] 1. Experimental Methods

[0078] All cells tested were divided into 4 × 10 3 The cells were inoculated at a density of 100 μL / well in a 96-well plate and left overnight. The tested compound was dissolved in DMF (concentration less than 0.2%) and diluted to the required concentration with phosphate buffered saline (PBS, pH=7.4), while CDDP was dissolved in physiological saline. As shown in Tables 1 and 2, after 72 hours of action of different concentrations of compounds, the cells were incubated with 10 μL MTT (5 mg / mL) at 37°C for 4 hours and with 150 μL dimethyl sulfoxide (DMSO) for 1 hour, and the absorbance at 570 nm was recorded with a microtiter plate photometer. The IC50 value was calculated using GraphPad Prism software to analyze the cell antiproliferative activity.

[0079] Table 1 Antiproliferative effects of compounds on tumor cell lines

[0080]

[0081]

[0082] Table 1 shows the results of the in vitro antitumor activity of the compounds on various cancer cell lines determined by the MTT method in the examples of the present invention. The cells were treated three times with the tested compounds at 37°C in the dark for 72 hours; the MTT method was used to determine the b IC 50 Values, expressed as mean ± SD; c CDDP, cisplatin; dCDDP+14a, CDDP:14a=1:1 (equimolar mixture, mol / mol).

[0083] Table 2 Antiproliferative effects of compounds on cisplatin-resistant cancer cell lines

[0084]

[0085] Table 2 shows the results of the antiproliferative activity of the compounds on various cisplatin-resistant cancer cell lines and normal cells measured by the MTT method in the examples of the present invention. Three independent experiments were performed on the cells at 37°C with different compounds, and the cells were treated in the dark for 72 hours. The results were taken as the average of the three experiments and expressed as mean±SD.

[0086] 2. Experimental results

[0087] As shown in Table 1 , among the target complexes, compound 16a has the strongest ability to inhibit cancer cell proliferation, and its inhibitory effect on PANC-1 cells is 4.7 times and 2.7 times greater than that of CDDP and 14a, respectively.

[0088] As shown in Table 2, the IC values ​​of 14a and 16a for A549 / CDDP and PANC-1 / CDDP 50 The values ​​were 3.99, 2.98, 2.07 and 1.02 μM, respectively, which were better than CDDP in terms of inhibitory effect. In addition, the RF1 and RF2 values ​​of 14a and 16a in A549 / CDDP and PANC-1 / CDDP were 3.87 and 4.47 times lower than those of CDDP, respectively, indicating that compounds 14a and 16a can effectively overcome the drug resistance of CDDP. At the same time, the RF value of the mixed group of 14a and CDDP was lower than that of CDDP (RF1, 1.72; RF2, 1.94), but the selectivity index (SI) for normal human cells (HUEVC and L02) was lower than that of 16a. In summary, 16a showed good antitumor activity against both CDDP-sensitive and resistant PANC-1 cells in vitro, and had strong tumor selectivity, which is worthy of further study of the antitumor mechanism.

[0089] Example 7: In vivo anticancer experiment:

[0090] 1. Model building

[0091] Twenty-five male athymic (Balb / c-nu) mice, 4 weeks old, were purchased from Zhejiang Charles River Laboratory Animal Co., Ltd. and raised under pathogen-free conditions. PANC-1 / CDDP cells (1×10 7 / mouse) were subcutaneously inoculated on the right side of mice to establish the PANC-1 / CDDP xenograft model. Figure 1 As shown, when the tumor volume reaches about 150mm 3At the time of the experiment, mice were randomly divided into five groups: saline treatment group, CDDP (5.0 mg / kg), 14a (5.92 mg / kg), 14a + CDDP (5.92 + 5.0 mg / kg) and 16a (11.51 mg / kg) treatment group. 14a and 16a were dissolved in a mixture containing 0.1% Tween-80 and 2.0% DMF and diluted to the required concentration with saline. CDDP was dissolved in saline. All mice were intravenously injected once every 7 days for 28 consecutive days. According to the formula 0.5 × length × width 2 To calculate tumor volume, body weight was measured every other day for 28 days.

[0092] 2. Experimental results

[0093] like Figure 1 As shown, CDDP showed only moderate tumor inhibitory activity with a tumor growth inhibition (TGI) rate of 33.5% compared with the saline-treated group. 14a was superior to CDDP in inhibiting tumor growth (TGI rate of 45.7%). However, when CDDP and 14a were mixed (1:1, mol / mol) or treated with a novel entity (16a), the TGI rates increased by 57.2% and 68.3%, respectively, indicating that 14a and CDDP jointly inhibited tumor growth. In addition, CDDP caused body weight loss within 12 days after two administrations compared with the saline-treated group, indicating the potential toxicity of CDDP. However, 14a or 16a monotherapy showed almost no weight loss in xenograft mice, while the combination treatment group lost weight after two administrations, indicating that the drug combination in the mixed group did not alleviate the systemic toxicity of CDDP. Therefore, 16a with 14a in the axial position of platinum (IV) can obtain better in vivo antitumor activity and lower toxicity than CDDP, and is a promising therapeutic approach.

[0094] The above embodiments are only for illustrating the technical concept and features of the present invention, and their purpose is to enable people familiar with the technology to understand the content of the present invention and implement it accordingly, and they cannot be used to limit the protection scope of the present invention. Any equivalent transformation or modification made according to the spirit of the present invention should be included in the protection scope of the present invention.

Claims

1. A cisplatin (IV) prodrug with Chuanxiong-chalcone as a ligand, characterized in that: The general structural formula is shown below: Where n=1–5.

2. A cisplatin (IV) prodrug with Chuanxiong-chalcone as a ligand according to claim 1, characterized in that: In the formula, n=1, 3, 5.

3. The method for preparing a cisplatin (IV) prodrug with Chuanxiong-chalcone as a ligand according to claim 1, characterized in that: The following steps are involved: (1) reacting compound 5 with compound 6 or 7 under the catalysis of an inorganic base to synthesize compound 8 or 9; (2) reacting compound 8 or 9 with compound 10 in the presence of an inorganic base and a catalyst to synthesize compound 11 or 12; (3) hydrolyzing compound 11 or 12 under the action of an inorganic base to obtain compound 13 or 14; (4) reacting compound 13 or 14 with a platinum (IV) intermediate in the presence of a condensing agent and an organic base to obtain compound 15 or 16; The synthetic route is as follows:

4. The method for preparing a cisplatin (IV) prodrug with Chuanxiong-chalcone as a ligand according to claim 3, characterized in that: The inorganic base in step (1) is a 50wt% potassium hydroxide aqueous solution; the inorganic base in step (2) is K2CO3, and the catalyst is KI; the inorganic base in step (3) is lithium hydroxide monohydrate; the condensing agent in step (4) is O-benzotriazole-N,N,N,N-tetramethyluronium tetrafluoroborate, and the organic base is triethylamine.

5. The method for preparing a cisplatin (IV) prodrug with Chuanxiong-chalcone as a ligand according to claim 3, characterized in that: In terms of molar amount, in step (1), compound 5: compound 6 or 7: inorganic base = 1: 1-3: 10-20; in step (2), compound 8 or 9: compound 10: inorganic base: catalyst = 1: 1-3: 2-4: 0.08-0.2; in step (3), compound 11 or 12: inorganic base = 1: 3-6; in step (4), compound 13 or 14: platinum (IV) intermediate: condensation agent: organic base = 1: 0.8-1.2: 1-2: 1-2.

6. The method for preparing a cisplatin (IV) prodrug with Chuanxiong-chalcone as a ligand according to claim 3, characterized in that: The reaction temperature of step (1) is 20-30°C and the time is 8-16h; the reaction temperature of step (2) is 50-60°C and the time is 8-16h; the reaction temperature of step (3) is 20-30°C and the time is 3-5h; the reaction temperature of step (4) is 35-50°C and the time is 1-3h.

7. The method for preparing a cisplatin (IV) prodrug with Chuanxiong-chalcone as a ligand according to claim 3, characterized in that: Steps (1) to (4) also include a purification process.

8. Use of a cisplatin (IV) prodrug with Chuanxiong-chalcone as a ligand as claimed in claim 1 in the preparation of an anti-pancreatic cancer drug.

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