13-O-decanoyl-phorbol ester derivative, preparation method thereof and application of 13-O-decanoyl-phorbol ester derivative in preparation of antitumor drugs

Abstract

The invention discloses a 13-O-decanoyl-phorbol ester derivative, a preparation method thereof and application of the 13-O-decanoyl-phorbol ester derivative in preparation of antitumor drugs, and belongs to the technical field of drugs, and the 13-O-decanoyl-phorbol ester derivative is shown as a general formula (I). According to the invention, nine phorbol ester derivatives are designed and synthesized, and the derivatives have better anti-proliferative activity on three tumor cells, namely A2780 (human ovarian cancer cells), Hela (cervical cancer cell line) and HepG2 (human liver cancer cells), and have the potential of being developed into novel anti-tumor drugs.

Description

Technical Field

[0001] The present invention relates to the technical field of medicines, and in particular to a 13-O-decanoyl-phorbol ester derivative, a preparation method thereof and an application thereof in the preparation of anti-tumor drugs. Background Art

[0002] Cancer is one of the most common and dangerous diseases that affect human health worldwide. According to statistics, more than 14 million people suffer from cancer every year, making it one of the main causes of death. At present, the treatment of tumors mainly adopts a combination of surgery, chemotherapy, radiotherapy and biological therapy, but the treatment effect is not satisfactory. Therefore, researchers have been working hard to find new tumor treatment targets and develop new therapeutic drugs, especially natural small molecule drugs. In addition, natural products have huge scaffold diversity, complexity and biological activity, and have long played a vital role in drug discovery and development, especially as anticancer and anti-infective agents.

[0003] Phorbol ester is a tetracyclic diterpenoid compound, which is widely distributed in plants of the Thymelaeaceae and Euphorbiaceae families, such as Sapium sebiferum, Glechoma longituba, Euphorbia lactea, Ephedra densiflora, Jatropha curcas, Syringa xylostellaria, Lacquercus ovata, and Populus australis. It has structural diversity due to the presence of multiple oxygen-containing functional groups in its polycyclic skeleton and has multiple biological activities, such as tumor-promoting activity, anti-tumor activity, anti-HIV activity, and anti-viral activity.

[0004] Protein kinase C (PKC) is a class of serine / threonine protein kinases. According to their cofactor requirements, PKC is divided into three subfamilies, including traditional (α, β, γ), novel (δ, ε, η, θ) and atypical (ζ, λ, ι) subfamilies. The activation of traditional PKC depends on diacylglycerol (DAG) and Ca 2+ , while the novel PKC only depends on DAG, and the atypical PKC does not depend on DAG and Ca 2 + Phorbol esters, as potent and long-lasting DAG analogs, can activate both conventional and novel PKCs, affecting the activities of multiple enzymes, protein biosynthesis, DNA, polyamines, cell differentiation processes, and gene expression. In addition, recent studies have shown that cancer-associated PKC mutations are often loss-of-function, suggesting that PKC has a tumor suppressor role, giving rise to the hypothesis that restoring rather than inhibiting PKC activity may be a new strategy to combat cancer.

[0005] So far, there is no report on the preparation method and application of 13-O-decanoyl-phorbol ester derivatives in the prior art. Summary of the invention

[0006] The present invention provides a 13-O-decanoyl-phorbol ester derivative and a preparation method thereof and application of the 13-O-decanoyl-phorbol ester derivative in the preparation of an anti-tumor drug. The present invention finds through creative research that the 13-O-decanoyl-phorbol ester derivative has a potential therapeutic effect on human ovarian cancer cells (A2780), cervical cancer cells (Hela) and human liver cancer cells (HepG2).

[0007] In order to achieve the above purpose, the embodiment of the present invention provides the following technical solutions:

[0008] A 13-O-decanoyl-phorbol ester derivative, the structure of which is shown in general formula (I): Wherein R is H (compound 4), (Compound 6a), (Compound 6b), (Compound 6c), (Compound 6d), (Compound 6e), (Compound 6f), (Compound 6g) or (Compound 6h).

[0009] The preparation method of the above-mentioned 13-O-decanoyl-phorbol ester derivatives, the synthetic route of the compound represented by general formula (I) is as follows:

[0010]

[0011] The synthesis process is as follows:

[0012] (1) Compound 1 (phorbol) reacts with tert-butyldimethylsilyl chloride to selectively protect 20-OH to obtain compound 2;

[0013] (2) Compound 2 is condensed with n-decanoic acid to form an ester to obtain compound 3;

[0014] (3) Compound 3 is subjected to the treatment of perchloric acid to remove the tert-butyldimethylsilyl (TBS) protecting group to obtain compound 4;

[0015] (4) Compound 3 was condensed with different carboxylic acids to form esters to prepare compounds 5a-h;

[0016] (5) Compounds 5a-h were treated with perchloric acid to remove the TBS protecting group to obtain compounds 6a-h.

[0017] Furthermore, the specific preparation process of compound 4 is as follows:

[0018] S1. Dissolve phorbol in pyridine, add tert-butyldimethylsilyl chloride at -5°C to 5°C, stir at room temperature until the reaction is complete, add saturated NaHCO3 solution to quench the reaction, extract with ethyl acetate, combine the organic layers, rinse the organic layers with saturated CuSO4 solution, separate and dry with anhydrous sodium sulfate, filter, concentrate under reduced pressure, and separate and purify by column chromatography to obtain compound 2;

[0019] S2. Dissolve compound 2 and n-decanoic acid in dichloromethane, add EDCI at -5°C to 5°C under nitrogen protection, stir for 10 to 20 min, then add DMAP, stir for 5 to 15 min, move to room temperature, stir until the reaction is complete, add saturated NaHCO3 solution to quench the reaction, extract with dichloromethane, separate the liquids and dry with anhydrous sodium sulfate, filter, and concentrate under reduced pressure to obtain a crude product of compound 3, add HClO4 in MeOH solution, stir at room temperature until the reaction is complete, add saturated NaHCO3 solution to quench the reaction, extract with ethyl acetate, separate the liquids and dry with anhydrous sodium sulfate, filter, and concentrate under reduced pressure, separate and purify by column chromatography to obtain compound 4.

[0020] The molar ratio of phorbol to tert-butyldimethylsilyl chloride is 1:5, the molar ratio of compound 2, n-decanoic acid, EDCI and DMAP is 1:(1.0-1.5):(1.0-1.5):1; and the molar ratio of compound 2 to perchloric acid is 1:25.

[0021] Furthermore, the specific preparation process of compounds 6a~h is as follows:

[0022] S1. Dissolve phorbol in pyridine, add tert-butyldimethylsilyl chloride at -5°C to 5°C, stir at room temperature until the reaction is complete, add saturated NaHCO3 solution to quench the reaction, extract with ethyl acetate, combine the organic layers, rinse the organic layers with saturated CuSO4 solution, separate and dry with anhydrous sodium sulfate, filter, concentrate under reduced pressure, and separate and purify by column chromatography to obtain compound 2;

[0023] S2, compound 2 and n-decanoic acid were dissolved in dichloromethane, EDCI was added at -5°C to 5°C under nitrogen protection, DMAP was added after stirring for 10 to 20 minutes, the mixture was stirred for 5 to 15 minutes and then moved to room temperature, stirred for complete reaction, saturated NaHCO3 solution was added to quench the reaction, the mixture was extracted with dichloromethane, the liquid was separated and dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and separated and purified by column chromatography to obtain compound 3;

[0024] S3. Dissolve compound 3 and acetic acid, (S)-2-methylbutyric acid, benzoic acid, 4-chlorobenzoic acid, 4-fluorobenzoic acid, 4-methoxybenzoic acid, 3-picolinic acid or 2-pyrazinecarboxylate in dichloromethane, add EDCI at -5°C to 5°C, stir for 10 to 20 min, then add DMAP, stir for 5 to 15 min, move to room temperature, stir until the reaction is complete, add saturated NaHCO3 solution to quench the reaction, extract with dichloromethane, separate the liquids and dry with anhydrous sodium sulfate, filter, concentrate under reduced pressure, add HClO4 in MeOH solution, stir at room temperature until the reaction is complete, add saturated NaHCO3 solution to quench the reaction, extract with ethyl acetate, separate the liquids and dry with anhydrous sodium sulfate, filter, concentrate under reduced pressure, separate and purify by column chromatography to obtain compounds 6a to 6h.

[0025] Among them, the molar ratio of phorbol and tert-butyldimethylsilyl chloride is 1:5, the molar ratio of compound 2, n-decanoic acid, EDCI and DMAP is 1:(1.0-1.5):(1.0-1.5):1; the molar ratio of compound 3 and acetic acid, (S)-2-methylbutyric acid, benzoic acid, 4-chlorobenzoic acid, 4-fluorobenzoic acid, 4-methoxybenzoic acid, 3-pyridinecarboxylic acid or 2-pyrazinecarboxylate and EDCI, DMAP is 1:3:3.5:3; the molar ratio of compound 3 and perchloric acid is 1:25.

[0026] The use of the aforementioned 13-O-decanoyl-phorbol ester derivatives in the preparation of anti-tumor drugs, wherein the tumor refers to human ovarian cancer, cervical cancer or human liver cancer.

[0027] Furthermore, the 13-O-decanoyl-phorbol ester derivatives are used in the preparation of drugs for inhibiting tumor cell proliferation, inducing pyroptosis or promoting apoptosis, and the tumor cells are A2780, Hela or HepG2.

[0028] The present invention uses the MTT method to carry out in vitro activity test on the 13-O-decanoyl-phorbol ester derivatives. The activity test shows that the compound synthesized by the present invention has good anti-proliferation activity against human ovarian cancer cells (A2780), cervical cancer cells (Hela) and human liver cancer cells (HepG2). The IC of compound 6b is 50 The lowest value is 13.58μM.

[0029] The present invention has the following advantages:

[0030] 1. The present invention designs and synthesizes 9 13-O-decanoyl-phorbol ester derivatives. The research results show that the preferred compound 6b has good inhibitory effects on A2780, HepG2 and Hela cells, IC 50 The values ​​were 13.58, 19.79 and 16.94 μM, respectively, which were stronger than the positive control group TPA.

[0031] 2. The synthetic route designed by the present invention has not been reported.

[0032] 3. Compound 6b exists in natural products. DETAILED DESCRIPTION

[0033] For a better understanding of the present invention, the present invention is further described in detail below in conjunction with embodiments, but the scope of protection claimed by the present invention is not limited to the scope represented by the embodiments.

[0034] Example 1: Synthesis of Compound 4

[0035] Compound phorbol (72.9 mg, 0.2 mmol) was dissolved in 5 mL of pyridine, tert-butyldimethylsilyl chloride (150.7 mg, 1 mmol) was slowly added at 0°C, and the reaction was allowed to proceed for 1 h at room temperature. TLC showed that the reaction was complete. Saturated NaHCO3 solution was added to quench the reaction, and the mixture was extracted with ethyl acetate (50 mL×3). The organic layers were combined, and the organic layers were washed with saturated CuSO4 solution. After separation, the organic layers were dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and separated and purified by column chromatography (CH2Cl2:MeOH=7:1, v / v) to obtain compound 2 (78.2 mg, white solid) with a yield of 82%.

[0036] Compound 2 (47.8 mg, 0.1 mmol) and n-decanoic acid (20.7 mg, 0.12 mmol) were dissolved in 10 mL of anhydrous dichloromethane, nitrogen was protected, EDCI (28.8 mg, 0.15 mmol) was slowly added at 0°C, stirred for 15 min, DMAP (12.2 mg, 0.1 mmol) was slowly added, stirred for 10 min and then moved to room temperature, reacted at room temperature overnight, TLC showed that the reaction was complete. Saturated NaHCO3 solution was added to quench the reaction, extracted with dichloromethane (50 mL × 3), dried with anhydrous sodium sulfate after separation, filtered, and concentrated under reduced pressure. 10 mL of methanol was added, 70% HClO4 (214 μL, 2.5 mmol) was added dropwise during stirring, and reacted at room temperature for 1 h, TLC showed that the reaction was complete. Saturated NaHCO3 solution was added to quench the reaction, and the mixture was extracted with ethyl acetate (50 mL×3). After separation, the mixture was dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and separated and purified by column chromatography (CH2Cl2:MeOH=15:1, v / v) to obtain compound 4 (45.6 mg, white solid) with a yield of 88%.

[0037] HRMS(ESI):C 30 H 46 O7Na[M+Na] + Theoretical value: 541.3142, actual value: 541.3137.

[0038] 1 H NMR(400MHz,Chloroform-d)δ7.55(d,J=2.3Hz,1H),5.63(d,J=5.4Hz,1H),4.05~3.91(m ,3H),3.37~3.23(m,1H),3.23~3.11(m,2H),2.48(q,J=19.0Hz,2H),2.34(t,J=7.5Hz,2H) ,1.99(dt,J=9.5,6.4Hz,2H),1.89(d,J=16.6Hz,1H),1.78~1.71(m,3H),1.61(q,J=7.2Hz ,2H),1.24(dd,J=22.4,10.6Hz,19H),1.01(dd,J=6.2,4.1Hz,4H),0.87(t,J=6.8Hz,3H).

[0039] Example 2: Synthesis of Compound 6a

[0040] Compound phorbol (72.9 mg, 0.2 mmol) was dissolved in 5 mL of pyridine, tert-butyldimethylsilyl chloride (150.7 mg, 1 mmol) was slowly added at 0°C, and the reaction was allowed to proceed for 1 h at room temperature. TLC showed that the reaction was complete. Saturated NaHCO3 solution was added to quench the reaction, and the mixture was extracted with ethyl acetate (50 mL×3). The organic layers were combined, and the organic layers were washed with saturated CuSO4 solution. After separation, the organic layers were dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and separated and purified by column chromatography (CH2Cl2: MeOH=7:1, v / v) to obtain compound 2 (78.2 mg, white solid) with a yield of 82%.

[0041] Compound 2 (78.2 mg, 0.16 mmol) and n-decanoic acid (32.7 mg, 0.19 mmol) were dissolved in 10 mL of anhydrous dichloromethane, nitrogen was protected, EDCI (46.0 mg, 0.24 mmol) was slowly added at 0°C, stirred for 15 min, DMAP (19.5 mg, 0.16 mmol) was slowly added, stirred for 10 min and then moved to room temperature, reacted at room temperature overnight, TLC showed that the reaction was complete. Saturated NaHCO3 solution was added to quench the reaction, extracted with dichloromethane (50 mL×3), dried with anhydrous sodium sulfate after separation, filtered, concentrated under reduced pressure, and separated and purified by column chromatography (CH2Cl2: MeOH=15:1, v / v) to obtain compound 3 (100.2 mg, white solid), with a yield of 97%.

[0042] Compound 3 (63.2 mg, 0.1 mmol) and acetic acid (18.0 mg, 0.3 mmol) were dissolved in 10 mL of anhydrous dichloromethane, nitrogen was protected, EDCI (67.1 mg, 0.35 mmol) was slowly added at 0°C, stirred for 15 min, DMAP (36.7 mg, 0.3 mmol) was slowly added, stirred for 10 min and then moved to room temperature, reacted at room temperature overnight, TLC showed that the reaction was complete. Saturated NaHCO3 solution was added to quench the reaction, extracted with dichloromethane (50 mL × 3), dried with anhydrous sodium sulfate after separation, filtered, and concentrated under reduced pressure. 10 mL of methanol was added, 70% HClO4 (214 μL, 2.5 mmol) was added dropwise during stirring, and reacted at room temperature for 1 h, TLC showed that the reaction was complete. Saturated NaHCO3 solution was added to quench the reaction, and the mixture was extracted with ethyl acetate (50 mL×3). After separation, the mixture was dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and separated and purified by column chromatography (CH2Cl2:MeOH=15:1, v / v) to obtain compound 6a (46.5 mg, white solid) with a yield of 83%.

[0043] HRMS(ESI):C 32 H 48 O8Na[M+Na] + Theoretical value: 583.3247, actual value: 583.3248.

[0044] 1 H NMR (400MHz, Chloroform-d) δ7.56 (dd, J=2.5, 1.4Hz, 1H), 5.74~5.63 (m, 2H), 5.36 (d, J=10. 3Hz, 1H), 3.99 (q, J = 12.9Hz, 2H), 3.28~3.21 (m, 2H), 2.99 (s, 1H), 2.62~2.43 (m, 2H), 2.39~2. 23(m,3H),2.19~2.09(m,1H),2.06(s,3H),1.74(dd,J=2.9,1.3Hz,3H),1.59(q,J=7.3Hz,2H) ,1.31~1.24(m,12H),1.23(s,3H),1.20(s,3H),1.05(d,J=5.2Hz,1H),0.87(t,J=6.8Hz,6H); 13C NMR(101MHz,Chloroform-d)δ209.19,176.45,170.88,160.84,140.60,132.84,129.20,78.27,73.68,68.02,65.36,56.07,42 .98,38.49,36.39,34.36,31.84,29.38,29.25,29.21,29.08,25.86,24.52,23.86,22.65,21.01,16.77,14.43,14.10,10.08.

[0045] Example 3: Synthesis of Compound 6b

[0046] The difference from Example 2 is that the acetic acid in Example 2 is replaced by (S)-2-methylbutyric acid (30.6 mg, 0.3 mmol), and the rest of the contents are consistent with Example 2, to obtain compound 6b (43.6 mg, light yellow solid) with a yield of 72.4%.

[0047] HRMS(ESI):C 35 H 54 O8Na[M+Na] + Theoretical value: 625.3717, actual value: 625.3712.

[0048] 1 H NMR(400MHz,Chloroform-d)δ7.58(t,J=1.9Hz,1H),5.74~5.65(m,2H),5.40(d,J=10.3Hz,1H),4.00(q ,J=12.9Hz,2H),3.30~3.21(m,2H),2.96(d,J=17.3Hz,1H),2.61~2.44(m,2H),2.42~2.24(m,4H),2.14( dq,J=10.1,6.4Hz,1H),1.77~1.70(m,3H),1.70~1.54(m,3H),1.47(ddd,J=13.5,7.5,6.0Hz,1H),1.32 ~1.23(m,15H),1.19(s,3H),1.14(d,J=7.0Hz,3H),1.05(d,J=5.1Hz,1H),0.89(dt,J=16.2,7.3Hz,9H); 13C NMR(101MHz,Chloroform-d)δ209.22,176.45,176.18,160.90,140.57,132.86,129.29,78.39,76.33,73.69,68.02,65.41,56.09,42.86,41.83 ,38.99,38.49,36.33,34.39,31.84,29.37,29.26,29.23,29.07,26.72 ,25.65,24.50,23.85,22.65,17.01,16.87,14.40,14.09,11.61,10.08.

[0049] Example 4: Synthesis of Compound 6c

[0050] The difference from Example 2 is that the acetic acid in Example 2 is replaced by benzoic acid (36.6 mg, 0.3 mmol), and the rest of the contents are consistent with Example 2, to obtain compound 6c (30.0 mg, light yellow solid) with a yield of 48.2%.

[0051] HRMS(ESI):C 37 H 50 O8Na[M+Na] + Theoretical value: 645.3404, actual value: 645.3401.

[0052] 1 H NMR (400MHz, Chloroform-d) δ8.05~8.00(m,2H),7.63~7.53(m,2H),7.45(dd,J=8.4,7.0Hz,2H),5.80(s,1H),5.7 3(dd,J=5.8,2.1Hz,1H),5.66(d,J=10.2Hz,1H),4.04(q,J=12.9Hz,2H),3.34(t,J=5.6Hz,1H),3.32~3.26(m,1H) ,2.75(s,1H),2.65~2.47(m,2H),2.42~2.29(m,3H),1.76(dd,J=3.0,1.3Hz,3H),1.63(q,J=7.4Hz,2H),1.38(s,3 H),1.27(q,J=5.6,3.4Hz,13H),1.21(s,3H),1.13(d,J=5.2Hz,1H),0.94(d,J=6.4Hz,3H),0.87(d,J=7.0Hz,3H); 13C NMR(101MHz,Chloroform-d)δ209.11,176.56,166.25,160.87,140.54,133.05,132.91,130.16,129.74,129.26,128.44,78.36,77.69,73.74,68. 08,65.44,56.18,43.32,39.13,38.62,36.69,34.42,31.86,29.39,29.2 8,29.25,29.09,26.02,24.58,23.85,22.67,17.07,14.53,14.12,10.11.

[0053] Example 5: Synthesis of Compound 6d

[0054] The difference from Example 2 is that the acetic acid in Example 2 is replaced by 4-chlorobenzoic acid (46.9 mg, 0.3 mmol), and the rest of the contents are consistent with Example 2, to obtain compound 6d (28.3 mg, light yellow solid) with a yield of 43.1%.

[0055] HRMS(ESI):C 37 H 49 ClO8Na[M+Na] + Theoretical value: 679.3014, actual value: 679.3019.

[0056] 1 H NMR (400MHz, Chloroform-d) δ7.97~7.92(m,2H),7.60(dd,J=2.5,1.5Hz,1H),7.45~7.38(m,2H),5.77(s,1H),5.7 2(dd,J=5.8,2.0Hz,1H),5.64(d,J=10.3Hz,1H),4.03(q,J=12.9Hz,2H),3.34(t,J=5.5Hz,1H),3.28(t,J=2.9Hz, 1H),2.83(d,J=4.1Hz,1H),2.65~2.47(m,2H),2.43~2.28(m,3H),1.76(dd,J=2.9,1.3Hz,3H),1.63(p,J=7.4Hz,2 H),1.36(s,3H),1.32~1.23(m,13H),1.20(s,3H),1.13(d,J=5.2Hz,1H),0.93(d,J=6.5Hz,3H),0.89~0.84(m,3H); 13C NMR(101MHz,Chloroform-d)δ209.11,176.54,165.45,160.80,140.61,139.57,132.95,131.10,129.16,128.80,128.56,78.35,78.15,73.72,68. 06,65.36,56.15,43.28,39.10,38.60,36.72,34.40,31.86,29.39,29.2 8,29.24,29.09,26.07,24.57,23.84,22.67,17.08,14.54,14.12,10.11.

[0057] Example 6: Synthesis of Compound 6e

[0058] The difference from Example 2 is that the acetic acid in Example 2 is replaced by 4-fluorobenzoic acid (42.0 mg, 0.3 mmol), and the rest of the contents are consistent with Example 2, to obtain compound 6e (27.5 mg, white solid) with a yield of 42.9%.

[0059] HRMS(ESI):C 37 H 49 FO8Na[M+Na] + Theoretical value: 663.3309, actual value: 663.3297.

[0060] 1 H NMR (400MHz, Chloroform-d) δ8.06~8.00(m,2H),7.60(dd,J=2.4,1.4Hz,1H),7.15~7.07(m,2H),5.79(s,1H),5.7 2(dd,J=5.8,2.0Hz,1H),5.63(d,J=10.3Hz,1H),4.03(q,J=12.9Hz,2H),3.34(t,J=5.6Hz,1H),3.28(p,J=2.9Hz, 1H),2.99~2.82(m,1H),2.67~2.47(m,2H),2.44~2.26(m,3H),1.84(s,1H),1.78~1.73(m,3H),1.63(p,J=7.3Hz,2 H),1.36(s,3H),1.33~1.24(m,12H),1.21(s,3H),1.12(d,J=5.2Hz,1H),0.93(d,J=6.5Hz,3H),0.90~0.84(m,3H); 13C NMR(101MHz,Chloroform-d)δ209.13,176.56,167.10,165.33,164.58,160.80,1 40.63,132.94,132.30,132.21,129.20,126.37,126.34,115.71,115.49,78.37,7 7.96,73.72,68.05,65.39,56.14,43.30,39.09,38.58,36.70,34.41,31.86,29.38,29.27,29.24,29.09,26.05,24.58,23.84,22.67,17.09,14.53,14.11,10.11.

[0061] Example 7: Synthesis of Compound 6f

[0062] The difference from Example 2 is that the acetic acid in Example 2 is replaced by 4-methoxybenzoic acid (45.6 mg, 0.3 mmol), and the rest of the contents are consistent with Example 2, to obtain compound 6f (23.6 mg, white solid) with a yield of 36.2%.

[0063] HRMS(ESI):C 38 H 52 O9Na[M+Na] + Theoretical value: 675.3509, actual value: 675.3502.

[0064] 1 H NMR (400MHz, Chloroform-d) δ8.00~7.95(m,2H),7.61(dd,J=2.5,1.5Hz,1H),6.95~6.90(m,2H),5.81(s,1H),5. 72(dd,J=5.9,2.0Hz,1H),5.62(d,J=10.2Hz,1H),4.03(q,J=12.9Hz,2H),3.86(s,3H),3.36~3.24(m,2H),2.77(d ,J=10.3Hz,1H),2.55(q,J=19.1Hz,2H),2.43~2.25(m,3H),1.76(dd,J=2.9,1.3Hz,3H),1.63(p,J=7.3Hz,2H),1. 37(s,3H),1.27(d,J=13.9Hz,13H),1.20(s,3H),1.11(d,J=5.2Hz,1H),0.93(d,J=6.5Hz,3H),0.89~0.84(m,3H); 13C NMR(101MHz,Chloroform-d)δ209.14,176.58,165.99,163.46,160.91,140.52,132.88,131.76,129.33,122.55,113.68,78.38,73.74,68.08,65. 49,56.17,55.48,43.36,39.11,38.60,36.64,34.42,31.86,29.39,29.2 8,29.25,29.09,25.95,24.57,23.85,22.67,17.08,14.53,14.12,10.11.

[0065] Example 8: Synthesis of Compound 6g

[0066] The difference from Example 2 is that the acetic acid in Example 2 is replaced by 3-pyridinecarboxylic acid (36.9 mg, 0.3 mmol), and the rest of the contents are consistent with Example 2, to obtain compound 6g (40.1 mg, white solid) with a yield of 64.3%.

[0067] HRMS(ESI):C 36 H 49 NO8Na[M+Na] + Theoretical value: 646.3356, actual value: 646.3353.

[0068] 1 H NMR(400MHz,Chloroform-d)δ9.10(d,J=2.1Hz,1H),8.98(dd,J=4.9,1.8Hz,1H),8.34(dt,J=8.0,1.9Hz,1H),7.58~ 7.54(m,1H),7.47(dd,J=8.0,4.9Hz,1H),5.74~5.65(m,3H),5.16(s,1H),4.03(q,J=13.1Hz,2H),3.43(t,J=5.6Hz, 1H),3.25(q,J=2.9Hz,1H),2.61(q,J=19.0Hz,2H),2.48~2.26(m,4H),1.75(dd,J=2.9,1.4Hz,3H),1.64(p,J=7.5Hz ,2H),1.40(s,3H),1.34~1.24(m,12H),1.21(s,3H),1.15(d,J=5.1Hz,1H),0.95(d,J=6.4Hz,3H),0.89~0.83(m,3H); 13C NMR(101MHz,Chloroform-d)δ209.14,176.49,164.83,159.96,153.35,150 .10,141.05,137.92,133.04,128.89,126.14,123.79,78.75,78.31,73.56 ,67.91,65.31,56.09,43.26,39.03,38.31,36.76,34.39,31.84,29.38,29 .26,29.23,29.08,26.10,24.58,23.85,22.66,17.08,14.60,14.11,10.16.

[0069] Example 9: Synthesis of Compound 6h

[0070] The difference from Example 2 is that the acetic acid in Example 2 is replaced by 2-pyrazine carboxylate (37.2 mg, 0.3 mmol), and the rest of the contents are consistent with Example 2, to obtain compound 6h (42.2 mg, white solid) with a yield of 67.7%.

[0071] HRMS(ESI):C 35 H 48 N2O8Na[M+Na] + Theoretical value: 647.3309, actual value: 647.3300.

[0072] 1 H NMR(400MHz,Chloroform-d)δ9.22(d,J=1.5Hz,1H),8.84~8.75(m,2H),7.61~7.56(m,1H),5. 80~5.68(m,3H),4.03(q,J=13.0Hz,2H),3.40~3.25(m,3H),2.56(q,J=19.0Hz,2H),2.46~2.2 8(m,3H),1.94~1.82(m,1H),1.75(dd,J=3.0,1.3Hz,3H),1.68~1.58(m,2H),1.38(s,3H),1.3 3~1.23(m,12H),1.21(s,3H),1.16(d,J=5.2Hz,1H),0.97(d,J=6.5Hz,3H),0.89~0.83(m,3H); 13C NMR(101MHz,Chloroform-d)δ209.07,176.63,163.58,160.46,147.55,145.93,144.99,143.51,140.83,133.06,128.95,79.77,78.38,73.64,67. 97,65.19,56.14,43.09,39.07,38.52,36.84,34.34,31.84,29.38,29.2 5,29.22,29.09,26.23,24.50,23.85,22.66,16.96,14.60,14.10,10.13.

[0073] Example 10: Study on the anti-tumor activity of 13-O-decanoyl-phorbol ester derivatives

[0074] Using phorbol 12-myristate-13-acetate (TPA) as a positive control, the in vitro antiproliferative activities of the nine synthesized compounds on A2780, Hela, and HepG2 cells were determined. The specific process is as follows:

[0075] Collect logarithmic phase cells, prepare single cell suspension with culture medium containing 10% fetal bovine serum (A2780: 1640 culture medium, Hela, HepG2: DMEM high glucose culture medium), inoculate 5000-13000 cells per well into 96-well plates, and incubate in a 37°C, 5% carbon dioxide incubator for 24 hours. After the cells adhere to the wall, discard the old culture medium, add different concentrations of test compounds, set seven concentration gradients for each compound (1, 5, 10, 20, 30, 40, 50 μM), set three sets of duplicate wells for each concentration, and culture for 48 hours. Remove the supernatant, add 20 μL of MTT solution (5 mg / mL) and 100 μL of serum-free culture medium to each well, and incubate in the dark for 4-6 hours. Then add 100 μL of triple dissolution solution to each well, put it in a carbon dioxide incubator and continue incubation for 6-12 hours to fully dissolve the formazan crystals. Finally, the absorbance of each well at 570 nm was measured using an enzyme reader to calculate the cell proliferation inhibition rate. The cell proliferation inhibition rate calculation formula is as follows:

[0076] Cell proliferation inhibition rate (%) = 1-(OD value of drug-treated group-OD value of blank group) / (OD value of control group-OD value of blank group) × 100%

[0077] The half inhibitory concentration (IC 50 ), each group of experiments was repeated three times independently and the average value was taken to obtain the final data. 50 The test results are shown in the following table:

[0078]

[0079] The results showed that the compounds of the present invention had different degrees of inhibitory activity against three tumor cell lines: HepG2, Hela and A2780. Overall, the inhibitory activity of the compounds against A2780 ovarian cancer cells was stronger than that against HepG2 liver cancer cells and Hela cervical cancer cells. Among them, compound 6b had the strongest activity, with an IC of 1.34 for A2780, HepG2 and Hela cells. 50 The values ​​were 13.58, 19.79 and 16.94 μM, respectively, which were stronger than the positive control group TPA.

[0080] Although the present invention has been disclosed as above in the form of a preferred embodiment, it is not intended to limit the present invention. Anyone familiar with this technology can make various changes and modifications without departing from the spirit and scope of the present invention. Therefore, the scope of protection of the present invention should be based on the definition of the claims.

Claims

1. A 13-O-decanoyl-phorbol ester derivative, characterized in that: The structure of the derivative is shown in general formula (I): Where R is H, 2. The method for preparing the 13-O-decanoyl-phorbol ester derivatives according to claim 1, characterized in that: The synthetic route is as follows: The synthesis process is as follows: (1) Compound 1 (phorbol) reacts with tert-butyldimethylsilyl chloride to selectively protect 20-OH to obtain compound 2; (2) Compound 2 is condensed with n-decanoic acid to form an ester to obtain compound 3; (3) Compound 3 is subjected to the treatment of perchloric acid to remove the tert-butyldimethylsilyl (TBS) protecting group to obtain compound 4; (4) Compound 3 was condensed with different carboxylic acids to form esters to prepare compounds 5a-h; (5) Compounds 5a-h were treated with perchloric acid to remove the TBS protecting group to obtain compounds 6a-h.

3. The method for preparing 13-O-decanoyl-phorbol ester derivatives according to claim 2, characterized in that: The specific preparation process of compound 4 is as follows: S1. Dissolve phorbol in pyridine, add tert-butyldimethylsilyl chloride at -5°C to 5°C, stir at room temperature until the reaction is complete, add saturated NaHCO3 solution to quench the reaction, extract with ethyl acetate, combine the organic layers, rinse the organic layers with saturated CuSO4 solution, separate and dry with anhydrous sodium sulfate, filter, concentrate under reduced pressure, and separate and purify by column chromatography to obtain compound 2; S2. Compound 2 and n-decanoic acid were dissolved in dichloromethane. Under nitrogen protection, 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDCI) was added at -5°C to 5°C. After stirring for 10 to 20 minutes, 4-dimethylaminopyridine (DMAP) was added. After stirring for 5 to 15 minutes, the mixture was cooled to room temperature and stirred until the reaction was complete. A saturated NaHCO3 solution was added to quench the reaction. The mixture was extracted with dichloromethane. After separation, the mixture was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product of compound 3. A MeOH solution of HClO4 was added to the mixture. The reaction was stirred at room temperature until the reaction was complete. A saturated NaHCO3 solution was added to quench the reaction. The mixture was extracted with ethyl acetate. After separation, the mixture was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The mixture was separated and purified by column chromatography to obtain compound 4.

4. The method for preparing 13-O-decanoyl-phorbol ester derivatives according to claim 2, characterized in that: The specific preparation process of compounds 6a~h is as follows: S1. Dissolve phorbol in pyridine, add tert-butyldimethylsilyl chloride at -5°C to 5°C, stir at room temperature until the reaction is complete, add saturated NaHCO3 solution to quench the reaction, extract with ethyl acetate, combine the organic layers, rinse the organic layers with saturated CuSO4 solution, separate and dry with anhydrous sodium sulfate, filter, concentrate under reduced pressure, and separate and purify by column chromatography to obtain compound 2; S2, compound 2 and n-decanoic acid were dissolved in dichloromethane, EDCI was added at -5°C to 5°C under nitrogen protection, DMAP was added after stirring for 10 to 20 minutes, the mixture was stirred for 5 to 15 minutes and then moved to room temperature, stirred for complete reaction, saturated NaHCO3 solution was added to quench the reaction, the mixture was extracted with dichloromethane, the liquid was separated and dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and separated and purified by column chromatography to obtain compound 3; S3. Dissolve compound 3 and acetic acid, (S)-2-methylbutyric acid, benzoic acid, 4-chlorobenzoic acid, 4-fluorobenzoic acid, 4-methoxybenzoic acid, 3-picolinic acid or 2-pyrazinecarboxylate in dichloromethane, add EDCI at -5°C to 5°C, stir for 10 to 20 min, then add DMAP, stir for 5 to 15 min, move to room temperature, stir until the reaction is complete, add saturated NaHCO3 solution to quench the reaction, extract with dichloromethane, separate the liquids and dry with anhydrous sodium sulfate, filter, concentrate under reduced pressure, add HClO4 in MeOH solution, stir at room temperature until the reaction is complete, add saturated NaHCO3 solution to quench the reaction, extract with ethyl acetate, separate the liquids and dry with anhydrous sodium sulfate, filter, concentrate under reduced pressure, separate and purify by column chromatography to obtain compounds 6a to 6h.

5. The method for preparing 13-O-decanoyl-phorbol ester derivatives according to claim 3, characterized in that: The molar ratio of phorbol alcohol to tert-butyldimethylsilyl chloride is 1:5, the molar ratio of compound 2, n-decanoic acid, EDCI and DMAP is 1:(1.0-1.5):(1.0-1.5):1; the molar ratio of compound 2 to perchloric acid is 1:

25.

6. The method for preparing 13-O-decanoyl-phorbol ester derivatives according to claim 4, characterized in that: The molar ratio of phorbol and tert-butyldimethylsilyl chloride is 1:5, the molar ratio of compound 2, n-decanoic acid, EDCI and DMAP is 1:(1.0-1.5):(1.0-1.5):1; the molar ratio of compound 3 and acetic acid, (S)-2-methylbutyric acid, benzoic acid, 4-chlorobenzoic acid, 4-fluorobenzoic acid, 4-methoxybenzoic acid, 3-pyridinecarboxylic acid or 2-pyrazinecarboxylate and EDCI, DMAP is 1:3:3.5:3; the molar ratio of compound 3 and perchloric acid is 1:

25.

7. The use of the 13-O-decanoyl-phorbol ester derivatives according to claim 1 in the preparation of anti-tumor drugs, characterized in that: The tumor refers to human ovarian cancer, cervical cancer or human liver cancer.

8. The use according to claim 7, characterized in that: The 13-O-decanoyl-phorbol ester derivatives are used in the preparation of drugs for inhibiting tumor cell proliferation, inducing pyroptosis or promoting apoptosis, wherein the tumor cells are A2780, Hela or HepG2.

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