苄基膦酸二乙酯乙酰胺类熊果酸衍生物的合成及其体外抗肝癌活性研究

Synthesis and Anti-hepatocellular Carcinoma In Vitro of Ursolic Acid Derivatives Bearing Benzylphosphonic Acid Diethyl Ester Acetamide Moieties

关键词:  熊果酸  衍生物  苄基膦酸二乙酯  合成  抗肝癌

Key words:  ursolic acid  derivatives  benzylphosphonic acid diethyl ester  synthesis  anti-hepatoma

摘要:

目的 考察新型熊果酸衍生物的体外抗肝癌活性及其作用机制。方法 依据"Topliss决策法"将熊果酸与不同取代的苄基膦酸二乙酯片段通过取代反应得到目标化合物,并采用MTT法考察其体外抗肝癌活性;通过分子对接试验分析化合物可能的作用靶点并通过Western blotting试验加以验证。结果 目标化合物4a~4e的化学结构经过¹H-NMR﹑ ¹³C-NMR以及HRMS的联合确证;化合物4b及4e对BEL-7402及HepG2 2种肝癌细胞株的抑制活性优于熊果酸及阳性对照药5-氟尿嘧啶,同时对正常肝细胞L02的毒性显著降低;化合物4e呈浓度依赖性下调p-AKT蛋白的表达。结论 化合物4e的体外抗肝癌活性最为理想(针对HepG2的IC₅₀值为2.69 μmol·L^-1),可作为AKT蛋白抑制剂进行深入研究。

Abstract:

OBJECTIVE To investigate the anti-hepatocellular carcinoma efficacy in vitro and mechanism of novel ursolic acid derivatives. METHODS The target compounds were designed according to the “Topliss decision method”, and obtained by substitution reaction between ursolic acid and different substituted diethyl benzyl phosphonic acid, then the anti-hepatocellular activity in vitro of these compounds was studied by MTT. The possible target of the compound was predicted by molecular docking study and verified by Western blotting. RESULTS Target compounds 4a-4e were characterized via ¹H-NMR, ¹³C-NMR, and HRMS. Compound 4b and 4e showed higher antiproliferative activity against the BEL-7402 and HepG2 cell lines compared with the positive-control drug 5-fluorouracil and ursolic acid. All the target compounds exhibited low cytotoxic activities against human normal liver cells(L02). In a concentration-dependent manner, compound 4e decreased p-AKT protein level. CONCLUSION Compound 4e exhibited the most potent antiproliferative activity and commendable selectivity between cancer and normal cells(IC₅₀=2.69 μmol·L^–1 against the HepG2 cell line), and could act as a potential inhibitor of AKT protein.