延胡索乙素对映体对人肝微粒体细胞色素P450酶抑制作用机制研究

Inhibitory mechanism of tetrahydropalmatine enantiomers on cytochrome P450 in human liver microsomes

目的考察延胡索乙素(tetrahydropalmatine,THP)对映体对人肝微粒体中主要细胞色素P450酶(cytochrome P450,CYP450)的抑制作用及其机制。方法采用Cocktail探针药物法分别考察左旋延胡索乙素[(-)-tetrahydropalmatine,(-)-THP]和右旋延胡索乙素[(+)-tetrahydropalmatine,(+)-THP]对人肝微粒体中主要I相药物代谢酶CYP1A2、CYP2C9、CYP2C19、CYP3A4、CYP2E1和CYP2D6活性的影响;采用两步孵育法考察(-)-THP对人肝微粒体中CYP2D6的底物右美沙芬脱甲基活性的影响,研究其对CYP2D6的抑制机制;采用时间依赖性实验考察(-)-THP对CYP2D6的酶动力学参数。结果 (+)-THP对CYP450各亚型无明显抑制作用,而(-)-THP对CYP2D6的抑制作用强(IC50=0.46μmol/L);在加或不加NADPH[(+)NADPH/(-)NADPH]预孵育体系中,(-)-THP对CYP2D6的IC50值分别为2.40、0.46μmol/L,即IC50(-)NADPH/IC50(+)NADPH=5.22;(-)-THP对CYP2D6的酶动力学参数Ki和Kinact分别为0.690μmol/L和0.084 6 min-1。结论 (-)-THP对CYP2D6的抑制作用强,抑制类型为基于机制抑制(MBI),提示在临床应用中需注意(-)-THP可能引起显著的代谢性药物相互作用。

Objective To investigate the inhibition of tetrahydropalmatine（THP） enantiomers on main cytochrome P450（CYP450） in human liver microsomes and the mechanism. Methods The effects of（-）-THP and（＋）-THP on the activies of main phase I metabolic enzymes in human liver microsomes, CYP1A2, CYP2C9, CYP2C19, CYP3A1, CYP2E1, and CYP2D6 were investigated using Cocktail probe drugs method; The effect of preincubation with（-）-THP on the CYP2D6 substrate dextromethorphan demethylation activity in human liver microsomes were investigated using a two-step incubation scheme and study its inhibitory mechanism was studied; The enzyme inhibitory kinetic parameters of CYP2D6 by（-）-THP were investigated using time-dependent incubation with human liver microsomes. Results The inhibitory effect of（＋）-THP on CYP450 subtype was not significant, while.CYP2D6 activity was significantly inhibited by（-）-THP（IC50 = 0.46 μmol/L）. The value of IC50 preincubation with or without NADPH were 2.40 and 0.46 μmol/L, respectively, IC50（-） NADPH / IC50（＋） NADPH = 5.22. And the enzyme inhibitory kinetic parameters of Ki and Kinact were 0.690 μmol/L and 0.084 6 min-1, respectively. Conclusion（-）-THP has a strong inhibition on CYP2D6, and the type of inhibition is mechanism-based inhibiton（MBI）, which should indicate a potential metabolic drug-drug interaction mediated by（-）-THP in clinical application.