CYP2D6基因多态性对阿托伐他汀代谢的影响

The influence of CYP2D6 gene polymorphism on atorvastatin metabolism

目的研究中国人群CYP2D6基因多态性对阿托伐他汀(A)在体外人肝微粒体代谢的影响,探讨CYP2D6基因在阿托伐他汀代谢过程的作用。方法收集肝胆手术患者手术旁正常肝组织样本,并提取人肝微粒体(HLM)及肝组织DNA。使用Taqman基因分型技术测定CYP2D6*3、*4、*5、*6、*9、*10和*41这7个位点多态性。采用HLM孵育A使其代谢,并用LC-MS/MS方法测定A及其代谢产物2-羟基阿托伐他汀(2A)和4-羟基阿托伐他汀(4A)的浓度,分析CYP2D6基因多态性对A体外代谢的影响。按照相关文献及分型结果将样本的CYP2D6代谢表型分为强代谢型(EMs)和中等代谢型(IMs)表现型。结果 A在人肝微粒体中生成2A和4A的酶反应速率常数分别为53.7及48.5μmol/L;最大反应速率分别为140.2及181.7 pmol/(mg·min)。A在55例人肝微粒体中的代谢呈现多态性,最小和最大速率分别为109.9和9.3 pmol/(mg·min)。本研究中CYP2D6*3、*4、*5、*6、*9、*10、*41等位基因突变频率分别为0%、1.82%、2.73%、0%、0%、69.09%、2.73%。CYP2D6*4、*5、*10、*41等位基因突变对阿托伐他汀在人肝微粒体反应体系中的代谢无明显影响。A的减少速率在EMs组比在IMs组高,但差异无统计学意义(P>0.05)。2A的生成速率在EMs和IMs组分别为(14.68±7.62)vs.(8.70±3.50)pmol/(mg·min),P=0.06。4A的生成速率在EMs和IMs组分别为(20.32±13.10)vs.(11.04±4.88)pmol/(mg·min),P=0.07。结论 CYP2D6*4、*5、*10、*41等位基因突变对阿托伐他汀代谢影响较小,而CYP2D6*4、*5、*10、*41多等位基因突变引起的代谢表型的差异对阿托伐他汀体外代谢生成2-羟基阿托伐他汀、4-羟基阿托伐他汀有一定的影响。

Objective To detect the influence of CYP2D6 gene polymorphism on atorvastatin metabolism in Chinese.Methods Human liver tissues from distant noncancerous liver tissues were obtained surgically or from liver resection for benign liver diseases at Sun Yat-Sen Memorial Hospital（Guangzhou, Guangdong, China） with the consent of patients between September 2012 and May 2015. Microsome and DNA were extracted from liver tissues. Atorvastatin（A） were metabolized on human liver microsome and its metabolites were measured by LC-MS / MS. Taq Man genotyping method were used to test CYP2D6＊3,＊4,＊5,＊6,＊9,＊10,＊41 alleles. Results Michaelis-Menten constant（Km） values of 2A and 4A were 53.7 and 48.5 μmol / L, respectively. The maximum rate of formation（Vmax） values of orthohydroxyatorvastatin and para-hydroxyatorvastatin formation from atorvastatin were 140.2 and 181.7 pmol /（mg·min）,respectively. The reduction rates of A and 4A varied were from 109.9 to 9.3 pmol /（mg·min）. Large variability in the hydroxylation of AT was observed in the 55 liver samples studied. The allele mutation frequency for CYP2D6 ＊ 3, ＊ 4,＊5, ＊6, ＊9, ＊10, ＊41 were 0%, 1.82%, 2.73%, 0%, 0%, 69.09% and 2.73%, respectively. There were no significant differences between the wild-type and the CYP2D6 ＊ 4, ＊ 5, ＊ 10 or ＊ 41 alleles variant in vitro. The reduction rate of atorvastatin in EMs was not significantly higher than that of IMs. Formation rate of 2A / 4A atorvastatin in IMs were both lower than EMs（2A, 8.70±3.50 pmol /（mg·min） for IMs versus 14.68±7.62 pmol /（mg·min） for EMs, P=0.06）;（4A, 11.04±4.88 pmol /（mg·min） for IMs versus 20.32±13.10 pmol /（mg·min）for EMs, P=0.07）.Conclusion There were no obvious effects of CYP2D6 ＊ 4, ＊ 5, ＊ 10 or ＊ 41 allele mutation on A metabolism.However, CYP2D6 gene polymorphism affects the CYP2D6 phenotype with reducing the metabolism of atorvastatin, that may be one of the mechanism for CYP2D6＊ 4 effecting atorvastatin-induced myopathy. However, more experiments must be performed to make sure the effect of CYP2D6 on metabolism of atorvastatin in vitro, and more researches must be needed to be verified in vivo.