CYP3A4和CYP2B6药物诱导剂体外筛选体系的建立

Establishment of in vitro screening system for CYP3A4 and CYP2B6 inducers

目的构建CYP酶药物诱导剂的体外筛选技术平台。方法利用双荧光素酶报告基因系统,将包含hPXR蛋白识别和结合调控元件的CYP3A4和2B6启动子序列插入报告基因上游,将表达载体和报告载体共转染HepG2细胞,用含有利福平或DMSO培养基培养48h后裂解进行双荧光素酶活性检测。结果经利福平处理的细胞裂解物荧光素酶比活性值与阴性对照组和溶剂组差异显著。结论本研究构建的CYP3A4和2B6体外筛选平台,可以有效地用于体外诱导剂的筛选。

[Objective] To establish an in vitro system to screen drug inducers of CYP450 enzymes. [Method] Clanging the promoters of CYP3A4 and CYP2B6 which contained the elements that hPXR, a kind of nuclear receptor, can recognize and bind, and inserting the trans-element to the upstream of firefly lueiferase reporter gene of the pGIA.17 vectors, Cotransfeeting the report vector and hPXR expression vector to HepG2 cell line, and analyzing luciferase activity after the cell treated with fifampin or DMSO. [Result] The fold activation value of luciferase of fifampin group was statistically different from control group and DMSO group. [Conclusion] We successfully estab-lished an in vitro system which can be used for screening drug agonist for hPXR and increasing the expression of CYP450 enzymes.