Pearson Josh T, Wahlstrom Jan L, Dickmann Leslie J, Kumar Santosh, Halpert James R, Wienkers Larry C, Foti Robert S, Rock Dan A
Department of Pharmacokinetics and Drug Metabolism, Amgen, Inc. Seattle, WA 98119-3105, USA.
Chem Res Toxicol. 2007 Dec;20(12):1778-86. doi: 10.1021/tx700207u. Epub 2007 Nov 15.
The role of C239 as the active-site residue responsible for forming the covalent linkage with raloxifene during P450 3A4 time-dependent inactivation (TDI) was recently identified. The corresponding residue in CYP3A5 is S239, and when the potential for TDI in P450 3A5 was investigated, only reversible inhibition was observed against midazolam and testosterone, with median inhibitory concentration (IC50) values of 2.4 and 2.9 microM, respectively. In a similar fashion, when C239 was replaced with alanine in P450 3A4, TDI was successfully engineered out, and the reversible inhibition was characterized by IC50 values of 3.7 and 3.5 microM against midazolam and testosterone, respectively. Metabolism studies confirmed that the reactive diquinone methide intermediate required for P450 3A4 inactivation formed in all of the P450 3A enzymes investigated. Furthermore, the absence of TDI in P450 3A5 led to an increase in the formation of GSH-related adducts of raloxifene compared with that for P450 3A4. Consequently, the absence of the nucleophilic cysteine leads to differential TDI and generation of reactive metabolites in the P450 3A enzyme, providing the foundation for pharmacogenetics that contributes to individual differences in susceptibility to adverse drug reactions.
最近确定了C239在细胞色素P450 3A4（CYP3A4）时间依赖性失活（TDI）过程中作为负责与雷洛昔芬形成共价键的活性位点残基的作用。CYP3A5中的相应残基是S239，当研究CYP3A5中TDI的可能性时，仅观察到对咪达唑仑和睾酮的可逆抑制作用，其半数抑制浓度（IC50）值分别为2.4和2.9微摩尔。同样，当在CYP3A4中将C239替换为丙氨酸时，成功消除了TDI，并且对咪达唑仑和睾酮的可逆抑制作用的IC50值分别为3.7和3.5微摩尔。代谢研究证实，在所研究的所有CYP3A酶中均形成了CYP3A4失活所需的反应性二甲醌中间体。此外，与CYP3A4相比，CYP3A5中不存在TDI导致雷洛昔芬的谷胱甘肽相关加合物的形成增加。因此，亲核半胱氨酸的缺失导致CYP3A酶中TDI的差异和反应性代谢物的产生，为药物遗传学提供了基础，这有助于解释个体对药物不良反应易感性的差异。
