Huskey S W, Dean D C, Miller R R, Rasmusson G H, Chiu S H
Department of Drug Metabolism, Merck Research Laboratories, Rahway 07065, USA.
Drug Metab Dispos. 1995 Oct;23(10):1126-35.
Finasteride, a prescription drug for the treatment of benign prostatic hypertrophy and alleviation of symptoms associated with benign prostatic hypertrophy and alleviation of symptoms associated with benign prostatic hypertrophy, has been shown to be metabolized in rat hepatic microsomes by hydroxylation at the t-butyl group (omega-OH finasteride), followed by further oxidation to the corresponding acid (omega-oic acid finasteride), with omega-aldehyde finasteride as an intermediate. In this study, we identified specific human cytochrome P450 (CYP) isozyme(s) involved in the in vitro metabolism of [14C]finasteride using CYP isozyme-selective inhibitors and microsomes containing specific recombinant human CYP isozymes (expressed in human AHH-1 TK+/-cells). Each of the three steps of the oxidative pathway was examined separately by using [14C]finasteride and its consecutive metabolites (omega-OH finasteride and omega-aldehyde finasteride) as substrates, and human liver microsomes or expressed recombinant CYP isozymes as the enzyme source. Gestodene, a mechanism-based inhibitor of CYP3A isozymes, showed a concentration-dependent inhibition of the oxidative metabolism of [14C]finasteride. In addition, the respective omega-OH finasteride and omega-oic acid finasteride metabolites were generated only by microsomes containing recombinant CYP3A4, but not the other isozymes (CYP1A1, CYP2B6, CYP2C8, CYP2C9, CYP2D6, and CYP2E1). Similar results were obtained for the oxidation of omega-OH finasteride to omega-aldehyde finasteride, suggesting that human CYP3A isozymes were involved in the oxidation of omega-OH finasteride. When omega-aldehyde finasteride was incubated with human liver microsomes in the presence of an NADPH regenerating system, both the omega-oic acid finasteride and the omega-OH finasteride were detected, suggesting that oxidative and reductive reactions were occurring simultaneously and that they were NADPH- or NADP-dependent. Inhibitors of CYP3A isozymes inhibited the oxidation of omega-aldehyde finasteride in a concentration-dependent manner; an increase in the reduction was also observed, presumably caused by inhibition of the competitive oxidative reaction. Other selective CYP inhibitors for CYP1A1/2 (alpha-naphthoflavone), CYP2C8-10 (sulfaphenazole), CYP2D6 (quinidine), and CYP2E1 (diallylsulfone) showed minor or no effects on both reactions. Consistent with these results, only microsomes containing human recombinant CYP3A4 catalyzed the oxidation of omega-aldehyde finasteride to omega-oic acid finasteride. These results indicate that the oxidation of omega-aldehyde finasteride was NADPH-dependent and was mediated at least in part by CYP3A4. In addition, NAD-dependent enzymes in cytosolic, microsomal, and mitochondrial fractions were capable of oxidizing omega-aldehyde finasteride to omega-oic acid finasteride. Other cellular fractions, particularly mitochondria, were shown to convert finasteride to omega-oic acid finasteride in a similar fashion.
非那雄胺是一种用于治疗良性前列腺增生并缓解其相关症状的处方药，已表明它在大鼠肝微粒体中通过叔丁基的羟基化作用（ω-羟基非那雄胺）进行代谢，随后进一步氧化为相应的酸（ω-羧酸非那雄胺），以ω-醛基非那雄胺作为中间体。在本研究中，我们使用CYP同工酶选择性抑制剂和含有特定重组人CYP同工酶（在人AHH-1 TK+/-细胞中表达）的微粒体，确定了参与[14C]非那雄胺体外代谢的特定人细胞色素P450（CYP）同工酶。氧化途径的三个步骤分别使用[14C]非那雄胺及其连续代谢物（ω-羟基非那雄胺和ω-醛基非那雄胺）作为底物，用人肝微粒体或表达的重组CYP同工酶作为酶源进行研究。孕二烯酮是一种基于机制的CYP3A同工酶抑制剂，显示出对[14C]非那雄胺氧化代谢的浓度依赖性抑制作用。此外，各自的ω-羟基非那雄胺和ω-羧酸非那雄胺代谢物仅由含有重组CYP3A4的微粒体产生，而不由其他同工酶（CYP1A1、CYP2B6、CYP2C8、CYP2C9、CYP2D6和CYP2E1）产生。对于ω-羟基非那雄胺氧化为ω-醛基非那雄胺也获得了类似结果，表明人CYP3A同工酶参与了ω-羟基非那雄胺的氧化。当在NADPH再生系统存在下将ω-醛基非那雄胺与人肝微粒体一起孵育时，检测到了ω-羧酸非那雄胺和ω-羟基非那雄胺，表明氧化和还原反应同时发生，且它们是NADPH或NADP依赖性的。CYP3A同工酶抑制剂以浓度依赖性方式抑制ω-醛基非那雄胺的氧化；还观察到还原作用增加，推测是由竞争性氧化反应的抑制引起的。其他针对CYP1A1/2（α-萘黄酮）、CYP2C8 - 10（磺胺苯吡唑）、CYP2D6（奎尼丁）和CYP2E1（二烯丙基砜）的选择性CYP抑制剂对这两种反应显示出轻微或无影响。与这些结果一致，只有含有重组人CYP3A4的微粒体催化了ω-醛基非那雄胺氧化为ω-羧酸非那雄胺。这些结果表明ω-醛基非那雄胺的氧化是NADPH依赖性的，并且至少部分由CYP3A4介导。此外，胞质、微粒体和线粒体部分中的NAD依赖性酶能够将ω-醛基非那雄胺氧化为ω-羧酸非那雄胺。其他细胞部分，特别是线粒体，也显示出以类似方式将非那雄胺转化为ω-羧酸非那雄胺。
