Stresser D M, Kupfer D
Department of Pharmacology and Molecular Toxicology, University of Massachusetts Medical Center, Shrewsbury 01545, USA.
Biochem Pharmacol. 1998 Jun 1;55(11):1861-71. doi: 10.1016/s0006-2952(98)00060-4.
This study demonstrates interplay among human hepatic cytochrome P450 (CYP) isoforms in transforming aromatic compounds from being prosubstrates of CYP3A4 into phenolic substrates. Incubation of methoxychlor with CYP2C19 yields the phenolic monodemethylated derivative (mono-OH-M). Additionally, CYP2C19 catalyzes the ortho-hydroxylation of mono-OH-M and of residual methoxychlor. CYP3A4 does not catalyze the O-demethylation or hydroxylation of methoxychlor, but does hydroxylate mono-OH-M (ortho to the phenolic hydroxyl) (Stresser DM and Kupfer D, Biochemistry 36: 2203-2210, 1997). A combination of reconstituted CYP2C19 and 3A4 in the same vessel elicits stimulation of the ortho-hydroxylation of mono-OH-M compared with 2C19 alone. It is unlikely that stimulation of hydroxylation was due to protein-protein interactions, generating more active P450(s), because progression of the stimulation was time-dependent. When reconstituted CYP3A4 was added to an ongoing incubation containing reconstituted 2C19, stimulation of catechol formation occurred. In another experiment, stimulatory activity was similar when 2C19 and 3A4 were reconstituted together in the same vesicles or separately. Cumulative evidence demonstrates that the stimulation of catechol formation resulted from CYP3A4-mediated ortho-hydroxylation of the phenolic metabolite(s) generated by CYP2C19. Similarly, estradiol 3-methyl ether is demethylated by CYP2C19 into estradiol, a CYP3A4 substrate for ortho-hydroxylation; there was significant stimulation of hydroxylation by combined 2C19 and 3A4. These findings demonstrate that pro-phenolic compounds (methoxychlor and estradiol 3-methyl ether) are prosubstrates of CYP3A4. Because catalysis may become evident only after prosubstrate conversion (by a different P450) into a substrate, caution is warranted when concluding a lack of catalytic involvement by a particular P450 isoform, based solely on data from the use of individual cDNA-expressed P450s.
本研究证明了人类肝脏细胞色素P450（CYP）同工型之间的相互作用，这种相互作用将芳香族化合物从CYP3A4的前体底物转化为酚类底物。甲氧滴滴涕与CYP2C19一起孵育会产生酚类单去甲基化衍生物（单羟基甲氧滴滴涕）。此外，CYP2C19催化单羟基甲氧滴滴涕和残留甲氧滴滴涕的邻位羟基化反应。CYP3A4不催化甲氧滴滴涕的O-去甲基化或羟基化反应，但会催化单羟基甲氧滴滴涕（酚羟基邻位）的羟基化反应（斯特雷瑟·D·M和库普弗·D，《生物化学》36: 2203 - 2210，1997）。与单独的CYP2C19相比，在同一容器中重组的CYP2C19和3A4组合会引发单羟基甲氧滴滴涕邻位羟基化反应的刺激作用。羟基化反应的刺激不太可能是由于蛋白质 - 蛋白质相互作用产生了更具活性的P450，因为刺激作用的进展是时间依赖性的。当将重组的CYP3A4添加到含有重组CYP2C19的正在进行的孵育体系中时，会发生儿茶酚形成的刺激作用。在另一项实验中，当CYP2C19和3A4在同一囊泡中一起重组或分别重组时，刺激活性相似。累积证据表明，儿茶酚形成的刺激作用是由CYP3A4介导的CYP2C19产生的酚类代谢物的邻位羟基化反应所致。同样，雌二醇3 - 甲醚被CYP2C19去甲基化为雌二醇，雌二醇是CYP3A4进行邻位羟基化反应的底物；CYP2C19和3A4组合对羟基化反应有显著的刺激作用。这些发现表明，前酚类化合物（甲氧滴滴涕和雌二醇3 - 甲醚）是CYP3A4的前体底物。由于催化作用可能仅在将前体底物（由另一种P450）转化为底物后才变得明显，因此仅基于使用单个cDNA表达的P450的数据来推断特定P450同工型缺乏催化参与时，需要谨慎。
