CYP2C19 participates in tolbutamide hydroxylation by human liver microsomes.
CYP2C19 participates in tolbutamide hydroxylation by human liver microsomes.
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发表时间：
2000-03
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通讯作者：
M. R. Wester;J. Lasker;Eric F Johnson;J. Raucy
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作者：
M. R. Wester;J. Lasker;Eric F Johnson;J. Raucy
Tolbutamide 是一种磺酰脲类口服降血糖药，其作用通过人类 CYP2C 亚家族的细胞色素 P-450 (CYP) 酶催化的甲苯磺酰甲基部分的羟基化而终止。尽管大多数研究表明 CYP2C9 是人类肝脏甲苯磺丁脲羟基化的唯一催化剂，但有证据表明其他 CYP2C 酶（例如 CYP2C19）也可能参与其中。为此，我们使用免疫化学方法来评估单个 CYP2C 在微粒体甲苯磺丁脲代谢中的作用。针对从人肝脏纯化的 CYP2C9 产生多克隆抗体，然后针对与固相支持物偶联的重组 CYP2C19 进行反吸附。 Western blotting显示，吸收的抗人CYP2C9制剂仅与重组CYP2C9和肝微粒体中相应的天然蛋白发生反应，并且不再识别CYP2C19和CYP2C8。单特异性抗 CYP2C9 不仅保留了抑制 CYP2C9 催化反应的能力，如纯化 CYP2C9 和人肝微粒体对双氯芬酸 4'-羟基化的显着 (90%) 抑制所证明的那样，而且还表现出代谢特异性，如其对纯化 CYP2C19 或人肝微粒体对 S-美芬妥英 4'-羟基化的可忽略不计的 (<15%) 抑制作用所证明。含有 CYP2C19 的肝微粒体。还发现单特异性抗 CYP2C9 在 CYP2C19 缺陷和含有 CYP2C19 的人肝微粒体中分别抑制甲苯磺丁脲羟基化率 93 +/- 4 和 78 +/- 6%。综上所述，我们的结果表明 CYP2C9 和 CYP2C19 均参与人肝微粒体对甲苯磺丁脲的羟基化，并且 CYP2C19 参与至少 14% 至 22% 的甲苯磺丁脲代谢。尽管人肝脏中 CYP2C19 的表达低于 CYP2C9，但它可能在表达高水平 CYP2C19 或催化缺陷的 CYP2C9 酶的受试者中的甲苯磺丁脲处置中发挥重要作用。
Tolbutamide is a sulfonylurea-type oral hypoglycemic agent whose action is terminated by hydroxylation of the tolylsulfonyl methyl moiety catalyzed by cytochrome P-450 (CYP) enzymes of the human CYP2C subfamily. Although most studies have implicated CYP2C9 as the exclusive catalyst of hepatic tolbutamide hydroxylation in humans, there is evidence that other CYP2C enzymes (e.g., CYP2C19) may also participate. To that end, we used an immunochemical approach to assess the role of individual CYP2Cs in microsomal tolbutamide metabolism. Polyclonal antibodies were raised to CYP2C9 purified from human liver, and were then back-adsorbed against recombinant CYP2C19 coupled to a solid-phase support. Western blotting revealed that the absorbed anti-human CYP2C9 preparation reacted with only recombinant CYP2C9 and the corresponding native protein in hepatic microsomes, and no longer recognized CYP2C19 and CYP2C8. Monospecific anti-CYP2C9 not only retained the ability to inhibit CYP2C9-catalyzed reactions, as evidenced by its marked (90%) inhibition of diclofenac 4'-hydroxylation by purified CYP2C9 and by human liver microsomes, but also exhibited metabolic specificity, as indicated by its negligible (<15%) inhibitory effect on S-mephenytoin 4'-hydroxylation by purified CYP2C19 or hepatic microsomes containing CYP2C19. Monospecific anti-CYP2C9 was also found to inhibit rates of tolbutamide hydroxylation by 93 +/- 4 and 78 +/- 6% in CYP2C19-deficient and CYP2C19-containing human liver microsomes, respectively. Taken together, our results indicate that both CYP2C9 and CYP2C19 are involved in tolbutamide hydroxylation by human liver microsomes, and that CYP2C19 underlies at least 14 to 22% of tolbutamide metabolism. Although expression of CYP2C19 in human liver is less than that of CYP2C9, it may play an important role in tolbutamide disposition in subjects expressing either high levels of CYP2C19 or a catalytically deficient CYP2C9 enzyme.