Stevens Jeffrey C, Hines Ronald N, Gu Chungang, Koukouritaki Sevasti B, Manro Jason R, Tandler Peter J, Zaya Matthew J
Pfizer Pharmacokinetics, Dynamics, and Metabolism, 301 Henrietta St., 7265-300-306, Kalamazoo, MI 49007, USA.
J Pharmacol Exp Ther. 2003 Nov;307(2):573-82. doi: 10.1124/jpet.103.054841. Epub 2003 Sep 15.
The human cytochrome P4503A forms show expression patterns subject to developmental influence. CYP3A7 and CYP3A4 are generally classified as the major fetal and adult liver forms, respectively. However, characterization of CYP3A4, -3A5, and -3A7 developmental expression has historically been confounded by the lack of CYP3A isoform-specific antibodies or marker enzyme activities. Therefore, the objective of this study was to characterize the developmental expression of hepatic CYP3A forms from early gestation to 18 years of age using up to 212 fetal and pediatric liver samples. Based on immunoquantitation, CYP3A5 protein expression was found to be highly variable, generally independent of age, and more frequently observed for African-American individuals. For differentiation of CYP3A4 and -3A7 levels, dehydroepiandrosterone metabolite patterns for expressed CYP3A forms were characterized and used for simultaneous quantitation of protein levels within liver microsome samples. The major metabolite formed by CYP3A4, 7beta-hydroxy-dehydroepiandrosterone, was identified based on cochromatography and mass spectra matching with the authentic standard. Kinetic analysis showed a 34-fold greater intrinsic clearance of 7beta-hydroxy-dehydroepiandrosterone by CYP3A4 versus -3A7, whereas CYP3A7 showed the highest 16alpha-hydroxy-dehydroepiandrosterone intrinsic clearance. Metabolite profiles for the expressed enzymes were fit to a multiple response model and CYP3A4 and -3A7 levels in fetal and pediatric liver microsome samples were calculated. Fetal liver microsomes showed extremely high CYP3A7 levels (311-158 pmol/mg protein) and significant expression through 6 months postnatal age. Low CYP3A4 expression was noted for fetal liver (< or =10 pmol/mg), with mean levels increasing with postnatal age.
人类细胞色素P4503A亚型呈现出受发育影响的表达模式。CYP3A7和CYP3A4通常分别被归类为主要的胎儿型和成人肝脏型。然而，由于缺乏CYP3A亚型特异性抗体或标记酶活性，CYP3A4、-3A5和-3A7发育表达的特征描述在历史上一直受到困扰。因此，本研究的目的是使用多达212份胎儿和儿科肝脏样本，来描述从妊娠早期到18岁肝脏中CYP3A亚型的发育表达情况。基于免疫定量分析，发现CYP3A5蛋白表达高度可变，通常与年龄无关，并且在非裔美国人个体中更常见。为了区分CYP3A4和-3A7的水平，对表达的CYP3A亚型的脱氢表雄酮代谢物模式进行了表征，并用于同时定量肝脏微粒体样本中的蛋白水平。通过与真实标准品的共色谱和质谱匹配，鉴定出由CYP3A4形成的主要代谢物7β-羟基-脱氢表雄酮。动力学分析表明，CYP3A4对7β-羟基-脱氢表雄酮的内在清除率比CYP3A7高34倍，而CYP3A7对16α-羟基-脱氢表雄酮的内在清除率最高。将表达酶的代谢物谱拟合到多反应模型中，并计算胎儿和儿科肝脏微粒体样本中CYP3A4和-3A7的水平。胎儿肝脏微粒体显示出极高的CYP3A7水平（311-158 pmol/mg蛋白），并且在出生后6个月内都有显著表达。胎儿肝脏中CYP3A4表达较低（≤10 pmol/mg），其平均水平随出生后年龄增加。
