Uehara Shotaro, Uno Yasuhiro, Tomioka Etsuko, Inoue Takashi, Sasaki Erika, Yamazaki Hiroshi
Laboratory of Drug Metabolism and Pharmacokinetics, Showa Pharmaceutical University, Machida, Tokyo, 194-8543, Japan.
Pharmacokinetics and Bioanalysis Center, Shin Nippon Biomedical Laboratories, Ltd., Kainan, Wakayama, Japan.
Biopharm Drug Dispos. 2017 Sep;38(6):394-397. doi: 10.1002/bdd.2080. Epub 2017 Jun 28.
Common marmosets (Callithrix jacchus) have attracted increasing attention as a useful small non-human primate model in preclinical research. However, studies on marmoset cytochrome P450 (P450) 2E enzyme have scarcely been conducted. In this study, the full-length cDNA encoding P450 2E1 enzyme was isolated from marmoset livers by reverse transcription (RT)-polymerase chain reaction (PCR). Marmoset P450 2E1 amino acid sequences were highly identical (>88%) to those of cynomolgus monkey and human P450 2E1 enzymes. Phylogenetic analysis indicated a close evolutionary relationship among marmoset, cynomolgus monkey, and human P450 2E1 enzymes. The tissue expression pattern analyzed by real-time RT-PCR and immunoblotting demonstrated that marmoset P450 2E1 mRNA and proteins were predominantly expressed in livers. Marmoset P450 2E1 enzyme heterologously expressed in Escherichia coli catalyzed the hydroxylation of p-nitrophenol, chlorzoxazone, and theophylline, similar to cynomolgus monkey and human P450 2E1 enzymes. By kinetic analyses, those P450 2E1 enzymes catalyzed p-nitrophenol hydroxylation with similar affinities and relatively high intrinsic clearance efficiencies. These results indicated that tissue distribution and enzyme-substrate specificity of marmoset P450 2E1 were similar to cynomolgus monkey and human P450 2E1 enzymes, suggesting that marmosets are a suitable primate model for P450 2E1-dependent drug and xenobiotic metabolism.
普通狨猴（Callithrix jacchus）作为一种有用的小型非人类灵长类动物模型，在临床前研究中受到了越来越多的关注。然而，关于狨猴细胞色素P450（P450）2E酶的研究却很少。在本研究中，通过逆转录（RT）-聚合酶链反应（PCR）从狨猴肝脏中分离出编码P450 2E1酶的全长cDNA。狨猴P450 2E1氨基酸序列与食蟹猴和人类P450 2E1酶的氨基酸序列高度同源（>88%）。系统发育分析表明，狨猴、食蟹猴和人类P450 2E1酶之间存在密切的进化关系。通过实时RT-PCR和免疫印迹分析的组织表达模式表明，狨猴P450 2E1 mRNA和蛋白质主要在肝脏中表达。在大肠杆菌中异源表达的狨猴P450 2E1酶催化对硝基苯酚、氯唑沙宗和茶碱的羟基化反应，类似于食蟹猴和人类P450 2E1酶。通过动力学分析，这些P450 2E1酶以相似的亲和力和相对较高的内在清除效率催化对硝基苯酚羟基化反应。这些结果表明，狨猴P450 2E1的组织分布和酶-底物特异性与食蟹猴和人类P450 2E1酶相似，表明狨猴是P450 2E1依赖性药物和外源性物质代谢的合适灵长类动物模型。
