Showa Pharmaceutical University, Machida, Tokyo 194-8543, Japan.
Department of Marmoset Research, Central Institute for Experimental Animals, Kawasaki, Japan.
Biochem Pharmacol. 2018 Jun;152:272-278. doi: 10.1016/j.bcp.2018.04.002. Epub 2018 Apr 5.
Common marmosets (Callithrix jacchus) are potentially useful nonhuman primate models for preclinical studies. An anti-inflammatory drug, diclofenac is reportedly metabolized mainly by human cytochrome P450 (P450) 2C9 to 4'-hydroxydiclofenac and minorly by P450 3A4 to 5-hydroxydiclofenac that leads to reactive intermediates. In this study, in vivo pharmacokinetics in six marmosets and in vitro metabolism in nine marmoset liver microsomes were analyzed using diclofenac to evaluate marmosets as preclinical drug metabolism models. In wild-type marmosets genotyped for P450 2C19 p.[(Phe7Leu; Ser254Leu; Ile469Thr)], plasma levels of 5-hydroxydiclofenac and 4'-hydroxydiclofenac were roughly similar, but the homozygotes showed approximately three-times higher plasma levels of 5-hydroxydiclofenac than those of 4'-hydroxydiclofenac after oral administrations of diclofenac (50 mg/kg). Nine marmoset liver microsomes catalyzed mainly diclofenac 5-hydroxylation with no significant effects of the the P450 2C19 genotype, and partly diclofenac 4'-hydroxylation that depended on the P450 2C19 genotype, at substrate concentrations of 10 µM and 100 µM. Both Michaels-Menten constant K values for diclofenac 4'-hydroxylation in human and marmoset liver microsomes were ∼30 μM and those for diclofenac 5-hydroxylation were ∼120 μM. Ketoconazole significantly suppressed only diclofenac 5-hydroxylation in marmoset or human liver microsomes at 0.030 μM, indicating main contribution of P450 3A enzymes, which were found to be P450 3A5/90 using recombinant marmoset P450 3A enzymes. These results suggest that marmosets would be a functional animal model for in vivo and in vitro metabolites likely generated via arene oxide intermediates of diclofenac by P450 3A enzymes in humans, primarily because marmosets lack the ortholog of human P450 2C9.
普通狨猴（Callithrix jacchus）是一种潜在的有用的非人类灵长类动物模型，可用于临床前研究。据报道，一种抗炎药物双氯芬酸主要通过人细胞色素 P450（P450）2C9 代谢为 4'-羟基双氯芬酸，次要通过 P450 3A4 代谢为 5-羟基双氯芬酸，导致反应性中间体。在这项研究中，使用双氯芬酸在六只狨猴体内进行药代动力学分析，并在九只狨猴肝微粒体中进行体外代谢，以评估狨猴作为临床前药物代谢模型。在 P450 2C19 p.[（Phe7Leu; Ser254Leu; Ile469Thr）]基因型为野生型的狨猴中，5-羟基双氯芬酸和 4'-羟基双氯芬酸的血浆水平大致相似，但杂合子口服双氯芬酸（50mg/kg）后 5-羟基双氯芬酸的血浆水平约为 4'-羟基双氯芬酸的三倍。九只狨猴肝微粒体主要催化双氯芬酸 5-羟化，P450 2C19 基因型无显著影响，部分催化双氯芬酸 4'-羟化，这取决于 P450 2C19 基因型，在 10μM 和 100μM 的底物浓度下。人肝微粒体和狨猴肝微粒体中双氯芬酸 4'-羟化的米氏常数 K 值均约为 30μM，双氯芬酸 5-羟化的 K 值均约为 120μM。酮康唑在 0.030μM 时仅显著抑制狨猴或人肝微粒体中的双氯芬酸 5-羟化，表明主要贡献来自 P450 3A 酶，使用重组狨猴 P450 3A 酶发现这些酶为 P450 3A5/90。这些结果表明，狨猴将成为一种功能性动物模型，用于体内和体外代谢物的研究，这些代谢物可能是通过 P450 3A 酶在人体内生成的二氯芬酸芳基氧化物中间体产生的，主要是因为狨猴缺乏与人 P450 2C9 同源的酶。
