School of Pharmacy and Pharmaceutical Sciences, University of Manchester, Stopford Building, Oxford Road, Manchester, M13 9PT, UK.
Drug Metab Dispos. 2012 Jul;40(7):1279-89. doi: 10.1124/dmd.112.045286. Epub 2012 Mar 26.
Repaglinide is presently recommended by the U.S. Food and Drug Administration as a clinical CYP2C8 probe, yet current in vitro and clinical data are inconsistent concerning the role of this enzyme in repaglinide elimination. The aim of the current study was to perform a comprehensive investigation of repaglinide metabolic pathways and assess their contribution to the overall clearance. Formation of four repaglinide metabolites was characterized using in vitro systems with differential complexity. Full kinetic profiles for the formation of M1, M2, M4, and repaglinide glucuronide were obtained in pooled cryopreserved human hepatocytes, human liver microsomes, human S9 fractions, and recombinant cytochrome P450 enzymes. Distinct differences in clearance ratios were observed between CYP3A4 and CYP2C8 for M1 and M4 formation, resulting in a 60-fold M1/M4 ratio in recombinant (r) CYP3A4, in contrast to 0.05 in rCYP2C8. Unbound K(m) values were within 2-fold for each metabolite across all in vitro systems investigated. A major system difference was seen in clearances for the formation of M2, which is suggested to be a main metabolite of repaglinide in vivo. An approximately 7-fold higher unbound intrinsic clearance was observed in hepatocytes and S9 fractions in comparison to microsomes; the involvement of aldehyde dehydrogenase in M2 formation was shown for the first time. This systematic analysis revealed a comparable in vitro contribution from CYP2C8 and CYP3A4 to the metabolism of repaglinide (<50%), whereas the contribution of glucuronidation ranged from 2 to 20%, depending on the in vitro system used. The repaglinide M4 metabolic pathway is proposed as a specific CYP2C8 probe for the assessment of drug-drug interactions.
瑞格列奈目前被美国食品和药物管理局推荐为一种临床 CYP2C8 探针，但目前关于该酶在瑞格列奈消除中的作用的体外和临床数据并不一致。本研究的目的是全面研究瑞格列奈的代谢途径，并评估其对总清除率的贡献。使用具有不同复杂程度的体外系统对瑞格列奈代谢产物的形成进行了特征描述。在冷冻保存的人肝细胞、人肝微粒体、人 S9 级分和重组细胞色素 P450 酶中获得了 M1、M2、M4 和瑞格列奈葡萄糖醛酸的形成的完整动力学谱。在 M1 和 M4 的形成中，CYP3A4 和 CYP2C8 之间观察到清除率比值的明显差异，导致重组（r）CYP3A4 中 M1/M4 比值为 60 倍，而 rCYP2C8 中为 0.05。在所研究的所有体外系统中，每种代谢产物的未结合 K(m) 值均相差 2 倍以内。在 M2 的形成中观察到一个主要的系统差异，M2 被认为是瑞格列奈体内的主要代谢产物。与微粒体相比，肝细胞和 S9 级分中 M2 形成的未结合固有清除率高约 7 倍；首次表明醛脱氢酶参与了 M2 的形成。这种系统分析显示，CYP2C8 和 CYP3A4 对瑞格列奈代谢的体外贡献相当（<50%），而葡萄糖醛酸化的贡献范围为 2%至 20%，具体取决于所用的体外系统。瑞格列奈 M4 代谢途径被提议作为评估药物相互作用的特异性 CYP2C8 探针。
