CEA, Pharmacology and Immunoanalysis Unit, DSV/iBiTecS, 91191 Gif-Sur-Yvette Cedex, France.
Rapid Commun Mass Spectrom. 2010 Aug 30;24(16):2407-19. doi: 10.1002/rcm.4641.
Phenotyping based on drug metabolism activity appears to be informative regarding mechanism-based interactions during drug development. We report here the first steps of the development of the innovative CIME cocktail. This cocktail is designed not only for the major cytochrome P450, with caffeine, amodiaquine, tolbutamide, omeprazole, dextromethorphan and midazolam as substrates of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6 and CYP3A, respectively, but also phase II enzymes UGT 1A1/6/9 with acetaminophen, P-gp and OATP1B1 with digoxin and rosuvastatin, and renal function with memantine. An assay combining ultra-performance liquid chromatography using a 1.7 microm particle size column with tandem mass spectrometry (UPLC/MS/MS) was set up for the simultaneous quantification of the 20 substrates and metabolites after extraction from human plasma using solid-phase extraction. The method was validated in the spirit of the FDA guidelines. Mean accuracy ranged from 87.7 to 115%, the coefficient of variance (CV%) of intra- and inter-run from 1.7 to 16.4% and from 1.6 to 14.9%, respectively, and for the limit of quantification (LOQ) with ten lots of plasma, accuracy ranged from 84 to 115% and CV% precision was <16%. Short-term stability was evaluated in eluate (4 h, room temperature), plasma (24 h, room temperature), the autosampler (24 h, 4 degrees C) and in three freeze/thaw cycles in plasma. All except three analytes were stable under these conditions. For the three others a specific process can be followed. This robust, fast and sensitive assay in human plasma provides an analytical tool for ten-probe drugs of the CIME cocktail. Clinical samples will be assayed in the near future using this new assay method.
基于药物代谢活性的表型分析似乎为药物开发过程中的基于机制的相互作用提供了信息。我们在此报告了创新的 CIME 鸡尾酒的开发的第一步。该鸡尾酒不仅设计用于主要细胞色素 P450，其中咖啡因、阿莫地喹、甲苯磺丁脲、奥美拉唑、右美沙芬和咪达唑仑分别为 CYP1A2、CYP2C8、CYP2C9、CYP2C19、CYP2D6 和 CYP3A 的底物，还设计用于 UGT1A1/6/9 与对乙酰氨基酚、P-糖蛋白和 OATP1B1 与地高辛和瑞舒伐他汀以及用于评估肾功能的美金刚。该方法采用 1.7 微米粒径色谱柱的超高效液相色谱与串联质谱（UPLC/MS/MS）相结合，通过固相萃取从人血浆中提取后，可同时定量 20 种底物和代谢物。该方法是按照 FDA 指南的精神进行验证的。平均准确度范围为 87.7%至 115%，内和日间的变异系数（CV%）分别为 1.7%至 16.4%和 1.6%至 14.9%，对于十个血浆样本的定量限（LOQ），准确度范围为 84%至 115%，CV%精度<16%。在洗脱液（4 小时，室温）、血浆（24 小时，室温）、自动进样器（24 小时，4 摄氏度）和三个血浆冷冻/解冻循环中评估短期稳定性。除了三个分析物外，所有分析物在这些条件下均稳定。对于其余三个分析物，可以遵循特定的处理过程。这种在人血浆中快速、灵敏的分析方法为 CIME 鸡尾酒的十种探针药物提供了分析工具。不久将使用这种新的分析方法检测临床样本。
