Global Proteomic Analysis of Human Liver Microsomes: Rapid Characterization and Quantification of Hepatic Drug-Metabolizing Enzymes
Global Proteomic Analysis of Human Liver Microsomes: Rapid Characterization and Quantification of Hepatic Drug-Metabolizing Enzymes
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DOI：
10.1124/dmd.116.074732
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发表时间：
2017-06-01
影响因子：
3.9
通讯作者：
Barber, Jill
中科院分区：
文献类型：
--
作者：
Achour, Brahim;Al Feteisi, Hajar;Barber, Jill
许多遗传和环境因素导致药物代谢和转运的个体差异，深刻影响疗效和毒性。精确给药，即将药物剂量靶向明确表征的亚群，取决于该亚群内药物代谢酶 (DME) 和转运蛋白概况的定量模型，并由定量蛋白质组学提供信息。我们报告了为此目的首次使用离子淌度质谱法，从而可以对来自不同个体的人肝微粒体（HLM）蛋白进行快速、稳健、无标记的定量。在四个样品中鉴定并定量了大约 1000 种蛋白质，其中平均包括 70 种 DME。技术变异和生物学变异是有区别的，技术变异约占总变异的10%。患者之间的生物学差异已被清楚地识别，样本显示了细胞色素 P450 和尿苷 5'-二磷酸葡萄糖醛酸基转移酶的一系列表达谱。我们的结果与之前的目标方法数据非常吻合。然而，无标记方法可以对体外系统进行更全面的表征，首次表明 HLM 具有显着的异质性。此外，DME 的传统测量单位（pmol mg(-1) HLM 蛋白）会因不相关的高丰度蛋白的变异性而引入误差。对这种变异性的模拟表明，表观 CYP3A4 丰度高达 1.7 倍的变化是人为的，DME 丰度之间的背景正相关性高达 0.2（斯皮尔曼相关系数）。我们建议，药代动力学预测和体内临床情况（基于生理学的药代动力学和体外-体内外推）中使用的蛋白质浓度应参考组织质量。
Many genetic and environmental factors lead to interindividual variations in the metabolism and transport of drugs, profoundly affecting efficacy and toxicity. Precision dosing, that is, targeting drug dose to a well characterized subpopulation, is dependent on quantitative models of the profiles of drug-metabolizing enzymes (DMEs) and transporters within that subpopulation, informed by quantitative proteomics. We report the first use of ion mobility-mass spectrometry for this purpose, allowing rapid, robust, label-free quantification of human liver microsomal (HLM) proteins from distinct individuals. Approximately 1000 proteins were identified and quantified in four samples, including an average of 70 DMEs. Technical and biological variabilities were distinguishable, with technical variability accounting for about 10% of total variability. The biological variation between patients was clearly identified, with samples showing a range of expression profiles for cytochrome P450 and uridine 5'-diphosphoglucuronosyltransferase enzymes. Our results showed excellent agreement with previous data from targeted methods. The label-free method, however, allowed a fuller characterization of the in vitro system, showing, for the first time, that HLMs are significantly heterogeneous. Further, the traditional units of measurement of DMEs (pmol mg(-1) HLM protein) are shown to introduce error arising from variability in unrelated, highly abundant proteins. Simulations of this variability suggest that up to 1.7-fold variation in apparent CYP3A4 abundance is artifactual, as are background positive correlations of up to 0.2 (Spearman correlation coefficient) between the abundances of DMEs. We suggest that protein concentrations used in pharmacokinetic predictions and scaling to in vivo clinical situations (physiologically based pharmacokinetics and in vitro-in vivo extrapolation) should be referenced instead to tissue mass.