Faucette Stephanie R, Wang Hongbing, Hamilton Geraldine A, Jolley Summer L, Gilbert Darryl, Lindley Celeste, Yan Bingfang, Negishi Masahiko, LeCluyse Edward L
Division of Drug Delivery and Disposition, School of Pharmacy, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599-7360, USA.
Drug Metab Dispos. 2004 Mar;32(3):348-58. doi: 10.1124/dmd.32.3.348.
The objectives of this study were to evaluate the ability of 14 compounds, which differentially activate human pregnane X receptor (hPXR), to induce CYP2B6 expression and to compare CYP2B6 and CYP3A4 concentration- and time-dependent induction by select inducers. Three primary human hepatocyte preparations were treated daily for 3 days with three concentrations of all compounds. Additional concentration- and/or time-response studies were conducted with clotrimazole, phenytoin, phenobarbital, and rifampin in six preparations. CYP2B6 and CYP3A4 protein and activities were assessed by Western blotting, bupropion hydroxylation, and testosterone 6beta-hydroxylation, respectively. To evaluate hPXR activation by the 14 compounds, reporter gene assays were conducted using Huh7 cells cotransfected with hPXR and a CYP2B6 (NR1)5-LUC reporter plasmid. Clotrimazole, phenobarbital, rifampin, and ritonavir strongly induced CYP2B6 and activated hPXR; dexamethasone t-butylacetate and sulfinpyrazone induced CYP2B6 weakly and activated hPXR moderately; paclitaxel strongly activated hPXR but did not increase CYP2B6 expression; carbamazepine and phenytoin moderately or strongly increased CYP2B6 expression but weakly activated hPXR; and dexamethasone, methotrexate, probenecid, sulfadimidine, and troleandomycin demonstrated weak or negligible effects on CYP2B6 and hPXR. EC50 values for CYP2B6 and CYP3A4 induction by clotrimazole, phenobarbital, phenytoin, and rifampin were strongly correlated (r2 = 0.99) and were statistically indistinguishable for clotrimazole, phenytoin, and rifampin. Kinetic constants governing time-dependent induction by phenobarbital and rifampin were also similar between CYP2B6 and CYP3A4. These results indicate that CYP2B6 is highly inducible by known CYP3A4 inducers and suggest that hPXR is a major determinant of CYP2B6-inducible expression for many, but not all, compounds evaluated in this study.
本研究的目的是评估14种能差异性激活人孕烷X受体（hPXR）的化合物诱导CYP2B6表达的能力，并比较选定诱导剂对CYP2B6和CYP3A4的浓度及时间依赖性诱导作用。用三种浓度的所有化合物对三种原代人肝细胞制剂进行为期3天的每日处理。对克霉唑、苯妥英、苯巴比妥和利福平在六种制剂中进行了额外的浓度和/或时间反应研究。分别通过蛋白质免疫印迹法、安非他酮羟基化反应和睾酮6β-羟基化反应评估CYP2B6和CYP3A4的蛋白及活性。为评估这14种化合物对hPXR的激活作用，使用与hPXR和CYP2B6（NR1）5-LUC报告质粒共转染的Huh7细胞进行报告基因检测。克霉唑、苯巴比妥、利福平和利托那韦强烈诱导CYP2B6并激活hPXR；醋酸地塞米松叔丁酯和磺吡酮弱诱导CYP2B6并中度激活hPXR；紫杉醇强烈激活hPXR但不增加CYP2B6表达；卡马西平和苯妥英中度或强烈增加CYP2B6表达但弱激活hPXR；地塞米松、甲氨蝶呤、丙磺舒、磺胺二甲嘧啶和醋竹桃霉素对CYP2B6和hPXR表现出微弱或可忽略不计的影响。克霉唑、苯巴比妥、苯妥英和利福平诱导CYP2B6和CYP3A4的半数效应浓度（EC50）值高度相关（r2 = 0.99），且克霉唑、苯妥英和利福平的EC50值在统计学上无显著差异。苯巴比妥和利福平对CYP2B6和CYP3A4时间依赖性诱导的动力学常数也相似。这些结果表明，已知的CYP3A4诱导剂可高度诱导CYP2B6，提示hPXR是本研究中评估的许多（但不是全部）化合物诱导CYP2B6表达的主要决定因素。
