Gupta Anshul, Mugundu Ganesh M, Desai Pankaj B, Thummel Kenneth E, Unadkat Jashvant D
Department of Pharmaceutics, University of Washington, Seattle, Washington 98195, USA.
Drug Metab Dispos. 2008 Jun;36(6):1172-80. doi: 10.1124/dmd.107.018689. Epub 2008 Mar 10.
Lack of an established cell line model to study induction of cytochromes P450 (P450s) and drug transporters poses a challenge in predicting in vivo drug-drug interactions. Although not well characterized, LS180 cells could be an excellent cell line to study induction of P450s and transporters because they express pregnane X receptor (PXR). Therefore, as part of a larger study of in vitro to in vivo prediction of inductive drug interactions, we determined induction of various P450s and drug transporters by the anti-human deficiency virus protease inhibitors (PIs) and the prototypic inducer, rifampin, in LS180 cells. Among these proteins, the various PIs significantly induced (n = 3-5) only CYP3A4 and multidrug resistance transporter 1 (MDR1) transcripts (2- to 50-fold). CYP3A4 activity (1'-hydroxymidazolam formation) was increased (2-fold) by rifampin (10 microM) but was reduced by the PIs (1.5- to 7-fold). Surprisingly, constitutive androstane receptor 1 (CAR1) was not found to be expressed in these cells. Additionally, using a reporter assay, we found that PIs did not activate CAR3 (the natural splice variant of CAR1) but significantly activated PXR (2- to 24-fold), which correlated well with induction of CYP3A4 and MDR1 transcripts (approximately r = 0.9). Furthermore, in a PXR-knockdown stable LS180 cell line, induction of CYP3A4 and MDR1 mRNA after treatment with PIs and rifampin was significantly reduced (1.4- to 5-fold) compared with that in PXR nonsilenced cells. Based on these data, we conclude that LS180 cells could be used as a readily available, high-throughput cell line to screen for PXR-mediated induction of CYP3A4 and MDR1 transcripts. These data also indicate that the majority of the PIs are likely to produce intestinal drug-drug interactions by inactivating or inhibiting CYP3A enzymes even though they induce CYP3A4 and MDR1 transcripts via PXR.
缺乏用于研究细胞色素P450（P450s）和药物转运蛋白诱导作用的成熟细胞系模型，这给预测体内药物相互作用带来了挑战。尽管特征尚不明确，但LS180细胞可能是研究P450s和转运蛋白诱导作用的优秀细胞系，因为它们表达孕烷X受体（PXR）。因此，作为体外到体内诱导性药物相互作用预测的一项更大规模研究的一部分，我们在LS180细胞中测定了抗人类免疫缺陷病毒蛋白酶抑制剂（PIs）和原型诱导剂利福平对各种P450s和药物转运蛋白的诱导作用。在这些蛋白中，各种PIs仅显著诱导（n = 3 - 5）CYP3A4和多药耐药转运蛋白1（MDR1）转录本（2至50倍）。利福平（10 microM）使CYP3A4活性（1'-羟基咪达唑仑形成）增加（2倍），但PIs使其降低（1.5至7倍）。令人惊讶的是，在这些细胞中未发现组成型雄甾烷受体1（CAR1）表达。此外，通过报告基因测定，我们发现PIs未激活CAR3（CAR1的天然剪接变体），但显著激活PXR（2至24倍），这与CYP3A4和MDR1转录本的诱导良好相关（约r = 0.9）。此外，在PXR敲低的稳定LS180细胞系中，与PXR未沉默的细胞相比，用PIs和利福平处理后CYP3A4和MDR1 mRNA的诱导显著降低（1.4至5倍）。基于这些数据，我们得出结论，LS180细胞可作为一种易于获得的高通量细胞系，用于筛选PXR介导的CYP3A4和MDR1转录本的诱导。这些数据还表明，大多数PIs可能通过使CYP3A酶失活或抑制来产生肠道药物相互作用，尽管它们通过PXR诱导CYP3A4和MDR1转录本。
