Downregulation of HepaRG Cell CYP Genes by Hypoxia Inducible Factor Prolyl Hydroxylase (HIF-PH) Inhibitor

Abstract

Metabolic enzymes are occasionally downregulated in in vitro induction studies. Recently, HepaRG cells have been used for CYP induction assays instead of human hepatocytes in the early drug discovery stage; however, there is limited information on CYP downregulation by drug stimulation. In this study, we evaluated the effect of hypoxia-inducible factor-prolyl hydroxylase (HIF-PH) inhibitors, which downregulate CYP in human hepatocytes, on CYP gene expression in HepaRG cells. Microarray analysis to determine the expression levels of pharmacokinetics-related enzymes and RT-PCR to determine the expression levels of CYP3A4, CYP2B6, CYP1A2, and their nuclear receptor mRNA were conducted in HepaRG cells treated with HIF-PH inhibitors. Treatment of HepaRG cells with HIF-PH inhibitors decreased the expression of several pharmacokinetics-related metabolic enzymes, whereas Erythropoietin (EPO) and Pyruvate Dehydrogenase Kinase1 (PDK1) genes were induced. The expression of CYP3A4 and CYP2B6 in HepaRG cells showed concentration- and time-dependent downregulation following treatment with the HIF-PH inhibitor. The downregulation of these enzymes was correlated with the decrease of PXR/RXRα and CAR/RXRα, respectively. CYP1A2 decreased transiently, but recovered with continued HIF-PH inhibitor treatment. CYP3A4 and CYP2B6 were downregulated by HIF-PH inhibitors in HepaRG cells and human hepatocytes. In contrast, CYP1A2 in HepaRG cells responded differently to HIF-PH inhibitors than in human hepatocytes. Since CYP downregulation is commonly observed with HIF-PH inhibitors, along with the induction of EPO and PDK1 genes, stabilizing HIF may be one of the factors involved in CYP downregulation.