Role of c-Jun-N-Terminal Kinase in Pregnane X Receptor-Mediated Induction of Human Cytochrome P4503A4 In Vitro.

Cytochrome P450 CYP3A4 is the most abundant drug-metabolizing enzyme and is responsible for the metabolism of ∼50% of clinically available drugs. Induction of CYP3A4 impacts the disposition of its substrates and leads to harmful clinical consequences, such as failure of therapy. To prevent such undesirable consequences, the molecular mechanisms of regulation of CYP3A4 need to be fully understood. CYP3A4 induction is regulated primarily by the xenobiotic nuclear receptor pregnane-X receptor (PXR). After ligand binding, PXR is translocated to the nucleus, where it binds to the CYP3A4 promoter and induces its gene expression. PXR function is modulated by phosphorylation(s) by multiple kinases. In this study, we determined the role of the c-Jun N-terminal kinase (JNK) in PXR-mediated induction of CYP3A4 enzyme in vitro. Human liver carcinoma cells (HepG2) were transfected with CYP3A4 luciferase and PXR plasmids, followed by treatment with JNK inhibitor (SP600125; SP) and PXR activators rifampicin (RIF) or hyperforin. Our results indicate that SP treatment significantly attenuated PXR-mediated induction of CYP3A4 reporter activity, as well as gene expression and enzyme activity. JNK knockdown by siRNA (targeting both JNK 1 and 2) also attenuated CYP3A4 induction by RIF. Interestingly, SP treatment attenuated JNK activation by RIF. Furthermore, treatment with RIF increased PXR nuclear levels and binding to the CYP3A4 promoter; SP attenuated these effects. This study shows that JNK is a novel mechanistic regulator of CYP3A4 induction by PXR.