Abstract
The effects of inhibitors of 2,3-oxidosqualene:lanosterol cyclase (cyclase) on cytochrome P450 expression were investigated in primary cultures of rat hepatocytes. Treatment of hepatocyte cultures for 24 h with either of the inhibitors [4′-(6-allyl-methyl-amino-hexyloxy)-2′-fluoro-phenyl]-(4-bromophenyl)-methanone fumarate (Ro 48-8071) or trans-N-(4-chlorobenzoyl)-N-methyl-(4-dimethylaminomethylphenyl)-cyclohexylamine (BIBX 79) selectively increased CYP3A mRNA and immunoreactive protein contents, with maximal accumulations occurring at 3 × 10-5 M Ro 48-8071 and 10-4 M BIBX 79. The abilities of Ro 48-8071, BIBX 79, and 3β-(2-diethylaminoethoxy)androst-5-en-17-one·HCl (U18666A) to induce murine CYP3A were abolished in hepatocyte cultures prepared from pregnane X receptor (PXR)-null mice, and cotransfection of primary cultured rat hepatocytes with a dominant-negative PXR prevented cyclase inhibitor-inducible luciferase expression from a PXR-responsive reporter plasmid. Cyclase inhibitor-mediated CYP3A mRNA induction was eliminated when primary cultured rat hepatocytes were cotreated with any of the following agents that inhibit steps upstream of cyclase in the cholesterol biosynthetic pathway: squalestatin 1 (squalene synthase inhibitor), (E)N-ethyl-N-(6,6-dimethyl-2-hepten-4-ynyl)-3-[(3,3′-bithiophen-5-yl)methoxy]benzenemethanamine (NB-598, squalene monooxygenase inhibitor), or pravastatin (HMG-CoA reductase inhibitor). Ro 48-8071-inducible CYP3A mRNA expression was restored when pravastatin-treated cultures were incubated with medium containing mevalonate. The concentration-dependence of Ro 48-8071-mediated CYP3A mRNA induction corresponded to the cellular contents of metabolically labeled squalene 2,3-oxide and squalene 2,3:22,23-dioxide, but not 24(S),25-epoxycholesterol. These results indicate that cyclase inhibitors are capable of inducing CYP3A expression in primary cultured rat and mouse hepatocytes and that the effect is mediated as a consequence of cyclase blockade through the evoked accumulation of one or more squalene metabolites that activate the PXR.
- Received September 4, 2003.
- Accepted February 17, 2004.
- The American Society for Pharmacology and Experimental Therapeutics
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