The long-term goals of this project are to determine the molecular factors that govern the drug-induced regulation of cytochrome P450 2B6 (CYP2B6) in human liver. Historically overlooked as an enzyme of consequence, recent evidence suggests that CYP2B6 plays a larger role in the elimination of drugs than previously believed and its specific content within the livers of some individuals can rival that of CYP3A4 when induced by potent agents, such as rifampicin or phenytoin. In the past several years, the nuclear receptor CAR (constitutive androstane receptor) has been implicated as a key mediator of xenobiotic regulation of CYP2B genes in many species. More recently, cross-talk between PXR and CAR for binding promoter sequences of both CYP3A and CYP2B genes suggests that both receptors can play a role in the regulation of either P450 gene. Notably, we have observed a very high correlation between the drug-induced expression of CYP2B6 in human hepatocytes and the activation of hPXR. By contrast, none of the potent inducers of CYP2B6 that we have identified to date show significant activation of hCAR. These results strongly suggest that there are significant and important species-specific differences in nuclear receptor regulation of CYP2B genes. The exact nature and dynamics of the relationship between these nuclear receptors in the regulation of CYP2B6 remains unclear, but we hypothesize that it is predominantly determined by hPXR activation, not hCAR. Accordingly, the SPECIFIC AIMS of this proposal are to test the following hypotheses: 1) induction of CYP2B6 in primary cultures of human hepatocytes correlates with activation of hPXR, and not hCAR, 2) hPXR transactivates CYP2B6 gene expression through the PBREM response element, 3) a distal XREM-Iike enhancer module is involved in the transactivation of CYP2B6 expression by hPXR, and 4) there are fundamental differences in the cellular disposition and biochemical activation steps of hCAR and mCAR. The data derived from the proposed studies should provide substantial insight into the molecular mechanisms of CYP2B6 expression in humans, and thus allow for better predictions of drug-drug interactions with CYP2B6 substrates and inducers.
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