Human cytochromes P4503A metabolize numerous therapeutic agents, bioactivate environmental xenobiotics, and are induced by drugs and pesticides. Because there is large interindividual variation in basal and inducible expression of CYP3A that affects therapeutic outcome and could serve as a susceptibility marker for environmentally caused diseases, the factors which regulate CYP3A are under intense investigation. Our recent studies have established that the product of the multidrug resistance gene (MDR1), the drug efflux transporter P-glycoprotein (Pgp), influences the pharmacological disposition of rifampicin and is a major determinant of the extent to which rifampicin induces cytochrome P4503A (CYP3A). Because Pgp transports steroids and other structurally diverse xenobiotics that are CYP3A substrates and inducers, this grant will extend our recent study to test hypothesis 1, that Pgp is a determinant of basal CYP3A expression and the magnitude of CYP3A induction by many other structurally unrelated xenobiotics. Drug disposition and drug-induction of CYP3A will analyzed in two model systems, an in vitro system (LS180 cells) in which human MDR1 Pgp is overexpressed and an in vivo system that lacks mdr1a/1b/Pgp (mdrla/lb (-/-) mice). The capabilities of mouse and human MDR/Pgp to transport some CYP3A inducers not previously demonstrated to be Pgp substrates will also be tested. To maximally capitalize on the mdrla/1b knockout mice and on other murine transgenic models, this grant will characterize CYP3A immunoreactive proteins, a representative CYP3A activity and CYP3A mRNAs in mouse liver. Hypothesis 2 will test whether Pgp, by influencing the intracellular concentration of CYP3A substrates, is a determinant of the extent of CYP3A-metabolites formed in the cell, and whether Pgp transports CYP3A generated metabolites.
This aim uses our novel cell lines which form polarized epithelium in culture and stably express functional CYP3A4 and Pgp. In total, this grant will explore the nature of the interaction between CYP3A substrates and inducers at the level of their common membrane transporter, Pgp, and the ramifications of these Pgp interactions on CYP3A mediated metabolism and the CYP3A inductive response.
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