Comparative analysis of CYP3A heteroactivation by steroid hormones and flavonoids in different in vitro systems and potential in vivo implications.

A systematic analysis of the heteroactivation of CYP3A-mediated carbamazepine 10,11-epoxidation has been investigated in three different in vitro systems, namely recombinant CYP3A4 and CYP3A5, human liver microsomes (HLMs) and cryopreserved human hepatocytes. The effect of 10 endogenous steroids and flavonoids was studied over a range of substrate and effector concentrations. A novel heteroactivation model was used to obtain the parameters EC(200) (concentration of effector required to produce 200% control) and heteroactivation ratio (the ratio of maximum observed reaction velocity to control). The EC(200) values obtained in HLMs and human hepatocytes were corrected for nonspecific binding. Heteroactivation of CYP3A5 has been demonstrated with mean heteroactivation ratios in CYP3A5, HLMs and hepatocytes on average 2-fold greater than in recombinant CYP3A4 for most of the effectors investigated. In recombinant CYP3A4, heteroactivation was greatest at substrate concentrations below K(m). Heteroactivation increased with effector concentration in a nonlinear manner and differed between effectors (mean heteroactivation ratios varied up to 12-fold). A greater extent of heteroactivation was observed in HLMs than in human hepatocytes for steroid effectors, but the opposite was true for flavonoid effectors. The observed heteroactivation of CYP3A in intact cells supports an in vivo relevance. From the in vitro heteroactivation data, a significant increase in clearance in vivo was predicted for substrates with a high dependence on CYP3A4 to the overall elimination, indicating that heteroactivation of CYP3A may be a potential source of interindividual variability.