Comparative effects of rifabutin and rifampicin on cytochromes P450 and UDP-glucuronosyl-transferases expression in fresh and cryopreserved human hepatocytes

The aim of this study was to evaluate rifabutin (RBT) and rifampicin (RIF) capabilities in inducing various xenobiotic metabolizing enzymes such as cy tochromes P450 (CYPs) and UDP-glucuronosyl-transferases (UGTs) in cultured fresh and cryopreserved human hepatocytes. Enzyme induction was assessed th rough the use of several diagnostic markers, i.e. testosterone, midazolam ( MDZ), diazepam (DZP) and 7-ethoxyresorufin for CYP-dependent enzyme reactio ns; and AZT for UGT-dependent enzyme reactions. RBT concentrations (0.118, 0.708 mu M) were selected according to previously published pharmacokinetic data in patients. The known CYP3A4 inducer in humans, RIF, was used as a p ositive control. At the concentrations used, no sign of cytotoxicity was ev idenced. Both compounds were able to dose-dependently induce the overall me tabolism of testosterone (similar to 2-fold for RBT, similar to 4-fold for RIF) and the formation of the 6 beta-hydroxylated-derivative (up to similar to 4-fold over control for RBT and similar to 10-fold for RIF), which is C YP3A4 dependent. The other hydroxylated metabolites (16 alpha-OH and 2 alph a-OH) were also enhanced. The metabolism of MDZ, which is specifically meta bolized by CYP3A4 in humans, was also investigated following drug's exposur e to hepatocytes. DZP one, which is governed by various CYPs, including CYP 3A, was also investigated. RBT was shown to increase the biotransformation of both benzodiazepines (similar to 1.9-fold over control). Moreover, the e ffects of both drugs on ethoxyresorufin O-deethylase activity (EROD), which is representative of CYP1A1/2 isoforms, were tested. Results showed only a moderate induction of this marker (similar to 2-fold over control) when co mpared to the high effect observed after hepatocyte exposure to 3-methylcho lantene (similar to 14-fold over control). Finally, the action of RBT and R IF on UGTs expression was investigated by using AZT as diagnostic substrate : glucuronides formation was not significantly affected by the two rifamyci n derivatives. On the whole, exposure of fresh or cryopreserved human hepat ocytes to RBT dose-dependently affected the levels of drug metabolizing enz ymes in a dose-dependent manner. However, as already demonstrated by in viv o pharmacokinetic studies, its inducing properties towards CYPs. CYP3A in p articular, are less pronounced than RIF. (C) 1999 Elsevier Science Ireland Ltd. All rights reserved.