Accelerated Communication INDUCTION OF CYTOCHROME P450 3A4 IN PRIMARY HUMAN HEPATOCYTES AND ACTIVATION OF THE HUMAN PREGNANE X RECEPTOR BY TAMOXIFEN AND 4-HYDROXYTAMOXIFEN

Tamoxifen is a widely utilized antiestrogen in the treatment and chemoprevention of breast cancer. Clinical studies document that tamoxifen administration markedly enhances the systemic elimination of other drugs. Additionally, tamoxifen enhances its own clearance following repeated dosing. The mechanisms that underlie these clinically important events remain unresolved. Here, we report that tamoxifen and its metabolite 4-hydroxytamoxifen markedly induce cytochrome P450 3A4, a drug-metabolizing enzyme of central importance, in primary cultures of human hepatocytes. Tamoxifen and 4-hydroxytamoxifen (1–10 M) significantly increased the CYP3A4 expression and activity (measured as the rate of testosterone 6 -hydroxylation). Maximal induction was achieved at the 5 M level. At this level, tamoxifen and 4-hydroxytamoxifen caused a 1.5to 3.3-fold (mean, 2.1-fold) and 3.4to 17-fold (mean, 7.5-fold) increase in the CYP3A4 activity, respectively. In comparison, rifampicin treatment resulted in a 6to 16fold (mean, 10.5-fold) increase. We also observed corresponding increase in the CYP3A4 immunoreactive protein and mRNA levels. Furthermore, tamoxifen and 4-hydroxytamoxifen efficaciously activated the human pregnane X receptor (hPXR; also known as the steroid xenobiotic receptor), a key regulator of CYP3A4 expression. The efficacy of tamoxifen and 4-hydroxytamoxifen relative to rifampicin for hPXR activation was 30 and 60%, respectively. Our results indicate that the mechanism of tamoxifen-mediated alteration in drug clearance pathways in humans may involve CYP3A4 induction by the parent drug and/or its metabolite. Furthermore, the CYP3A4 induction may be a result of hPXR activation. These findings have important implications for optimizing the use of tamoxifen and in the development of newer antiestrogens. Tamoxifen, a nonsteroidal triphenylethylene, is currently the endocrine therapeutic agent of choice for all stages of breast cancer. It was also recently approved for use as a chemopreventive agent in women with high risk of contracting this disease in the future. Despite its well documented beneficial effects, tamoxifen use is associated with several major problems including serious drug-drug interactions. Several clinical trials indicate that tamoxifen has the propensity to alter the drug elimination pathway(s), resulting in markedly reduced plasma levels of coadministered compounds. In this regard, recently completed clinical trials indicate that tamoxifen reduced the plasma levels of aromatase inhibitors letrozole and anastrozole by 37 and 27%, respectively (Dowsett et al., 1999; ATAC Trialists’ Group, 2001). Such drug-drug interactions are of special concern with tamoxifen since numerous women are required to take tamoxifen daily for an extended time period and as such are likely to be simultaneously exposed to many drugs and nutraceuticals. Another consequence of tamoxifen-induced changes in drug disposition is that tamoxifen pharmacokinetics exhibit time-dependent changes. There are marked differences in the single dose versus steady-state (when multiple doses have been taken) pharmacokinetics of tamoxifen (Etienne et al., 1989). The relative abundance of tamoxifen (as a fraction of the combined levels of tamoxifen and its metabolites) is lower at steady state compared with that following single dose. This is accompanied with increased relative abundance of tamoxifen metabolites at steady state. The extent of tamoxifen conversion to its metabolites has serious implications for its efficacy and toxicity since some of its metabolites are potent antiestrogens whereas some others are impliSupported by a grant from the American Cancer Society (Ohio Division), Women’s Health Program, University of Cincinnati Medical Center, the National Institutes of Health (DK53452), and the American Institute for Cancer Research. 1 Abbreviations used are: P450, cytochrome P450; hPXR, human pregnane X receptor; XREM, xenobiotic response element module; DMSO, dimethyl sulfoxide; HuH7, human hepatocellular carcinoma; HPLC, high-performance liquid