title: Effect of tamoxifen on the enzymatic activity of human cytochrome CYP2B6.
abstract: Tamoxifen is primarily used in the treatment of breast cancer. It has been approved as a chemopreventive agent for individuals at high risk for this disease. Tamoxifen is metabolized to a number of different products by cytochrome P450 enzymes. The effect of tamoxifen on the enzymatic activity of bacterially expressed human cytochrome CYP2B6 in a reconstituted system has been investigated. The 7-ethoxy-4-(trifluoromethyl)coumarin O-deethylation activity of purified CYP2B6 was inactivated by tamoxifen in a time- and concentration-dependent manner. Enzymatic activity was lost only in samples that were incubated with both tamoxifen and NADPH. The inactivation was characterized by a K(I) of 0.9 microM, a k(inact) of 0.02 min(-1), and a t(1/2) of 34 min. The loss in the 7-ethoxy-4-(trifluoromethyl)coumarin O-deethylation activity did not result in a similar percentage loss in the reduced carbon monoxide spectrum, suggesting that the heme moiety was not the major site of modification. The activity of CYP2B6 was not recovered after removal of free tamoxifen using spin column gel filtration. The loss in activity seemed to be due to a modification of the CYP2B6 and not reductase because adding fresh reductase back to the inactivated samples did not restore enzymatic activity. A reconstituted system containing purified CYP2B6, NADPH-reductase, and NADPH-generating system was found to catalyze tamoxifen metabolism to 4-OH-tamoxifen, 4'-OH-tamoxifen, and N-desmethyl-tamoxifen as analyzed by high-performance liquid chromatography analysis. Preliminary studies showed that tamoxifen had no effect on the activities of CYP1B1 and CYP3A4, whereas CYP2D6 and CYP2C9 exhibited a 25% loss in enzymatic activity.
authors.affiliation: Department of Pharmacology, University of Michigan, 1150 West Medical Center Drive, Ann Arbor, MI 48109-0632, USA.
authors.forename: Chitra
authors.initials: C
authors.lastname: Sridar
authors.forename: Ute M
authors.initials: UM
authors.lastname: Kent
authors.forename: Lisa M
authors.initials: LM
authors.lastname: Notley
authors.forename: Elizabeth M J
authors.initials: EM
authors.lastname: Gillam
authors.forename: Paul F
authors.initials: PF
authors.lastname: Hollenberg
mesh_terms: D000470:Alkaloids; D018931:Antineoplastic Agents, Hormonal / Q000737:chemistry / Q000494:pharmacology*; D001189:Aryl Hydrocarbon Hydroxylases; D044182:Benzylisoquinolines; D003374:Coumarins / Q000037:antagonists & inhibitors / Q000378:metabolism; D065702:Cytochrome P-450 CYP2B6; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000032:analysis / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004926:Escherichia coli / Q000187:drug effects / Q000201:enzymology; D006418:Heme / Q000378:metabolism; D006801:Humans; D010089:Oxidoreductases, N-Demethylating / Q000032:analysis / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013379:Substrate Specificity / Q000187:drug effects / Q000502:physiology; D013629:Tamoxifen / Q000494:pharmacology*
chemical_list: D000470:Alkaloids; D018931:Antineoplastic Agents, Hormonal; D044182:Benzylisoquinolines; D003374:Coumarins; D065607:Cytochrome P-450 Enzyme Inhibitors; D004791:Enzyme Inhibitors; D013629:Tamoxifen; C056033:7-ethoxy-4-trifluoromethylcoumarin; C069407:7-O-ethyl fangchinoline; D006418:Heme; D003577:Cytochrome P-450 Enzyme System; D001189:Aryl Hydrocarbon Hydroxylases; C585599:CYP2B6 protein, human; D065702:Cytochrome P-450 CYP2B6; D010089:Oxidoreductases, N-Demethylating