Metabolism of Tamoxifen by Recombinant Human Cytochrome P450 Enzymes: Formation of the 4-Hydroxy, 4′-Hydroxy andN-Desmethyl Metabolites and Isomerization oftrans-4-Hydroxytamoxifen

Abstract

ABSTRACT:The cytochrome P450 (P450)-mediated biotransformation of tamoxifen is important in determining both the clearance of the drug and its conversion to the active metabolite, trans-4-hydroxytamoxifen. Biotransformation by P450 forms expressed extrahepatically, such as in the breast and endometrium, may be particularly important in determining tissue-specific effects of tamoxifen. Moreover, tamoxifen may serve as a useful probe drug to examine the regioselectivity of different forms. Tamoxifen metabolism was investigated in vitro using recombinant human P450s. Forms CYP1A1, 1A2, 1B1, 2A6, 2B6, 2C9, 2C19, 2D6, 2E1, 3A4, 3A5, and 3A7 were coexpressed in Escherichia coli with recombinant human NADPHcytochrome P450 reductase. Bacterial membranes were harvested and incubated with tamoxifen or trans-4-hydroxytamoxifen under conditions supporting P450-mediated catalysis. CYP2D6 was the major catalyst of 4-hydroxylation at low tamoxifen concentrations (170 ؎ 20 pmol/40 min/0.2 nmol P450 using 18 M tamoxifen), but CYP2B6 showed significant activity at high substrate concentrations (28.1 ؎ 0.8 and 3.1 ؎ 0.5 nmol/120 min/0.2 nmol P450 for CYP2D6 and CYP2B6, respectively, using 250 M tamoxifen). These two forms also catalyzed 4-hydroxylation (13.0 ؎ 1.9 and 1.4 ؎ 0.1 nmol/120 min/0.2 nmol P450, respectively, for CYP2B6 and CYP2D6 at 250 M tamoxifen; 0.51 ؎ 0.08 pmol/40 min/0.2 nmol P450 for CYP2B6 at 18 M tamoxifen). Tamoxifen N-demethylation was mediated by CYP2D6, 1A1, 1A2, and 3A4, at low substrate concentrations, with contributions by CYP1B1, 2C9, 2C19 and 3A5 at high concentrations. CYP1B1 was the principal catalyst of 4-hydroxytamoxifen trans-cis isomerization but CYP2B6 and CYP2C19 also contributed.

Association of Human Cytochrome P450 1A1 (CYP1A1) and Sulfotransferase 1A1 (SULT1A1) Polymorphisms with Differential Metabolism and Cytotoxicity of Aminoflavone

“…Specifically, tamoxifen is metabolized predominantly by CYP2D6 to form two active metabolites, 4-hydroxytamoxifen (4-OH-TAM) and 4-hydroxy-Ndesmethyltamoxifen (endoxifen), which show much greater affinity for the estrogen receptor than tamoxifen (36,37); furthermore, 4-OH-TAM and endoxifen are metabolized by SULT1A1 to form highly reactive products, leading to DNA adducts (38,39). There is substantial evidence that functional polymorphisms (including SNPs and gene copy number variations) of CYP2D6 and SULT1A1 are associated with tamoxifen pharmacokinetics and clinical outcomes of efficacy and adverse effects (40)(41)(42)(43)(44).…”

Impact of CYP2D6, CYP3A5, CYP2C9 and CYP2C19 polymorphisms on tamoxifen pharmacokinetics in Asian breast cancer patients

“…In view of the greater prevalence of CYP2D6*5 (CYP2D6del) and *10 (100C>T; rs1065852) among Asian populations, these polymorphisms may be more important in influencing the pharmacokinetics of tamoxifen in Asian breast cancer patients. Apart from CYP2D6, other cytochrome P450 enzymes (CYP3A4/5, CYP2C9 and CYP2C19) also contribute to the overall metabolism of tamoxifen and its metabolites, albeit to different extents [4,[20][21][22][23].…”

Drug‐Metabolizing Enzymes and Transporters: Expression in the Human Prostate and Roles in Prostate Drug Disposition

“…Studies in nonhuman systems have strongly implicated CYP3A and CYP2C as targets for inhibition by tamoxifen metabolites (Reidy and Murray, 1989); similar inhibitory processes with human CYPs may give rise to clinically relevant drug interactions. There is a major role for CYP3A4 in the ␣-hydroxylation (genotoxic product) and N-demethylation (inactive product) of tamoxifen; CYP2D6 and CYP1A also contribute to N-demethylation, whereas 4-hydroxylation to the active product primarily involves CYP2D6 and CYP2B6 (Boocock et al, 2002;Crewe et al, 2002). Factors that influence CYP3A, CYP2B6, and CYP1A activity (including induction and inhibition) and CYP2D6 activity (such as genetic polymorphism and enzyme inhibition) greatly affect tamoxifen action.…”

“…Tamoxifen and toremifene are oxidized by CYPs to pharmacologically active and inactive metabolites and also to reactive products with genotoxic potential (Boocock et al, Huang et al, 2000;Murray et al, 1994 Long andDolan, 2001;Reid et al, 1999Cresteil et al, 2002Shou et al, 1998Takara et al, 2002Relling et al, 1994Kawashiro et al, 1998 Flavopiridol Ifosfamide 2002; Crewe et al, 2002). Several CYPs seem to be involved in these pathways, with the oxidation of tamoxifen as the prototypic agent in this class being particularly well studied; biotransformation of toremifene has been less well investigated (Kim et al, 2003).…”