Reduced genotoxicity of [D₅-ethyl]-tamoxifen implicates α-hydroxylation of the ethyl group as a major pathway of tamoxifen activation to a liver carcinogen

Abstract

A proposed mechanism for the metabolic activation of tamoxifen to electrophilic species that form DNA adducts leading to liver cancer involves alpha-hydroxylation of the ethyl group in the critical first step. This mechanism predicts that tamoxifen deuterated at the alpha-position would be less genotoxic than the non-deuterated compound owing to an isotope effect that would reduce the rate of oxidative metabolism at this position. This hypothesis has now been tested with experiments conducted in rats in vivo and in human cells in vitro. It was found that the deuterated compound [D5-ethyl]-tamoxifen is significantly less genotoxic than tamoxifen. Administration of 0.06 and 0.12 mmol/kg [D5-ethyl]-tamoxifen to female Fischer rats by gavage resulted in a 2.5- and 1.7-fold reduction respectively in the levels of hepatic DNA adducts present 24 h after treatment, compared with the non-deuterated compound. Treatment of MCL-5 cells with [D5-ethyl]-tamoxifen resulted in a 2- to 3-fold decrease, compared with tamoxifen, in the number of micronuclei induced in cells arrested in cytokinesis by cytochalasin-B. Further evidence is provided by UV irradiation of the major hepatic DNA adducts isolated from tamoxifen-treated rats, which caused a large shift in the chromatographic mobility of the adducts, consistent with the presence of a 1,2-olefinic linkage in the tamoxifen residue of the adduct leading to cyclization to a phenanthrenylic compound, and consistent with this adduct having arisen from reaction with DNA at the alpha-position of tamoxifen. Taken together, these data strongly suggest that tamoxifen is metabolized to a liver carcinogen by alpha-hydroxylation of its ethyl group.

The Use of Deuterium Isotope Effects to Probe the Active Site Properties, Mechanism of Cytochrome P450-Catalyzed Reactions, and Mechanisms of Metabolically Dependent Toxicity

“…Genotoxicity of the widely used antitumor drug, tamoxifen, was decreased 2-to 3-fold in vivo in rats and in vitro in a MCL-5 human cell line that retains cytochrome P450 activity by deuterium substitution for hydrogen in the allylic ethyl group (Table 3) (Phillips et al, 1994). These and other results suggest that liver carcinogenicity in rats caused by tamoxifen involves allylic ␣-carbon oxidation that may generate a reactive quinone methide.…”

New insights into the metabolism of tamoxifen and its role in the treatment and prevention of breast cancer

“…Although many candidates were described [75][76][77][78], the metabolite found to be responsible for the initiation of rat liver carcinogenesis is α-hydroxytamoxifen [79][80][81][82][83] (Figure 5) Alpha-hydroxytamoxifen has been resolved into R-(+) and S-(−) enantiomers. Metabolism by rat liver microsomes gave equal amounts of the two forms, but in hepatocytes the R form gave 8x the level of DNA adducts as the S form.…”

The gene expression of hepatic proteins responsible for DNA repair and cell proliferation in tamoxifen‐induced hepatocarcinogenesis

“…[14][15][16][17][18][19][20][21] Although N-desmethyltamoxifen, tamoxifen N-oxide, and 4-hydroxytamoxifen are major metabolites of tamoxifen, DNA adducts of these metabolites are hardly detected in the liver, 19,22) suggesting that these metabolites might be considered primarily as detoxification forms.…”

“…[8][9][10][11][12][13] α-(N 2 -Deoxyguanosinyl)tamoxifen is detected as a major DNA adduct of tamoxifen in the rat liver, and the adduct has been shown to be formed through metabolic activations, α-hydroxylation and O-sulfation, of tamoxifen. [14][15][16][17][18][19][20][21] Although N-desmethyltamoxifen, tamoxifen N-oxide, and 4-hydroxytamoxifen are major metabolites of tamoxifen, DNA adducts of these metabolites are hardly detected in the liver, 19,22) suggesting that these metabolites might be considered primarily as detoxification forms.…”