Abstract 3765: Induction of amineoxidases as a possible mechanism of action of tamoxifen metabolites

To elucidate the mechanism of action of endoxifen in breast cancer, we determined its effects on estradiol-mediated growth of ERalpha-positive MCF-7 cells. Endoxifen is the active metabolite of tamoxifen; the most widely used selective estrogen receptor modulator for anti-estrogen therapy for women with estrogen receptor alpha-positive breast cancer. Clinical trials recognized that tamoxifen was a prodrug and got converted to active metabolites in the human body. Earlier studies recognized 4-hydroxytamoxifen (4HT) as the active metabolite of tamoxifen, although recent studies showed that the concentration of endoxifen was up to 10-fold higher than that of 4HT in patients treated with tamoxifen. These metabolites are formed by the action of cytochrome P450 2D6 (CYP2D6). We used quantitative PCR analysis to examine the effects of endoxifen on estrogenic stimulation of early response genes, c-myc, c-fos and Tff1. Since the natural polyamines, putrescine, spermidine and spermine, play important roles in estrogenic function, we also measured the activities of enzymes in the polyamine biosynthetic/metabolic pathway: ornithine decarboxylase (ODC), S-adenosyl-L-methionine decarboxylase (AdoMetDC), spermidine/spermine N1-acetyltransferase (SSAT), spermine oxidase (SMO), and acetylpolyamine oxidase (APAO). Polyamine levels were quantified by high performance liquid chromatography. Our results showed that estradiol increased the proliferation of MCF-7 cells by 2- to 3-fold and endoxifen suppressed the effects of estradiol on cell growth. The expression of c-myc, c-fos and Tff1 genes was significantly increased by estradiol, while the addition of endoxifen suppressed estradiol-mediated increase in gene expression. Estradiol increased the activity of ODC and AdoMetDC, whereas endoxifen suppressed the activity of these enzymes. Endoxifen had no effect on SSAT activity, whereas it significantly increased activities of amineoxidases, SMO and APAO. Estradiol increased putrescine and spermidine levels and endoxifen inhibited this increase. There was no significant effect on spermine levels. These data suggest that endoxifen exerts its growth inhibitory effects on MCF-7 cells by suppressing the expression of estradiol-stimulated oncogenes and polyamine biosynthetic pathway. The antiestrogenic effect of endoxifen is also manifested through the stimulation of polyamine oxidase enzymes, SMO and APAO. These results indicate for the first time that the upregulation of SMO and APA