Effects of biologically active metabolites of tamoxifen on the proliferation kinetics of MCF-7 human breast cancer cells in vitro.

The effects of two major metabolites of tamoxifen, N-demethyltamoxifen (DMT) and 4-hydroxytamoxifen (4OHT), on MCF-7 cell proliferation and cell cycle kinetic parameters were compared with those of the parent compound. All three compounds produced dose-dependent decreases in the rate of cell proliferation which were accompanied by decreases in the percentage of S- and G2-M-phase cells. 4OHT was 100- to 167-fold more potent than both tamoxifen and DMT in producing these effects, and this was correlated with their relative binding affinities (RBAs) for the cytoplasmic estrogen receptor (ER) (17 beta-estradiol = 100, 4OHT = 41, tamoxifen = DMT = 2). At doses less than or equal to 2.5 microM, these effects were completely reversed by 17 beta-estradiol, but the required 17 beta-estradiol:antiestrogen concentration ratios differed, i.e., 1:10 to 1:1 for 4OHT compared with 1:1000 to 1:100 for tamoxifen and DMT. Although the concentrations of 17 beta-estradiol required for reversal were related to affinity of the metabolite for ER, they were 5- to 20-fold lower than predicted from the measured RBAs. When the rate of cell proliferation was measured over a range of concentrations of antiestrogen, in the presence or absence of 17 beta-estradiol, it was highly correlated (r2 = 0.96) with the percentage of S-phase cells. In addition to these 17 beta-estradiol-reversible events, all three compounds caused 17 beta-estradiol-irreversible cytotoxicity at higher concentrations (greater than or equal to 7.5 microM DMT and 4OHT, 10 microM tamoxifen). The order of potency in producing this effect was DMT greater than 4OHT greater than tamoxifen, which correlated with neither the RBAs for ER nor the RBAs for the high-affinity microsomal antiestrogen binding site. These data support the concept that estrogens and antiestrogens compete for a common event which regulates the rate of cell proliferation probably by controlling the proportion of cells entering S phase. Although it appears likely that ER is intimately involved in this regulatory process, 17 beta-estradiol-irreversible mechanisms are also involved in antiestrogen action in vitro.