Influence of antiestrogens on EGF- and IGF-I-Mediated proliferation of human breast cancer cells

120 It has been reported that antiestrogens inhibit the growth of estrogen receptor (ER)-positive MCF-7 cell growth by antagonizing the . ll0 mitogenie effects of estradiol, and thus should not inhibit prolifera- ~ tion in estrogen-free media. However, it has been demonstrated that ~ t00 antiestrogens inhibit MCF-7 cell growth in the absence of estradiol, suggesting that they operate in part through mechanisms other than I estradiol antagonism at the ER (15). Further, the activity of certain .." growth factors on cell proliferation is influenced by both estrogens ! and antiestrogens (4,5). To more completely define the antiprotifer- i ative mechanisms of action of antiestrogens, the objective of the pres- ~' ent study was to compare the antiproliferative activities of three antiestrogens [tamoxifen (TAM), ICI-182,780 (ICI) and Analog (AII)] which differ in chemical structure and biological mechanism on insulin-like growth factor-1 (IGF-1) or epidermal growth factor (EGF)induced proliferation of MCF-7 human breast cancer cells under serum-free conditions. The MCF-7 cells were obtained from the Michigan Cancer Foundation and grown as nmnolayer cultures in RPMI 1640 media as previously described (8). After harvesting, cells were seeded into 96-well tissue culture plates at a concentration of 10 ~ cells/well in RPMI containing 5% calf serum and allowed to attach overnight. The medium was replaced with RPMI + 5% TCH (defined serum replacement, Celox Corporation, Hopkins, MN) 24 h before treatment. Experimental treatment with growth factor (10 -1° M), antiestrogen (10 -6 M), estradiol (10 9 M) or a combination was initiated on Day I and medium containing treatments was replaced on alternate days. These experiments were conducted with 10 -t° M EGF or IGF-I, based on results of previous dose-related observations 140(13). Cell proliferation was determined with the neutral red dye 13amethod as previously described (2). Cell proliferation was quanti- -~ tared on treatment Day 7 and reported as a percentage of control- ~ 120treated cells. For statistical purposes, triplicate samples of each t3 ll0treatment were included in each experiment and the combined resuhs of three experiments were viewed as a two-factor analysis of variance having independent observations within treatment combinations.