Effects of 17 beta-estradiol metabolites on cell cycle events in MCF-7 cells.

Different cell growth effects were observed in MCF-7 cells after six daily exposures to either 17 beta-estradiol (E2), 2-hydroxyestradiol (2-OHE2), or 2-methoxyestradiol (2-MeOE2) at 10 nM levels. 2-OHE2 enhanced cell growth significantly (P < 0.05) more than did the parent compound, whereas 2-MeOE2 inhibited cell growth. To identify the estrogen-affected cellular processes involved in cell cycle progression, hydroxy urea-synchronized MCF-7 cells were studied. No effects on DNA synthesis in mid-S-phase or on mitotic indices were observed after E2 or 2-OHE2 treatment. 2-MeOE2, however, significantly (P < 0.05) inhibited DNA synthesis and mitosis. Synchronized cells were exposed for 1 h to E2, 2-OHE2, or 2-MeOE2 before cAMP levels were determined in early S-phase and mid-S-phase, as well as during mitosis. E2 and 2-OHE2 had no effect, but 2-MeOE2 caused a significant (P < 0.05) increase in cAMP concentration in early S-phase and a decrease during mitosis. Phosphorylation of S-phase proteins was also studied. [32P]Pi incorporation was significantly (P < 0.05) enhanced in many proteins in 2-MeOE2-exposed cells. Small proteins (M(r) < 25,000), as well as large proteins (M(r) > 220,000), were most prominently affected. In comparison, E2 and 2-OHE2 had little effect. We suggest that the enhanced 2-MeOE2-induced protein phosphorylation during S-phase may affect S-phase events, which subsequently causes inhibition of mitosis. Protein synthesis during G2/M transition was unexpectedly enhanced by 2-OHE2 and was not enhanced by E2. [35S]Methionine incorporation into proteins in the order of M(r) 32,000-46,000, 47,000-50,000, 58,000-67,000, and 83,000-89,000 was significantly (P < 0.05) increased. 2-MeOE2 had no effect. The results of this study indicate that 2-OHE2 may be the more potent mitogen, whereas 2-MeOE2 acts as a cytostatin.