Comparison of the proliferative effects of estradiol and conjugated equine estrogens on human breast cancer cells and impact of continuous combined progestogen addition

Abstract

Objectives: So far, most epidemiological studies investigating breast cancer risk and hormone replacement therapy have been conducted with conjugated equine estrogens (CEE). Recent trials indicate that the addition of progestogens may increase breast cancer risk. In the present study, we compared the effects of the human estrogen 17β-estradiol (E2) with those of the main equine components of CEE, i.e. equilin (Eq) and 17α-dihydroequilin (Dheq) on the proliferation of human breast cancer cells. The proliferative effect of progestogen addition was also investigated. Materials and methods: The well-established human breast cancer cell line MCF-7 was used as an in vitro model. The proliferative effect of E2, Eq and Dheq was tested in the concentration range 0.01-10 nmol/l. The progestogens progesterone, medroxyprogesterone acetate (MPA) and norethisterone (NET) were continuously combined with 0.1 nmol/l estrogen at concentrations of 0.01 nmol/l, 1 nmol/l, 0.1 μmol/l and 10 μmol/l. Proliferation was measured after 7 days by the adenosine triphosphate (ATP) chemosensitivity test. Results: All three estrogens increased the proliferation of MCF-7 cells by between 40 and 180%. The most proliferatively potent estrogen was E2, followed by Eq and Dheq, which showed a slightly lower proliferative activity than E2. The addition of progesterone inhibited E2-induced proliferation by about 30%, but only at the high non-physiological concentration of 10 μmol/l. All three progestogens inhibited Eq-induced proliferation, although their effect tended to be low, with values between 5 and 40%. No progestogen reduced Dheq-induced proliferation by more than 20%. In contrast, MPA slightly increased the proliferation rate by about 5% at the high physiological concentration of 0.1 μmol/l when combined with Dheq. The same held true when MPA and NET were added at the high pharmacological concentration of 10 μmol/l, causing increases of about 10%. Conclusions: Our results indicate that equine estrogens have a proliferative action similar to that of 17β-estradiol. Continuous addition of progestogens does not result in any major reduction of proliferative potency. Some progestogens may even enhance the estrogen-induced proliferation of pre-existing breast cancer cells, particularly when combined with certain equine estrogens. However, in none of the tested circumstances do progestogens increase the proliferative effect of estradiol, and progesterone has no deleterious effect even at pharmacological levels, in contrast to progestogens.