Carcinogenicity of estrogens in human breast epithelial cells

Epidemiological and clinical evidences indicate that breast cancer risk is associated with prolonged ovarian function that results in elevated circula ting levels of steroid hormones. Principal among these is estrogen, which i s associated with two important risk factors, early onset of menarche and l ate menopause. However, up to now there is no direct experimental evidence that estrogens are responsible of the initiation of human breast cancer. We postulate that if estrogens are causative agents of this disease, they sho uld elicit in human breast epithelial cells (HBEC) genomic alterations simi lar to those exhibited by human breast cancers, such as DNA amplification a nd loss of genetic material representing tumor suppressor genes. These effe cts could result from binding of the hormone to its nuclear receptors (ER) or from its metabolic activation to reactive metabolites. This hypothesis w as tested by treating with the natural estrogen 17 beta -estradiol (E-2) an d the synthetic steroid diethylstilbestrol (DES) MCF-10F cells, a HBEC line that is negative for ER. Cells treated with the chemical carcinogen benzo (a) pyrene (BP) served as a positive control of cell transformation. BP-, E -2-, and DES-treated MCF-10F cells showed increases in survival efficiency and colony efficiency in agar methocel, and loss of ductulogenic capacity i n collagen gel. The largest colonies were formed by BP-treated cells, becom ing progressively smaller in DES- and E-2-treated cells. The loss of ductul ogenic capacity was maximal in BP-, and less prominent in E2- and DES-treat ed cells. Genomic analysis revealed that E-2- and DES-treated cells exhibit ed loss of heterozygosity in chromosomes 3 and 11, at 3p21, 3p21-21.2, 3p21 .1-14.2, and 3p14.2-14.1, and at 11q23.3 and 11q23.1-25 regions, respective ly. It is noteworthy that these loci are also affected in breast lesions, s uch as ductal hyperplasia, carcinoma in situ, and invasive carcinoma. Our d ata are the first ones to demonstrate that estrogens induce in HBEC phenoty pic changes indicative of cell transformation and that those changes are as sociated with significant genomic alterations that might unravel new pathwa ys in the initiation of breast cancer.