The growth of both normal and transformed epithelial cells of the female reproductive system is stimulated by estrogens, mainly through the activation of estrogen receptor α (ERα), which is a ligand-regulated transcription factor. The selective ER modulator tamoxifen (TAM) has been widely used as an ER antagonist in breast tumor; however, long-term treatment is associated with an increased risk of endometrial cancer. To provide new insights into the potential mechanisms involved in the agonistic activity exerted by TAM in the uterus, we evaluated the potential of 4-hydroxytamoxifen (OHT), the active metabolite of TAM, to transactivate wild-type ERα and its splice variant expressed in Ishikawa and HEC1A endometrial tumor cells, respectively. OHT was able to antagonize only the activation of ERα by 17β-estradiol (E2) in Ishikawa cells, whereas it up-regulated c-fos expression in a rapid manner similar to E2 and independently of ERα in both cell lines. This stimulation occurred through the G protein-coupled receptor named GPR30 and required Src-related and epidermal growth factor receptor tyrosine kinase activities, along with the activation of both ERK1/2 and phosphatidylinositol 3-kinase/AKT pathways. Most importantly, OHT, like E2, stimulated the proliferation of Ishikawa as well as HEC1A cells. Transfecting a GPR30 antisense expression vector in both endometrial cancer cell lines, OHT was no longer able to induce growth effects, whereas the proliferative response to E2 was completely abrogated only in HEC1A cells. Furthermore, in the presence of the inhibitors of MAPK and phosphatidylinositol 3-kinase pathways, PD 98059 and wortmannin, respectively, E2 and OHT did not elicit growth stimulation. Our data demonstrate a new mode of action of E2 and OHT in endometrial cancer cells, contributing to a better understanding of the molecular mechanisms involved in their uterine agonistic activity.

The higher incidence of thyroid carcinoma (TC) in women during reproductive years compared with men and the increased risk associated with the therapeutic use of estrogens have suggested a pathogenetic role exerted by these steroids in the development of TC. In the present study, we evaluated the potential of 17β-estradiol (E2), genistein (G), and 4-hydroxyta-moxifen (OHT) to regulate the expression of diverse estrogen target genes and the proliferation of human WRO, FRO, and ARO thyroid carcinoma cells, which were used as a model system. We have ascertained that ARO cells are devoid of estrogen receptors (ERs), whereas both WRO and FRO cells express a single variant of ERα that was neither transactivated, modulated, nor translocated into the nucleus upon treatment with ligands. However, E2, G, and OHT were able either to induce the transcriptional activity of c-fos promoter constructs, including those lacking the estrogen-responsive elements, or to increase c-fos, cyclin A, and D1 expression. It is noteworthy that we have demonstrated that the G protein-coupled receptor 30 (GPR30) and the mitogen-activated protein kinase (MAPK) pathway mediate both the up-regulation of c-fos and the growth response to E2, G, and OHT in TC cells studied, because these stimulatory effects were prevented by silencing GPR30 and using the MEK inhibitor 2′-amino-3′-methoxyflavone (PD 98059). Our findings provide new insight into the molecular mechanisms through which estrogens may induce the progression of TC.

The prevalence of obesity has been steadily increasing over the past few decades in several developed and developing countries, with resultant hazardous health implications. Substantial epidemiological evidence has shown that excessive adiposity strongly influences risk, prognosis, and progression of various malignancies, including breast cancer. Indeed, it is now well recognized that obesity is a complex physiologic state associated with multiple molecular changes capable of modulating the behavior of breast tumor cells as well of the surrounding microenvironment. Particularly, insulin resistance, hyperactivation of insulin-like growth factor pathways, and increased levels of estrogen due to aromatization by the adipose tissue, inflammatory cytokines, and adipokines contribute to breast cancerogenesis. Among adipokines, leptin, whose circulating levels increase proportionally to total adipose tissue mass, has been identified as a key member of the molecular network in obesity. This review summarizes the current knowledge on the epidemiological link existing between obesity and breast cancer and outlines the molecular mechanisms underlying this connection. The multifaceted role of the obesity adipokine leptin in this respect is also discussed.

Purpose: The molecular mechanisms involved in the repressive effects exerted by estrogen receptors (ER) on peroxisome proliferator–activated receptor (PPAR) γ–mediated transcriptional activity remain to be elucidated. The aim of the present study was to provide new insight into the crosstalk between ERα and PPARγ pathways in breast cancer cells. Experimental Design: Using MCF7 and HeLa cells as model systems, we did transient transfections and electrophoretic mobility shift assay and chromatin immunoprecipitation studies to evaluate the ability of ERα to influence PPAR response element–mediated transcription. A possible direct interaction between ERα and PPARγ was ascertained by coimmunoprecipitation assay, whereas their modulatory role in the phosphatidylinositol 3-kinase (PI3K)/AKT pathway was evaluated by determining PI3K activity and AKT phosphorylation. As a biological counterpart, we investigated the growth response to the cognate ligands of both receptors in hormone-dependent MCF7 breast cancer cells. Results: Our data show for the first time that ERα binds to PPAR response element and represses its transactivation. Moreover, we have documented the physical and functional interactions of ERα and PPARγ, which also involve the p85 regulatory subunit of PI3K. Interestingly, ERα and PPARγ pathways have an opposite effect on the regulation of the PI3K/AKT transduction cascade, explaining, at least in part, the divergent response exerted by the cognate ligands 17β-estradiol and BRL49653 on MCF7 cell proliferation. Conclusion: ERα physically associates with PPARγ and functionally interferes with PPARγ signaling. This crosstalk could be taken into account in setting new pharmacologic strategies for breast cancer disease.

Obesity confers risks to cancer development and progression but the mechanisms underlying these risks remain unclear. In this study, we identify a role for the obesity cytokine leptin, which has been implicated previously in breast cancer development, as a determinant for the tumor-promoting activity of cancer-associated fibroblasts (CAF) in both wild-type (WT) and K303R mutant estrogen receptor-α (ERα)–expressing breast cancer cells. Human CAFs stimulated a greater increase in the proliferation and migration of breast cancer cells expressing the K303R-ERα hyperactive receptor than WT-ERα–expressing cells. A concomitant increase was seen in leptin receptor isoform expression and activation of the leptin signaling pathway in cells expressing K303R-ERα compared with WT-ERα, correlating with leptin effects on cell growth, motility, and invasiveness in mutant cells. Epidermal growth factor and other factors secreted by K303R-ERα cells stimulated CAF proliferation, migration, and subsequent leptin secretion. Moreover, K303R-ERα expression generated a leptin hypersensitive phenotype in vivo. Together, our results reveal a bidirectional cross-talk between breast cancer cells and “educated” CAFs that drives tumor progression via leptin signaling. In elucidating a mechanism that connects obesity and cancer, these findings reinforce the concept that blocking cancer–stromal cell communication may represent an effective strategy for targeted therapy of breast cancer. Cancer Res; 72(6); 1416–27. ©2012 AACR.