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The ERα signaling pathway is one of the most important and most studied pathways in human breast cancer, yet numerous questions still exist such as how hormonally responsive cancers progress to a more aggressive and hormonally independent phenotype. We have noted that human breast cancers exhibit a strong direct correlation between ERα and E-cadherin expression by immunohistochemistry, suggesting that ERα signaling might regulate E-cadherin and implying that this regulation might influence epithelial–mesenchymal transition (EMT) and tumor progression. To investigate this hypothesis and the mechanisms behind it, we studied the effects of ERα signaling in ERα-transfected ERα-negative breast carcinoma cell lines, the MDA-MB-468 and the MDA-MB-231 and the effects of ERα knockdown in naturally expressing ERα-positive lines, MCF-7 and T47D. When ERα was overexpressed in the ERα-negative lines, 17β-estradiol (E2) decreased slug and increased E-cadherin. Clones maximally exhibiting these changes grew more in clumps and became less invasive in Matrigel. When ERα was knocked down in the ERα-positive lines, slug increased, E-cadherin decreased, cells became spindly and exhibited increased Matrigel invasion. ERα signaling decreased slug expression by two different mechanisms: directly, by repression of slug transcription by the formation of a corepressor complex of ligand-activated ERα, HDAC inhibitor (HDAC1), and nuclear receptor corepressor (N-CoR) that bound the slug promoter in three half-site estrogen response elements (EREs); indirectly by phosphorylation and inactivation of GSK-3β through phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt). The GSK-3β inactivation, in turn, repressed slug expression and increased E-cadherin. In human breast cancer cases, there was a strong inverse correlation between slug and ERα and E-cadherin immunoreactivity. Our findings indicate that ERα signaling through slug regulates E-cadherin and EMT. [ABSTRACT FROM AUTHOR]

Estrogen receptor-α (ERα) and its ligand estradiol (E2) has critical roles in breast cancer growth and are key therapeutic targets. In this study, we report a novel dual role of the adenosine A1 receptor (Adora1) as an E2/ERα target and a regulator of ERα transcriptional activity. In ERα-positive breast cancer cells, E2 upregulated Adora1 messenger RNA (mRNA) and protein levels, an effect that was reversed by the E2 antagonist ICI 182 780. Small interference RNA ablation of Adora1 in ERα-positive cells reduced basal and E2-dependent proliferation, whereas Adora1 over-expression in an ERα-negative cell line induced proliferation. The selective Adora1 antagonist, DPCPX, reduced proliferation, establishing Adora1 as a mediator of E2/ERα-dependent breast cancer growth. Intriguingly, Adora1 ablation decreased both mRNA and protein levels of ERα and, consequently, estrogen-responsive element-dependent ERα transcriptional activity. Moreover, Adora1 ablation decreased binding activity of ERα to the promoter of its target gene TFF1 and led to reduced TFF1 promoter activity and mRNA levels, suggesting that Adora1 is required for full transcriptional activity of ERα on E2 stimulation. Taken together, we showed a short feed-forward loop involving E2, ERα and Adora1 that favors breast cancer growth. These data suggest that Adora1 may represent an important target for therapeutic intervention in hormone-dependent breast cancer. [ABSTRACT FROM AUTHOR]

Approximately 80% of breast cancers express the estrogen receptor-α (ERα) and are treated with anti-estrogens. Resistance to these agents is a major cause of mortality. We have shown that estrogen inhibits Notch, whereas anti-estrogens or estrogen withdrawal activate Notch signaling. Combined inhibition of Notch and estrogen signaling has synergistic effects in ERα-positive breast cancer models. However, the mechanisms whereby Notch-1 promotes the growth of ERα-positive breast cancer cells are unknown. Here, we demonstrate that Notch-1 increases the transcription of ERα-responsive genes in the presence or absence of estrogen via a novel chromatin crosstalk mechanism. Our data support a model in which Notch-1 can activate the transcription of ERα-target genes via IKKα-dependent cooperative chromatin recruitment of Notch–CSL–MAML1 transcriptional complexes (NTC) and ERα, which promotes the recruitment of p300. CSL binding elements frequently occur in close proximity to estrogen-responsive elements (EREs) in the human and mouse genomes. Our observations suggest that a hitherto unknown Notch-1/ERα chromatin crosstalk mediates Notch signaling effects in ERα-positive breast cancer cells and contributes to regulate the transcriptional functions of ERα itself. [ABSTRACT FROM AUTHOR]

The breast cancer susceptibility gene 1 (BRCA1) is mutated in approximately 50% of hereditary breast cancers, and its expression is decreased in 30–40% of sporadic breast cancers, suggesting a general role in breast cancer development. BRCA1 physically and functionally interacts with estrogen receptor-α (ERα) and several transcriptional regulators. We investigated the relationship between cellular BRCA1 levels and tamoxifen sensitivity. Decreasing BRCA1 expression in breast cancer cells by small interfering RNA alleviated tamoxifen-mediated growth inhibition and abolished tamoxifen suppression of several endogenous ER-targeted genes. ER-stimulated transcription and cytoplasmic signaling was increased without detectable changes in ER or ER coregulator expression. Co-immunoprecipitation studies showed that with BRCA1 knockdown, tamoxifen-bound ERα was inappropriately associated with coactivators, and not effectively with corepressors. Chromatin immunoprecipitation studies demonstrated that with tamoxifen, BRCA1 knockdown did not change ERα promoter occupancy, but resulted in increased coactivator and decreased corepressor recruitment onto the endogenous cyclin D1 promoter. Our results suggest that decreased BRCA1 levels modify ERα-mediated transcription and regulation of cell proliferation in part by altering ERα–coregulator association. In the presence of tamoxifen, decreased BRCA1 expression results in increased coactivator and decreased corepressor recruitment on ER-regulated gene promoters.Oncogene (2009) 28, 575–586; doi:10.1038/onc.2008.405; published online 10 November 2008 [ABSTRACT FROM AUTHOR]

We have previously shown that increased and deregulated estrogen receptor α expression in the mammary gland leads to the development of proliferative disease and cancer. To address the importance of cyclin D1 in ERα-mediated mammary tumorigenesis, we crossed ERα-overexpressing mice with cyclin D1 knockout mice. Mammary gland morphogenesis was completely interrupted in the ERα-overexpressing cyclin D1-deficient triple transgenic mice. In addition to a highly significant reduction in mammary epithelial cell proliferation, cyclin E was upregulated resulting in DNA damage checkpoint activation and apoptosis. This imbalance between proliferative and apoptotic rates in conjunction with remarkable structural defects and cellular disorganization in the terminal end buds interrupted ductal morphogenesis. Interestingly, the structure of the mammary fat pad was fundamentally altered by the consequences of overexpressing ERα in the epithelial cells in the absence of cyclin D1 illustrating how alterations in the epithelial compartment can impact surrounding stromal composition. Transplantation of embryonic ERα-overexpressing and cyclin D1-deficient mammary epithelium into the cleared fat pad of wild-type mice did not rescue the aberrant mammary gland phenotype indicating that it was intrinsic to the mammary epithelial cells. In conclusion, although cyclin D1 is not essential for proliferation of normal mammary epithelial cells, ERα-overexpressing cells are absolutely dependent on cyclin D1 for proliferation. This differential requirement for cyclin D1 in normal vs abnormal mammary epithelial cells supports the application of cyclin D1 inhibitors as therapeutic interventions in ERα-overexpressing breast cancers.Oncogene (2008) 27, 3186–3193; doi:10.1038/sj.onc.1210974; published online 10 December 2007 [ABSTRACT FROM AUTHOR]

BRCA1 can regulate estrogen receptor-α (ERα) activity. This study tested the hypotheses that Brca1 loss in mammary epithelium alters the estrogenic growth response and that exposure to increased estrogen or ERα collaborates with Brca1 deficiency to accelerate preneoplasia and cancer development. Longer ductal extension was found in mammary glands of Brca1f/f;MMTV-Cre mice during puberty as compared to wild-type mice. Terminal end bud differentiation was impaired in Brca1 mutant mice with preservation of prolactin-induced alveolar differentiation. Exogenous estrogen stimulated an abnormal sustained increase in mammary epithelial cell proliferation and the appearance of ERα-negative preneoplasia in postpubertal Brca1 mutant mice. Carcinogenesis was investigated using Brca1f/f;MMTV-Cre mice hemizygous for p53. Exogenous estrogen increased the percentage of mice with multiple hyperplastic alveolar nodules. Targeted conditional ERα overexpression in mammary epithelial cells of mice that were Brca1 mutant and hemizygous for p53 increased the percentage of mice exhibiting multiple hyperplastic nodules, invasive mammary cancers and cancer multiplicity. Significantly more than half of the preneoplasia and cancers were ERα negative even as their initiation was promoted by ERα overexpression.Oncogene (2008) 27, 794–802; doi:10.1038/sj.onc.1210674; published online 23 July 2007 [ABSTRACT FROM AUTHOR]

Estrogen and its receptor alpha (ERα) have been implicated in the tissue-specific tumorigenesis associated with BRCA1 mutations. However, the majority of breast cancers developed in human BRCA1 mutation carriers are ERα-negative, challenging the link between BRCA1 and estrogen/ERα in breast cancer formation. Using a mouse model lacking the full-length form of BRCA1, here we show that ERα is highly expressed in the premalignant mammary gland and initiation stages of tumorigenesis, although its expression is gradually diminished during mammary tumor progression. We demonstrate that the absence of full-length BRCA1 increases sensitivity of cells to estrogen-induced extracellular signal-regulated kinase 1/2 phosphorylation and cyclin D1 expression. The absence of BRCA1 turns the proliferation of ERα-positive cells from a paracrine fashion to an autocrine or endocrine fashion. Consequently, BRCA1-mutant cells are sensitized to estrogen-induced cell proliferation in vitro and mammary tumorigenesis in vivo. These findings illustrate a molecular mechanism for estrogen/ERα signals in BRCA1-associated tissue-specific tumor formation, and identify several key elements in the estrogen/ERα-signaling cascade that can serve as potential therapeutic targets for BRCA1-associated tumorigenesis.Oncogene (2007) 26, 7204–7212; doi:10.1038/sj.onc.1210527; published online 14 May 2007 [ABSTRACT FROM AUTHOR]

Expression microarray analysis identified over 930 genes regulated during puberty in the mouse mammary gland. Most prominent were genes whose expression increased in parallel with pubertal development and remained high thereafter. Members of the Wnt, transforming growth factor-β and oestrogen-signalling pathways were significantly overrepresented. Comparison to expression data from CITED1 knockout mice identified a subset of oestrogen-responsive genes displaying altered expression in the absence of CITED1. Included in this subset are stanniocalcin2 (Stc2) and amphiregulin (Areg). Chromatin immunoprecipitation revealed that ERα binds to oestrogen response elements in both the Stc2 and Areg genes in the mammary gland during puberty. Additionally, CITED1 and ERα localize to the same epithelial cells of the pubertal mammary gland, supporting a role for interaction of these two proteins during normal development. In a human breast cancer data set, expression of Stc2, Areg and CITED1 parallel that of ERα. Similar to ERα, CITED1 expression correlates with good outcome in breast cancer, implying that potential maintenance of the ERα–CITED1 co-regulated signalling pathway in breast tumours can indicate good prognosis.Oncogene (2007) 26, 6406–6419; doi:10.1038/sj.onc.1210468; published online 7 May 2007 [ABSTRACT FROM AUTHOR]

Tumor hypoxia is associated with cancer invasiveness, metastasis and treatment failure. Recent data suggest that the major target for endocrine treatment in breast cancer, ERα, is downregulated during hypoxia, but the mechanism behind this remains unknown. MAPK signaling as well as ERα regulation has earlier been independently linked to hypoxia and we now demonstrate HIF-1α and ERK1/2-activation in vivo towards the necrotic zone in DCIS of the breast, parallel with ERα downregulation. Hypoxia further caused transcriptional downregulation of ERα via activation of ERK1/2 in cell lines and, importantly, MEK1/2 inhibitors (U0126 or PD184352) or ERK1/2 suppression by siRNA partially restored the ERα expression. U0126 combined with tamoxifen accordingly produced an increased efficacy of the anti-estrogens during hypoxia. Based on these findings, we suggest a promising novel therapy for ERα-positive breast cancer where a combination of endocrine treatment and ERK1/2 inhibitors may increase treatment response by improved targeting of dormant hypoxic tumor cells.Oncogene (2005) 24, 6835–6842. doi:10.1038/sj.onc.1208830; published online 27 June 2005 [ABSTRACT FROM AUTHOR]