Positive Regulation of Estrogen Receptor Alpha in Breast Tumorigenesis

Estrogen receptor alpha (ERα, NR3A1) contributes through its expression in different tissues to a spectrum of physiological processes, including reproductive system development and physiology, bone mass maintenance, as well as cardiovascular and central nervous system functions. It is also one of the main drivers of tumorigenesis in breast and uterine cancer and can be targeted by several types of hormonal therapies. ERα is expressed in a subset of luminal cells corresponding to less than 10% of normal mammary epithelial cells and in over 70% of breast tumors (ER+ tumors), but the basis for its selective expression in normal or cancer tissues remains incompletely understood. The mapping of alternative promoters and regulatory elements has delineated the complex genomic structure of the ESR1 gene and shed light on the mechanistic basis for the tissue-specific regulation of ESR1 expression. However, much remains to be uncovered to better understand how ESR1 expression is regulated in breast cancer. This review recapitulates the current body of knowledge on the structure of the ESR1 gene and the complex mechanisms controlling its expression in breast tumors. In particular, we discuss the impact of genetic alterations, chromatin modifications, and enhanced expression of other luminal transcription regulators on ESR1 expression in tumor cells.

GPR30 Activation by 17β-Estradiol Promotes p62 Phosphorylation and Increases Estrogen Receptor α Protein Expression by Inducing Its Release from a Complex Formed with KEAP1

Estrogens can elicit rapid cellular responses via the G-protein-coupled receptor 30 (GPR30), followed by estrogen receptor α (ERα/ESR1)-mediated genomic effects. Here, we investigated whether rapid estrogen signaling via GRP30 may affect ESR1 expression, and we examined the underlying molecular mechanisms. The exposure of human endometrial cancer cells to 17β-estradiol promoted p62 phosphorylation and increased ESR1 protein expression. However, both a GPR30 antagonist and GPR30 silencing abrogated this phenomenon. GPR30 activation by 17β-estradiol elicited the SRC/EGFR/PI3K/Akt/mTOR signaling pathway. Intriguingly, unphosphorylated p62 and ESR1 were found to form an intracellular complex with the substrate adaptor protein KEAP1. Upon phosphorylation, p62 promoted ESR1 release from the complex, to increase its protein expression. Given the critical role played by p62 in autophagy, we also examined how this process affected ESR1 expression. The activation of autophagy by everolimus decreased ESR1 by promoting p62 degradation, whereas autophagy inhibition with chloroquine increased ESR1 expression. The treatment of female C57BL/6 mice with the autophagy inhibitor hydroxychloroquine—which promotes p62 expression—increased both phosphorylated p62 and ESR1 expression in uterine epithelial cells. Collectively, our results indicate that 17β-estradiol-mediated GPR30 activation elicits the SRC/EGFR/PI3K/Akt/mTOR signaling pathway and promotes p62 phosphorylation. In turn, phosphorylated p62 increased ESR1 expression by inducing its release from complexes that included KEAP1. Our findings may lead to novel pharmacological strategies aimed at decreasing ESR1 expression in estrogen-sensitive cells.

Circ_0040039 May Aggravate Intervertebral Disk Degeneration by Regulating the MiR-874-3p-ESR1 Pathway

The functional alteration of nucleus pulposus cells (NPCs) exerts a crucial role in the occurrence and progression of intervertebral disk degeneration (IDD). Circular RNAs and microRNAs (miRs) are critical regulators of NPC metabolic processes such as growth and apoptosis. In this study, bioinformatics tools, encompassing Gene Ontology pathway and Venn diagrams analysis, and protein–protein interaction (PPI) network construction were used to identify functional molecules related to IDD. PPI network unveiled that ESR1 was one of the most critical genes in IDD. Then, a key IDD-related circ_0040039-miR-874-3p-ESR1 interaction network was predicted and constructed. Circ_0040039 promoted miR-874-3p and repressed ESR1 expression, and miR-874-3p repressed ESR1 expression in NPCs, suggesting ESR1 might be a direct target of miR-874-3p. Functionally, circ_0040039 could enhance NPC apoptosis and inhibit NPC growth, revealing that circ_0040039 might aggravate IDD by stabilizing miR-874-3p and further upregulating the miR-874-3p-ESR1 pathway. This signaling pathway might provide a novel therapeutic strategy and targets for the diagnosis and therapy of IDD-related diseases.

The gene encoding the estrogen receptor, ESR1, is differentially expressed in the primary tumors and metastases of patients with breast cancer based on survival outcomes and its expression correlates with survival.

Breast cancer is a leading cause of cancer death for women in the United States (1). Metastatic breast cancer is a major public health problem with a 5-year survival rate of 27% (1). We mined published microarray datasets to identify genes whose expression was most significantly different when comparing primary and metastatic tumors based on survival outcomes at 24 months (2, 3). In the primary tumors of patients with breast cancer, we identified the estrogen receptor alpha, ESR1 (also known as ER) as one of the most significantly differentially expressed genes when comparing tumor transcriptomes based on disease-free survival at 24 months. When comparing the transcriptomes of metastases from patients with stage IV metastatic breast cancer, we again found that ESR1 was among the genes whose expression was most significantly different based on overall survival at 24 months. In metastatic tissues, expression of ESR1 positively correlated with overall patient survival. Stratifying patients based on ESR1 expression revealed that higher ESR1 expression in both primary and metastatic tissues was generally associated with enhanced patient survival. Though established immunohistochemical classification of breast cancers groups tumors in a binary fashion based on ER expression at the protein level as ER+ and ER-, these data suggest that graded interpretation of estrogen receptor expression at the mRNA level, in primary tumors and in metastatic tissues, may be useful for prognostic purposes.

Unliganded Progesterone Receptor Governs Estrogen Receptor Gene Expression by Regulating DNA Methylation in Breast Cancer Cells

Breast cancer prognosis and response to endocrine therapy strongly depends on the expression of the estrogen and progesterone receptors (ER and PR, respectively). Although much is known about ERα gene (ESR1) regulation after hormonal stimulation, how it is regulated in hormone-free condition is not fully understood. We used ER-/PR-positive breast cancer cells to investigate the role of PR in ESR1 regulation in the absence of hormones. We show that PR binds to the low-methylated ESR1 promoter and maintains both gene expression and DNA methylation of the ESR1 locus in hormone-deprived breast cancer cells. Depletion of PR reduces ESR1 expression, with a concomitant increase in gene promoter methylation. The high amount of methylation in the ESR1 promoter of PR-depleted cells persists after the stable re-expression of PR and inhibits PR binding to this genomic region. As a consequence, the rescue of PR expression in PR-depleted cells is insufficient to restore ESR1 expression. Consistently, DNA methylation impedes PR binding to consensus progesterone responsive elements. These findings contribute to understanding the complex crosstalk between PR and ER and suggest that the analysis of ESR1 promoter methylation in breast cancer cells can help to design more appropriate targeted therapies for breast cancer patients.

Adipocytes ESR1 Expression, Body Fat and Response to Testosterone Therapy in Hypogonadal Men Vary According to Estradiol Levels

Estradiol (E2), mainly produced from Testosterone (T) in men, promotes visceral lipolysis. However, high visceral fat and hyperestrogenemia are features of obese hypogonadal (HG) men. Our study objectives are to evaluate relationships between circulating E2 and: (1) fat mass; (2) Estrogen Receptor α (ESR1) expression in subcutaneous adipose tissue; (3) changes in body fat after 6 months (M) of T therapy in HG men. Hypotheses: (1) existence of a range of circulating E2 associated with better body composition; (2) serum E2 determines tissue E2 sensitivity which affects response to T therapy. Men 40–74 years old, T < 300 (ng/dL), given T-cypionate for 6 months. Subjects were divided into 4-E2 categories: (1) <10.0; (2) 10.0–15.9; (3) 16.0–19.9; (4) ≥20.0 (pg/mL). Body composition (DXA), fat biopsies (liposuction), gene expression (qPCR), serum E2 and T (LC/MS), at baseline and 6 months. We enrolled 105 men; 90 completed the study. Group 2 had lower total and truncal fat mass (p < 0.01) but higher % lean mass (p < 0.001). ESR1 mRNA was the highest in group 1 (p = 0.01). At 6 months, group 1 had higher reduction in total (p = 0.03) and truncal (p = 0.01) fat. In conclusion, serum E2 = 10–15.9 (pg/mL) is associated with the best body composition profile in HG men; however, those with E2 < 10 (pg/mL) had the best response (greater fat loss) to T replacement possibly because of greater E2 sensitivity.

Coamplification of miR-4728 protects HER2-amplified breast cancers from targeted therapy

HER2 (ERBB2) amplification is a driving oncogenic event in breast cancer. Clinical trials have consistently shown the benefit of HER2 inhibitors (HER2i) in treating patients with both local and advanced HER2+ breast cancer. Despite this benefit, their efficacy as single agents is limited, unlike the robust responses to other receptor tyrosine kinase inhibitors like EGFR inhibitors in EGFR-mutant lung cancer. Interestingly, the lack of HER2i efficacy occurs despite sufficient intracellular signaling shutdown following HER2i treatment. Exploring possible intrinsic causes for this lack of response, we uncovered remarkably depressed levels of NOXA, an endogenous inhibitor of the antiapoptotic MCL-1, in HER2-amplified breast cancer. Upon investigation of the mechanism leading to low NOXA, we identified a micro-RNA encoded in an intron of HER2, termed miR-4728, that targets the mRNA of the Estrogen Receptor α (ESR1). Reduced ESR1 expression in turn prevents ERα-mediated transcription of NOXA, mitigating apoptosis following treatment with the HER2i lapatinib. Importantly, resistance can be overcome with pharmacological inhibition of MCL-1. More generally, while many cancers like EGFR-mutant lung cancer are driven by activated kinases that when drugged lead to robust monotherapeutic responses, we demonstrate that the efficacy of targeted therapies directed against oncogenes active through focal amplification may be mitigated by coamplified genes.

Abnormal extracellular matrix remodelling in the cervix of pregnant relaxin-deficient mice is not associated with reduced matrix metalloproteinase expression or activity

Relaxin regulates cervical extracellular matrix (ECM) remodelling during pregnancy by modifying collagen and other ECM molecules by unknown mechanisms. We hypothesised that abnormal collagen remodelling in the cervix of pregnant relaxin-deficient (Rln1−/−) mice is due to excessive collagen (Col1a1 and Col3a1) and decreased matrix metalloproteinases (Mmp2, Mmp9, Mmp13 and Mmp7) and oestrogen receptors (Esr1 and Esr2). Quantitative polymerase chain reaction, gelatinase zymography, MMP activity assays and histological staining evaluated changes in ECM in pregnant wildtype (Rln1+/+) and Rln1−/− mice. Cervical Col1a1, Col3a1 and total collagen increased in Rln1−/− mice and were higher at term compared with Rln1+/+ mice. This was not correlated with a decrease in gelatinase (Mmp2, Mmp9) expression or activity, Mmp7 or Mmp13 expression, which were all significantly higher in Rln1−/− mice. In late pregnancy, circulating MMP2 and MMP9 were unchanged. Esr1 expression was highest in Rln1+/+ and Rln1−/− mice in late pregnancy, coinciding with a decrease in Esr2 in Rln1+/+ but not Rln1−/− mice. The relaxin receptor (Rxfp1) decreased slightly in late-pregnant Rln1+/+ mice, but was significantly higher in Rln1−/− mice. In summary, relaxin deficiency results in increased cervical collagen in late pregnancy, which is not explained by a reduction in Mmp expression or activity or decreased Rxfp1. However, an imbalance between Esr1 and Esr2 may be involved.