Estrogen Receptor Alpha Negative Breast Research Articles

TRIM3 facilitates estrogen signaling and modulates breast cancer cell progression

BackgroundBreast cancer is the most common cancer in women worldwide. More than 70% of breast cancers are estrogen receptor (ER) alpha positive. Compared with ER alpha-negative breast cancer, which is more aggressive and has a shorter survival time, ER alpha-positive breast cancer could benefit from endocrine therapy. Selective estrogen receptor modulators, such as tamoxifen, are widely used in endocrine therapy. Approximately half of ER alpha-positive breast cancer patients will eventually develop endocrine resistance, making it a major clinical challenge in therapy. Thus, decoding the throughput of estrogen signaling, including the control of ER alpha expression and stability, is critical for the improvement of breast cancer therapeutics.MethodsTRIM3 and ER alpha protein expression levels were measured by western blotting, while the mRNA levels of ER alpha target genes were measured by RT–PCR. A CCK-8 assay was used to measure cell viability. RNA sequencing data were analyzed by Ingenuity Pathway Analysis. Identification of ER alpha signaling activity was accomplished with luciferase assays, RT–PCR and western blotting. Protein stability assays and ubiquitin assays were used to detect ER alpha protein degradation. Ubiquitin-based immunoprecipitation assays were used to detect the specific ubiquitination modification on the ER alpha protein.ResultsIn our current study, we found that TRIM3, an E3 ligase, can promote ER alpha signaling activity and breast cancer progression. TRIM3 depletion inhibits breast cancer cell proliferation and migration, while unbiased RNA sequencing data indicated that TRIM3 is required for the activity of estrogen signaling on the -genome-wide scale. The immunoprecipitation assays indicated that TRIM3 associates with ER alpha and promotes its stability, possibly by inducing K63-linked polyubiquitination of ER alpha. In conclusion, our data implicate a nongenomic mechanism by which TRIM3 stabilizes the ER alpha protein to control ER alpha target gene expression linked to breast cancer progression.ConclusionOur study provides a novel posttranslational mechanism in estrogen signaling. Modulation of TRIM3 expression or function could be an interesting approach for breast cancer treatment.Graphical abstract2noEgdU1b5aSV4mMoG9UypVideo abstract

MicroRNA-497 downregulation contributes to cell proliferation, migration, and invasion of estrogen receptor alpha negative breast cancer by targeting estrogen-related receptor alpha

Metastasis has become the main challenge for treatment of estrogen receptor alpha (ERα) negative breast cancer. Here, we found a negative correlation between miR-497 and estrogen-related receptor alpha (ERRα), a nuclear receptor overexpressed in ERα negative breast cancer. Targeted inhibition of ERRα by si-RNA increased miR-497 expression while overexpression of ERRα inhibited miR-497 expression. Further investigation showed that miR-497 targeted ERRα by binding to the 3'UTR region of ERRα. Luciferase assay and ChIP assay confirmed that ERα directly regulated the transcription of miR-497, suggesting that loss of ERα lowered miR-497 level in ERα negative breast cancer. Further, overexpression of miR-497 not only inhibited ERRα expression but also reduced MIF level and MMP9 activity, which led to significant decreases in cell proliferation, migration, and invasion of ERα negative breast cancer. Taken together, our findings suggested that, in ERα negative breast cancer, the low level of ERα reduced miR-497 expression, which promoted ERRα expression that enhanced cell proliferation, migration, and invasion by increasing MIF expression and MMP9 activity.

Putative biomarkers and targets of estrogen receptor negative human breast cancer.

Breast cancer is a progressive and potentially fatal disease that affects women of all ages. Like all progressive diseases, early and reliable diagnosis is the key for successful treatment and annihilation. Biomarkers serve as indicators of pathological, physiological, or pharmacological processes. Her2/neu, CA15.3, estrogen receptor (ER), progesterone receptor (PR), and cytokeratins are biomarkers that have been approved by the Food and Drug Administration for disease diagnosis, prognosis, and therapy selection. The structural and functional complexity of protein biomarkers and the heterogeneity of the breast cancer pathology present challenges to the scientific community. Here we review estrogen receptor-related putative breast cancer biomarkers, including those of putative breast cancer stem cells, a minor population of estrogen receptor negative tumor cells that retain the stem cell property of self-renewal. We also review a few promising cytoskeleton targets for ER alpha negative breast cancer.

Detection of mRNA for Nuclear Receptor Co-Activators 1 and 3 in Archival Breast Cancer Tissue and Surrounding Stroma: A Tissue Expression Study~!2009-01-14~!2010-02-10~!2010-07-20~!

Before the age of 75 years, approximately 10% of women will be diagnosed with breast cancer, one of the most common malignancies and a leading cause of death among women. The objective of this study was to determine if expression of the nuclear receptor coactivators 1 and 3 (NCoA1 and NCoA3) varied in breast cancer grades. RNA was extracted from 25 breast tumours and transcribed into cDNA which underwent semi-quantitative polymerase chain reaction, normalised using 18S. Analysis indicated that an expression change for NCoA1 in cancer grades and estrogen receptor alpha negative tissue (P= 0.028 and 0.001 respectively). NCoA1 expression increased in grade 3 and estrogen receptor alpha negative tumours, compared to controls. NCoA3 showed a similar, but not significant, trend in grade and a non-significant decrease in estrogen receptor alpha negative tissues. Expression of NCoA1 in late stage and estrogen receptor alpha negative breast tumours may have implications to breast cancer treatment, particularly in the area of manipulation of hormone signalling systems in advanced tumours.

Induction of G1 phase arrest and apoptosis in MDA‐MB‐231 breast cancer cells by troglitazone, a synthetic peroxisome proliferator‐activated receptor γ (PPARγ) ligand

Peroxisome proliferator-activated receptor gamma (PPARgamma) ligands inhibit cell proliferation and induce apoptosis in cancer cells. Here we wished to determine whether the PPARgamma ligand induces apoptosis and cell cycle arrest of the MDA-MB-231 cell, an estrogen receptor alpha negative breast cancer cell line. The treatment of MDA-MB-231 cell with PPARgamma ligands was shown to induce inhibition of cell growth in a dose-dependent manner as determined by MTT assay. Cell cycle analysis showed a G1 arrest in MDA-MB-231 cells exposed to troglitazone. An apoptotic effect by troglitazone demonstrated that apoptotic cells elevated by 2.5-fold from the control level at 10 microM, to 3.1-fold at 50 microM and to 3.5-fold at 75 microM. Moreover, troglitazone treatment, applied in a dose-dependent manner, caused a marked decrease in pRb, cyclin D1, cyclin D2, cyclin D3, Cdk2, Cdk4 and Cdk6 expression as well as a significant increase in p21 and p27 expression. These results indicate that troglitazone causes growth inhibition, G1 arrest and apoptotic death of MDA-MB-231 cells.

A novel gene STYK1/NOK is upregulated in estrogen receptor-alpha negative estrogen receptor-beta positive breast cancer cells following estrogen treatment

The human STYK1/NOK protein is approximately 30-35% similar to mouse fibroblast growth factor receptor 3 and a kinase homologue in D. melanogaster in the tyrosine protein kinase region. STYK1/NOK was identified as being up regulated in MDA-MB-231, an estrogen receptor-alpha negative breast cancer cell line, following 12 h of estrogen treatment at 1x10(-9) M. On further investigation of STYK1/NOK in estrogen treated cell line MDA-MB-231, STYK1/NOK was up regulated at 6 h post treatment when compared to untreated cells. We also investigated the expression levels of STYK1/NOK in other breast cancer cell lines MCF-7, MDA-MB-231, BT-549, and MDA-MB-435S using QRT-PCR. In addition, the analysis of message accumulation was increased with other synthetic estrogen response modifiers. We propose that the regulation of STYK1/NOK is achieved independent of ERalpha and suggests further investigation to the relevance of this kinase in breast cancer progression.

The synergistic inhibitory effect of 5-aza-2-deoxycytidine and Tamoxifen on estrogen receptor alpha negative breast cancer cell lines in vitro

To observe if ER alpha gene can be induced by 5-aza-CdR in ER alpha negative human breast cancer cell lines (MDA-MB-231 and MDA-MB-435) and the synergistic inhibitory effects of 5-aza-CdR and Tamoxifen on these two cell lines in vitro. The status of 5'CpG island methylation of ER alpha gene in ER alpha negative (MDA-MB-231 and MDA-MB-435) human breast cancer cell lines and 20 cases of breast cancer tissue was studied by MSP, the expression of ER alpha mRNA was inspected by using RT-PCR after these two cell lines were treated with 5-aza-CdR. Cell proliferation was evaluated by MTT assay, distribution of cell cycle and rate of apoptosis were determined by flow cytometry after these two cell lines were treated with 5-aza-CdR or TAM alone, or in combination in vitro. The 5'CpG island is methylated in the core promotor of ER alpha gene in ER alpha negative (MDA-MB-231 and MDA-MB-435) human breast cancer cell lines and the methylating rate is 25.0%, 66.7%, 83.3%, 100% in 20 cases of breast cancer tissue of stage I, II, III, IV, respectively. The expression of ER alpha mRNA was induced in these two cell lines after treated with 5-aza-CdR, MTT array showed the proliferation activity of these two cell lines was obviously reduced in 5-aza-CdR group and the inhibitory effect on proliferation was enhanced when 5-aza-CdR combined with TAM compared with control group, the induction of apoptosis was 11.20% and 8.71% respectively by 5-aza-CdR, while the rate of apoptosis is 48.8% and 53.1% when these two cells were treated with 5-aza-CdR combined with TAM. 5-aza-CdR can re-express ER alpha by demethylating and sensitive ER alpha negative human breast cancer cell lines to TAM, 5-aza-CdR and TAM synergistically inhibit proliferation and induce apoptosis in ER alpha negative human breast cancer cell lines, thus offer a new way for the treatment of ER alpha negative breast cancer.

Study of the CpG methylation status of ER alpha gene in estrogen receptor alpha-negative breast cancer cell lines and the role of hydralazine demethylation

To detect the 5'CpG island methylation of estrogen receptor (ER) alpha gene promotor region in ER alpha-negative human breast cancer cell lines (MDA-MB-231 and MDA-MB-435) and breast cancer tissues; and to investigate the possibility of hydralazine in restoring the expression of ER alpha gene through demethylation. The CpG island methylation status of ER alpha gene promotor region-A, B and C in ER alpha-negative human breast cancer cell lines (MDA-MB-231 and MDA-MB-435) and 20 cases of breast cancer tissues were analyzed using methylation-specific polymerase chain reaction (MSP). The mRNA expression profile of ER alpha isoforms (ERalpha-A, ERalpha-B and ERalpha-C) and coding region in ER alpha gene were analyzed by using RT-PCR, after hydralazine treatment. In the two cell lines studied, CpG island was methylated in ERalpha-A and ERalpha-B but not ERalpha-C. Among them, methylation of CpG island in ERalpha-A was obtained in 13 breast tumor cases; methylation of ERalpha-B seen in 10 tumor cases; methylation of both ERalpha-A and ERalpha-B seen in 9 cases; and methylation of ERalpha-C was obtained in only 1 case. The positivity rates were 65%, 50%, 45% and 5% respectively. The ER alpha gene non-expression in breast cancer is probably associated with CpG island methylation in ER alpha gene promotor region A and B, and the level of methylation is enhanced as advance of tumors in clinical stage. Hydralazine, served as a demethylating agent enables to restore the expression of ER alpha gene.

The histone deacetylase inhibitor trichostatin A sensitizes estrogen receptor alpha-negative breast cancer cells to tamoxifen

The histone deacetylase inhibitor trichostatin A sensitizes estrogen receptor alpha-negative breast cancer cells to tamoxifen

The histone deacetylase inhibitor trichostatin A sensitizes estrogen receptor alpha-negative breast cancer cells to tamoxifen

The histone deacetylase inhibitor trichostatin A sensitizes estrogen receptor alpha-negative breast cancer cells to tamoxifen