Alpha Positive Breast Cancer Cell Research Articles

DNA aptamer probes for detection of estrogen receptor α positive carcinomas

Estrogen receptor alpha (ERα) also known as NR3A1 (nuclear receptor subfamily 3, group A, member 1) is a ligand-activated transcription factor. It is an important biomarker for breast cancer metastasis. In the present study, we report a novel DNA aptamer candidate against estrogen receptor (ER) alpha structure. Theenriched aptamercandidate was obtained after 14 iterative cycles of invitroprotein-SELEX process. Isothermal calorimetry study suggests the nanomolar sensitivity of the candidate ER_Apt1 to its target protein. Fluorescence- and chemiluminescence-binding assays confirm the specificity of the candidate aptamer to ER alpha positive breast cancer cell line. Comparative analysis of ER_Apt1 to ER alpha monoclonal antibody wasalso performed to analyze the expression of ER alpha in various malignant cancer cell line. Cytochemical and immunohistochemistry assay indicates its potential use as a diagnostic agent against ERα positive carcinomas. The nucleotide aptamer sequences described in the present study can be used for the detection, treatment, prophylaxis and diagnosis of ERα-related disorder.

P037 Ki67 assessment using a 5-grade scale revealed high reproducibility for luminal type breast cancer

Goals: Local application of estradiol (E2) to treat vulvovaginal atrophy in postmenopausal breast cancer patients receiving aromatase inhibitors is known to elevate serum estradiol levels and thereby might counteract breast cancer therapy. Thus, vaginal application of estriol (E3) has been recommended for these patients. However, it is unclear to what extent E3 stimulates breast cancer cell growth. In this study, we examined the effect of E3 on growth and gene expression of two human breast cancer cell lines. Methods: We used an established in vitro cell culture assay and compared the effect of E2 and E3 on growth of the estrogen receptor alpha-positive breast cancer cell lines MCF-7 and T-47D testing a wide range of hormone concentrations of 10–10M. E3 effects on gene expression were examined by means of reporter gene assays, RT-qPCR and Western blot analysis. Results: E3 acted as a potent estrogen and exerted a mitogenic effect on T-47D and MCF-7 cells at concentrations of 10M (288 pg/ml) and higher. With regard to activation of an estrogen response element (ERE) in breast cancer cells, effects of E3 were visible at 10M. The same concentrations of E3 activated expression of the estrogen-responsive gene PR and of the proliferation genes cyclin A2, cyclin B1, Ki-67, c-myc and b-myb, providing molecular mechanisms underlying the observed growth increase. Conclusion: Like E2, low levels of E3 were able to trigger a robust estrogenic response in breast cancer cells. Thus, our data suggest caution regarding use of E3 by breast cancer survivors. Disclosure of Interest: No significant relationships.

Effects of estriol on growth, gene expression and estrogen response element activation in human breast cancer cell lines

ObjectiveLocal application of estradiol (E2) to treat vulvovaginal atrophy in postmenopausal breast cancer patients receiving aromatase inhibitors is known to elevate serum estradiol levels and thereby might counteract breast cancer therapy. Thus, vaginal application of estriol (E3) has been recommended for these patients. However, it is unclear to what extent E3 stimulates breast cancer cell growth. In this study, we examined the effect of E3 on growth and gene expression of two human breast cancer cell lines. MethodsWe used an established in vitro cell culture assay and compared the effect of E2 and E3 on growth of the estrogen receptor alpha-positive breast cancer cell lines MCF-7 and T-47D testing a wide range of hormone concentrations of 10−12–10−7M. E3 effects on gene expression were examined by means of reporter gene assays, RT-qPCR and Western blot analysis. ResultsE3 acted as a potent estrogen and exerted a mitogenic effect on T-47D and MCF-7 cells at concentrations of 10−9M (288pg/ml) and higher. With regard to activation of an estrogen response element (ERE) in breast cancer cells, effects of E3 were visible at 10−10M. The same concentrations of E3 activated expression of the estrogen-responsive gene PR and of the proliferation genes cyclin A2, cyclin B1, Ki-67, c-myc and b-myb, providing molecular mechanisms underlying the observed growth increase. ConclusionsLike E2, low levels of E3 were able to trigger a robust estrogenic response in breast cancer cells. Thus, our data suggest caution regarding use of E3 by breast cancer survivors.

Abstract 2405: Loss of TBK1 induces an epithelial-mesenchymal transition through upregulation of Slug in the breast cancer cell lines

Abstract Estrogen receptor alpha (ER alpha) is a key regulator of proliferation and differentiation in mammary epithelium through their well-documented effects on transcription and represents a crucial prognostic indicator and therapeutic target in breast cancer. Here, we show that loss of TBK1 in ER alpha positive breast cancer cells causes an epithelial-mesenchymal transition (EMT). We found that TBK1 is downregulated in ER alpha negative breast tumors by analyzing published microarray data sets, suggesting that TBK1 might regulate ER alpha expression. To investigate this hypothesis, we made the ER alpha positive breast cancer cell lines stably expressing TBK1-specific shRNA. Knockdown of TBK1 decreased E-cadhrein expression (epithelial marker) and increased N-cadherin, vimentin, fibronectin expression (mesenchymal marker). Additionally, loss of TBK1 in ER alpha positive breast cancer cell lines leads to the downregulation of ER alpha expression. Interestingly, ER alpha repression by knockdown of TBK1 induced the upregulation of transcriptional repressor Slug, a master regulator of EMT. Because ER alpha expression is suppressed by loss of TBK1, it is suggested that TBK1 plays a role in the inverse correlation between Slug and ER alpha. Our findings indicate that loss of TBK1 expression is associated in between ER alpha expression status and invasive growth of breast cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2405. doi:1538-7445.AM2012-2405

Melatonin, environmental light, and breast cancer

Although many factors have been suggested as causes for breast cancer, the increased incidence of the disease seen in women working in night shifts led to the hypothesis that the suppression of melatonin by light or melatonin deficiency plays a major role in cancer development. Studies on the 7,12-dimethylbenz[a]anthracene and N-methyl-N-nitrosourea experimental models of human breast cancer indicate that melatonin is effective in reducing cancer development. In vitro studies in MCF-7 human breast cancer cell line have shown that melatonin exerts its anticarcinogenic actions through a variety of mechanisms, and that it is most effective in estrogen receptor (ER) alpha-positive breast cancer cells. Melatonin suppresses ER gene, modulates several estrogen dependent regulatory proteins and pro-oncogenes, inhibits cell proliferation, and impairs the metastatic capacity of MCF-7 human breast cancer cells. The anticarcinogenic action on MCF-7 cells has been demonstrated at the physiological concentrations of melatonin attained at night, suggesting thereby that melatonin acts like an endogenous antiestrogen. Melatonin also decreases the formation of estrogens from androgens via aromatase inhibition. Circulating melatonin levels are abnormally low in ER-positive breast cancer patients thereby supporting the melatonin hypothesis for breast cancer in shift working women. It has been postulated that enhanced endogenous melatonin secretion is responsible for the beneficial effects of meditation as a form of psychosocial intervention that helps breast cancer patients.

Leptin interferes with the effects of the antiestrogen ICI 182,780 in MCF-7 breast cancer cells.

Obesity is a risk factor for breast cancer development in postmenopausal women and correlates with shorter disease-free and overall survival in breast cancer patients, regardless of menopausal status. Adipose tissue is a major source of leptin, a cytokine regulating energy balance and controlling different processes in peripheral tissues, including breast cancer cell growth. Here, we investigated whether leptin can counteract antitumorigenic activities of the antiestrogen ICI 182,780 in breast cancer cells. Mitogenic response to leptin and the effects of leptin on ICI 182,780-dependent growth inhibition were studied in MCF-7 estrogen receptor alpha-positive breast cancer cells. The expression of leptin receptor and the activation of signaling pathways were studied by Western immunoblotting. The interference of leptin with ICI 182,780-induced estrogen receptor alpha degradation was probed by Western immunoblotting, fluorescence microscopy, and pulse-chase experiments. Leptin effects on estrogen receptor alpha-dependent transcription in the presence and absence of ICI 182,780 were studied by luciferase reporter assays and chromatin immunoprecipitation. MCF-7 cells were found to express the leptin receptor and respond to leptin with cell growth and activation the signal transducers and activators of transcription 3, extracellular signal-regulated kinase-1/2, and Akt/GSK3/pRb pathways. The exposure of cells to 10 nmol/L ICI 182,780 blocked cell proliferation, induced rapid estrogen receptor alpha degradation, inhibited nuclear estrogen receptor alpha expression, and reduced estrogen receptor alpha-dependent transcription from estrogen response element-containing promoters. All of these effects of ICI 182,780 were significantly attenuated by simultaneous treatment of cells with 100 ng/mL leptin. Leptin interferes with the effects of ICI 182,780 on estrogen receptor alpha in breast cancer cells. Thus, high leptin levels in obese breast cancer patients might contribute to the development of antiestrogen resistance.

The Anti-estrogenic Effect of All-trans-retinoic Acid on the Breast Cancer Cell Line MCF-7 Is Dependent on HES-1 Expression

All-trans-retinoic acid has been shown to have an antiproliferative effect in the estrogen receptor alpha-positive breast cancer cell line MCF-7. The mechanism of this effect is not well understood. We have previously shown that 17beta-estradiol down-regulates the basic helix-loop-helix factor Hairy and Enhancer of Split homologue-1 in MCF-7 and T47D cells (Ström, A., Arai, N., Leers, J., and Gustafsson, J. A. (2000) Oncogene 19, 5951-5953) and that this down-regulation is essential for proliferation in response to 17beta-estradiol. Treatment of the same cells with all-trans-retinoic acid prevented 17beta-estradiol-mediated down-regulation of the factor. The antiproliferative effect of all-trans-retinoic acid correlated well with the prevention of Hairy and Enhancer of Split homologue-1 down-regulation. Increasing concentrations of all-trans-retinoic acid, in the range of 1-1000 nm, produced a dose-dependent inhibition of proliferation and prevented 17beta-estradiol-mediated down-regulation of Hairy and Enhancer of Split homologue-1. By using a receptor-specific ligand we were able to show that the retinoic acid receptor alpha is important for regulation of the Hairy and Enhancer of Split homologue-1. Expression of a dominant negative form of Hairy and Enhancer of Split homologue-1 in MCF-7 cells abolished the growth-inhibitory effect of all-trans-retinoic acid in these cells. This finding indicates that Hairy and Enhancer of Split homologue-1 is a mediator of the antiproliferative effect of all-trans-retinoic acid in estrogen receptor alpha-positive breast cancer cell lines.

Antitumor and toxic effects of two derivatives of podophyliotoxin (GP 4 and GP 7) as compared with etopostde (VP 16)

Furanodiene is a natural product isolated from Rhizoma curcumae, and exhibits broad-spectrum anticancer activities in vitro and in vivo. Our previous study proved that furanodiene could increase growth inhibition of steroidal agent in ER alpha-positive breast cancer cells, but whether furanodiene can influence ER status is not clear. In this study, we confirmed that furanodiene down-regulated the ER alpha protein expression level and inhibited E2-induced estrogen response element (ERE)-driven reporter plasmid activity in ER alpha-positive MCF-7 cells. Actually, ERoc-knockdown cells were more sensitive than ER alpha positive cells to furanodiene on the cytotoxicity effect. So the anti-cancer effects of furanodiene and non steroidal agent in breast cancer cells still requires further investigation. Our results showed that furanodiene exposure could enhance growth inhibitory effects of doxorubicin in ERoc-negative MDA-MB-231 cells and ER alpha-low expression 4T1 cells. However, furanodiene did not increase the cytotoxicity of doxorubicin in ER alpha-positive breast cancer cells, non-tumorigenic breast epithelial cells, macrophage cells, hepatocytes cells, pheochromocytoma cells and cardiac myoblasts cells. Furanodiene enhances the anti-cancer effects of doxorubicin in ER alpha-negative breast cancer cells through suppressing cell viability via inducing apoptosis in mitochondria-caspases-dependent and reactive oxygen species-independent manners. These results indicate that furanodiene may be a promising and safety natural agent for cancer adjuvant therapy in the future. (C) 2016 Published by Elsevier B.V.