miR-200 affects tamoxifen resistance in breast cancer cells through regulation of MYB

AbstractResistance to tamoxifen is a major clinical challenge. Research in recent years has identified epigenetic changes as mediated by dysregulated miRNAs that can possibly play a role in resistance to tamoxifen in breast cancer patients expressing estrogen receptor (ER). We report here elevated levels of EMT markers (vimentin and ZEB1/2) and reduced levels of EMT-regulating miR-200 (miR-200b and miR-200c) in ER-positive breast cancer cells, MCF-7, that were resistant to tamoxifen, in contrast with the naïve parental MCF-7 cells that were sensitive to tamoxifen. Further, we established regulation of c-MYB by miR-200 in our experimental model. C-MYB was up-regulated in tamoxifen resistant cells and its silencing significantly decreased resistance to tamoxifen and the EMT markers. Forced over-expression of miR-200b/c reduced c-MYB whereas reduced expression of miR-200b/c resulted in increased c-MYB We further confirmed the results in other ER-positive breast cancer cells T47D cells where forced over-expression of c-MYB resulted in induction of EMT and significantly increased resistance to tamoxifen. Thus, we identify a novel mechanism of tamoxifen resistance in breast tumor microenvironment that involves miR-200-MYB signaling.

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Up-Regulation of Glioma-Associated Oncogene Homolog 1 Expression by Serum Starvation Promotes Cell Survival in ER-Positive Breast Cancer Cells

Cellular Physiology and Biochemistry ◽

10.1159/000430156 ◽

2015 ◽

Vol 36 (5) ◽

pp. 1862-1876 ◽

Cited By ~ 3

Author(s):

Juan Xu ◽

Gaoxiang Huang ◽

Zongjing Zhang ◽

Jieying Zhao ◽

Mingzhuo Zhang ◽

...

Keyword(s):

Breast Cancer ◽

Cell Survival ◽

Cancer Cells ◽

Microarray Analysis ◽

Breast Cancer Cells ◽

Serum Starvation ◽

Over Expression ◽

Er Positive ◽

Mcf 7 ◽

Positive Breast Cancer

Background/Aims: Cancer cells are resistant to ischemia and starvation. Glioma-associated oncogene homolog 1 (Gli1) is a positive transcriptional activator of Hedgehog (Hh) pathway and plays an essential role in the development of cancers, including breast cancer. However, how Gli1 promotes cell survival remains elusive. The main purpose of this study is to investigate the pro-survival effect of Gli1 under serum starvation and its molecular mechanism in ER-positive breast cancer cells. Methods: Gene expression was determined by quantitative real-time PCR (QRT-PCR) and Western blot. The survival of Gli1 stably transfected ER-positive breast cancer cell lines (Gli1-MCF-7 and Gli1-T47D cells) and their untransfected control cells was estimated by WST-8 assay. Microarray analysis was performed to screen downstream Hh/Gli1 target genes in Gli1-overexpressed MCF-7 cells. Transcriptional activities of NF-kappaB were measured by luciferase assays. ChIP analysis was performed to explore whether cIAP2 was a direct target gene of Gli1. Results: Serum starvation significantly up-regulated the expression of Gli1 gene through activating PI3K/AKT pathway. Over-expression of Gli1 markedly promoted cell survival under serum starvation. Microarray analysis revealed that 338 genes were differentially expressed in Gli1-MCF-7 cells compared with those in the control cells. Among these genes, cellular inhibitor of apoptosis 2 (cIAP2), coding an anti-apoptosis and pro-survival protein, was significantly up-regulated not only by Hh/Gli1 pathway, but also by serum starvation. However, ChIP assay revealed no binding of Gli1 to cIAP2 promoter at the region of -1792 to -1568bp. Moreover, over-expression of Gli1 resulted in enhanced trans-activation of transcriptional factor NF-κB. Suppression of NF-κB signaling with NF-κB inhibitor Bay11-7082, significantly reduced the expression of cIAP2 and the cell survival under serum starvation. Conclusion: Serum starvation significantly up-regulated the expression of Gli1, which in turn increased its key target cIAP2 expression and enhanced NF-κB/cIAP2 pathway, resulting in promoting cell survival under serum starvation. These findings may provide new insights into the pro-survival mechanisms of Gli1 in breast cancer.

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miR-449a Suppresses Tamoxifen Resistance in Human Breast Cancer Cells by Targeting ADAM22

Cellular Physiology and Biochemistry ◽

10.1159/000493964 ◽

2018 ◽

Vol 50 (1) ◽

pp. 136-149 ◽

Cited By ~ 13

Author(s):

Jun Li ◽

Mingjie Lu ◽

Jiao Jin ◽

Xiyi Lu ◽

Tongpeng Xu ◽

...

Keyword(s):

Breast Cancer ◽

Cancer Cells ◽

Breast Cancer Cells ◽

Tamoxifen Resistance ◽

Luciferase Assay ◽

Protein Protein Interaction ◽

Er Positive ◽

Tamoxifen Resistant ◽

Mcf 7 ◽

Positive Breast Cancer

Background/Aims: Most of estrogen receptor positive breast cancer patients respond well initially to endocrine therapies, but often develop resistance during treatment with selective estrogen receptor modulators (SERMs) such as tamoxifen. Altered expression and functions of microRNAs (miRNAs) have been reportedly associated with tamoxifen resistance. Thus, it is necessary to further elucidate the function and mechanism of miRNAs in tamoxifen resistance. Methods: Tamoxifen sensitivity was validated by using Cell Counting Kit-8 in tamoxifen-sensitive breast cancer cells (MCF-7, T47D) and tamoxifen-resistant cells (MCF-7/TAM, T47D/ TAM). Quantitative real-time polymerase chain reaction (qRT-PCR) was performed to detect the expression level of miR-449a in tamoxifen-sensitive/-resistant cells and patient serums. Dual-luciferase assay was used to identify the binding of miR-449a and predicted gene ADAM22. The expression level of ADAM22 was determined by qRT-PCR and western blotting in miR-449a +/- breast cancer cells. Subsequently, rescue experiments were carried out to identify the function of ADAM22 in miR-449a-reduced tamoxifen resistance. Finally, Gene ontology (GO) and Protein-protein interaction analyses were performed to evaluate the potential mechanisms of ADAM22 in regulating tamoxifen resistance. Results: MiR-449a levels were downregulated significantly in tamoxifen-resistant breast cancer cells when compared with their parental cells, as well as in clinical breast cancer serum samples. Overexpression of miR-449a re-sensitized the tamoxifen-resistant breast cancer cells, while inhibition of miR-449a conferred tamoxifen resistance in parental cells. Luciferase assay identified ADAM22 as a direct target gene of miR-449a. Additionally, silencing of ADAM22 could reverse tamoxifen resistance induced by miR-449a inhibition in ER-positive breast cancer cells. GO analysis results showed ADAM22 was mainly enriched in the biological processes of cell adhesion, cell differentiation, gliogenesis and so on. Protein-protein interaction analyses appeared that ADAM22 might regulate tamoxifen resistance through PPARG, LGI1, KRAS and LYN. Conclusion: Decreased miR-449a causes the upregulation of ADAM22, which induces tamoxifen resistance of breast cancer cells. These results suggest that miR-449a, functioning by targeting ADAM22, contributes to the mechanisms underlying breast cancer endocrine resistance, which may provide a potential therapeutic strategy in ER-positive breast cancers.

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Inducible upregulation of oestrogen receptor-β1 affects oestrogen and tamoxifen responsiveness in MCF7 human breast cancer cells

Journal of Molecular Endocrinology ◽

10.1677/jme.1.01688 ◽

2005 ◽

Vol 34 (2) ◽

pp. 553-566 ◽

Cited By ~ 48

Author(s):

L C Murphy ◽

B Peng ◽

A Lewis ◽

J R Davie ◽

E Leygue ◽

...

Keyword(s):

Breast Cancer ◽

Cancer Cells ◽

Oestrogen Receptor ◽

Breast Cancer Cells ◽

Breast Cancer Patients ◽

Growth Responses ◽

Over Expression ◽

Er Positive ◽

Positive Breast Cancer ◽

Tamoxifen Sensitivity

To investigate the effect of altered oestrogen receptor (ER)α and ERβ expression on oestrogen and anti-oestrogen action in breast cancer, we have stably expressed an inducible ERβ1 in MCF7 breast cancer cells. Stably expressing clones were isolated and over-expression of ERβ1 correlated with increased levels of specific radiolabelled oestradiol (E2) binding. Increased ERβ1 did not affect endogenous levels of ERα but increased progesterone receptor (PR) levels. Over-expression of ERβ1 reduced growth responses to E2 in contrast to little if any effect of over-expression of ERα. In oestrogen-replete conditions, over-expression of ERβ1 but not ERα reduced proliferation. Over-expression of ERβ1 did not result in anti-oestrogen resistance but was associated with increased sensitivity to 4-hydroxytamoxifen. Our results suggested that over-expression of ERβ1 in the presence of an endogenously expressed ERα was associated with tamoxifen sensitivity but may negatively modulate ERα-mediated growth. However, not all ERα activities were inhibited since endogenous PR expression was increased by both ERα and ERβ1 over-expression. These data paralleled those seen in some in vivo studies showing a relationship between PR and ERβ expression as well as ERβ expression and tamoxifen sensitivity of ER-positive breast cancer patients. These models are relevant and will be useful for dissecting the role of ERβ1 expression in ER-positive breast cancer.

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Abstract 4002: Adipose tissue supernatant of overweight ER positive breast cancer patients promotes migration of MCF-7 breast cancer cells

10.1158/1538-7445.am2020-4002 ◽

2020 ◽

Author(s):

Jie Ma ◽

Hetty Timmer-Bosscha ◽

Paul M. Werker ◽

Carolina P. Schröder ◽

Marlous Arjaans

Keyword(s):

Breast Cancer ◽

Adipose Tissue ◽

Cancer Cells ◽

Cancer Patients ◽

Breast Cancer Cells ◽

Breast Cancer Patients ◽

Er Positive ◽

Mcf 7 ◽

Positive Breast Cancer

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SGK3 Is an Estrogen-Inducible Kinase Promoting Estrogen-Mediated Survival of Breast Cancer Cells

Molecular Endocrinology ◽

10.1210/me.2010-0294 ◽

2011 ◽

Vol 25 (1) ◽

pp. 72-82 ◽

Cited By ~ 35

Author(s):

Yuanzhong Wang ◽

Dujin Zhou ◽

Sheryl Phung ◽

Selma Masri ◽

David Smith ◽

...

Keyword(s):

Breast Cancer ◽

Protein Kinase ◽

Cell Survival ◽

Cancer Cells ◽

Breast Cancer Cells ◽

Dependent Manner ◽

Binding Regions ◽

Er Positive ◽

Mcf 7 ◽

Positive Breast Cancer

Serum- and glucocorticoid-inducible kinase 3 (SGK3) is a protein kinase of the AGC family of protein kinase A, protein kinase G, and protein kinase C and functions downstream of phosphatidylinositol 3-kinase (PI3K). Recent study revealed that SGK3 plays a pivotal role in Akt/protein kinase B independent signaling downstream of oncogenic PI3KCA mutations in breast cancer. Here we report that SGK3 is an estrogen receptor (ER) transcriptional target and promotes estrogen-mediated cell survival of ER-positive breast cancer cells. Through a meta-analysis on 22 microarray studies of breast cancer in the Oncomine database, we found that the expression of SGK3 is significantly higher (5.7-fold, P < 0.001) in ER-positive tumors than in ER-negative tumors. In ER-positive breast cancer cells, SGK3 expression was found to be induced by 17β-estradiol (E2) in a dose- and time-dependent manner, and the induction of SGK3 mRNA by E2 is independent of newly synthesized proteins. We identified two ERα-binding regions at the sgk3 locus through chromatin immunoprecipitation with massively parallel DNA sequencing. Promoter analysis revealed that ERα stimulates the activity of sgk3 promoters by interaction with these two ERα-binding regions on E2 treatment. Loss-of-function analysis indicated that SGK3 is required for E2-mediated cell survival of MCF-7 breast carcinoma cells. Moreover, overexpression of SGK3 could partially protect MCF-7 cells against apoptosis caused by antiestrogen ICI 182,780. Together, our study defines the molecular mechanism of regulation of SGK3 by estrogen/ER and provides a new link between the PI3K pathway and ER signaling as well as a new estrogen-mediated cell survival mechanism mediated by SGK3 in breast cancer cells.

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Abstract 2104: MicroRNA-101 promotes estrogen independent growth and confers tamoxifen resistance in ER positive breast cancer cells

10.1158/1538-7445.am10-2104 ◽

2010 ◽

Cited By ~ 1

Author(s):

Mohit Sachdeva ◽

Hailong Wu ◽

Peng Ru ◽

Larn Hwang ◽

Vuong Trieu ◽

...

Keyword(s):

Breast Cancer ◽

Cancer Cells ◽

Breast Cancer Cells ◽

Tamoxifen Resistance ◽

Er Positive ◽

Positive Breast Cancer

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Identification of estrogenically synthesized LRP16 as an ERα coactivator and its role in ERα-positive breast cancer

Journal of Clinical Oncology ◽

10.1200/jco.2006.24.18_suppl.10676 ◽

2006 ◽

Vol 24 (18_suppl) ◽

pp. 10676-10676

Author(s):

W. Han ◽

Y. Zhao ◽

Z. Wu ◽

Y. Mu ◽

L. Yu ◽

...

Keyword(s):

Breast Cancer ◽

Cancer Cells ◽

Northern Blot ◽

Breast Cancer Cells ◽

Tumor Diameter ◽

Er Positive ◽

Mcf 7 ◽

Positive Breast Cancer

10676 Background: Aberrant ERα activity is linked to genesis and malignant progression of breast cancer through direct target gene activation or repression. A complex network of coregulatory proteins is largely believed to determine the transcriptional activity of ERα. LRP16 was identified previously to be an estrogen (E2) responsive gene, but its function involving in conferring estrogen signalling pathway is not clear. Methods: Endogenous LRP16 expression in MCF-7 cells was stably suppressed by retrovirus-mediated small interference RNA (siRNA). The effects of LRP16 expression on E2-stimulated growth and invasive ability of MCF-7 cells were determined in vitro and in vivo assays. The effects of LRP16 expression on ERα transactivation were determined by luciferase assays. The interaction of LRP16 and ERα was examined by GST pull-down and coimmunopricipitation (CoIP) assays. Northern blot and Western blot were used to detect the mRNA and protein levels of ER target genes in LRP16-inhibited MCF-7 cells. The LRP16 expression levels in primary breast cancer were detected by Northern blot. Results: Fristly, LRP16 expression was characterized to be dependent on estrogen activities. Then, LRP16 was identified to be an estrogen-independent ERα cofactor in ER-positive breast cancer cells and demonstrate that LRP16 is an essential coactivator to ERα-mediated transactivation in an estrogen-dependent manner. Suppression of LRP16 expression in ER-positive breast cancer cells specifically inhibits the transcription of ER upregulated genes, results in the increase of E-cadherin expression through ER mediation. In vitro and in vivo data demonstrate that suppression of LRP16 inhibits the ability of estrogen-stimulated proliferation and invasiveness of ER-positive breast cancer cells. The pathological and clinical characteristics of human breast cancer includining ER/PR-positiveness, tumor diameter and the involvement of axillary lymphoid nodes were tightly linked with the LRP16 gene expression level. Conclusions: These results establish a mechanistic link between estrogen receptor status, its coactivator LRP16, and progression of ER-positive breast cancers, and may provide a novel antiestrogenic target for the therapy of ER positive breast cancer. No significant financial relationships to disclose.

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Noninvasive identification of lineage-specific circular RNA for ER-positive breast cancer.

Journal of Clinical Oncology ◽

10.1200/jco.2020.38.15_suppl.3543 ◽

2020 ◽

Vol 38 (15_suppl) ◽

pp. 3543-3543 ◽

Cited By ~ 1

Author(s):

Jason Brown ◽

Palak Shah ◽

Josh Vo ◽

Lanbo Xiao ◽

Yashar Niknafs ◽

...

Keyword(s):

Breast Cancer ◽

Cancer Cells ◽

Cancer Patients ◽

Breast Cancer Cells ◽

Circular Rna ◽

Breast Cancer Cell Lines ◽

Breast Cancer Patients ◽

Non Invasive ◽

Er Positive ◽

Positive Breast Cancer

3543 Background: Non-invasive testing in plasma using RNA biomarkers has been limited by exoribonuclease-mediated degradation of RNA. Circular RNA (circRNA) are covalently closed RNA structures that resist this degradation due to their circular structure. Therefore circRNA are more stable than their linear counterparts. CircRNA are formed by alternative backsplicing of the 3’ end of a downstream exon to the 5’ end of an upstream exon. Here, we propose a novel method for non-invasive identification of circRNA and demonstrate circularized forms of several lineage and cancer specific targets for estrogen receptor-positive breast cancer. Methods: Capture RNA sequencing on cancer tissue was previously performed to determine the relative expression of potential circRNA isoforms in breast cancer patients. These isoforms as well as those predicted by intron length were screened using a quantitative PCR-based assay on ER-positive breast cancer cells. RNA extracted from breast cancer cells are exposed to ribonuclease R to demonstrate stability of circRNA. CircRNA derived from targets with known universal expression are used as positive controls as well as for analysis on plasma. Results: We identify the circRNA isoforms with highest expression for five genes, including ESR1, that are differentially expressed in ER-positive breast cancer compared to other cancers and normal breast tissue. We determine that the circRNA corresponding to all five targets is specifically expressed in breast cancer cell lines with at least 1000-fold higher expression than in non-ER positive breast cancer cell lines. We demonstrate that the highest expressing circRNA isoforms are resistant to degradation by ribonuclease R, whereas corresponding linear mRNA is susceptible. We also demonstrate the presence and stability of positive control circRNA in plasma from patients without cancer. Conclusions: CircRNA are promising biomarkers for early non-invasive detection of cancer due to their stability in plasma. This assay reliably detects ER-positive breast cancer specific circRNA, and exoribonuclease resistance has been validated. Application of this diagnostic assay to plasma from breast cancer patients is underway.

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