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Silencing MED1 sensitizes breast cancer cells to pure anti-estrogen fulvestrant in vitro and in vivo.

ABSTRACT: Pure anti-estrogen fulvestrant has been shown to be a promising ER antagonist for locally advanced and metastatic breast cancer. Unfortunately, a significant proportion of patients developed resistance to this type of endocrine therapy but the molecular mechanisms governing cellular responsiveness to this agent remain poorly understood. Here, we've reported that knockdown of estrogen receptor coactivator MED1 sensitized fulvestrant resistance breast cancer cells to fulvestrant treatment. We found that MED1 knockdown further promoted cell cycle arrest induced by fulvestrant. Using an orthotopic xenograft mouse model, we found that knockdown of MED1 significantly reduced tumor growth in mice. Importantly, knockdown of MED1 further potentiated tumor growth inhibition by fulvestrant. Mechanistic studies indicated that combination of fulvestrant treatment and MED1 knockdown is able to cooperatively inhibit the expression of ER target genes. Chromatin immunoprecipitation experiments further supported a role for MED1 in regulating the recruitment of RNA polymerase II and transcriptional corepressor HDAC1 on endogenous ER target gene promoter in the presence of fulvestrant. These results demonstrate a role for MED1 in mediating resistance to the pure anti-estrogen fulvestrant both in vitro and in vivo.

SUBMITTER: Zhang L

PROVIDER: S-EPMC3728322 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Silencing MED1 sensitizes breast cancer cells to pure anti-estrogen fulvestrant in vitro and in vivo.

Zhang Lijiang L Cui Jiajun J Leonard Marissa M Nephew Kenneth K Li Yongquan Y Zhang Xiaoting X

PloS one 20130730 7

Pure anti-estrogen fulvestrant has been shown to be a promising ER antagonist for locally advanced and metastatic breast cancer. Unfortunately, a significant proportion of patients developed resistance to this type of endocrine therapy but the molecular mechanisms governing cellular responsiveness to this agent remain poorly understood. Here, we've reported that knockdown of estrogen receptor coactivator MED1 sensitized fulvestrant resistance breast cancer cells to fulvestrant treatment. We foun ...[more]

PMID: 23936234

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Project description:We have previously demonstrated that endoxifen is the most important tamoxifen metabolite responsible for eliciting the anti-estrogenic effects of this drug in breast cancer cells expressing estrogen receptor-alpha (ER?). However, the relevance of ER? in mediating endoxifen action has yet to be explored. Here, we characterize the molecular actions of endoxifen in breast cancer cells expressing ER? and examine its effectiveness as an anti-estrogenic agent in these cell lines.MCF7, Hs578T and U2OS cells were stably transfected with full-length ER?. ER? protein stability, dimer formation with ER? and expression of known ER target genes were characterized following endoxifen exposure. The ability of various endoxifen concentrations to block estrogen-induced proliferation of MCF7 parental and ER?-expressing cells was determined. The global gene expression profiles of these two cell lines was monitored following estrogen and endoxifen exposure and biological pathway analysis of these data sets was conducted to identify altered cellular processes.Our data demonstrate that endoxifen stabilizes ER? protein, unlike its targeted degradation of ER?, and induces ER?/ER? heterodimerization in a concentration dependent manner. Endoxifen is also shown to be a more potent inhibitor of estrogen target genes when ER? is expressed. Additionally, low concentrations of endoxifen observed in tamoxifen treated patients with deficient CYP2D6 activity (20 to 40 nM) markedly inhibit estrogen-induced cell proliferation rates in the presence of ER?, whereas much higher endoxifen concentrations are needed when ER? is absent. Microarray analyses reveal substantial differences in the global gene expression profiles induced by endoxifen at low concentrations (40 nM) when comparing MCF7 cells which express ER? to those that do not. These profiles implicate pathways related to cell proliferation and apoptosis in mediating endoxifen effectiveness at these lower concentrations.Taken together, these data demonstrate that the presence of ER? enhances the sensitivity of breast cancer cells to the anti-estrogenic effects of endoxifen likely through the molecular actions of ER?/? heterodimers. These findings underscore the need to further elucidate the role of ER? in the biology and treatment of breast cancer and suggest that the importance of pharmacologic variation in endoxifen concentrations may differ according to ER? expression.

Project description:Endocrine therapies include aromatase inhibitors and the selective estrogen receptor (ER) downregulator fulvestrant. This study aimed to determine whether the reported efficacy of fulvestrant over anastrozole, and high- over low-dose fulvestrant, reflect distinct transcriptional responses.Global gene expression profiles from ER?-positive breast carcinomas before and during presurgical treatment with fulvestrant (n = 22) or anastrozole (n = 81), and corresponding in vitro models, were compared. Transcripts responding differently to fulvestrant and estrogen deprivation were identified and integrated using Gene Ontology, pathway and network analyses to evaluate their potential significance.The overall transcriptional response to fulvestrant and estrogen deprivation was correlated (r = 0.61 in presurgical studies, r = 0.87 in vitro), involving downregulation of estrogen-regulated and proliferation-associated genes. The transcriptional response to fulvestrant was of greater magnitude than estrogen deprivation (slope = 0.62 in presurgical studies, slope = 0.63 in vitro). Comparative analyses identified 28 genes and 40 Gene Ontology categories affected differentially by fulvestrant. Seventeen fulvestrant-specific genes, including CAV1/2, SNAI2, and NRP1, associated with ER?, androgen receptor (AR), and TP53, in a network regulating cell cycle, death, survival, and tumor morphology. Eighteen genes responding differently to fulvestrant specifically predicted antiproliferative response to fulvestrant, but not anastrozole. Transcriptional effects of low-dose fulvestrant correlated with high-dose treatment, but were of lower magnitude (ratio = 0.29).The transcriptional response to fulvestrant has much in common with estrogen deprivation, but is stronger with distinctions potentially attributable to arrest of estrogen-independent ER? activity and involvement of AR signaling. Genes responding differently to fulvestrant may have predictive utility. These data are consistent with the clinical efficacy of fulvestrant versus anastrozole and higher dosing regimens.

Project description:IntroductionWe have previously reported that induction of epidermal growth factor receptor and ErbB2 in response to antihormonal agents may provide an early mechanism to allow breast cancer cells to evade the growth-inhibitory action of such therapies and ultimately drive resistant cell growth. More recently, the other two members of the ErbB receptor family, ErbB3 and ErbB4, have been implicated in antihormone resistance in breast cancer. In the present study, we have investigated whether induction of ErbB3 and/or ErbB4 may provide an alternative resistance mechanism to antihormonal action in a panel of four oestrogen receptor (ER)-positive breast cancer cell lines.MethodsMCF-7, T47D, BT474 and MDAMB361 cell lines were exposed to fulvestrant (100 nM) for seven days, and effects on ErbB3/4 expression and signalling, as well as on cell growth, were assessed. Effects of heregulin ?1 (HRG?1) were also examined in the absence and presence of fulvestrant to determine the impact of ER blockade on the capacity of this ErbB3/4 ligand to promote signalling and cell proliferation.ResultsFulvestrant potently reduced ER expression and transcriptional activity and significantly inhibited growth in MCF-7, T47D, BT474 and MDAMB361 cells. However, alongside this inhibitory activity, fulvestrant also consistently induced protein expression and activity of ErbB3 in MCF-7 and T47D cells and ErbB4 in BT474 and MDAMB361 cell lines. Consequently, fulvestrant treatment sensitised all cell lines to the actions of the ErbB3/4 ligand HRG?1 with enhanced ErbB3/4-driven signalling activity, reexpression of cyclin D1 and significant increases in cell proliferation being observed when compared to untreated cells. Indeed, in T47D and MDAMB361 HRG?1 was converted from a ligand having negligible or suppressive growth activity into one that potently promoted cell proliferation. Consequently, fulvestrant-mediated growth inhibition was completely overridden by HRG?1 in all four cell lines.ConclusionsThese findings suggest that although antihormones such as fulvestrant may have potent acute growth-inhibitory activity in ER-positive breast cancer cells, their ability to induce and sensitise cells to growth factors may serve to reduce and ultimately limit their inhibitory activity.

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Silencing MED1 sensitizes breast cancer cells to pure anti-estrogen fulvestrant in vitro and in vivo.

Publications

Silencing MED1 sensitizes breast cancer cells to pure anti-estrogen fulvestrant in vitro and in vivo.

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