Abstract P1-07-01: Histone deacetylase inhibitor entinostat reverses epithelial to mesenchymal transition of breast cancer cells by reversing the repression of E-cadherin

Abstract

Abstract Loss of ERa in breast cancer correlates with poor prognosis, increased recurrence rates and higher incidence of metastasis. In our previous studies, we have shown that histone deacetylase (HDAC) inhibitor entinostat (ENT) can upregulate ERα and aromatase in ER-negative cells and tumors, making them sensitive to aromatase inhibitors (AIs). In the current study, we are showing that ENT can also reverse epithelial to mesenchymal transition (EMT), which is considered to be a first step in the process of metastases formation. EMT is characterized by loss of intracellular adhesion (loss of E-cadherin); loss of epithelial markers such as cytokeratins and upregulation of mesenchymal markers such as vimentin; acquisition of fibroblast-like spindle morphology and increased motility. Various carcinomas undergo varying degrees of EMT and capacity to undergo EMT correlates inversely with levels of E-Cadherin. It is widely accepted that loss of E-cadherin is associated with more invasive phenotype. Epigenetic silencing of E-cadherin has been implicated in metastatic cell lines and invasive breast cancers. Triple negative breast cancer cells such as MDA-MB-231 and Hs578T show a basal phenotype characterized by loss of E-cadherin expression and higher expression of mesenchymal markers such as N-cadherin, vimentin along with transcriptional repressors such as twist and snail. In this study, we measured the effect of entinostat on the EMT. When MDA-MB-231 and Hs578T cells were treated with ENT, E-cadherin transcription was increased along with reduction in N-cadherin mRNA expression. Similar results were also seen in tumors of MDA-MB-231 and Hs578T xenografts treated with ENT (for 5 weeks and 2 weeks respectively). A dose dependent increase in E-cadherin was seen along with a dose dependent decrease in N-cadherin mRNA. Although, we did not observe any reduction in vimentin protein, phosphorylation of vimentin was increased and vimentin remodeling was changed as seen by immunofluorescence. We performed chromatin immunoprecipitation (ChIP) assay to measure the activation of E-cadherin promoter. Treatment of MDA-MB-231 and Hs578T cells increased the activation of E-cadherin promoter as seen by increased acetyl histones at the promoter region of E-cadherin. Twist and snail are known repressors of E-cadherin gene and we saw that ENT treatment reduced the association of twist and snail with the E-cadherin promoter. ENT was also able to downregulate twist, which may be responsible for reduced twist association with the E-cadherin promoter. In summary, these findings suggest that HDAC inhibitor ENT can reverse EMT and may help reduce the formation of metastasis. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P1-07-01.