PO-289 Dual inhibition of WNT/β-catenin signalling and histone deacetylation as a new strategy to eliminate breast cancer stem cells by augmentation of apoptosis

Abstract

IntroductionEpigenetic changes play a critical role in the regulation of cancer stem cell (CSC) properties and the development of drug resistance. Modulation of histone acetylation program is closely related to differentiation and apoptosis process. CSCs, a subset of tumour cells, are responsible for disease relapse because of an acquired resistance to apoptosis and the Wnt signalling which is associated with cell survival/self-renewal and differentiation, is re-activated in these cells. Therefore, in the present study, we focused on a possible cytotoxic/apoptotic effect of the combination of niclosamide (Wnt/β-catenin pathway inhibitor) and Valproic acid (VPA, histone deacetylase inhibitor) on breast CSCs.Material and methodsThe effect of niclosamide (1 µM, 24 hour pre-treatment) and VPA (0.63–5 mM) combination on the viability of MCF-7s cells (CSCs-enriched population) were demonstrated by the ATP assay. Acetylated histone H3 levels at selected doses for the combination were assessed by ELISA. Protein levels associated with the Wnt/β-catenin signalling pathway, EMT, and histone modifications were shown by western blotting. Cell death mode was investigated via Hoechst 33342/PI double staining, M30 ELISA, real-time PCR (gene levels associated with apoptosis and autophagy) and western blotting (protein levels associated with apoptosis, autophagy and ER stress).Results and discussionsWe found that combination therapy exhibited a marked decrease in cell viability by inducing extrinsic apoptosis along with the stronger Wnt inhibition and increased histone H3 acetylation in MCF-7s cells. Furthermore, it was found that mesenchymal markers (fibronectin, N-cadherin, and ZEB1) were decreased at 72 hour and cytokeratin 18 as an epithelial marker was re-expressed in which H3K9ac and H3K4me3 were also increased. In addition, ER stress and blockade of autophagic flux have also been shown to be involved in this process.ConclusionIn conclusion, the future success of this combination approach in targeting CSCs and converted CSCs to non-CSCs may hold significant promise for successful treatment of breast cancer.