Investigation of the Role of CpG Methylation in Epithelial Mesenchymal Transition Master Genes in a Chemoresistant Ovarian Cancer Cell line

Abstract

Ovarian cancer is one of the commonly diagnosed cancers among women. Chemoresistant is an essential reason for treatment failure and high mortality. Emerging evidence connects epithelial mesenchymal transition (EMT) like changes and acquisition of chemoresistance in cancer. DNA methylation influences cellular processes including EMT. Here, we investigate EMT like changes in cisplatin-resistant A2780 ovarian cancer cells (A2780cis), and we study the DNA methylation role in EMT master genes regulation. Cell viability assay was carried to test the sensitivity of A2780, and A2780cis human cancer cell lines to cisplatin compared to other cancer cell lines. Differential mRNA expression of EMT markers using qPCR was conducted to investigate EMT like changes. The role of CpGs methylation in gene expression regulation was investigated by 5-azacytidine (5-aza) treatment. DNA methylation changes in EMT genes were identified using Methylscreen assay between A2780 and A2780cis cells. A2780cis maintains its cisplatin tolerance ability and exhibits phenotypic changes congruent with EMT. Methylscreen assay and qPCR study revealed DNA hypermethylation in promoters of epithelial adhesion molecules CDH1 and EPCAM in A2780cis compared to the cisplatin-sensitive parental cells, these changes were concomitant with gene expression down-regulation. DNA hypomethylation associated with transcription up-regulation of the mesenchymal marker TWIST2 was observed in the resistant cells. Azacytidine treatment confirmed the DNA methylation role in the regulation of gene expression of CDH1, EPCAM and TWIST2 genes. A2780cis cell line undergoes EMT like changes, and EMT master genes are regulated by DNA methylation. A better perception of the molecular alterations which correlate with chemoresistance may lead to therapeutic benefits such as chemosensitivity restoration.