Investigation of role of CpG methylation in some epithelial mesenchymal transition gene in a chemoresistant ovarian cancer cell line

Abstract

Ovarian cancer is one of the lethal gynecologic cancers. Chemoresistance is an essential reason for treatment failure and high mortality. Emerging evidence connects epithelial-mesenchymal transition (EMT) like changes and acquisition of chemoresistance in cancers. Including EMT, DNA methylation influences cellular processes. Here, EMT-like changes were investigated in cisplatin-resistant A2780 ovarian cancer cells (A2780cis), wherein role of DNA methylation in some EMT genes regulations was studied. Cell viability assay was carried out to test the sensitivity of A2780, and A2780cis human cancer cell lines to cisplatin. Differential mRNA expression of EMT markers using qPCR was conducted to investigate EMT like changes. CpG methylation role 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. In order to evaluate if DNA methylation changes are causally underlying EMT, treatment with 5-aza followed by Cisplatin was done on A2780cis cells. Accordingly, morphological changes were studied under the microscope, whereas EMT marker’s gene expression changes were investigated using qPCR. In this respect, A2780cis cell line has maintained its cisplatin tolerance ability and exhibits phenotypic changes congruent with EMT. Methylscreen assay and qPCR study have 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 DNA methylation role in regulating gene expression of CDH1, EPCAM and TWIST2 genes. A2780cis cell line undergoes EMT like changes, and EMT genes are regulated by DNA methylation. To that end, a better understanding of the molecular alterations that correlate with chemoresistance may lead to therapeutic benefits such as chemosensitivity restoration.

Tumor-derived apoptotic extracellular vesicle-mediated intercellular communication promotes metastasis and stemness of lung adenocarcinoma

“…4 B). The aberrant reactivation of the EMT is a prerequisite for the acquisition of a malignant phenotype in tumor cells, including increased migratory and invasive capabilities, as well as resistance to therapeutic agents [ 29 , 30 ], and reactivation of the EMT also has a complicated relationship with cancer stem cells [ 31 ]. Furthermore, selected genes that have been implicated in EMT and stemness were selected to individually verify their expression patterns via real-time quantitative PCR ( Fig.…”

Epigallocatechin Gallate for the Treatment of Benign and Malignant Gynecological Diseases—Focus on Epigenetic Mechanisms

“…Therefore, it was suggested that DNMT inhibitors might become part of a chemosensitization strategy [ 240 ]. A recently published paper confirmed that EMT genes involved in the metastasis mechanism were regulated by DNA methylation in cisplatin-resistant ovarian cancer cells and, therefore, could be a target of epigenetic-based therapy [ 241 , 242 ]. A significant relationship was reported between promoter methylation of the RAS association domain family protein 1a gene (RASSF1A), one of the tumor suppressor genes, and the risk of ovarian cancer [ 243 ].…”

Drug resistance in ovarian cancer: from mechanism to clinical trial

“…For instance, hypermethylation of ABCB1 and demethylation of the ABCG2 promoter may affect therapeutic efficacy and lead to chemoresistance in ovarian carcinoma, effects attributed to upregulation of P-gp [159,160]. Abnormal methylation of genes involved in the PI3K-AKT, MAPK, and Wnt pathways and in EMT confers resistance on HGSOC cells [161][162][163]. In addition, loss of RAD51C promoter methylation and a low level of BRCA1 methylation have been verified to cause drug resistance.…”