Sa1771 Role of DNA Methylation on Epithelial Mesenchymal Transition (EMT)-Like Phenotype of Colon Cancer Cells

Abstract

Background: Epigenetic regulation of gene expression plays a key role in cancer biology. It is well known that gene-specific hypermethylation and genomic global hypomethylation is related to progression of cancer, and the methylation status is regulated by DNA methyltransferase (DNMT). DNMT1 is not only responsible for the maintenance of global DNA methylation pattern during cell proliferation, DNMT1 is also necessary for aberrant CpG island methylation in human cancer cells. Several researches demonstrate the therapeutic potentials of genetic or pharmacologic inhibition of DNMT1 by reactivating silenced tumor-suppressor genes. On the other hand, some literatures show that decreasing DNMT1 levels or activity can potentially enhance invasiveness of cancer cells, indicating that the effect of DNMT1 inhibition is cell context-specific. Aim: We evaluated the impact of DNMT1 inhibition or demethylation drug on epithelial mesenchymal transition (EMT)-like phenotype, including the determination of key molecule on the induction of EMT-like phenotype. Materials and Methods: Wild type and DNMT1 knock-out (KO) colon cancer HCT-116 cells were used in this study. Alternatively, small interfering RNA (siRNA), and 5-aza-2'-deoxycytidine (5aza) was used to silence DNMT1 expression in HCT116 cells. The resulting cell lines were validated by reverse transcriptase-polymerase chain reaction and Western blotting. Proliferation, migration and invasion assays were done in engineered cells to evaluate the effect of DNMT1 silencing on cellular phenotype. Result: Our data shows that DNMT1 KO and 5-aza treatment on HCT-116 cells resulted in EMT-like gene expression change. Genes specific to epithelial cells, such as E-cadherin, were down regulated, and genes related to mesenchymal markers, such as vimentin, were up regulated. Those engineered cells also exhibit higher cellular motility and acquired anoikis resistance. Those EMT-like phenotype were not correlated to the expression of Snail or Twist, but had propensity for the involvement of Zeb1 expression. Conclusion: Inhibiting DNMT1 on cancer cells, at least in some particular context, result in acquisition of EMT-like property. Our data indicates the necessity of selecting situation or patient when applying DNMT inhibition as treatment sensitizing agents.