Diverse effects of DNMT1 inhibition and MBD2 knockdown on gene expression in Hep3B and HepG2 cells

Abstract

Cancer cell DNA is globally hypomethylated, but has regions of hypermethylation. Such regional hypermethylation, mediated by methylated DNA binding (MBD) proteins, inhibits gene expression. Demethylation of DNA or inhibition of MBD proteins may prove beneficial in treating cancers. We are investigating the effects on gene expression of 5‐aza‐2′‐deoxycytidine (ADC), an inhibitor of DNA methyltransferase 1 (DNMT1), and siRNA directed against MBD protein 2 (MBD2) in the human liver cancer cell lines, Hep3B and HepG2. Cells are treated in vitro for 72h with 1 µM ADC in dimethylsulfoxide (DMSO) or with DMSO alone, or are transfected for 72h with siRNA directed against MBD2 or with scrambled siRNA. RNA is then isolated for microarray analysis (Affymetrix). ADC‐treated Hep3B and HepG2 cells exhibited increased expression (≥2‐fold) of 168 and 166 genes, respectively. Only 10 of these genes were the same in both cell lines. There was no overlap between the two cell lines in classes of genes that were induced by ADC. MBD2 siRNA induced in Hep3B cells increased expression (≥2‐fold) of 351 genes. Only 5 of these genes were also induced by ADC in Hep3B cells. Inhibition of DNMT1 by ADC and knockdown of MBD2 by siRNA lead to unique molecular responses in liver cancer cells. The ramifications of these findings with respect to therapies that target epigenetic factors remain to be determined. American Cancer Society