17-DMAG has the Potentiated Anticancer Effects Against Hepatocellular Carcinoma Cells by Transfection of the Gene Encoding Hepatitis B Viral X Protein

ABSTRACT

BACKGROUND/AIMS: Hepatitis B viral protein X (HBx) is implicated in the pathogenesis of hepatocellular carcinoma (HCC) as well as the elevation of heat shock proteins (HSPs) after hepatitis B virus (HBV) infection. We thus investigated the anticancer effects of an HSP90 inhibitor 17-Dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG) in HBx-transfected hepatocellular carcinoma cells. METHODS: pcDNA-HBx was made by inserting the HBx gene derived from the HBV-infected patient into pcDNA3.1 using the restriction enzymes (XbaI/HindIII). HBx-expressing HepG2 cells were then generated by transfecting HepG2 cells with pcDNA containing HBx gene. To compare the anticancer effects of 17-DMAG between pcDNA-HBx transfected HepG2 cells and the control cells (pcDNA-transfected HepG2 cells), we performed various molecular studies, including Ez-cytox proliferation assay, Western blot analysis, and flow cytometry. RESULTS: 17-DMAG inhibited the proliferation of pcDNA-HBx transfected HepG2 cells better than control cells (P<0.05). After treating with a various concentration of 17-DMAG (50–1,000 nM), pcDNA-HBx transfected HepG2 cells exhibited higher expression of pro-apoptotic proteins (c-caspase-3, c-caspase-8, and c-caspase-9) than did control cells (P<0.05). pcDNA-HBx transfected HepG2 cells showed higher activities of caspase-3, caspase-8, and caspase-9 than did control cells (P<0.05). Finally, we found that the expression of pro-apoptotic proteins (PARP and c-caspase-3) was considerably decreased by the use of a caspase inhibitor suggesting that 17-DMAG induces the cell death of HepG2 cells caspase-dependently. CONCLUSIONS: Our study strongly suggests that 17-DMAG has antiviral effects against HBV as well as anticancer effects against HepG2 cells. Thus, the application of 17-DMAG appears to be particularly advantageous to the HCC patients related with HBV infection.