Identification of the Genes Chemosensitizing Hepatocellular Carcinoma Cells to Interferon-α/5-Fluorouracil and Their Clinical Significance

The incidence of advanced hepatocellular carcinoma (HCC) is increasing worldwide, and its prognosis is extremely poor. Interferon-alpha (IFN-alpha)/5-fluorouracil (5-FU) therapy is reportedly effective in some HCC patients. In the present study, to improve HCC prognosis, we identified the genes that are sensitizing to these agents. The screening strategy was dependent on the concentration of ribozymes that rendered HepG2 cells resistant to 5-FU by the repeated transfection of ribozymes into the cells. After 10 cycles of transfection, which was initiated by 5,902,875 sequences of a ribozyme library, three genes including protein kinase, adenosine monophosphate (AMP)-activated, gamma 2 non-catalytic subunit (PRKAG2); transforming growth factor-beta receptor II (TGFBR2); and exostosin 1 (EXT1) were identified as 5-FU-sensitizing genes. Adenovirus-mediated transfer of TGFBR2 and EXT1 enhanced IFN-alpha/5-FU-induced cytotoxicity as well as 5-FU, although the overexpression of these genes in the absence of IFN-alpha/5-FU did not induce cell death. This effect was also observed in a tumor xenograft model. The mechanisms of TGFBR2 and EXT1 include activation of the TGF-beta signal and induction of endoplasmic reticulum stress, resulting in apoptosis. In HCC patients treated with IFN-alpha/5-FU therapy, the PRKAG2 mRNA level in HCC tissues was positively correlated with survival period, suggesting that PRKAG2 enhances the effect of IFN-alpha/5-FU and serves as a prognostic marker for IFN-alpha/5-FU therapy. In conclusion, we identified three genes that chemosensitize the effects of 5-FU and IFN-alpha/5-FU on HCC cells and demonstrated that PRKAG2 mRNA can serve as a prognostic marker for IFN-alpha/5-FU therapy.