We investigated the effects of interferon-beta (IFN-beta) on the growth of human liver cancer cells. The effects of IFN-beta with or without 5-fluorouracil (5-FU) on the proliferation of 13 liver cancer cell lines were investigated in vitro. Chronologic change in IFN-alpha receptor 2 (IFNAR-2) expression was monitored in hepatocellular carcinoma (HCC) cells (HAK-1B) cultured with IFN-beta. After HAK-1B cells were transplanted into nude mice, various doses of IFN-beta were administered, and the tumor volume, weight, histology, tumor blood vessel, and angiogenesis factor expression were examined. IFN-beta inhibited the growth of 11 cell lines with apoptosis in a dose-dependent and time-dependent manner. With IFN-beta, IFNAR-2 expression in HAK-1B cells was significantly downregulated from 6 to 12 h. IFN-beta induced a dose-dependent decrease in tumor volume and weight and a significant increase of apoptosis in the tumor. Both basic fibroblast growth factor (bFGF) and blood vessel number in the tumor decreased only in mice receiving the lowest dose (1000 IU) of IFN-beta. IFN-beta with 10 muM of 5-FU frequently induced synergistic antiproliferative effects. IFN-beta with or without 5-FU induces strong antitumor effects in HCC cells, and we conclude that IFN-beta is useful for the prevention and treatment of HCC.