The therapeutic effect of the fibroblast-mediated human interferon (IFN alpha) gene therapy in combination with interleukin-2 (IL-2) activated killer cells (AK)/doxorubicin (i.e., adoptive chemoimmunotherapy) on nude mice bearing the human hepatocellular carcinoma (HCC) was investigated. A fibroblast cell clone (NIH3T3-IFN alpha+) secreting 1024 U/ml human IFN alpha was obtained from 14 positive clones by BMGNeo-IFN alpha DNA transfection, G418-resistant selection, limiting dilution and assay of IFN alpha activity. After i.p. implantation of NIH3T3-IFN alpha+ encapsulated into collagen, serum human IFN alpha activity could be detected from 12 h to day 15 with a peak at 72 h. AK were prepared from human peripheral mononuclear cells costimulated in vitro by IL-2 and inactivated human SMMC 7721 HCC cells. When the NIH3T3-IFN alpha+ cells were i.p. implanted into the HCC-bearing nude mice, the grown of HCC was inhibited and the survival time of the mice was extended. The growth of HCC was inhibited more obviously when AK was i.v. injected and IL-2 was i.p. injected after the NIH3T3-IFN alpha+ cells had been implanted. The best therapeutic effect was achieved when NIH3T3-IFN alpha+ cells were used in combination with IL-2/AK/doxorubicin. All these results suggested that the fibroblast-mediated human IFN alpha gene therapy could be used to treat the human hepatocellular carcinoma effectively and that when used in combination with IL-2-based adoptive chemoimmunotherapy, the therapeutic effect would be better.