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Apoptin is a chicken anemia virus-derived, p53-independent, bcl-2-insensitive apoptotic protein with the ability to specifically induce apoptosis in tumor cells. To explore the use of theApoptingene in cancer gene therapy, we constructed a recombinant fowlpox virus expressing the Apoptin protein (vFV-Apoptin) and compared the tumor-killing activity of the recombinant virus with that of wild-type fowlpox virus in the human hepatoma cell line HepG2. We found that although cells were somewhat resistant to the basal cytotoxic effect of wild-type fowlpox virus, infection with vFV-Apoptincaused a pronounced, additional cytotoxic effect. Furthermore, cell death and disruption of tumor integrity were apparent in the vFV-Apoptin-infected cells. We also tested whether fowlpox virus-mediated expression of Apoptin in tumor cells could stimulate an antitumor effect by injecting aggressive subcutaneous tumors derived from H22 mouse hepatoma cells in C57BL/6 mice with vFV-Apoptin. We found that fowlpox virus-mediated intratumoral expression of theApoptingene can induce protective and therapeutic antitumor effects and significantly increase survival. Taken together, these data indicate that infection of tumors with fowlpox virus expressing Apoptin inhibits tumor growth, induces apoptosis and may be an effective cancer treatment.