Background : Human carcinoembryonic antigen (CEA) is a 180-kd glycoprotein expressed in human colorectal, gastric, pancreatic, breast, and non–small–cell lung carcinomal. Previous studies have demonstrated enhanced immune responses to other antigens presented with vaccinia virus proteins via a recombinant CEA-vacciniavirus construct, designated rV (WR)-CEA, and have demonstrated humoral anti-CEA responses in mice after immunization with that virus. Purpose : The goals of this study were (a) to construct a recombinant CEA-vaccinia vaccine in a less virulent vacinia strain that is potentially safe and effective for treatment of patients whose tumors express CEA abd (b) to evaluate the ability of the recombinant CEA- vaccinia vaccine to prevent and reverse tumor growth in mice and to elicit cell-mediated and humoral anti-CEA immune responses. Methods : Using the New York city strain of vaccinia virus, which is used in smallpox vaccination and is more attenuated for humans than rV(WR), we derived a recombinant CEA-vaccinia construct, designated rV(NYC)-CEA. The ability of this construct to induce antitumor immunity was evaluated in mice receiving sub-cutanneous injections of murine colon adenocarcinoma cells expressing the human CEA gene. Results : Administration of rV(NYC)-CEA in mice induced strong anti-CEA antibody responses, as well as CEA-specific cell-mediated responses, including delayed-type hypersensitivity, lymphoproliferative, and cytotoxic responses. Vaccination of mice with the rV(NYC)-CEA rendered them resistant to the growth of subsequently transplanted CEA -expressing tumors. Moreover, when mice were vaccinated 7 days after tumor cell injection, tumor growth was either greatly reduced or eliminated. No toxic effects were observed in any of the mice. Conclusion: These studies demonstrate that antitumor activity can be induced with the use of a recombinant CEA - vaccinia virus construct derived from an attenuated vaccinia strain, and they reveal the range of cell-mediated and humoral responses induced by this recombinant vaccine.[J Natl Cancer Inst 84:1084 –1091, 1992]