Diversified prime and boost protocols using recombinant vaccinia virus and recombinant non-replicating avian pox virus to enhance T-cell immunity and antitumor responses.

Recombinant vaccinia viruses containing tumor associated genes represent an attractive vector to induce immune responses to weak immunogens in cancer immunotherapy protocols. The property of intense immunogenicity of vaccinia proteins, however, also serves to limit the number of inoculations of recombinant vaccinia viruses. Host immune responses to the first immunization have been shown to limit the replication of subsequent vaccinations and thus reduce effectiveness of boost inoculations. The use of recombinant avian pox viruses (avipox) such as the canarypox (ALVAC) or fowlpox are potential candidates for immunization protocols in that they can infect mammalian cells and express the inserted transgene, but do not replicate in mammalian cells. We report here the construction and characterization of a canarypox (ALVAC) recombinant expressing the human carcinoembryonic antigen (CEA) gene (designated ALVAC-CEA). Antibody, lymphoproliferative and cytolytic T-cell responses as well as tumor inhibition were shown to be elicited by the ALVAC-CEA recombinant in a murine model. The utilization of a diversified immunization scheme using a recombinant vaccinia virus followed by recombinant avian pox virus was shown to be far superior than the use of either one alone in eliciting CEA-specific T-cell responses. Experiments were conducted to determine if the use of a diversified immunization scheme using a recombinant vaccinia virus (rV-CEA) and ALVAC-CEA would be superior to the use of either one alone in eliciting CEA-specific T-cell responses. When mice were immunized with rV-CEA and then ALVAC-CEA. CEA-specific T-cell responses were at least four times greater, and for superior to those achieved with three immunizations of ALVAC-CEA. Multiple boosts of ALVAC-CEA following rV-CEA immunization further potentiated anti-tumor effects and CEA specific T-cell responses. These studies demonstrate the proof of concept of the advantage of diversified immunization protocols employing both recombinant vaccinia and recombinant avipox vectors.