Objectives: To study memory T cell proliferative responses and cytokine profiles induced in HIV-1 seronegative volunteers immunized with a live recombinant canarypox vector expressing HIV-1 antigens (ALVAC-HIV) and boosted with a recombinant gp120 subunit vaccine. Design: HIV-specific T cell proliferative responses and cytokines were measured 2 weeks after vaccination. Cytokines secreted by T helper 1 cells (Th1) [interleukin (IL)-2 and interferon-γ (IFN-γ)] and T helper 2 (Th2) cells (IL-4, IL-5, IL-6, and IL-10) were assessed both at the mRNA and the protein level. Methods: Peripheral blood mononuclear cells (PBMC) were stimulated in vitro with HIV antigens. Subsequently, T cell proliferation was measured in a standard lymphoproliferation assay; secreted cytokines were measured using an enzyme-linked immunosorbent assay and upregulation of cytokine mRNA was measured using reverse transcriptase polymerase chain reaction. Results: All individuals who had received ALVAC-HIV followed by the protein vaccine exhibited HIV-1-specific T cell proliferative responses. Moreover, the PBMC of all prime-boost vaccinated individuals produced detectable IFN-γ and IL-10 in response to stimulation with HIV-1 envelope glycoprotein antigens; 83% also had detectable levels of IL-2 and IL-6, 71% had detectable levels of IL-4, and 86% had detectable levels of IL-5. Conclusions: These data indicate that this vaccination regimen was inducing both Th1- and Th2-type responses to HIV-1 envelope antigens. This prime-boost vaccination approach elicited T cell help for the generation of cytotoxic T lymphocyte responses as well as help for antibody production and so promises to generate a broad HIV-1-specific immune response. (C) 2000 Lippincott Williams and Wilkins.