Background: DNA vaccines can elicit both humoral and cellular immune responses, and offer a useful approach to preventing HIV-1 infection. However, results from clinical trials to date suggest that DNA-based vaccines are only weakly immunogenic in humans. We sought to improve the immunogenicity of a DNA vaccine by targeting antigen-presenting cells (APC) through fusion of SIV gag with the extracellular domain of mouse CTLA-4.

Methods: We constructed a DNA vaccine encoding a fusion gene, combining the ecto-domain of mouse CTLA-4 with a synthetic, “optimized” SIV gag. As a control, we made a similar construct, CTLA4mut-gag, wherein the B7-binding domain in CTLA-4 was mutated. We vaccinated mice intramuscularly with these vaccines using various dosages and schedules. Cell-mediated immune responses were assessed by studying splenocytes in antigen-specific IFN-γ ELISpot assays, and humoral responses were measured by ELISA.

Results: Overall, fusion of wild-type CTLA-4 to gag significantly enhanced Gag-specific IgG titers by ~100-fold, as compared to responses elicited against CTLA4mut-gag. Moreover, Gag-specific cell-mediated responses were consistently increased by ~2-fold. Subsequent flow-cytometric, microscopic, and immunologic studies showed that CTLA-4-gag, but not the mutant form, enhanced the binding of Gag to dendritic cells in vitro, as well as improved antigen internalization and presentation to T-cells. Ex vivo studies demonstrated that in mice vaccinated with CTLA4-gag, dendritic cells from draining lymph nodes served as better APC than dendritic cells from non-draining lymph nodes. These findings from mechanistic studies collectively suggest that the increased immunogenicity is indeed the result of better APC targeting mediated by a functional CTLA-4.

Conclusions: Based on these observations, we propose that a strategy using CTLA-4 to target APC should be pursued to enhance the immunogenicity of HIV-1 vaccines.