816. Development of Potent DNA Vaccines by Coupling of Linear MIDGE Vectors to Peptides

Plasmid vectors currently used for DNA vaccination contain many sequences including antibiotic resistance genes which are not needed for the vaccine and that can potentially be dangerous for the host, i.e. in inducing allergic responses. We have developed linear, covalently-closed minimalistic vectors (MIDGE) containing only the expression cassette. MIDGE are a safe and more efficient alternative to plasmids at inducing protective immune responses in DNA vaccination protocols (i.e. hepatitis or leishmania models). More importantly, MIDGE vectors can be chemically modified at both ends by the specific linking of peptides, proteins, sugars, antibodies, biotin etc. We performed experiments to evaluate the effect of the covalent attachment of a short peptide of 11 aminoacids (Pro-Lys-Lys-Lys-Arg-Lys-Val-Glu-Asp-Pro-Tyr) to the MIDGE, a new vector type termed MIDGE-TH 1P. MIDGE-TH1P led to a drastic enhancement of antigenicity, lowered the necessary amount of DNA for detection of antibody response and an earlier onset of measurable antibody titers in vivo. In further mice experiments a pronounced shift of the cytokine profile accompanying the response towards the TH1 type was detected: MIDGE-TH1P resulted in a higher IgG2a/IgG1 ratio, a strong induction of IFN-γ producing antigen-specific CD8+ and more IFN-γ producing CD4+ T-cells compared to plain MIDGE and plasmid vectors. In vitro stimulation of both mouse and human PBMCs confirmed the TH1 shift as measured by the increased production of IL12p40 induced by MIDGE-TH1P vectors (in comparison to plain MIDGE and plasmid as well). These advantages are recently being exploited in several applications where a quicker immune response and/or a TH1-type response are required. Although the mechanism underlying these effects is still unknown, preliminary evidence suggests a direct activation of dendritic cells by this linear DNA-peptide structure as the most likely candidate.