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184 HIV Gag DNA Vaccine: Mutation in the Gag Increases Its Immunogenicity
H. Zhang and L. Qiao*
Loyola Univ. Chicago, Maywood, IL, USA
Background: A plasmid DNA encoding Gag normally produces Gag particles. The particles are taken up by antigen-presenting cells, and then Gag peptides are presented to specific cytotoxic T cells. To enhance the antigen presentation, we hypothesize that if Gag is not stable and the particle formation is blocked, the Gag will be degraded in MHC class I processing pathway and effectively presented to CTLs, leading to enhanced immunogenicity.
Methods: Four amino acids (LELE) were inserted between 21 and 22 of the wild-type (wt) Gag by PCR. The Gag degradation was determined by pulse-chase analysis. Particle formation was determined by EM. The immunogenicity was determined in Balb/c mice by subcutaneous injection with the plasmids encoding the wt Gag or mutant Gag. CTL generation in spleen was determined by ELISPOT.
Results: The mice were immunized with a DNA vaccine encoding the wt Gag or the mutant Gag that is not stable and cannot form particles. We found that the mutant Gag induced significantly more specific cytotoxic T cells than the vaccine encoding wt Gag.
Conclusions: Instablization of Gag and blockade of formation of particles leads to an enhanced immunogenicity.
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