DNA IMMUNIZATION WITH VACCINIA VIRUS L3 CODING PROTEIN PROMOTES STRONG T- CYTOTOXIC IMMUNE

Poxviruses are complex dsDNA viruses with over 200 genes; many of them with unidentified roles in the stimulation of immune responses. L3L ORF is highly conserved among all poxviruses and encodes a virion protein required in the virus transcription. The main purpose of this study is to uncover new possible vaccine candidates in order to develop a vaccine suitable for every member of the population including immunosuppressed individuals and cancer patients, the latter being the second leading cause of death in the Caribbean Region. We hypothesize that administration of a plasmid encoding for L3 protein (pL3L) will induce strong cell-mediated immune responses in a mouse model. To validate our hypothesis, we measured the production of IFN-g and IL-4 by ELISPOT. Our IFN-g ELISPOT analysis shows mean frequencies of spot-forming cells (SFC) per million splenocytes of 459.1± 59.6, 98.9±37.0 and 9.6±4.1 (p ≤ 0.001) in groups immunized with pL3L, pVAX1 and Naïve, respectively. Moreover, our CD8+ depletion assay shows a significant decrease in the mean frequency of SFC per million splenocytes of 84.7± 25.9 for CD8+ depleted fractions versus 459.1± 59.6 (p ≤ 0.0001) for non-depleted. Flow cytometry analysis of both fractions confirmed that production of IFN-g was triggered by T-cytotoxic cells. In contrast, IL-4 ELISPOT assays show mean frequencies of 517.8, 462.2 and 497.2 for animals immunized with pL3L, pVAX1 and Naïve groups, showing no statistical significance. Furthermore, production of both cytokines analyzed by ELISA showed similar results. These data demonstrate that our formulation induces a strong Th1 immune response.