556. Toward the Development of Electroporation for Delivery of DNA Vaccines to Humans

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The objectives of this study were: 1) to evaluate the potency of plasmid DNA vaccines encoding two different HIV antigens in Rhesus macaques when delivered intramuscularly with electroporation (EP), and 2) to develop EP electrodes and pulse conditions suitable for human clinical applications. Two groups of monkeys (n=5) were immunized with DNA vaccines encoding HIV Gag and an oligomeric form of HIV Env by three subsequent i.m. injections at weeks 0, 4, and 14. In one group (EP group), injection was followed by six EP pulses delivered with a 6-needle array electrode at a nominal field strength of 233V/cm (Widera et al., J. Immunol. 164, 4635–40, 2000). All animals received a boost with rSF2 Gag protein (week 29) and rSF162 Env protein (weeks 38 and 75). During the DNA priming period (up to week 29) the EP group showed faster onset, higher peak titers (>100-fold for Gag) and longer duration of the antibody response than the non-EP group. Protein boosts restored (Gag) or enhanced (Env) peak Ab titers obtained after the third DNA immunization. Cell-mediated responses were also consistently stronger with EP. Helper T cell responses were measured by a lymphoproliferation assay (LPA) and intracellular cytokine staining (ICS). Anti-Gag LPA responses in animals given EP produced stable stimulation indices (SIs) of 12–19 throughout the DNA priming period, compared with a maximum SI of 5 without EP. For Env the corresponding SIs were 8 –10 vs. a maximum of 4. Also, IFN-gamma production in CD4 T cells, as measured by ICS, was 5 to 9-fold higher in the EP group during the entire DNA priming period for Gag, compared with a 2 to 5-fold increase for Env. Cytolytic T cell activity (CTL response) was measured after in vitro culture of PBMC. EP treatment modestly increased the response rate to anti-Gag CTL (20 vs. 13 cumulative responses). However, anti-Env CTLs were markedly enhanced by EP (16 vs. 2 cumulative responses), with 3 out of 5 animals consistently responding throughout the DNA priming period, vs. only one animal responding transiently in the non-EP group. These data demonstrate that in vivo EP is effective at increasing the potency of DNA vaccines against two different HIV antigens in Rhesus by triggering faster, stronger and longer antibody, helper T cell and CTL responses. Naked DNA injection, although effective in rodents, is known to be ineffective in non-human primates. Since EP is a physical delivery system that consistently results in significantly increased immune responses in a spectrum of species from mice to monkeys, it is reasonable to hypothesize that EP could enable effective DNA vaccination in humans as well. In order to make EP more patient-friendly, we developed EP instrumentation and pulse conditions more compatible with routine clinical applications than those used for the treatments described above. We reduced the number of needle electrodes from 6 to 4, reduced electrode toxicity, and limited the number of pulses to two. Under these clinically superior conditions, gene expression levels and antibody responses in rabbits were unaffected. This encourages further non-human primate studies in preparation for clinical trials.