Plasmid DNA vaccines have been licensed for use in domesticated animals because of their excellent immunogenicity, but none have yet been licensed for use in humans. Here we report a retrospective analysis of 1218 healthy human volunteers enrolled in 10 phase I clinical trials in which DNA plasmids encoding HIV antigens were administered. Elicited T-cell immune responses were quantified by validated intracellular cytokine staining (ICS) stimulated with HIV peptide pools. HIV-specific binding and neutralizing antibody activities were also analyzed using validated assays. Results showed that, in the absence of adjuvants and boosting with alternative vaccines, DNA vaccines elicited CD8+ and CD4+ T-cell responses in an average of 13.3% (95% CI: 9.8–17.8%) and 37.7% (95% CI: 31.9–43.8%) of vaccine recipients, respectively. Three vaccinations (vs. 2) improved the proportion of subjects with antigen-specific CD8+ responses (p = 0.02), as did increased DNA dosage (p = 0.007). Furthermore, female gender and participants having a lower body mass index were independently associated with higher CD4+ T-cell response rate (p = 0.001 and p = 0.008, respectively). These vaccines elicited minimal neutralizing and binding antibody responses. These findings of the immunogenicity of HIV DNA vaccines in humans can provide guidance for future clinical trials.