The potential utility of plasmid DNA as an HIV-1 vaccination modality currently is an area of active investigation. However, recent studies have raised doubts as to whether plasmid DNA alone will elicit immune responses of sufficient magnitude to protect against pathogenic AIDS virus challenges. We therefore investigated whether DNA vaccine-elicited immune responses in rhesus monkeys could be augmented by using either an IL-2͞Ig fusion protein or a plasmid expressing IL-2͞Ig. Sixteen monkeys, divided into four experimental groups, were immunized with (i) sham plasmid, (ii) HIV-1 Env 89.6P and simian immunodeficiency virus mac239 Gag DNA vaccines alone, (iii) these DNA vaccines and IL-2͞Ig protein, or (iv) these DNA vaccines and IL-2͞Ig plasmid. The administration of both IL-2͞Ig protein and IL-2͞Ig plasmid induced a significant and sustained in vivo activation of peripheral T cells in the vaccinated monkeys. The monkeys that received IL-2͞Ig plasmid generated 30-fold higher Env-specific antibody titers and 5-fold higher Gag-specific, tetramer-positive CD8؉ T cell levels than the monkeys receiving the DNA vaccines alone. IL-2͞Ig protein also augmented the vaccine-elicited immune responses, but less effectively than IL-2͞Ig plasmid. Augmentation of the immune responses by IL-2͞Ig was evident after the primary immunization and increased with subsequent boost immunizations. These results demonstrate that the administration of IL-2͞Ig plasmid can substantially augment vaccine-elicited humoral and cellular immune responses in higher primates.

A critical priority for human immunodeficiency virus type 1 (HIV-1) vaccine development is standardization of reagents and assays for evaluation of immune responses elicited by candidate vaccines. To provide a panel of viral reagents from multiple vaccine trial sites, 60 international HIV-1 isolates were expanded in peripheral blood mononuclear cells and characterized both genetically and biologically. Ten isolates each from clades A, B, C, and D and 10 isolates each from CRF01_AE and CRF02_AG were prepared from individuals whose HIV-1 infection was evaluated by complete genome sequencing. The main criterion for selection was that the candidate isolate was pure clade or pure circulating recombinant. After expansion in culture, the complete envelope (gp160) of each isolate was verified by sequencing. The 50% tissue culture infectious dose and p24 antigen concentration for each viral stock were determined; no correlation between these two biologic parameters was found. Syncytium formation in MT-2 cells and CCR5 or CXCR4 coreceptor usage were determined for all isolates. Isolates were also screened for neutralization by soluble CD4, a cocktail of monoclonal antibodies, and a pool of HIV-1-positive patient sera. The panel consists of 49 nonsyncytium-inducing isolates that use CCR5 as a major coreceptor and 11 syncytium-inducing isolates that use only CXCR4 or both coreceptors. Neutralization profiles suggest that the panel contains both neutralization-sensitive and -resistant isolates. This collection of HIV-1 isolates represents the six major globally prevalent strains, is exceptionally large and well characterized, and provides an important resource for standardization of immunogenicity assessment in HIV-1 vaccine trials.