This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. We reported that mutations in N-linked glycosylation sites in V2 (aa. 188 and 198) and V3 (aa. 303) of HIV-1 89.6 gp120 increased neutralization sensitivity of the mutant viruses. In this study, we compared the ability of the wild-type or the mutant Env to induce neutralizing antibodies (NtAb) and protection. Pig-tailed macaques (6/group) were primed with two recombinant vaccinia viruses (VV), one expressing SIVmac239 Gag/Pol, and the other, HIV89.6 Env gp160, in one of three forms: wild-type (Group 1), single mutant in V2 (aa. 198) (Group 2), or triple mutant in V2 and V3 (Group 3). Animals were boosted 12-14 mo later with SIV gag/pol DNA and the cognate gp140 Env protein in alum. Control animals were primed with VV and boosted with vector plasmid and adjuvant only. Immunization with recombinant VV alone elicited low but persistent gp120-specific antibodies. After boosting, titers increased 30-50-fold to ~105 in all three groups. However, their NtAb activities differed. Group 1 animals generated only low NtAb against SHIV89.6, SHIV89.6P and HIV-1SF162, with mean titers of 35, 55, and 37, respectively, as defined by serum dilutions resulting in 50% reduction of viral infectivity in TZM-bl cell assays. Group 2 animals had significantly higher NtAb against the same viruses, with mean titers of 800, 600 and 1200 respectively. Sera from Group 2 animals also neutralized a panel of subtype B primary isolates (SS1196.1, QH0692.42, SC422661.8, PVO.4, AC10.0.29, RHPA.7, and REJO.67) at low to moderate levels (50%-80% or 80%-90% reduction of infectivity, respectively, at 1:15 serum dilution). Group 3 animals failed to generate any detectable NtAb. Significant protection against intrarectal SHIV89.6P challenge was observed in Group 1 and 2 animals, which was correlated in part with NtAb. These results indicate potential advantages for glycan modification in vaccine design, although the role of specific glycans requires further examinations.
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