Background: Replication competent Adenovirus 5 host range mutant (Ad5hr)-SIVenv/rev priming and SIV gp120 boosting has partially protected against mucosal SIV_mac251 challenge. Here, the impact of multi-component SIV recombinants on protective efficacy was examined.

Methods: Rhesus macaques, 8/group, were primed intranasally and orally (0 weeks [wks]) and intra-tracheally (12 wks). Groups I-V received Ad5hr-SIVenv/rev. In addition, group II received Ad5hr-SIVgag, group III, Ad5hr-SIVnef, and group IV, Ad5hr-SIVgag plus Ad5hr-SIVnef. Groups I-IV were boosted intramuscularly at 24 and 36 wks with SIVgp120 in MPL-SE adjuvant, and group V with peptomer, an SIV polypeptide mimicking the CD4 binding region of gp120. Group VI controls received vector/adjuvant only. Cellular immunity was monitored by ELISpot, using overlapping Env, Gag, Nef, and Rev peptides, and by tetramer staining on 2 Mamu A*01 macaques/group. Neutralizing antibody activity was evaluated. An intra-rectal SIV_mac251 challenge was administered at 42 wks. Viral loads were assayed by NASBA, proviral DNA by PCR, and virus isolation by co-culture.

Results: All SIV antigens were immunogenic; the frequency of macaques positive by ELISpot was Rev, 31%; Nef, 44%; Gag, 67%; and Env, 82%. Peak mean IFN-(-secreting cells were approximately 270 spot forming cells/10⁶ PBMC for Rev, 350 for Nef, 1000 for Gag, and 950 for Env. Responses persisted > 38 wks post-initial immunization. Tetramer staining confirmed the presence of SIV-specific memory cells. All immunized macaques except those in group V developed neutralizing antibodies. Post-challenge control viremia peaked at 2 x 10⁸ SIV RNA copies/ml plasma, with a set point of 2 x 10⁷. All immunized groups had significantly reduced mean acute viral loads. Group II-IV macaques also had significant 20-fold reductions of mean viral loads at set point. Four (4) macaques had undetectable viremia over 24 wks of monitoring and were negative for virus isolation, but positive for proviral DNA. Four (4) macaques cleared viremia to undetectable levels, and 4 others controlled viral loads to the threshold of detection. These 12 macaques were distributed in groups II (2/7), III (4/8), IV (3/8), and V (3/8).

Conclusions: Replication competent Ad-recombinant priming and sub-unit boosting elicited potent, persistent immunity and conferred significant protection against challenge with the highly pathogenic SIV_mac251. The solid protection was not associated with a single immune response.