Potent, persistent induction and modulation of cellular immune responses in rhesus macaques primed with Ad5hr-simian immunodeficiency virus (SIV) env/rev, gag, and/or nef vaccines and boosted with SIV gp120

Abstract

Immunity elicited by multicomponent vaccines delivered by replication-competent Ad5hr-simian immunodeficiency virus (SIV) recombinants was systematically investigated. Rhesus macaques were immunized mucosally at weeks 0 and 12 with Ad5hr-SIV(smH4) env/rev, with or without Ad5hr-SIV(mac239) gag or Ad5hr-SIV(mac239) nef, or with all three recombinants. The total Ad5hr dosage was comparably adjusted among all animals with empty Ad5hr-DeltaE3 vector. The macaques were boosted with SIV gp120 in monophosphoryl A-stable emulsion adjuvant at 24 and 36 weeks. Controls received Ad5hr-DeltaE3 vector or adjuvant only. By ELISPOT analysis, all four SIV gene products elicited potent cellular immune responses that persisted 42 weeks post-initial immunization. Unexpectedly, modulation of this cellular immune response was observed among macaques receiving one, two, or three Ad5hr-SIV recombinants. Env responses were significantly enhanced throughout the immunization period in macaques immunized with Ad5hr-SIV env/rev plus Ad5hr-SIV gag and tended to be higher in macaques that also received Ad5hr-SIV nef. Macaques primed with all three recombinants displayed significant down-modulation in numbers of gamma interferon (IFN-gamma)-secreting cells specific for SIV Nef, and the Env- and Gag-specific responses were also diminished. Modulation of antibody responses was not observed. Down-modulation was seen only during the period of Ad5hr-recombinant priming, not during subunit boosting, although SIV-specific IFN-gamma-secreting cells persisted. The effect was not attributable to Ad5hr replication differences among immunization groups. Vaccine delivery via replication-competent live vectors, which can persistently infect new cells and continuously present low-level antigen, may be advantageous in overcoming competition among complex immunogens for immune recognition. Effects of current multicomponent vaccines on individual immune responses should be evaluated with regard to future vaccine design.

FIG. 1.

Immunization schedule and outline of immunization groups. The immunogens, dosages, and adjuvant used are described in Materials and Methods.

FIG. 2.

Construction and expression of Ad5hr-SIV nef_Δ1-13. (A) Schematic showing Ad5hr-SIV nef_Δ1-13 recombinant design and construction. (B) Western blot of Ad5hr-SIV nef_Δ1-13 expression on human 293 (lane 1) and monkey-derived VERO (lane 3) cell lines. Negative controls include uninfected VERO cells (lane 7) and VERO cells infected with Ad5hrΔE3 vector without the nef insert (lane 5).

FIG. 2.

Construction and expression of Ad5hr-SIV nef_Δ1-13. (A) Schematic showing Ad5hr-SIV nef_Δ1-13 recombinant design and construction. (B) Western blot of Ad5hr-SIV nef_Δ1-13 expression on human 293 (lane 1) and monkey-derived VERO (lane 3) cell lines. Negative controls include uninfected VERO cells (lane 7) and VERO cells infected with Ad5hrΔE3 vector without the nef insert (lane 5).

FIG. 3.

Modulation of IFN-γ secretion in response to Env peptide stimuli. Ad5hr-SIV recombinants administered to the various immunization groups are denoted by the inserted SIV genes. Line graphs reflect average SFC per 10⁶ PBMC ± the standard error of the mean at each time point among all macaques in the indicated group. Arrows denote Ad5hr-SIV recombinant immunizations at weeks 0 and 12 and gp120 boosts at weeks 24 and 36.

FIG. 4.

Modulation of IFN-γ secretion in response to Nef peptide stimuli. Data are illustrated as described in the legend to Fig. 3.

FIG. 5.

Modulation of IFN-γ secretion in response to Gag peptide stimuli. Data are illustrated as described in the legend to Fig. 3.

FIG. 6.

Lack of modulation of vaccine-induced humoral immune responses. (A) Serum binding titers to SIV gp120. Mean titers ± standard errors of the means are shown for each immunization group, tested 2 weeks after each indicated time point. (B) Neutralizing antibody titers ± standard errors of the means against T-cell line-adapted SIV_mac251 are shown for each immunization group at 2 weeks following the gp120 boosts.

FIG. 7.

Ad replication within immunization groups. Ad replication was assessed by determining the duration of viral shedding into nasal secretions, as described in Materials and Methods. The mean duration ± the standard error of the mean is shown.