Rapid SIV Env-specific mucosal and serum antibody induction augments cellular immunity in protecting immunized, elite-controller macaques against high dose heterologous SIV challenge

Abstract

Three Indian rhesus macaques, Ad-SIV primed/protein boosted and exposed twice to high-dose mucosal SIV(mac251) challenges, exhibited elite control of viremia over 6.5 years. They were negative for host factors associated with control of SIV infection. After a third intrarectal challenge with SIV(smE660), all controlled viremia, with one (macaque #5) maintaining undetectable viremia in blood. Acquisition was not blocked, but virus was contained in the jejunum and draining lymph nodes. Polyfunctional memory T cell responses and high-titered neutralizing and non-neutralizing serum and mucosal antibodies were present before and maintained post-challenge. The level of protection seen for animal #5 was predicted from analyses of gene transcription in jejunum 2 weeks post-challenge. Macaques #7 and #9, exhibiting lower pre-challenge cellular and humoral immunity, partially controlled the SIV(smE660) challenge. Initial vaccine-induced control by macaque #5 extended to the SIV(smE660) challenge due to multiple immune mechanisms that were boosted and augmented by cryptic SIV exposure.

Figure 1

Immunization and challenge history of elite controller macaques (31, 40). (A) Macaques #5, #7, and #9 were originally immunized with an Ad-SIV_smH4 env/rev recombinant vector and boosted with a conformational CD4 binding site peptomer. Two priming immunizations given first orally and intranasally and second intratracheally were followed by two intramuscular boosts of peptomer in PBS. A high dose SIV_mac251 rectal challenge was administered 6 weeks later. The times of second and third intrarectal challenges with SIV_mac251 and SIV_smE660, respectively, and an intermediate in vivo CD8 depletion are indicated. (B) SIV RNA copies/ml plasma are shown for all three macaques, exhibiting elite control of the first high dose SIV_mac251 rectal challenge and durable protection from a identical, second SIV_mac251 rectal challenge one year later. Control group represents the geometric mean of 8 and 2 control macaques for the first and second challenges, respectively. (C) In vivo peripheral CD8⁺ T cell depletion resulted in reemergence of virus for macaque #5. Virus remained undetectable for #7 and #9 (not shown). (D) Outcome of a third heterologous rectal SIV_E660 challenge, 3.5 years after the CD8 depletion study and 8 years since initial immunization. Macaques 858 and 870 are naïve controls.

Figure 2

The sum percent of IFN-γ⁺, + IL-2⁺, + TNF-α⁺ total memory T cells in PBMCs for macaque #5. (A) Shown are the total cytokine positive cells among total memory CD4 or CD8 T cells at weeks −2, 2 and 36 post SIV_smE660 challenge after stimulation with three separate peptide pools. (B) Pie charts for each flow sample from panel A, representing the percent triple, double and single cytokine secreting cells. Neg = negative response.

Figure 3

The sum percent of IFN-γ⁺, + IL-2⁺, + TNF-α⁺ total memory T cells in BAL samples for macaques #5, #7 and #9 over time. (A) Shown are the total cytokine positive cells among total memory CD4 or CD8 T cells at weeks -2, 2 and 36 post SIV_smE660 challenge after stimulation with peptide pools noted. Cell numbers were lower at week 2 post-challenge, so peptide pools were combined for stimulations. For ease in interpretation, responses were summed at weeks -2 and 36 as well. (B) Pie charts for each flow sample from panel A, representing the percent triple, double and single cytokine secreting cells. Neg = negative response. ND = denotes not done.

Figure 4

SIV_mac251 serum binding antibody titers (log₁₀) throughout the three SIV challenges.

Figure 5

Neutralizing antibody titers in plasma after SIV_smE660 challenge. Neutralization titers to (A) TCLA-SIV _mac251 and (B) primary SIV_mac251. CD4-induced antibody neutralization of HIV-2 without sCD4 added (C) and with sCD4 added (D). In (A), neutralizing titers for macaque #5 were >4.4×10⁴ at all time points.

Figure 6

Non-neutralizing antibody titers in plasma after SIV_smE660 challenge. ADCC titers of plasma using SIV₂₅₁ gp120- (A) and SIV_smE660 gp120-coated target cells (B). (C) % ADCVI of SIV_smE660 infection of rhesus PBMC using plasma diluted 1:200.

Figure 7

Mucosal antibody binding titers and transcytosis inhibition by rectal secretions post SIV_smE660 challenge. SIV_mac251 Env specific IgG (A) and IgA (B) titers shown as specific activity values (ratio of specific divided by total IgG or IgA in each sample). Transcytosis inhibition of SIV_mac251 (C) and SIV_smE660 (D) across a tight epithelial cell barrier.

Figure 8

Transcriptional profile of jejunal biopsies week 2 post challenge. (A) Up-regulated genes involved in barrier integrity, epithelial repair and regeneration. (B) Up-regulated immune response genes for macaque #5.

Figure 9

Changes in gene expression levels in jejunal tissue in vaccinated and control macaques 2 weeks post SIV_smE660 challenge. (A) Heat diagram of changes in gene expression in each vaccinated animal as compared to control animals (#858 and #870) following SIV_smE660 challenge. Relative increase (red) or decrease (green) of mRNA levels are shown. (B) Following hierarchical clustering, gene expression levels for the elite controller macaque #5 were then compared to the two control macaques (genes within red box). Pathways statistically over represented in the gene lists are shown to the right of the red arrow with the genes and P value indicated.