Deficiency of HIV-Gag-specific T cells in early childhood correlates with poor viral containment

Abstract

Perinatal HIV infection is characterized by a sustained high-level viremia and a high risk of rapid progression to AIDS, indicating a failure of immunologic containment of the virus. We hypothesized that age-related differences in the specificity or function of HIV-specific T cells may influence HIV RNA levels and clinical outcome following perinatal infection. In this study, we defined the HIV epitopes targeted by 76 pediatric subjects (47 HIV infected and 29 HIV exposed, but uninfected), and assessed the ability of HIV-specific CD8 and CD4 T cells to degranulate and produce IFN-gamma, TNF-alpha, and IL-2. No responses were detected among HIV-uninfected infants, whereas responses among infected subjects increased in magnitude and breadth with age. Gag-specific responses were uncommon during early infancy, and their frequency was significantly lower among children younger than 24 mo old (p = 0.014). Importantly, Gag responders exhibited significantly lower HIV RNA levels than nonresponders (log viral load 5.8 vs 5.0; p = 0.005). Both the total and Gag-specific T cell frequency correlated inversely with viral load after correction for age, whereas no relationship with targeting of other viral proteins was observed. Functional assessment of HIV-specific T cells by multiparameter flow cytometry revealed that polyfunctional CD8 cells were less prevalent in children before 24 mo of age, and that HIV-specific CD4 cell responses were of universally low frequency among antiretroviral-naive children and absent in young infants. These cross-sectional data suggest that qualitative differences in the CD8 response, combined with a deficiency of HIV-specific CD4 cells, may contribute to the inability of young infants to limit replication of HIV.

FIGURE 1. Absence of HIV-1 specific T cell responses among uninfected infants exposed to HIV-1 in utero

Twenty-nine HIV-exposed but uninfected infants and young children were screened by IFNγ ELISPOT for responses to overlapping peptides spanning all HIV-1 proteins. None had a detectable response, in contrast to HIV-infected subjects, 77% of whom had a detectable response (36 of 47).

FIGURE 2. The magnitude and breadth of HIV-1-specific T cells increase with age

IFNγ ELISPOT assays from 25 antiretroviral-naïve subjects and 9 subjects with VL≥ 50,000 copies/ml despite therapies were analyzed (n=34). There was a positive correlation between age and (A) the total magnitude (Spearman's r=0.40, p=0.02) and (B) the breadth (Spearman's r=0.52, p=0.002) of the HIV-1-specific T cell response.

FIGURE 3. Proteins preferentially targeted by HIV-1-specific T cells differ by age

(A) Prevalence of IFNγ ELISPOT responses to each of the 9 HIV-1 proteins among 34 antiretroviral-naïve or highly viremic subjects, stratified by age. Recognition of HIV-1-Gag increased with age (Chi-square test for trend, p=0.011). No statistically significant trend was observed for the other viral proteins. (B) The same data are displayed as the relative proportion of the total HIV-specific response targeting each individual viral protein (calculated as the protein-specific SFC/million divided by the total; Chi-square test for trend, p<0.001). (C) The total magnitude of the Gag-specific response in subjects older than 24 months was significantly higher than in those <24 months old (Wilcoxon rank-sum Test, p=0.014). Bars indicate median values.

FIGURE 4. Total and Gag-specific T cell responses correlate inversely with viral load

Cross-sectional analysis of HIV-1-specific T cell responses among 25 antiretroviral-naïve subjects demonstrated a highly significant inverse correlation between HIV viral load and (A) the total breadth of the HIV-specific T cell response (Spearman's r=−0.71, p<0.001), as well as (B) the total magnitude of this response (Spearman's r=−0.63, p<0.001). The breadth and magnitude of the HIV-specific T cell immune response remained inversely correlated with viral load after controlling for age in a multivariate regression model (p = 0.027 for the aggregate effect of breadth and magnitude). (C) The total magnitude of the Gag-specific T cell response correlated inversely with viral load after adjusting for known covariates (p=0.037).

FIGURE 5. Responders to HIV-1 Gag exhibit lower viral loads

Subjects with Gag-specific T cell responses (n=14, median VL=703,000 copies/ml) had significantly lower viral loads than Gag-nonresponders (n=11, median VL=102,090 copies/ml; p=0.005). Bars indicate median values.

FIGURE 6. Multiple effector functions of HIV-specific T cells assessed by 9-parameter flow cytometry

(A) Representative scheme for identification of Env-specific CD8 T cell responses is shown for a 12-month old antiretroviral-naïve subject. Gates for each of the 3 functions were established based on the negative control tube (i.e. cytokine vs. CD8 plot) for each individual subject. (B) The frequency of CD8 T cells producing IFNγ, producing TNFα, or degranulating in response to pooled overlapping peptides spanning Gag, Nef, Pol, RTVVV (Rev, Tat, Vif, Vpr, and Vpu), and Env. The number of responding subjects (among 15 antiretroviral-naïve subjects) is indicated in parentheses for each pool. Bars indicate median values. (C) A strong linear correlation was observed between CD8 T cell degranulation and production of IFNγ (Spearman's r=0.87; p<0.0001). (D) The frequency of IFNγ-producing CD4 cells specific for HIVGag increases linearly with log-age (Spearman's r=0.586; p=0.014; dotted line indicates 95% confidence band).