Virological outcome after structured interruption of antiretroviral therapy for human immunodeficiency virus infection is associated with the functional profile of virus-specific CD8+ T cells

Abstract

A clear understanding of the antiviral effects of CD8(+) T cells in the context of chronic human immunodeficiency virus (HIV) infection is critical for the development of prophylactic vaccines and therapeutics designed to support T-cell-mediated immunity. However, defining the potential correlates of effective CD8(+) T-cell immunity has proven difficult; notably, comprehensive analyses have demonstrated that the size and shape of the CD8(+) T-cell response are not necessarily indicative of efficacy determined by measures of plasma viral load. Here, we conducted a detailed quantitative and qualitative analysis of CD8(+) T-cell responses to autologous virus in a cohort of six HIV-infected individuals with a history of structured interruption of antiretroviral therapy (ART) (SIT). The magnitude and breadth of the HIV-specific response did not, by themselves, explain the changes observed in plasma virus levels after the cessation of ART. Furthermore, mutational escape from targeted epitopes could not account for the differential virological outcomes in this cohort. However, the functionality of HIV-specific CD8(+) T-cell populations upon antigen encounter, determined by the simultaneous and independent measurement of five CD8(+) T-cell functions (degranulation and gamma interferon, macrophage inflammatory protein 1beta, tumor necrosis factor alpha, and interleukin-2 levels) reflected the emergent level of plasma virus, with multiple functions being elicited in those individuals with lower levels of viremia after SIT. These data show that the quality of the HIV-specific CD8(+) T-cell response, rather than the quantity, is associated with the dynamics of viral replication in the absence of ART and suggest that the effects of SIT can be assessed by measuring the functional profile of HIV-specific CD8(+) T cells.

FIG. 1.

Immunological and virological characteristics of HIV-infected individuals during SIT and definitive cessation of ART. A cohort of six individuals with chronic HIV infection received SIT comprised of 8 weeks on ART followed by 4 weeks off ART for a period of up to 92 weeks. At week 92 of the study, individuals voluntarily stopped ART. Viral load (HIV RNA copies/ml plasma) and absolute numbers of CD4⁺ and CD8⁺ T cells per mm³ of blood were monitored throughout treatment interruption and cessation periods. Gray shading depicts periods on ART. Patients 103, 119, and 124 restarted ART at weeks 100, 104, and 117, respectively.

FIG. 2.

Magnitude of the total HIV-specific T-cell response. The total magnitudes of the CD4⁺ (A) and CD8⁺ (B) T-cell responses determined using mixes of overlapping peptides specific for Gag, Pol, Env, Nef, and Rev and Tat, Vpu, Vpr, and Vif (RTVV) are shown for each subject as the percent cytokine production during the first week of the last cycle on ART (week 84) and subsequent time points after the cessation ART. Levels of pVL are shown as log₁₀ HIV RNA copies/ml. Overall, the magnitude of the total HIV-specific T-cell response was not predictive of virological outcome.

FIG. 3.

Functional characterization of HIV-specific CD8⁺ T-cell populations during the cessation of ART. PBMCs from patients 101 (week 124), 102 (week 112), and 114 (week 124) (A) and from patients 119 (week 100) and 124 (week 100) (B) were stimulated for 6 h with the relevant peptides and stained with a panel of monoclonal antibodies to examine degranulation (CD107a), cytokine production (IFN-γ, TNF-α, and IL-2), and chemokine production (MIP1β). The 31 possible positive responses that can be discerned from the simultaneous examination of these five functional parameters are shown on the x axis and are color-coded according to the number of functions expressed per group. The total frequency of CD8⁺ T cells displaying each particular functional profile is shown on the y axis.

FIG. 4.

Functional profiles of HIV-specific CD8⁺ T-cell populations. Individual combinations specific for each functional category (5+, 4+, and 3+, etc.) are expressed as a percentage of the total response and ordered from highest functionality to lowest. The majority of Gag- and Nef-specific CD8⁺ T cells from the immunodominant populations in patients 101 and 114, respectively, expressed predominantly 4+ and 3+ functions, as did patient 102. In contrast, Gag-specific CD8⁺ T cells from patients 119 and 124 had more limited functionality; the subdominant Gag-specific CD8⁺ T-cell population from patient 114 exhibited similarly restricted functionality.

FIG. 5.

Sequence evolution and recognition of targeted epitopes by HIV-specific CD8⁺ T cells. Longitudinal sequence analyses of immunogenic regions showed limited evolution within viral populations during SIT and the post-SIT period off ART. HIV-specific CD8⁺ T-cell recognition of viral epitopes was determined using overlapping 11-mer peptides offset by 1 amino acid from consensus and autologous sequences for each targeted region. Only the dominant viral epitopes are shown for clarity. CD8⁺ T cells from patients 101 and 114 recognized multiple HIV epitopes restricted by different HLA class I alleles. Of note, the highest-frequency responses consistently targeted autologous over consensus peptides.