High-functional-avidity cytotoxic T lymphocyte responses to HLA-B-restricted Gag-derived epitopes associated with relative HIV control

Abstract

Virus-specific cytotoxic T lymphocytes (CTL) with high levels of functional avidity have been associated with viral clearance in hepatitis C virus infection and with enhanced antiviral protective immunity in animal models. However, the role of functional avidity as a determinant of HIV-specific CTL efficacy remains to be assessed. Here we measured the functional avidities of HIV-specific CTL responses targeting 20 different, optimally defined CTL epitopes restricted by 13 different HLA class I alleles in a cohort comprising 44 HIV controllers and 68 HIV noncontrollers. Responses restricted by HLA-B alleles and responses targeting epitopes located in HIV Gag exhibited significantly higher functional avidities than responses restricted by HLA-A or HLA-C molecules (P = 0.0003) or responses targeting epitopes outside Gag (P < 0.0001). The functional avidities of Gag-specific and HLA-B-restricted responses were higher in HIV controllers than in noncontrollers (P = 0.014 and P = 0.018) and were not restored in HIV noncontrollers initiating antiretroviral therapy. T-cell receptor (TCR) analyses revealed narrower TCR repertoires in higher-avidity CTL populations, which were dominated by public TCR sequences in HIV controllers. Together, these data link the presence of high-avidity Gag-specific and HLA-B-restricted CTL responses with viral suppression in vivo and provide new insights into the immune parameters that mediate spontaneous control of HIV infection.

Fig. 1.

CTL responses targeting HLA-B-restricted and Gag-derived epitopes exhibit high levels of functional avidity. Epitope-specific responses were analyzed for 112 HIV-infected individuals (68 noncontrollers and 44 controllers). (A) The median functional avidity of 99 HLA-B-restricted responses was compared with the median functional avidities of 76 HLA-A-restricted and 24 HLA-C-restricted responses. (B and C) Stratification for epitope localization by HIV protein. The median functional avidity of responses targeting Gag-derived epitopes was compared with the median functional avidities of responses directed against epitopes derived from Pol, Nef, and Env. The box plots represent the functional avidities of all epitopes tested in each group; median values, 25th and 75th percentiles, and ranges are displayed. The Kruskal-Wallis test followed by Dunn's posttest for multiple comparisons was used to assess significance. ns, not significant; *, P = 0.01 to 0.05; **, P = 0.001 to 0.01; ***, P < 0.001.

Fig. 2.

HIV controllers mount CTL responses with higher functional avidities than noncontrollers. (A) Stacked bars indicate the numbers of individuals (controllers or noncontrollers) tested for each epitope. (B) Comparing all responses, controllers exhibited significantly higher functional avidities than noncontrollers. (C) Functional avidities are shown for controllers and noncontrollers after stratification of responses according to the restricting HLA molecule. Epitope-specific responses restricted by HLA-B but not HLA-A or HLA-C showed higher functional avidities with controllers (gray-shaded boxes) than with noncontrollers (white boxes). (D) Gag-derived epitopes but not non-Gag-derived epitopes elicited responses with higher functional avidities with controllers (gray-shaded boxes) than with noncontrollers (white boxes). ns, not significant.

Fig. 3.

HAART-mediated suppression of viral replication does not select for high-avidity CTL responses. Functional avidities and magnitudes for a total of 14 different epitope-specific CTL responses were quantified for 12 individuals with chronic HIV infection before and after the initiation of HAART. For epitopes tested for more than one individual, mean values and standard deviations are indicated. (A) CTL response magnitudes were measured in IFN-γ ELISpot assays before and after the initiation of HAART; fold change between the untreated and treated time points is displayed in each case. (B) Changes in CTL response magnitudes versus time after the initiation of HAART (Spearman rank test). (C) Functional avidities of CTL responses were measured before and after the initiation of HAART; log change between the untreated and treated time points is displayed in each case. (D) Changes in the functional avidities of CTL responses versus time after the initiation of HAART (Spearman rank test).

Fig. 4.

Responses restricted by HLA alleles associated with superior control of HIV infection exhibit high levels of functional avidity. The functional avidities of CTL responses specific for two Gag-derived epitopes restricted by the protective alleles HLA-B27 and HLA-B57 were compared either with those measured for all other tested epitope-specific responses, regardless of allelic restriction, or with those for all other HLA-B-restricted responses. CTL responses to B27-KK10 (A) or B57-KF11 (B) showed higher median functional avidities than either other HLA-B-restricted responses or all other responses studied. The box plots represent the functional avidities of all epitopes tested in each group; median values, 25th and 75th percentiles, and ranges are displayed.

Fig. 5.

Responses to promiscuously presented epitopes exhibit higher functional avidities in the context of HLA class I alleles associated with better disease outcome. The functional avidities of CTL responses to epitopes presented in the context of different HLA class I alleles were determined for untreated individuals with chronic HIV infection. (A to E) CTL responses in individuals expressing HLA class I alleles associated with better control (“good alleles,” red lines) or accelerated disease progression (“bad alleles,” blue lines) were compared with responses to the corresponding epitopes presented in the context of alleles with neutral disease outcome (black lines). (F to H) Epitopes promiscuously presented on alleles associated with average rate of HIV disease progression were included as controls. (I) To allow exploratory statistical comparisons, avidities of the response restricted by the “more beneficial” allele were compared to the avidities of those for the “less beneficial” allele for each epitope tested in different HLA contexts. Responses to epitopes restricted by HLA class I alleles associated with a more beneficial disease outcome showed a strong trend toward higher functional avidities than those of the corresponding responses restricted by less beneficial alleles. Horizontal red bars in panel I represent median values.

Fig. 6.

High-avidity CTL responses in controllers are associated with narrow TCR repertoires. CDR3β amino acid sequence, TRBV and TRBJ usage, and relative frequency are shown for all clonotypes within each epitope-specific response. For each epitope, one controller and one noncontroller pair was studied. Public clonotypes are color coded in the CDR3 sequence column and were identified as TCRβ amino acid sequences observed for more than one individual with reference to an extensive database including unpublished data and data from previous studies (4, 12, 34, 49, 50, 58, 80, 85). The IMGT nomenclature is used (48).