HLA-A*7401-mediated control of HIV viremia is independent of its linkage disequilibrium with HLA-B*5703

Abstract

The potential contribution of HLA-A alleles to viremic control in chronic HIV type 1 (HIV-1) infection has been relatively understudied compared with HLA-B. In these studies, we show that HLA-A*7401 is associated with favorable viremic control in extended southern African cohorts of >2100 C-clade-infected subjects. We present evidence that HLA-A*7401 operates an effect that is independent of HLA-B*5703, with which it is in linkage disequilibrium in some populations, to mediate lowered viremia. We describe a novel statistical approach to detecting additive effects between class I alleles in control of HIV-1 disease, highlighting improved viremic control in subjects with HLA-A*7401 combined with HLA-B*57. In common with HLA-B alleles that are associated with effective control of viremia, HLA-A*7401 presents highly targeted epitopes in several proteins, including Gag, Pol, Rev, and Nef, of which the Gag epitopes appear immunodominant. We identify eight novel putative HLA-A*7401-restricted epitopes, of which three have been defined to the optimal epitope. In common with HLA-B alleles linked with slow progression, viremic control through an HLA-A*7401-restricted response appears to be associated with the selection of escape mutants within Gag epitopes that reduce viral replicative capacity. These studies highlight the potentially important contribution of an HLA-A allele to immune control of HIV infection, which may have been concealed by a stronger effect mediated by an HLA-B allele with which it is in linkage disequilibrium. In addition, these studies identify a factor contributing to different HIV disease outcomes in individuals expressing HLA-B*5703.

FIGURE 1

LD between HLA-A*7401 and other class I alleles in southern African populations. A, All HLA-B and HLA-Cw alleles that are significantly linked to HLA-A*7401 in a pooled cohort from South Africa (Durban, Bloemfontein, and Kimberley cohorts, n = 1510) and in Botswana (n = 514) are shown. The p values were calculated by Fisher’s exact test and corrected for multiple comparisons using the Bonferroni approach, using the online tool at: http://www.hiv.lanl.gov/content/immunology/hla/hla_linkage.html. No significant linkage associations were identified for HLA-A*7401 among Zimbabweans recruited via the Thames Valley cohort (n = 73, data not shown). All subjects with HLA-B*35 are pooled together as HLA-B*35(01). B, Percentage of HLA-A*7401⁺ subjects with HLA-B*5703 in each of five southern African cohorts. Statistically significant LD between these alleles is present only in the Durban cohort.

FIGURE 2

Viral load (VL) and CD4⁺ T cell count in the presence and absence of HLA-A*7401 in adults with chronic HIV-1 C-clade infection in southern African cohorts. A, VL in Durban, South Africa. B, VL in pooled southern Africa cohort (Durban, South Africa; Gaborone, Botswana; Thames Valley subjects from southern Africa [Botswana, Malawi, South Africa, and Zimbabwe]). C, CD4⁺ T cell count in Gaborone, Botswana. D, CD4⁺ T cell count in pooled southern Africa cohort (as above). Median VL (RNA copies/ml plasma) or CD4⁺ T cell count (cells/mm³) for each group is stated. Boxes show median, 25th and 75th percentile; whiskers 10th–90th percentile. The p values by Mann–Whitney U test.

FIGURE 3

Effect of combinations of HLA-A*7401 and HLA-B*5703 on viral load. A and B, Viral load (RNA copies/ml plasma) of C-clade–infected adult study subjects from all study subjects (A) and Durban (B), in the presence and absence of HLA-B*5703 and HLA-A*7401. Boxes show median, 25th and 75th percentile; whiskers 10th–90th percentile. The p values were by Mann–Whitney U test. C and D, Subjects with HLA-B*5703 ranked by viral load and divided by quartiles into four interquartile (IQ) pools for all study subjects (n = 97) (C) and Durban (n = 55) (D). The proportion of each of the four subgroups with HLA-A*7401 is shown, demonstrating that HLA-A*7401 is enriched in the IQ pool with the lowest viral loads. The p values were by Fisher’s exact test.

FIGURE 4

Proportion of subjects with and without HLA-A*7401 making IFN-γ ELISPOT responses to HIV-1 C-clade OLPs. IFN-γ ELI-SPOT responses to OLP-19 and OLP-100 are significantly enriched among subjects with HLA-A*7401 (Durban cohort; n = 1010; p values by Fisher’s exact test). Subjects with HLA-B*1510 are removed from the analysis of responses to OLP-100 to avoid potential confounding because of the presence of an overlapping HLA-B*1510 epitope IL-9 (IHSISERIL, Rev 52–60) (29, 56), accounting for higher than expected rate of responses in the HLA-A*7401⁻ population (method as previously described [13]). n = number making response/total number of subjects.

FIGURE 5

In vitro data to support three new HLA-A*7401–restricted epitopes: Gag-GR11, RT-QR9, and Rev-RR9. A, Magnitude of IFN-γ ELISPOT responses to Gag-GR11 (GQMVHQAISPR), the alternative epitope QR10 (QMVHQAISPR), and other peptide truncations in a chronically infected adult subject from South Africa (subject ID SK362: HLA-A*0301/A*7401/B*1503/B*1510/Cw*0210/Cw*0401). B, Magnitude of IFN-γ ELISPOT responses to RT-QR9 (QIYPGIKVR) and peptide truncations in a chronically infected adult subject from Zimbabwe (subject ID H005: HLA-A*7401/A*3004/B*3501/B*5802/Cw*0401/Cw*0602). C, Magnitude of IFN-γ ELISPOT responses to Rev-RR9 (RQIHSISER), the alternative epitope RR11 (RQRQIHSISER), and the OLP containing these epitopes (OLP-100) in a chronically infected adult subject from Uganda (subject ID R070: HLA-A*0201/A*7401/B*4016/B*4901/Cw*0701/Cw*0802). D, FACS plot gated on live CD3⁺ population of PBMCs from subject R070, stained with MHC class I tetramer HLA-A*7401-Rev-RR9 conjugated to PE fluorochrome, showing a tetramer-positive population of CD3⁺ CD8⁺ T lymphocytes.

FIGURE 6

Binding of HLA-A*7401 to putative optimal epitopes, including overlapping variants of epitopes in p17, RT, and Rev and sequence variants of the epitope in Nef. Binding affinity KD (nM) of HLA-A*7401 to optimal peptides assessed by LOCI (35). Strength of binding is classified on a log scale in accordance with previous methods (35).

FIGURE 7

Relation between HLA-A*7401 status and frequency of HLA-B*5703–restricted CD8⁺ T cell mutations/IFN-γ ELISPOT responses. A, Frequency of HLA-B*5703–selected Gag polymorphisms [defined by previous studies (3, 6, 16, 17)], in 64 subjects with HLA-B*5703 according to the presence/absence of HLA-A*7401. p24-Gag sequences from Bloemfontein (n = 8), Durban (n = 38), Gaborone (n = 11), and Thames Valley cohorts (n = 7), with sequence data available for TW10 (n = 63), KF11 (n = 64), ISW9 (n = 63), and all three epitopes (n = 62). B, Frequency of IFN-γ ELISPOT response to three HLA-B*5703 epitopes in p24-Gag in 46 subjects with HLA-B*5703 according to the presence or absence of HLA-A*7401. The p values were by Fisher’s exact test. Data from Durban (n = 31) and Thames Valley cohorts (n = 15).