Control of HIV-1 in elite suppressors despite ongoing replication and evolution in plasma virus

Abstract

A subset of HIV-1-infected patients known as elite controllers or suppressors (ES) control the virus naturally. We have previously demonstrated sequence discordance between proviral and plasma gag clones in ES, much of which can be attributed to selective pressure from the host (J. R. Bailey, T. M. Williams, R. F. Siliciano, and J. N. Blankson, J. Exp. Med. 203:1357-1369, 2006). However, it is not clear whether ongoing viral replication continues in ES once the control of viremia has been established or whether selective pressure impacts this evolution. The cytotoxic T-lymphocyte (CTL) response in ES often targets Gag and frequently is superior to that of HIV-1 progressors, partially due to the HLA class I alleles B*57/5801 and B*27, which are overrepresented in ES. We therefore examined longitudinal plasma and proviral gag sequences from HLA-B*57/5801 and -B*27 ES. Despite the highly conserved nature of gag, we observed clear evidence of evolution in the plasma virus, largely due to synonymous substitutions. In contrast, evolution was rare in proviral clones, suggesting that ongoing replication in ES does not permit the significant reseeding of the latent reservoir. Interestingly, there was little continual evolution in CTL epitopes, and we detected de novo CTL responses to autologous viral mutants. Thus, some ES control viremia despite ongoing replication and evolution.

FIG. 1.

Phylogenetic analysis of gag in elite suppressor patients. Phylogenies were estimated by using a classical approach, functioning under a maximum-likelihood (ML) optimality criterion. All sequences are clonal, and APOBEC-mediated hypermutated sequences were removed from analysis. Bootstrap values of 50 and higher are displayed. Protective HLA B alleles are noted beneath the patient number. Patients are not related in any way but are placed on the same tree for display purposes. Colors indicate time, with the scale below in years. Triangles represent clonal plasma sequences, squares represent proviral sequences from HLA-DR⁺ CD4⁺ T cells, and circles represent clonal proviral sequences from HLA-DR⁻ CD4⁺ T cells. To the right, escape mutations in HLA-B*57- and -B*27-restricted epitopes are denoted in black or gray and aligned with the appropriate symbol on the tree. Escape mutations are listed above the grid, with the epitopes in which the mutations occurred listed above the mutations.

FIG. 2.

Phylogenetic analysis of gag in ES8. Phylogenies were constructed as described for Fig. 1; ES8 is isolated due to the large number of sequences obtained for this patient. Colors indicate time, with the scale below in years. Triangles represent plasma sequences, squares represent proviral sequences from HLA-DR⁺ CD4⁺ T cells, and circles represent proviral sequences from HLA-DR⁻ CD4⁺ T cells. To the right, escape mutations in HLA B*57- and -B*27-restricted epitopes are denoted in black or gray and aligned with the appropriate symbol on the tree. Escape mutations are listed above the grid, with the epitopes in which the mutations occurred listed above the mutations.

FIG. 3.

Analysis of synonymous and nonsynonymous mutation in the plasma virus and proviral compartments. Shown are p-distance values for plasma (A) and proviral (B) sequences as determined by comparing early and late samples for each patient utilizing the Nei-Gojobori method. The numbers of differences also were calculated for plasma (C) and proviral (D) sequences using the Nei-Gojobori method.

FIG. 4.

Relevant sequence regions from clonal, near-full-length gag amplified from plasma (pl) or from either resting (HLA-DR⁻) or activated (HLA-DR⁺) CD4⁺ T cells. The date of sample acquisition and number of clones that are identical to the displayed sequences are noted. The HLA-B*27-restricted epitope KK10 (Gag 263-272) and HLA-B*57-restricted epitopes IW9 (Gag 147-155), KF11 (Gag 162-172), TW10 (Gag 240-249), and QW9 (Gag 308-316) all are denoted in shaded boxes. Sites of compensatory mutations for KK10 (S173) (42) and TW10 (H219, I223, M228) escape mutants also are shaded, and for ES7 the DL15 epitope is displayed. Sequences from 2004 and 2005 for ES3, ES7, ES8, and ES9 have been previously reported and are shown for comparative purposes only.

FIG. 5.

(A) Intracellular IFN-γ staining analysis of ES1 HLA-B27 tetramer-positive cells to autologous and wild-type KK10 (KRWIILGLNK; Gag 263-272) peptides. (B) IFN-γ ELISPOT analysis of ES8 CD8⁺ T cells to autologous TW10 variants and wild-type TW10 peptide (TSTLQEQIGW; Gag 240-249). (C) IFN-γ ELISPOT analysis of ES31 CD8⁺ T cells to autologous TW10 variant and wild-type TW10 peptides. (D) IFN-γ ELISPOT analysis of the ES7 CD8⁺ T cells to the wild-type and the autologous peptide containing the K335R mutation in DL15 (DCKTILKALGPAATL; Gag 329-343). (E) Magnitude of IFN-γ response by ES8, ES3, ES7, and ES9 to four epitopes in which evolution occurred in the plasma virus of either ES7 or ES8. Data reflect the IFN-γ response to the autologous variant of the epitope of each patient at the latest time point available. Open symbols indicate SFC < 50. Circles indicate the response by patients in whom no evolution was seen in this epitope, while diamonds indicate the response by the patient in whom the evolution of the plasma virus occurred in this epitope (ES8 for TW10 and ES7 for IW9, QW9, and KF11).