Determinant of HIV-1 mutational escape from cytotoxic T lymphocytes.

CD8+ class I-restricted cytotoxic T lymphocytes (CTLs) usually incompletely suppress HIV-1 in vivo, and while analogous partial suppression induces antiretroviral drug-resistance mutations, epitope escape mutations are inconsistently observed. However, escape mutation depends on the net balance of selective pressure and mutational fitness costs, which are poorly understood and difficult to study in vivo. Here we used a controlled in vitro system to evaluate the ability of HIV-1 to escape from CTL clones, finding that virus replicating under selective pressure rapidly can develop phenotypic resistance associated with genotypic changes. Escape varied between clones recognizing the same Gag epitope or different Gag and RT epitopes, indicating the influence of the T cell receptor on pressure and fitness costs. Gag and RT escape mutations were monoclonal intra-epitope substitutions, indicating limitation by fitness constraints in structural proteins. In contrast, escape from Nef-specific CTL was more rapid and consistent, marked by a polyclonal mixture of epitope point mutations and upstream frameshifts. We conclude that incomplete viral suppression by CTL can result in rapid emergence of immune escape, but the likelihood is strongly determined by factors influencing the fitness costs of the particular epitope targeted and the ability of responding CTL to recognize specific epitope variants

Determinants of HIV-1 Mutational Escape From Cytotoxic T Lymphocytes

CD8+ class I–restricted cytotoxic T lymphocytes (CTLs) usually incompletely suppress HIV-1 in vivo, and while analogous partial suppression induces antiretroviral drug-resistance mutations, epitope escape mutations are inconsistently observed. However, escape mutation depends on the net balance of selective pressure and mutational fitness costs, which are poorly understood and difficult to study in vivo. Here we used a controlled in vitro system to evaluate the ability of HIV-1 to escape from CTL clones, finding that virus replicating under selective pressure rapidly can develop phenotypic resistance associated with genotypic changes. Escape varied between clones recognizing the same Gag epitope or different Gag and RT epitopes, indicating the influence of the T cell receptor on pressure and fitness costs. Gag and RT escape mutations were monoclonal intra-epitope substitutions, indicating limitation by fitness constraints in structural proteins. In contrast, escape from Nef-specific CTL was more rapid and consistent, marked by a polyclonal mixture of epitope point mutations and upstream frameshifts. We conclude that incomplete viral suppression by CTL can result in rapid emergence of immune escape, but the likelihood is strongly determined by factors influencing the fitness costs of the particular epitope targeted and the ability of responding CTL to recognize specific epitope variants

P04-15. Prevalence of Broadly Neutralizing Antibody Responses During Acute/Early HIV Infection

Background: Determining how cross-reactive neutralizing antibody (NAb) responses develop during natural HIV-1 infection may provide key information to understand the role they play in controlling the infection and in disease progression. Here we investigated the frequency and breadth of broadly NAb responses during acute and early infection in a well controlled cohort, and attempted to characterize the factors associated with the development of such responses. Methods: The plasma of 38 clade B acutely-infected, antiretroviral-naive subjects from two cohorts was screened for breadth of neutralization against a panel of heterologous isolates in an Env pseudovirus neutralization assay. Clade A, B and C variants were chosen from reference panels of viruses, created to evaluate the NAb responses elicited during infection or immunization. Results: Our preliminary screening strategy demonstrated that broadly neutralizing antibodies can be detected in a third of the infected subjects. Breadth of neutralization can develop as early as one year during natural HIV infection, however the majority of breadth was observed at over two years post infection. Conclusion: Cross-reactive NAbs are developed more frequently during early infection than previously thought. The consequences of this 'early' development of cross-reactive NAbs on plasma viremia and disease progression are under investigation. The work described was supported by National Institute of Health grant number R01 AI047708-11

Identification of a CCR5-Expressing T Cell Subset That Is Resistant to R5-Tropic HIV Infection

Infection with HIV-1 perturbs homeostasis of human T cell subsets, leading to accelerated immunologic deterioration. While studying changes in CD4(+) memory and naïve T cells during HIV-1 infection, we found that a subset of CD4(+) effector memory T cells that are CCR7(−)CD45RO(−)CD45RA(+) (referred to as T(EMRA) cells), was significantly increased in some HIV-infected individuals. This T cell subset displayed a differentiated phenotype and skewed Th1-type cytokine production. Despite expressing high levels of CCR5, T(EMRA) cells were strikingly resistant to infection with CCR5 (R5)–tropic HIV-1, but remained highly susceptible to CXCR4 (X4)–tropic HIV-1. The resistance of T(EMRA) cells to R5-tropic viruses was determined to be post-entry of the virus and prior to early viral reverse transcription, suggesting a block at the uncoating stage. Remarkably, in a subset of the HIV-infected individuals, the relatively high proportion of T(EMRA) cells within effector T cells strongly correlated with higher CD4(+) T cell numbers. These data provide compelling evidence for selection of an HIV-1–resistant CD4(+) T cell population during the course of HIV-1 infection. Determining the host factors within T(EMRA) cells that restrict R5-tropic viruses and endow HIV-1–specific CD4(+) T cells with this ability may result in novel therapeutic strategies against HIV-1 infection

T Cell Activation Markers and African Mitochondrial DNA Haplogroups among Non-Hispanic Black Participants in AIDS Clinical Trials Group Study 384

Introduction: Mitochondrial function influences T cell dynamics and is affected by mitochondrial DNA (mtDNA) variation. We previously reported an association between African mtDNA haplogroup L2 and less robust CD4 cell recovery on antiretroviral therapy (ART) in non-Hispanic black ACTG 384 subjects. We explored whether additional T cell parameters in this cohort differed by mtDNA haplogroup. Methods: ACTG 384 randomized ART-naïve subjects to two different nucleoside regimens with efavirenz, nelfinavir, or both. CD4 and CD8 memory and activation markers were available at baseline and week 48 on most subjects. mtDNA sequencing was performed on whole blood DNA, and haplogroups were determined. We studied non-Hispanic black subjects with HIV RNA <400 copies/mL at week 48. Analyses included Wilcoxon ranksum test and linear regression. Results: Data from 104 subjects were included. Major African mtDNA haplogroups included L1 (N = 25), L2 (N = 31), and L3 (N = 32). Baseline age, HIV RNA, and CD4 cells did not differ between L2 and non-L2 haplogroups. Compared to non-L2 haplogroups, L2 subjects had lower baseline activated CD4 cells (median 12% vs. 17%; p = 0.03) and tended toward lower activated CD8 cells (41% vs. 47%; p = 0.06). At 48 weeks of ART, L2 subjects had smaller decreases in activated CD4 cells (−4% vs. −11%; p = 0.01), and smaller CD4 cell increases (+95 vs. +178; p = 0.002). In models adjusting for baseline age, CD4 cells, HIV RNA, and naïve-to-memory CD4 cell ratio, haplogroup L2 was associated with lower baseline (p = 0.04) and 48-week change in (p = 0.01) activated CD4 cells. Conclusions: Among ART-naïve non-Hispanic blacks, mtDNA haplogroup L2 was associated with baseline and 48-week change in T cell activation, and poorer CD4 cell recovery. These data suggest mtDNA variation may influence CD4 T cell dynamics by modulating T cell activation. Further study is needed to replicate these associations and identify mechanisms

Limited T Cell Receptor Diversity of HCV-specific T Cell Responses Is Associated with CTL Escape

Escape mutations are believed to be important contributors to immune evasion by rapidly evolving viruses such as hepatitis C virus (HCV). We show that the majority of HCV-specific cytotoxic T lymphocyte (CTL) responses directed against viral epitopes that escaped immune recognition in HCV-infected chimpanzees displayed a reduced CDR3 amino acid diversity when compared with responses in which no CTL epitope variation was detected during chronic infection or with those associated with protective immunity. Decreased T cell receptor (TCR) CDR3 amino acid diversity in chronic infection could be detected long before the appearance of viral escape mutations in the plasma. In both chronic and resolved infection, identical T cell receptor clonotypes were present in liver and peripheral blood. These findings provide a deeper understanding of the evolution of CTL epitope variations in chronic viral infections and highlight the importance of the generation and maintenance of a diverse TCR repertoire directed against individual epitopes

Deep sequence analysis of HIV adaptation following vertical transmission reveals the impact of immune pressure on the evolution of HIV

Human immunodeficiency virus (HIV) can adapt to an individual’s T cell immune response via genomic mutations that affect antigen recognition and impact disease outcome. These viral adaptations are specific to the host’s human leucocyte antigen (HLA) alleles, as these molecules determine which peptides are presented to T cells. As HLA molecules are highly polymorphic at the population level, horizontal transmission events are most commonly between HLA-mismatched donor/recipient pairs, representing new immune selection environments for the transmitted virus. In this study, we utilised a deep sequencing approach to determine the HIV quasispecies in 26 mother-to-child transmission pairs where the potential for founder viruses to be pre-adapted is high due to the pairs being haplo-identical at HLA loci. This scenario allowed the assessment of specific HIV adaptations following transmission in either a non-selective immune environment, due to recipient HLA mismatched to original selecting HLA, or a selective immune environment, mediated by matched donor/recipient HLA. We show that the pattern of reversion or fixation of HIV adaptations following transmission provides insight into the replicative cost, and likely compensatory networks, associated with specific adaptations in vivo. Furthermore, although transmitted viruses were commonly heavily pre-adapted to the child’s HLA genotype, we found evidence of de novo post-transmission adaptation, representing new epitopes targeted by the child’s T cell response. High-resolution analysis of HIV adaptation is relevant when considering vaccine and cure strategies for individuals exposed to adapted viruses via transmission or reactivated from reservoirs

Cellular Immune Responses and Viral Diversity in Individuals Treated during Acute and Early HIV-1 Infection

Immune responses induced during the early stages of chronic viral infections are thought to influence disease outcome. Using HIV as a model, we examined virus-specific cytotoxic T lymphocytes (CTLs), T helper cells, and viral genetic diversity in relation to duration of infection and subsequent response to antiviral therapy. Individuals with acute HIV-1 infection treated before seroconversion had weaker CTL responses directed at fewer epitopes than persons who were treated after seroconversion. However, treatment-induced control of viremia was associated with the development of strong T helper cell responses in both groups. After 1 yr of antiviral treatment initiated in acute or early infection, all epitope-specific CTL responses persisted despite undetectable viral loads. The breadth and magnitude of CTL responses remained significantly less in treated acute infection than in treated chronic infection, but viral diversity was also significantly less with immediate therapy. We conclude that early treatment of acute HIV infection leads to a more narrowly directed CTL response, stronger T helper cell responses, and a less diverse virus population. Given the need for T helper cells to maintain effective CTL responses and the ability of virus diversification to accommodate immune escape, we hypothesize that early therapy of primary infection may be beneficial despite induction of less robust CTL responses. These data also provide rationale for therapeutic immunization aimed at broadening CTL responses in treated primary HIV infection

Abacavir-Reactive memory T Cells are present in drug naïve individuals

Background Fifty-five percent of individuals with HLA-B*57:01 exposed to the antiretroviral drug abacavir develop a hypersensitivity reaction (HSR) that has been attributed to naïve T-cell responses to neo-antigen generated by the drug. Immunologically confirmed abacavir HSR can manifest clinically in less than 48 hours following first exposure suggesting that, at least in some cases, abacavir HSR is due to re-stimulation of a pre-existing memory T-cell population rather than priming of a high frequency naïve T-cell population. Methods To determine whether a pre-existing abacavir reactive memory T-cell population contributes to early abacavir HSR symptoms, we studied the abacavir specific naïve or memory T-cell response using HLA-B*57:01 positive HSR patients or healthy controls using ELISpot assay, intra-cellular cytokine staining and tetramer labelling. Results Abacavir reactive CD8+ T-cell responses were detected in vitro in one hundred percent of abacavir unexposed HLA-B*57:01 positive healthy donors. Abacavir-specific CD8+ T cells from such donors can be expanded from sorted memory, and sorted naïve, CD8+ T cells without need for autologous CD4+ T cells. Conclusions We propose that these pre-existing abacavir-reactive memory CD8+ T-cell responses must have been primed by earlier exposure to another foreign antigen and that these T cells cross-react with an abacavir-HLA-B*57:01-endogenous peptide ligand complex, in keeping with the model of heterologous immunity proposed in transplant rejection