Jason M. Brenchley

HIV preferentially infects HIV-specific CD4+ T cells

HIV infection is associated with the progressive loss of CD4+ T cells through their destruction or decreased production. A central, yet unresolved issue of HIV disease is the mechanism for this loss, and in particular whether HIV-specific CD4+ T cells are preferentially affected. Here we show that HIV-specific memory CD4+ T cells in infected individuals contain more HIV viral DNA than other memory CD4+ T cells, at all stages of HIV disease. Additionally, following viral rebound during interruption of antiretroviral therapy, the frequency of HIV viral DNA in the HIV-specific pool of memory CD4+ T cells increases to a greater extent than in memory CD4+ T cells of other specificities. These findings show that HIV-specific CD4+ T cells are preferentially infected by HIV in vivo. This provides a potential mechanism to explain the loss of HIV-specific CD4+ T-cell responses, and consequently the loss of immunological control of HIV replication. Furthermore, the phenomenon of HIV specifically infecting the very cells that respond to it adds a cautionary note to the practice of structured therapy interruption.

A novel approach to the analysis of specificity, clonality, and frequency of HIV-specific T cell responses reveals a potential mechanism for control of viral escape.

Escape from the CD8+ T cell response through epitope mutations can lead to loss of immune control of HIV replication. Theoretically, escape from CD8+ T cell recognition is less likely when multiple TCRs target individual MHC/peptide complexes, thereby increasing the chance that amino acid changes in the epitope could be tolerated. We studied the CD8+ T cell response to six immunodominant epitopes in five HIV-infected subjects using a novel approach combining peptide stimulation, cell surface cytokine capture, flow cytometric sorting, anchored RT-PCR, and real-time quantitative clonotypic TCR tracking. We found marked variability in the number of clonotypes targeting individual epitopes. One subject recognized a single epitope with six clonotypes, most of which were able to recognize and lyse cells expressing a major epitope variant that arose. Additionally, multiple clonotypes remained expanded during the course of infection, irrespective of epitope variant frequency. Thus, CD8+ T cells comprising multiple TCR clonotypes may expand in vivo in response to individual epitopes, and may increase the ability of the response to recognize virus escape mutants.

HIV-Specific Cytolytic CD4 T Cell Responses During Acute HIV Infection Predict Disease Outcome

HIV-specific CD4 T cell responses during acute infection show robust cytolytic activity and correlate with a lower viral set point and better clinical outcome. T Cells Take Center Stage With more than 33 million infected people worldwide, the HIV/AIDS pandemic is the most devastating infectious disease in recent history. The virus infects and kills one of the central players in the immune system—the CD4 T cell. CD4 T cells provide critical helper signals to B lymphocytes, enabling B cells to produce antibodies, and they also aid another key immune cell, CD8 T cells, which kill virally infected cells. However, CD4 T cells specific for HIV are preferentially infected with this deadly virus and therefore are presumed to be unable to help the host immune system combat HIV. Given the importance of CD4 T cell responses in other viral infections, Soghoian et al. sought to revisit the role of HIV-specific CD4 T cells in the control of HIV infection. They followed a group of HIV-positive patients starting almost immediately after the individuals became infected. The patients who were able to better control HIV showed a robust and early expansion of their HIV-specific CD4 T cells compared to those subjects who were not able to control the virus. Surprisingly, these T cells comprised not only classical CD4 helper cells but also cytolytic “killer” CD4 T cells. Indeed, these CD4 T cells were able to kill HIV-infected cells directly, suggesting that they are involved in the control of HIV. The researchers also made another key observation. At the earliest point during the acute phase of HIV infection, they found that patients who had HIV-specific CD4 T cells containing lots of the death protein granzyme A progressed much more slowly (1 year or more) to full-blown disease than did those patients with T cells containing much less granzyme A. The quality of T cell response in acute HIV infection was able to predict better or worse disease outcomes later on. The unexpected expansion of CD4 T cells with the ability to directly kill HIV-infected cells observed in this study not only demonstrates the key role that cytolytic CD4 T cells play during HIV infection but also sheds new light on the general immunobiology of these cells and raises questions about their roles in other viral infections. Early immunological events during acute HIV infection are thought to fundamentally influence long-term disease outcome. Whereas the contribution of HIV-specific CD8 T cell responses to early viral control is well established, the role of HIV-specific CD4 T cell responses in the control of viral replication after acute infection is unknown. A growing body of evidence suggests that CD4 T cells—besides their helper function—have the capacity to directly recognize and kill virally infected cells. In a longitudinal study of a cohort of individuals acutely infected with HIV, we observed that subjects able to spontaneously control HIV replication in the absence of antiretroviral therapy showed a significant expansion of HIV-specific CD4 T cell responses—but not CD8 T cell responses—compared to subjects who progressed to a high viral set point (P = 0.038). Markedly, this expansion occurred before differences in viral load or CD4 T cell count and was characterized by robust cytolytic activity and expression of a distinct profile of perforin and granzymes at the earliest time point. Kaplan-Meier analysis revealed that the emergence of granzyme A+ HIV-specific CD4 T cell responses at baseline was highly predictive of slower disease progression and clinical outcome (average days to CD4 T cell count <350/μl was 575 versus 306, P = 0.001). These data demonstrate that HIV-specific CD4 T cell responses can be used during the earliest phase of HIV infection as an immunological predictor of subsequent viral set point and disease outcome. Moreover, these data suggest that expansion of granzyme A+ HIV-specific cytolytic CD4 T cell responses early during acute HIV infection contributes substantially to the control of viral replication.

The Functional Profile of Primary Human Antiviral CD8+ T Cell Effector Activity Is Dictated by Cognate Peptide Concentration

Antiviral CD8+ T cells can elaborate at least two effector functions, cytokine production and cytotoxicity. Which effector function is elaborated can determine whether the CD8+ T cell response is primarily inflammatory (cytokine producing) or antiviral (cytotoxic). In this study we demonstrate that cytotoxicity can be triggered at peptide concentrations 10- to 100-fold less than those required for cytokine production in primary HIV- and CMV-specific human CD8+ T cells. Cytolytic granule exocytosis occurs at peptide concentrations insufficient to cause substantial TCR down-regulation, providing a mechanism by which a CD8+ T cell could engage and lyse multiple target cells. TCR sequence analysis of virus-specific cells shows that individual T cell clones can degranulate or degranulate and produce cytokine depending on the Ag concentration, indicating that response heterogeneity exists within individual CD8+ T cell clonotypes. Thus, antiviral CD8+ T cell effector function is determined primarily by Ag concentration and is not an inherent characteristic of a virus-specific CD8+ T cell clonotype or the virus to which the response is generated. The inherent ability of viruses to induce high or low Ag states may be the primary determinant of the cytokine vs cytolytic nature of the virus-specific CD8+ T cell response.

Public clonotype usage identifies protective Gag-specific CD8+ T cell responses in SIV infection

Despite the pressing need for an AIDS vaccine, the determinants of protective immunity to HIV remain concealed within the complexity of adaptive immune responses. We dissected immunodominant virus-specific CD8+ T cell populations in Mamu-A*01+ rhesus macaques with primary SIV infection to elucidate the hallmarks of effective immunity at the level of individual constituent clonotypes, which were identified according to the expression of distinct T cell receptors (TCRs). The number of public clonotypes, defined as those that expressed identical TCR β-chain amino acid sequences and recurred in multiple individuals, contained within the acute phase CD8+ T cell population specific for the biologically constrained Gag CM9 (CTPYDINQM; residues 181–189) epitope correlated negatively with the virus load set point. This independent molecular signature of protection was confirmed in a prospective vaccine trial, in which clonotype engagement was governed by the nature of the antigen rather than the context of exposure and public clonotype usage was associated with enhanced recognition of epitope variants. Thus, the pattern of antigen-specific clonotype recruitment within a protective CD8+ T cell population is a prognostic indicator of vaccine efficacy and biological outcome in an AIDS virus infection.

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.

Preferential Infection Shortens the Life Span of Human Immunodeficiency Virus-Specific CD4+ T Cells In Vivo

ABSTRACT CD4+ T-cell help is essential for effective immune responses to viruses. In human immunodeficiency virus (HIV) infection, CD4+ T cells specific for HIV are infected by the virus at higher frequencies than other memory CD4+ T cells. Here, we demonstrate that HIV-specific CD4+ T cells are barely detectable in most infected individuals and that the corresponding CD4+ T cells exhibit an immature phenotype compared to both cytomegalovirus (CMV)-specific CD4+ T cells and other memory CD4+ T cells. However, in two individuals, we observed a rare and diametrically opposed pattern in which HIV-specific CD4+ T-cell populations of large magnitude exhibited a terminally differentiated immunophenotype; these cells were not preferentially infected in vivo. Clonotypic analysis revealed that the HIV-specific CD4+ T cells from these individuals were cross-reactive with CMV. Thus, preferential infection can be circumvented in the presence of cross-reactive CD4+ T cells driven to maturity by coinfecting viral antigens, and this physical proximity rather than activation status per se is an important determinant of preferential infection based on antigen specificity. These data demonstrate that preferential infection reduces the life span of HIV-specific CD4+ T cells in vivo and thereby compromises the generation of effective immune responses to the virus itself; further, this central feature in the pathophysiology of HIV infection can be influenced by the cross-reactivity of responding CD4+ T cells.

Expansion of activated human naïve T‐cells precedes effector function

Naïve T‐cells divide and mature, both functionally and phenotypically, upon stimulation through the T‐cell receptor. Although much is known about the overall changes that occur in naïve cells upon TCR stimulation, and the different memory/effector populations that arise following stimulation, the relationship between cell division and functional and phenotypical changes that occur after activation is poorly understood. Here, we examine the early stages of human naïve and antigen‐experienced T‐cell activation, and the relationship between cell division and acquisition of effector function during the transition from resting antigen‐experienced or naïve T‐cells into effector cells. Stimulated naïve T‐cells proliferate prior to acquisition of effector function, as measured by cytokine production and expression of effector‐associated cell surface molecules. Additionally, we show that interlukin‐7 (IL‐7) can drive proliferation of naïve T‐cells without TCR:MHC peptide interactions. IL‐7 alone does not, however, drive the proliferation of antigen‐experienced T‐cells. Memory T‐cells will divide in response to exogenous IL‐7 but only in the presence of naïve T‐cells and IL‐2. This study contributes to the current understanding of the mechanistic differences between naïve and memory T‐cell responses by defining the functional and phenotypic changes that occur to T‐cells after stimulation.

Evaluation of the Pathogenesis of Decreasing CD4+ T Cell Counts in Human Immunodeficiency Virus Type 1–Infected Patients Receiving Successfully Suppressive Antiretroviral Therapy

Most human immunodeficiency virus (HIV)-infected individuals experience increases in peripheral CD4(+) T cell counts with suppressive antiretroviral therapy (ART) that achieves plasma HIV RNA levels that are less than the limit of detection. However, some individuals experience decreasing CD4(+) T cell counts despite suppression of plasma viremia. We evaluated 4 patients with a history of CD4(+) T cell decline despite successfully suppressive ART, from a median of 719 cells/mm(3) (range, 360-1141 cells/mm(3)) to 227 cells/mm(3) (range, 174-311 cells/mm(3)) over a period of 18-24 months; 3 of the patients were receiving tenofovir and didanosine, which may have contributed to this decrease. There was no evidence of HIV replication, nor of antiretroviral drug resistance in the blood or lymphoid tissue, or increased proliferation or decreased thymic production of naive CD4(+) T cells. All 4 patients had significant fibrosis of the T cell zone of lymphoid tissue, which appeared to be an important factor in the failure to reconstitute T cells.

Lack of in vivo compartmentalization among HIV-1 infected naive and memory CD4(+) T cell subsets

Viral compartmentalization between naïve and memory CD4(+) T cell subsets has been described, but only for individuals who were receiving antiretroviral therapy (ART). We present here an extensive analysis of the viral quasispecies residing in the naïve, central and effector memory CD4(+) T cell subsets in a number of therapy naïve individuals and representing an array of HIV-1 subtypes. We longitudinally analyzed subset-specific infection and evolution in a subtype B infected individual who switches from CCR5 to dual CCR5/CXCR4 coreceptor usage. We show that the central memory subset, the predominantly infected subset, harbors a more diverse viral population compared to the others. Through sequence analysis of the env C2V3 region we demonstrate a lack of viral compartmentalization among all subsets. Upon coreceptor switch we observe a pronounced increase in the infection level of the naïve population. Our findings emphasize the importance of all CD4(+) T cell subsets to viral evolution.