Jason D. Barbour

Virologic and immunologic consequences of discontinuing combination antiretroviral-drug therapy in HIV-infected patients with detectable viremia

BACKGROUND In many patients with human immunodeficiency virus (HIV) infection, therapy with potent antiretroviral drugs does not result in complete suppression of HIV replication. The effect of cessation of therapy in these patients is unknown. METHODS Sixteen patients who had a plasma HIV RNA level of more than 2500 copies per milliliter during combination antiretroviral-drug therapy were randomly assigned, in a 2:1 ratio, to discontinue or continue therapy. Plasma HIV RNA levels, CD4 cell counts, and drug susceptibility were measured weekly. Viral replicative capacity was measured at base line and at week 12. RESULTS Discontinuation of therapy for 12 weeks was associated with a median decrease in the CD4 cell count of 128 cells per cubic millimeter and an increase in the plasma HIV RNA level of 0.84 log copies per milliliter. Virus from all patients with detectable resistance at entry became susceptible to HIV-protease inhibitors within 16 weeks after the discontinuation of therapy. Drug susceptibility began to increase a median of six weeks after the discontinuation of therapy and was temporally associated with increases in plasma HIV RNA levels and decreases in CD4 cell counts. Viral replicative capacity, measured by means of a recombinant-virus assay, was low at entry into the study and increased after therapy was discontinued. Despite the loss of detectable resistance in plasma, resistant virus was cultured from peripheral-blood mononuclear cells in five of nine patients who could be evaluated. Plasma HIV RNA levels, CD4 cell counts, and drug susceptibility remained stable in the patients who continued therapy. CONCLUSIONS Despite the presence of reduced drug susceptibility, antiretroviral-drug therapy can provide immunologic and virologic benefit. This benefit reflects continued antiviral-drug activity and the maintenance of a viral population with a reduced replicative capacity.

Tim-3 expression defines a novel population of dysfunctional T cells with highly elevated frequencies in progressive HIV-1 infection

Progressive loss of T cell functionality is a hallmark of chronic infection with human immunodeficiency virus 1 (HIV-1). We have identified a novel population of dysfunctional T cells marked by surface expression of the glycoprotein Tim-3. The frequency of this population was increased in HIV-1–infected individuals to a mean of 49.4 ± SD 12.9% of CD8+ T cells expressing Tim-3 in HIV-1–infected chronic progressors versus 28.5 ± 6.8% in HIV-1–uninfected individuals. Levels of Tim-3 expression on T cells from HIV-1–infected inviduals correlated positively with HIV-1 viral load and CD38 expression and inversely with CD4+ T cell count. In progressive HIV-1 infection, Tim-3 expression was up-regulated on HIV-1–specific CD8+ T cells. Tim-3–expressing T cells failed to produce cytokine or proliferate in response to antigen and exhibited impaired Stat5, Erk1/2, and p38 signaling. Blocking the Tim-3 signaling pathway restored proliferation and enhanced cytokine production in HIV-1–specific T cells. Thus, Tim-3 represents a novel target for the therapeutic reversal of HIV-1–associated T cell dysfunction.

Critical loss of the balance between Th17 and T regulatory cell populations in pathogenic SIV infection.

Chronic immune activation and progression to AIDS are observed after SIV infection in macaques but not in natural host primate species. To better understand this dichotomy, we compared acute pathogenic SIV infection in pigtailed macaques (PTs) to non-pathogenic infection in African green monkeys (AGMs). SIVagm-infected PTs, but not SIVagm-infected AGMs, rapidly developed systemic immune activation, marked and selective depletion of IL-17-secreting (Th17) cells, and loss of the balance between Th17 and T regulatory (Treg) cells in blood, lymphoid organs, and mucosal tissue. The loss of Th17 cells was found to be predictive of systemic and sustained T cell activation. Collectively, these data indicate that loss of the Th17 to Treg balance is related to SIV disease progression.

Tryptophan Catabolism by Indoleamine 2,3-Dioxygenase 1 Alters the Balance of TH17 to Regulatory T Cells in HIV Disease

Patients with AIDS have fewer immune cells to defend against microbial invasion through the gut, a critical loss that may be caused by a tryptophan metabolite produced by other immune cells. Loss of the Defenders at the Gate Like archers stationed along the walls of a medieval castle, the immune system patrols the vulnerable parts of our body to keep pathogens at bay. One of these susceptible areas is the mucosa of the gastrointestinal tract, which is continually exposed to ingested and resident pathogens. This defense breaks down in patients with AIDS, in which sentinel immune cells [T helper 17 (TH17) cells] are missing from the gastrointestinal lining, potentially accounting for some secondary infections acquired by these patients. Favre and colleagues present evidence that the loss of these cells (and a parallel increase in immune suppressor cells) is caused by a metabolite of the amino acid tryptophan, new understanding that should help to prevent this serious consequence of HIV infection. HIV disease is in part an inflammatory disease, and activated T cells and cytokines circulate in patients’ blood, along with pathogen-derived molecules that trigger the innate immune system. The authors show that, in patients with serious AIDS, who are in this inflammatory state, the enzyme indoleamine 2,3-dioxygenase 1 (IDO1), which catabolizes tryptophan, is elevated in dendritic cells (DCs)—agents that present antigen to the immune system—from the blood, lymph nodes, and mucosa of the lower gastrointestinal tract. The inflammation-related molecules interferon γ and bacterial lipopolysaccharide can induce IDO1 in isolated DCs. This excess IDO1 activation increased blood concentrations of tryptophan catabolites in patients, and two of the catabolites increased the proportion of TH17 (activating) immune cells and decreased the proportion of T regulatory (Treg) (suppressing) immune cells in culture. In patients with serious disease, the authors found that the ratio of TH17 to Treg cells was much lower than normal, which hampers the ability of the body to raise an effective immune defense against pathogens. This dysfunctional system would set up a reinforcing loop that progressively depletes vulnerable tissues of their immune protection. Paradoxically, it seems, activation of the immune system by HIV may be contributing to the decline in immune function that is the hallmark of the disease. IDO1 inhibitors are being tested for their efficacy in interfering with this dangerous depletion of defenses. The pathogenesis of human and simian immunodeficiency viruses is characterized by CD4+ T cell depletion and chronic T cell activation, leading ultimately to AIDS. CD4+ T helper (TH) cells provide protective immunity and immune regulation through different immune cell functional subsets, including TH1, TH2, T regulatory (Treg), and interleukin-17 (IL-17)–secreting TH17 cells. Because IL-17 can enhance host defenses against microbial agents, thus maintaining the integrity of the mucosal barrier, loss of TH17 cells may foster microbial translocation and sustained inflammation. Here, we study HIV-seropositive subjects and find that progressive disease is associated with the loss of TH17 cells and a reciprocal increase in the fraction of the immunosuppressive Treg cells both in peripheral blood and in rectosigmoid biopsies. The loss of TH17/Treg balance is associated with induction of indoleamine 2,3-dioxygenase 1 (IDO1) by myeloid antigen-presenting dendritic cells and with increased plasma concentration of microbial products. In vitro, the loss of TH17/Treg balance is mediated directly by the proximal tryptophan catabolite from IDO metabolism, 3-hydroxyanthranilic acid. We postulate that induction of IDO may represent a critical initiating event that results in inversion of the TH17/Treg balance and in the consequent maintenance of a chronic inflammatory state in progressive HIV disease.

Sustained CD4+ T Cell Response after Virologic Failure of Protease Inhibitor-Based Regimens in Patients with Human Immunodeficiency Virus Infection

The relationship between plasma human immunodeficiency virus (HIV) RNA levels and peripheral CD4+ T cell counts was examined in 380 HIV-infected adults receiving long-term protease inhibitor therapy. Patients experiencing virologic failure (persistent HIV RNA >500 copies RNA/mL) generally had CD4+ T cell counts that remained greater than pretherapy baseline levels, at least through 96 weeks of follow-up. The CD4+ T cell response was directly and independently related to degree of viral suppression below the pretreatment baseline. For any given HIV RNA level measured 12 weeks after virologic failure, subsequent CD4+ T cell decline was slower in patients receiving a protease inhibitor-based regimen than in a historical control group of untreated patients. These observations suggest that transient or partial declines in plasma HIV RNA levels can have sustained effects on CD4+ T cell levels.

Poor CD4 T cell restoration after suppression of HIV-1 replication may reflect lower thymic function.

ObjectiveTo characterize immune phenotype and thymic function in HIV-1-infected adults with excellent virologic and poor immunologic responses to highly active antiretroviral therapy (HAART). MethodsCross-sectional study of patients with CD4 T cell rises of ⩾ 200 × 106 cells/l (CD4 responders; n = 10) or < 100 × 106 cells/l (poor responders; n = 12) in the first year of therapy. ResultsPoor responders were older than CD4 responders (46 versus 38 years;P < 0.01) and, before HAART, had higher CD4 cell counts (170 versus 35 × 106 cells/l;P = 0.11) and CD8 cell counts (780 versus 536 × 106 cells/l ; P = 0.02). After a median of 160 weeks of therapy, CD4 responders had more circulating naive phenotype (CD45+CD62L+) CD4 cells (227 versus 44 × 106 cells/l ; P = 0.001) and naive phenotype CD8 cells (487 versus 174 × 106 cells/l ; P = 0.004) than did poor responders (after 130 weeks). Computed tomographic scans showed minimal thymic tissue in 11/12 poor responders and abundant tissue in 7/10 responders (P = 0.006). Poor responders had fewer CD4 cells containing T cell receptor excision circles (TREC) compared with CD4 responders (2.12 versus 27.5 × 106 cells/l ; P = 0.004) and had shorter telomeres in CD4 cells (3.8 versus 5.3 kb ; P = 0.05). Metabolic labeling studies with deuterated glucose indicated that the lower frequency of TREC-containing lymphocytes in poor responders was not caused by accelerated proliferation kinetics. ConclusionPoor CD4 T cell increases observed in some patients with good virologic response to HAART may be caused by failure of thymic T cell production.

Evolution of Phenotypic Drug Susceptibility and Viral Replication Capacity during Long-Term Virologic Failure of Protease Inhibitor Therapy in Human Immunodeficiency Virus-Infected Adults

ABSTRACT Continued use of antiretroviral therapy despite the emergence of drug-resistant human immunodeficiency virus (HIV) has been associated with the durable maintenance of plasma HIV RNA levels below pretherapy levels. The factors that may account for this partial control of viral replication were assessed in a longitudinal observational study of 20 HIV-infected adults who remained on a stable protease inhibitor-based regimen despite ongoing viral replication (plasma HIV RNA levels consistently >500 copies/ml). Longitudinal plasma samples (n = 248) were assayed for drug susceptibility and viral replication capacity (measured by using a single-cycle recombinant-virus assay). The initial treatment-mediated decrease in plasma viremia was directly proportional to the reduction in replicative capacity (P = 0.01). Early virologic rebound was associated the emergence of a virus population exhibiting increased protease inhibitor phenotypic resistance, while replicative capacity remained low. During long-term virologic failure, plasma HIV RNA levels often remained stable or increased slowly, while phenotypic resistance continued to increase and replicative capacity decreased slowly. The emergence of primary genotypic mutations within protease (particularly V82A, I84V, and L90M) was temporally associated with increasing phenotypic resistance and decreasing replicative capacity, while the emergence of secondary mutations within protease was associated with more-gradual changes in both phenotypic resistance and replicative capacity. We conclude that HIV may be constrained in its ability to become both highly resistant and highly fit and that this may contribute to the continued partial suppression of plasma HIV RNA levels that is observed in some patients with drug-resistant viremia.

Increased thymic mass and circulating naive CD4 T cells in HIV-1-infected adults treated with growth hormone

Objective To determine whether treatment with growth hormone (GH) enhances thymopoiesis in individuals infected with HIV-1. Methods Five HIV-1-infected adults were treated with GH for 6–12 months in a prospective open-label study. Immunological analyses were performed before GH treatment and repeated at 3 month intervals after GH initiation. Thymic mass was analysed using computed tomography with quantitative density and volume analysis. Analysis of circulating lymphocytes, including naive and memory T cell subsets, was performed using multiparameter flow cytometry. Results GH treatment was associated with a marked increase in thymic mass in all GH recipients. Circulating naive CD4 T cells also increased significantly in all patients during GH therapy, suggesting an enhancement of thymopoiesis. Conclusion GH has significant effects on the human immune system, including the reversal of thymic atrophy in HIV-1-infected adults. De-novo T cell production may thus be inducible in immunodeficient adults.

Tim-3 marks human natural killer cell maturation and suppresses cell-mediated cytotoxicity

Natural killer (NK) cells are innate lymphocytes that play an important role against viral infections and cancer. This effect is achieved through a complex mosaic of inhibitory and activating receptors expressed by NK cells that ultimately determine the magnitude of the NK-cell response. The T-cell immunoglobulin- and mucin domain-containing (Tim)-3 receptor was initially identified as a T-helper 1-specific type I membrane protein involved in regulating T-cell responses. Human NK cells transcribe the highest amounts of Tim-3 among lymphocytes. Tim-3 protein is expressed on essentially all mature CD56(dim)CD16(+) NK cells and is expressed heterogeneously in the immature CD56(bright)CD16(-) NK-cell subset in blood from healthy adults and in cord blood. Tim-3 expression was induced on CD56(bright)CD16(-) NK cells after stimulation with IL-15 or IL-12 and IL-18 in vitro, suggesting that Tim-3 is a maturation marker on NK cells. Whereas Tim-3 has been used to identify dysfunctional T cells, NK cells expressing high amounts of Tim-3 are fully responsive with respect to cytokine production and cytotoxicity. However, when Tim-3 was cross-linked with antibodies it suppressed NK cell-mediated cytotoxicity. These findings suggest that NK-cell responses may be negatively regulated when NK cells encounter target cells expressing cognate ligands of Tim-3.

CD4+ T Cell Kinetics and Activation in Human Immunodeficiency Virus—Infected Patients Who Remain Viremic Despite Long-Term Treatment with Protease Inhibitor—Based Therapy

T cell dynamics were studied in human immunodeficiency virus-infected patients who continued using antiretroviral therapy despite detectable plasma viremia (RNA copies >2500 /mL). CD4(+) cell fractional replacement rates, measured by the deuterated glucose technique, were lower in treated patients with detectable viremia than in untreated patients and were similar to those in patients with undetectable viremia. Cell cycle and activation markers exhibited similar trends. For any level of viremia, CD4(+) cell fractional replacement rates were lower in patients with drug-resistant virus than in patients with wild-type virus, which suggests that the resistant variant was less virulent. Interruption of treatment in patients with drug-resistant viremia resulted in increased CD4(+) cell activation, increased CD4(+) cell turnover, and decreased CD4(+) cell counts. These data indicate that partial virus suppression reduces CD4(+) cell turnover and activation, thereby resulting in sustained CD4(+) cell gains, and that measurements of T cell dynamics may provide an in vivo marker of viral virulence.