[show abstract][hide abstract] ABSTRACT: Mechanisms by which CD4+ regulatory T cells (T(regs)) mediate suppression of virus-specific responses remain poorly defined. Adenosine, mediated via CD39 and CD73, has been shown to play a role in the action of murine T(regs) . In this study we investigate the phenotype of T(regs) in the context of human immunodeficiency virus (HIV)-1 infection, and the function of these cells in response to HIV-1-Gag and cytomegalovirus (CMV) peptides. Phenotypic data demonstrate a decrease in forkhead box transcription factor 3 (FoxP3+) T(reg) numbers in the peripheral blood of HIV-1+ individuals compared to healthy controls, which is most pronounced in those with high HIV-1 RNA plasma load. Due to aberrant expression of CD27 and CD127 during HIV-1 disease, these markers are unreliable for T(reg) identification. The CD3+ CD4+ CD25(hi) CD45RO+ phenotype correlated well with FoxP3 expression in both the HIV-1+ and seronegative control cohorts. We observed expression of CD39 but not CD73 on T(regs) from HIV-1+ and healthy control cohorts. We demonstrate, through T(reg) depletion, the suppressive potential of T(regs) over anti-CMV responses in the context of HIV-1 infection; however, no recovery of the HIV-1-specific T cell response was observed indicating a preferential loss of HIV-1-specific T(reg) function. We propose that before immunotherapeutic manipulation of T(regs) is considered, the immunoregulatory profile and distribution kinetics of this population in chronic HIV-1 infection must be elucidated fully.
[show abstract][hide abstract] ABSTRACT: INITIO is an open-labelled randomized trial evaluating first-line therapeutic strategies for human immunodeficiency virus-1 (HIV-1) infection. In an immunology substudy a tetanus toxoid booster (TTB) immunization was planned for 24 weeks after initiation of highly active antiretroviral therapy (HAART). All patients had received tetanus toxoid immunization in childhood. Generation of proliferative responses to tetanus toxoid was compared in two groups of patients, those receiving a protease inhibitor (PI)-sparing regimen (n = 21) and those receiving a PI-containing (n = 54) regimen. Fifty-two participants received a TTB immunization [PI-sparing (n = 15), PI-containing (n = 37)] and 23 participants did not [PI-sparing (n = 6) or PI-containing (n = 17)]. Cellular responses to tetanus antigen were monitored by lymphoproliferation at time of immunization and every 24 weeks to week 156. Proportions with a positive response (defined as stimulation index > or = 3 and Delta counts per minute > or = 3000) were compared at weeks 96 and 156. All analyses were intent-to-treat. Fifty-two participants had a TTB immunization at median 25 weeks; 23 patients did not. At weeks 96 and 156 there was no evidence of a difference in tetanus-specific responses, between those with or without TTB immunization (P = 0.2, P = 0.4). There was no difference in the proportion with response between those with PI-sparing or PI-containing regimens at both time-points (P = 0.8, P = 0.7). The proliferative response to tetanus toxoid was unaffected by initial HAART regimen. Anti-tetanus responses appear to reconstitute eventually in most patients over 156 weeks when treated successfully with HAART, irrespective of whether or not a TTB immunization has been administered.
[show abstract][hide abstract] ABSTRACT: To evaluate functional potential and phenotypic markers in HIV-1-infected patients immunized with HIV-1 rgp160.
We assessed changes in T-cell phenotype and immune function in 12 HIV-1-infected individuals that were part of a therapeutic vaccine study from 1992 to 1995 [Sandstrom E, Wahren B. Therapeutic immunisation with recombinant gp160 in HIV-1 infection: a randomised double-blind placebo-controlled trial. Nordic VAC-04 Study Group. Lancet 1999;353(9166):1735-42]. The patients received 160 microg HIV-1 rgp160 or placebo i.m. at baseline (day 0), and months 1, 2, 3, 4, 6, and thereafter every 3 months. Frozen peripheral blood mononuclear cells (PBMC) were retrieved from time points 0, 9, 12 and 24 months for phenotypic analysis utilizing flow cytometry.
Up-regulation of immune activation markers HLA-DR and CD38 was observed at baseline and throughout the monitoring period on both CD4+ and CD8+ T cells in all patients, reflecting immune activation due to persistent high viral load. Further enhanced expression of activation markers was observed over time in the vaccine group, but not the placebo group. We also observed a consistent long-term increase of the CD4+ central memory population (CD3+CD4+CD45RA-CCR7+) in the vaccinated group.
Administration of eight doses of rgp160 in a year appeared to partially reverse some of the defects exerted by HIV-1 on the immune system. A combination of vaccination with effective antiretroviral therapy (ART) may thus represent an immunotherapeutic intervention for treatment of chronic HIV-1 infection. The improvement of a HIV-1-specific central memory population and HIV-1 antigen-specific CD4+ lymphoproliferative responses may have contributed to the short-term improved survival reported in the vaccinated group.
[show abstract][hide abstract] ABSTRACT: Anergy and defective immune responses are characteristic of chronic HIV-1 infection. The programmed death 1 (PD-1)/PD-1 ligand (PD-L1) pathway appears to be involved in downregulating T-cell functionality. We found raised levels of PD-L1 in aviraemic chronically infected HIV-1-positive patients, which may contribute to incomplete immune reconstitution after treatment with HAART.
AIDS 07/2007; 21(10):1379-81. · 6.41 Impact Factor
[show abstract][hide abstract] ABSTRACT: The long-term immunological benefit of protease inhibitor (PI)-sparing antiretroviral therapy (ART) using non-nucleoside reverse transcriptase inhibitors (NNRTIs) remains poorly investigated.
A total of 120 ART-naive, HIV-1-infected participants were included in the immunology substudy of INITIO, an international randomized trial comparing two NRTIs (didanosine + stavudine) combined with either: one NNRTI (efavirenz; EFV), one non-boosted PI (nelfinavir; NFV), or one NNRTI + one PI (EFV/NFV). CD4+ T-cell counts, HIV-1 plasma RNA load (VL), T-cell phenotype, T-cell proliferation and IFN-gamma production against opportunistic/recall and HIV-1 antigens/peptides were compared at baseline and at week (W) 96 and W156.
Participants (37 EFV, 44 NFV, 39 EFV/NFV) had similar baseline VL; median CD4+ T-cell counts/mm3 were: 144 (64-303) EFV, 212 (42-313) NFV and 257 (86-331) EFV/NFV. At W156, the proportion of patients with VL < or =50 copies/ml was not different between the arms (P=0.3). From baseline to W156 there was a significant increase in CD4+ T-cell counts (P<0.001) and in naive CD4+ T cells (P<0.001), with no difference between arms and percentages of total and activated CD8+ T cells decreased significantly (P<0.001) in all arms. The decrease in activated memory CD4+ T-cells was significantly greater in the EFV arm at W96 (P=0.03) and W156 (P=0.01), but did not persist after adjusting for baseline CD4+ T-cell counts. During follow-up, responses to opportunistic pathogens increased in all patients while specific T-cell responses to HIV-1-p24 and gp160 recombinant proteins or to Gag and Nef peptides were not restored.
Regimens using/sparing PIs provide similar levels of long-term immune reconstitution even in patients with low CD4+ T-cell counts.
[show abstract][hide abstract] ABSTRACT: The enzyme-linked immunospot (ELIspot) assay is a highly sensitive and valuable tool for determining the frequency of cytokine-secreting T cells. It is essential to determine both frequencies and functional capabilities of antigen-specific T cells, including cytokine secretion, degranulation, and cytotoxicity in order to obtain a fuller picture of the immune status of an individual. We describe here for the first time a perforin-release ELIspot assay which, when used in combination with IFN-gamma and IL-4 ELIspots, permits rapid assessment of these functional parameters for antigen-specific T cells. Whole antigen or peptides from HIV-1, recall and other viral antigens were used for in vitro stimulation. Anti-HIV-1 responses in treated chronically infected individuals were weak, both in terms of perforin and IFN-gamma production. Tetanus toxoid stimulation was associated with moderate perforin release and a predominantly type-2 IL-4 producing response, whilst herpes simplex virus antigen stimulation resulted in perforin release but only a weak type-1 IFN-gamma response. Anti-cytomegalovirus responses generated high levels of perforin in conjunction with IFN-gamma. Cytokines IL-2 and IL-12/IL-15 induced perforin release coupled with an IFN-gamma type-1 response. Perforin release strongly correlated with IFN-gamma production to individual influenza, Epstein-Barr virus or cytomegalovirus MHC class I restricted peptides, in an HIV-1 sero-negative cohort, indicating a cytolytic type-1 CD8+ T-cell response. Evaluation of immunogenicity and putative efficacy of candidate vaccines using IFN-gamma will not be as informative alone as when combined with perforin and IL-4 evaluations, which allow assessment of specific cytotoxic potential without extensive cell culture.
Journal of Immunological Methods 02/2006; 308(1-2):216-30. · 2.23 Impact Factor
[show abstract][hide abstract] ABSTRACT: Efficacious protection for future generations from HIV-1 infection through the development of prophylactic vaccines is the best hope for the millions of individuals living with the threat of HIV-1 infection. Advances in the development of non-curative chemotherapy for those already infected have changed the course of the epidemic for those with access to the drugs. However in the ten years since the advent of highly active anti-retroviral therapy, the expectancy of curative chemotherapy has been quashed, and the constant need for a next generation of drugs is evident. As our understanding of HIV-1 pathogenesis increases, it is becoming apparent that novel approaches and strategies will be required to halt the global progression of HIV-1. Immune-based therapies are being considered in the context of effective antiretroviral therapy. Such immune-based therapy must allow the induction or regeneration of HIV-1-specific T-cell responses with the potential to control viremia and purge viral reservoirs. Studies of therapy substitution, treatment interruption, therapeutic vaccines and/or cytokines and/or hormones have been carried out and are briefly summarised in this review.
Current Medicinal Chemistry 02/2006; 13(26):3203-11. · 4.07 Impact Factor
[show abstract][hide abstract] ABSTRACT: Increasing numbers of patients are choosing to interrupt highly active antiretroviral therapy (HAART). We describe the effect of patient-directed treatment interruption (PDTI) on plasma viral loads (pVL), proviral DNA (pDNA), lymphocyte subsets and immune responses in 24 chronically HIV-1 infected individuals. Patients were divided into group A with pVL > 50 copies/ml and group B with pVL < 50 copies/ml, prior to the PDTI. pVL rose significantly in group B during the first month off HAART and was associated with a significant decrease in CD4 T-cell count. At baseline there was a significant difference in HIV-1 pDNA levels between groups A and B, however, levels significantly increased in group B, but not in group A during PDTI becoming equivalent after 1 month PDTI. We have previously shown no increase in pDNA over the time of substitution in patients switching HAART regimens despite a small rebound in pVL. These observations indicate that to protect low pDNA levels PDTI should be discouraged and that changing regimen at the first sign of failure should be advised where possible. Only transient, no longer than 4 week, HIV-1-specific responses were observed during PDTI in 5/24 patients, 2 from group A and 3 from group B. The low numbers of responders and the transient nature of the anti-HIV-1 immune responses do not favour the auto-vaccination hypothesis.
[show abstract][hide abstract] ABSTRACT: HIV-1 infection is characterized by increase in inhibitory receptors and loss of activating receptors on natural killer (NK) cells, resulting in loss of cell activity. Exceptionally, for an inhibitory receptor, the proportion of NK cells bearing CD94-NKG2A decreases during HIV-1 infection. It is not understood whether HIV-1 itself or other concomitant infections drive these changes.
To investigate the relationship between HIV-1 viraemia and changes in C-type lectin-like receptor expression in NK cells and to investigate the effect of highly active antiretroviral therapy (HAART) on these changes.
Three cohorts of patients were studied: (1) before, during and after treatment interruption in aviraemic and viraemic patients receiving HAART (n = 15); (2) HIV-1-positive treatment-naive individuals (n = 13); and (3) HIV-1-positive individuals receiving successful HAART for a minimum of 1 year without interruption (n = 11). Flow cytometry was used to study the expression of NKG2A before and after treatment interruption and to define expanded populations of NK cells in untreated and treated HIV-1-positive individuals. Assays were performed in vitro to assess the cytotoxicity of the expanded populations.
Increases in plasma HIV-1 RNA during treatment interruption in aviraemic HAART-treated individuals did not influence the proportion of NK cells carrying the complex CD94-NKG2A. Loss of NKG2A NK cells corresponded to the dramatic expansion of a distinct population of cells expressing a functional activating CD94-NKG2C receptor with skewed expression of killer cell immunoglobulin-like receptor family and natural cytotoxicity receptors.
Changes in the NK cell repertoire during HIV-1 infection were not a result of HIV-1 viraemia alone but resembled those associated with concomitant infections.
[show abstract][hide abstract] ABSTRACT: Chemokine receptor genetic mutations are among the factors which have been shown to influence human susceptibility to HIV-1 infection and progression. The CCR2-64I mutation has been shown to delay HIV-1 disease progression in some studies. Here we show evidence of delayed disease progression, reflected in maintenance of a stable viral load and a slow CD4 T-cell decline, in a patient with the CCR2-64I gene. We then consider the potential value of identifying these genetic defects in the era of fusion/entry inhibiting therapeutics.
International Journal of STD & AIDS 06/2005; 16(5):392-4. · 1.00 Impact Factor
[show abstract][hide abstract] ABSTRACT: Gary Woodnutt – Diversa Corp., San Diego, CA, USA
There has been considerable effort in attempting to develop effective HIV vaccines but the overall goal has remained elusive. Animal and clinical data suggest that disease progression can be slowed, but most vaccine attempts have resulted in marginal benefit in the long term. Newer strategies incorporating the need to generate CD4-helper T-lymphocyte responses as well as CD-8 are discussed and appear, at least in preliminary results, to be of more benefit. Recent data have provided understanding of the interaction between virus and the immune system and, as more insight is obtained around HIV-1 responses in animal and human systems, increasingly effective vaccines and immunotherapeutics will be developed. Imami et al. discuss some of the, not inconsiderable, hurdles that need to be overcome and look positively at the novel approaches that will lead to the identification of successful vaccine strategies in the future.
Drug Discovery Today Therapeutic Strategies 01/2004; 1(4):461-467.
[show abstract][hide abstract] ABSTRACT: Reconstitution of functional CD4(+) T cell responsiveness to in vitro stimuli is associated with continuous highly active antiretroviral therapy (HAART). Thirty-six antiretroviral naive patients received HAART over 16 weeks. Antigen-specific, mitogen and interleukin (IL)-2 induced lymphocyte proliferative responses and specific IL-2 and IL-4 production were assessed at each time-point, together with quantification of HIV-1 RNA load and lymphocyte populations. Reconstitution of recall responses was limited largely to persistent antigens such as Herpes simplex virus and Candida, rather than to HIV-1 or neo-antigens. Recall antigens, mitogens and IL-2-induced renewed responses were associated with in-vitro production of IL-2, but not IL-4. Differential responsiveness to low versus high concentration IL-2 stimulus increases in a stepwise manner, suggesting normalization of IL-2 receptor expression and improved functionality. These increases in in-vitro proliferative responses thus probably reflect short lived effector clones, driven by ongoing antigenic stimulus associated with persisting long-term organisms. In this context non-responsiveness to HIV-1 antigens suggests ongoing HIV-1 specific clonal T cell anergy.
[show abstract][hide abstract] ABSTRACT: The effect of altering antiretroviral therapy (ART) on responses to viral, recall and human immunodeficiency virus (HIV)-1-specific recombinant antigens and interleukin-2 (IL-2) in HIV-1-infected patients was assessed. A longitudinal cohort study in eight HIV-1 infected individuals following a clinically indicated therapy change (seven for drug intolerance and one for virological failure) from protease inhibitor (PI) to non-nucleoside reverse transcriptase inhibitor (NNRTI)-based antiretroviral regimens was performed. CD4 T-cell counts, viral loads, lymphoproliferative responses, cytokine production and latent proviral deoxyribonucleic acid (DNA) were measured at baseline and at weeks 12 and 24 after therapy substitution. Following therapy-switch there was a 33% proportional increase in mitogen response (95% confidence interval (CI), 3-33%) and a 31% increase (95% CI, 15-48%) in viral and recall-antigen responses. Six patients developed proliferative responses to low concentration IL-2 stimulation. All patients demonstrated an increase in median HIV-1-specific responses, as three had detectable virus at baseline (two being viral rebound); this may reflect an autovaccination effect. Proviral DNA changes largely reflected plasma HIV-1 ribonucleic acid (RNA). In conclusion, NNRTI substitution for a PI may favour immune reconstitution with an improvement in HIV-1-specific responses, which may reflect differential effects on antigen processing and presentation, an autovaccination effect or alternatively a potential suppressive effect of the PI.
Scandinavian Journal of Immunology 07/2003; 57(6):600-7. · 2.20 Impact Factor
[show abstract][hide abstract] ABSTRACT: The thymic output of patients receiving highly active antiretroviral therapy (HAART) was assessed by sjTREC (signal joint T cell receptor rearrangement excision circle) analysis to determine the thymic contribution to CD4(+) T cell reconstitution during initial therapy and during interleukin 2 (IL-2) and/or Remune supplementation of HAART. Levels of sjTRECs were observed to decline dramatically after the first 4 weeks of HAART and then increased without significant associated changes in CD4(+) T cell counts. HAART supplementation with IL-2 was observed to lead to rapid increases in CD4(+) T cells that were accompanied by sjTREC decreases. No notable changes in CD4(+) T cell counts and sjTRECs were seen in patients receiving HAART supplemented with Remune alone. The results indicate CD4(+) T cell maintenance during initial treatment of HIV-1 with HAART and early CD4(+) T cell reconstitution of patients receiving IL-2 with HAART is largely due to thymus-independent mechanisms, with the thymus making a limited contribution.
AIDS Research and Human Retroviruses 03/2003; 19(2):103-9. · 2.71 Impact Factor
[show abstract][hide abstract] ABSTRACT: Interleukin-16 (IL-16) and the beta-chemokines (RANTES, monocyte chemotactic protein-1 (MCP-1), macrophage inhibitory protein (MIP)-1alpha and (MIP)-1beta) are soluble in vitro suppressors of macrophage tropic HIV-1 strains. The reduction of HIV-1 RNA plasma levels in late-stage patients receiving protease inhibitors has been associated with increased concentrations of MIP-1alpha, MIP-1beta, RANTES and IL-16 and a decrease in levels of MCP-1. We determined plasma levels of MCP-1, MIP-1alpha, MIP-1beta, RANTES and IL-16 during the first 16 weeks of highly active antiretroviral therapy (HAART) in chronic HIV-1-infected patients. Patients were administered one of two therapeutic regimens based on either a protease inhibitor (PI) or a non-nucleoside reverse transcriptase inhibitor (NNRTI). No differences were seen in the levels of RANTES and IL-16 over the first 16 weeks of HAART in either treatment group. MCP-1 decreased significantly in the PI-treated group over the first 16 weeks of HAART (P = 0.0003). A significant increase was observed in the levels of MIP-1alpha and MIP-1beta in the NNRTI cohort (P = 0.0010 and P = 0.0012, respectively). A significant decrease in levels of MIP-1alpha and MIP-1beta (P = 0.0015 and P = 0.0299, respectively) was observed over the 16 weeks in the PI cohort. A significant difference was seen when the levels of MIP-1alpha and MIP-1beta were compared between the NNRTI and the PI cohorts at week 16 (P = 0.04 and P = 0.05, respectively). Evaluation of CCR5 expression ex vivo revealed no difference between the two treatment groups. Patients were genotyped for CCR5 Delta32 and the incidence of heterozygosity was lower than in the HIV-1 seronegative controls (3% compared to 19%).
[show abstract][hide abstract] ABSTRACT: HIV-1-specific CD8 cytotoxic and CD4 helper T-lymphocytes, which are respectively the central effector and regulatory cells in viral infections, together with fully functional antigen-presenting cells, are essential at all stages of HIV-1 infection to control viral activity. Recent studies indicate that such protective HIV-1-specific immune responses can be preserved/induced in HIV-1-infected individuals, utilizing strategies such as treatment interruption after early HAART. Despite successful combination antiretroviral drug therapy, strong anti-HIV-1 T-cell responses are often not apparent in chronic HIV-1 infection, diminishing the probability of viral eradication. Thus, the therapeutic use of immunization and cytokines are required to induce and steer immunity towards a desirable outcome. Here, we review and discuss therapeutic immunization and immunotherapy with regard to their potential use in the treatment of chronic HIV-1 infection.
Current opinion in investigational drugs (London, England: 2000) 09/2002; 3(8):1138-45. · 3.55 Impact Factor
[show abstract][hide abstract] ABSTRACT: Most patients with chronic HIV-1 infection lack functional CD4(+) and CD8(+) HIV-1-specific T cells with proliferative and cytolytic capacity, respectively. This is despite being able to produce intracellular cytokines in response to viral antigens. Protease inhibitor (PI)-based highly active anti-retroviral therapy (HAART) is unable to completely eradicate virus and fails to enable total restoration of immunity including induction of anti-HIV-1 responses. We have taken novel approaches towards the treatment of chronic HIV-1 disease with the aim of instigating long-term non-progressor status and depletion of virus reservoirs. HIV-1-specific CD4(+) and CD8(+) T cell responses were measured following the administration of cytokines, during therapeutic vaccination, and following treatment interruption (TI) or drug therapy change. Administration of cytokines, with or without therapeutic vaccination, in HAART treated patients, improved both CD4(+) and CD8(+) HIV-1-specific T cell responses even in late-stage disease. Virus-specific T cell responses were also seen during TI or when transient viraemia was apparent, and following therapy change from a PI- to a non-nucleoside-based HAART regimen. Reconstitution of HIV-1-specific immune responses was found to be transient and reversal to the previous anergic state was rapid. Viral reservoirs in the latently infected resting CD4(+) T cells, on follicular dendritic cells of germinal centers or even in infected thymic epithelium may be involved in clonal suppression and anergy. These may present major obstacles to the maintenance of HIV-1-specific responses and the eventual eradication of HIV-1.
[show abstract][hide abstract] ABSTRACT: The CD45 antigen is essential for normal antigen receptor-mediated signalling in lymphocytes, and different patterns of splicing of CD45 are associated with distinct functions in lymphocytes. Here we show that abnormal CD45 splicing caused by a C77G transversion in exon A of the gene encoding CD45 (PTPRC) is associated with increased susceptibility to HIV-1 infection.