Toll-like receptor agonists enhance HIV-specific T cell response mediated by plasmacytoid dendritic cells in diverse HIV-1 disease progression phenotypes.

BACKGROUND: Plasmacytoid dendritic cells (pDCs) sense viral and bacterial products through Toll-like receptor (TLR)-7 and -9 and translate this sensing into Interferon-α (IFN-α) production and T-cell activation. The understanding of the mechanisms involved in pDCs stimulation may contribute to HIV-cure immunotherapeutic strategies. The objective of the present study was to characterize the immunomodulatory effects of TLR agonist stimulations in several HIV-1 disease progression phenotypes and in non HIV-1 infected donors. METHODS: pDCs, CD4 and CD8 T-cells were isolated from 450 ml of whole blood from non HIV-1 infected donors, immune responders (IR), immune non responders (INR), viremic (VIR) and elite controller (EC) participants. pDCs were stimulated overnight with AT-2, CpG-A, CpG-C and GS-9620 or no stimuli. After that, pDCs were co-cultured with autologous CD4 or CD8 T-cells and with/without HIV-1 (Gag peptide pool) or SEB (Staphylococcal Enterotoxin B). Cytokine array, gene expression and deep immunophenotyping were assayed. FINDINGS: pDCs showed an increase of activation markers levels, interferon related genes, HIV-1 restriction factors and cytokines levels after TLR stimulation in the different HIV-disease progression phenotypes. This pDC activation was prominent with CpG-C and GS-9620 and induced an increase of HIV-specific T-cell response even in VIR and INR comparable with EC. This HIV-1 specific T-cell response was associated with the upregulation of HIV-1 restriction factors and IFN-α production by pDC. INTERPRETATION: These results shed light on the mechanisms associated with TLR-specific pDCs stimulation associated with the induction of a T-cell mediated antiviral response which is essential for HIV-1 eradication strategies. FUNDING: This work was supported by Gilead fellowship program, the Instituto de Salud Carlos III (Fondo Europeo de Desarrollo Regional, FEDER, "a way to make Europe") and the Red Temática de Investigación Cooperativa en SIDA and by the Spanish National Research Council (CSIC).

Persistent HIV-controllers are more prone to spontaneously clear HCV: a retrospective cohort study.

INTRODUCTION: HIV-controllers have the ability to spontaneously maintain viraemia at low or undetectable levels in the absence of antiretroviral treatment. Furthermore, HIV-controllers seem to have a superior capacity to spontaneously clear hepatitis C virus (HCV) compared to non HIV-controllers. Some of these subjects eventually lose HIV-controller status (transient controllers), whereas some HIV-controllers show a persistent natural HIV control (persistent controllers). We aimed to analyse whether persistent controllers have superior capacity to spontaneously clear HCV compared to transient controllers. METHODS: We recruited HIV-controllers from January 1981 up to October 2016 with available antibodies to HCV (anti-HCV) data (n = 744). Factors associated with HIV spontaneous control in relation to HCV status were analysed in persistent and transient HIV-controllers with anti-HCV positive (n = 202 and n = 138 respectively) in comparison with 1700 HCV positive non HIV-controllers recruited from January 1981 up to March 2018, bivariate and multivariate analyses, following a logistic regression model, were applied. In addition, the factors related to the loss and time to lose HIV-controller status were explored (n = 744) using Log rank test and Kaplan-Meier curves, in this case the multivariate analysis consisted in a Cox regression model. RESULTS: A higher frequency of HCV spontaneous clearance was found in persistent HIV-controllers (25.5%) compared to non-controllers (10.2%). After adjusting for potential confounders, as sex, age, HIV transmission risk, CD4+ T-cell nadir and time of follow-up, HCV clearance was independently associated with persistent HIV spontaneous control (p = 0.002; OR (95% CI) = 2.573 (1.428 to 4.633)), but not with transient spontaneous control (p = 0.119; 1.589 (0.888 to 2.845)). Furthermore, persistent HIV-controllers were more likely to spontaneously clear the HCV in comparison with transient controllers (p = 0.027; 0.377 (0.159 to 0.893). Finally, not to lose or lengthen the time of losing this control was independently associated with HCV spontaneous clearance (p = 0.010; 0.503 (0.297 to 0.850). CONCLUSIONS: This study shows an association between spontaneous persistent HIV-control and HCV spontaneous clearance. The study findings support the idea of preserved immune mechanisms in persistent HIV control implicated in HCV spontaneous clearance. These results highlight persistent HIV-controllers but not transient controllers as a good model of functional HIV cure.

High CD8 T cell percentage and HCV replication control are common features in HIV-1 controllers and HTLV-2-co-infected patients with a history of injection drug use.

HTLV-2/HIV-1-coinfected patients and HIV-infected patients with natural HIV-1 control show an immune capacity that allows some control of viral infections. These two groups of patients have showed an immune capacity that allows them to have some control over viral infections, very strong control of HIV-1 replication in the case of HIV-1 controllers. The purpose of this retrospective cross-sectional study was to compare viral and immunologic parameters between three cohorts of Caucasian adult HIV-1-infected patients, including HIV-1 controllers (29 patients), HTLV-2/HIV-1 chronic progressors (56 patients), and HIV-1 chronic progressors (101 patients), followed in two different tertiary University Hospitals in Spain. Demographic parameters, nadir CD4 T cell count, CD4 and CD8 T cell counts and percentage, anti-HCV antibodies, HCV RNA load, HCV genotype, HIV-1 RNA loads, and anti-HTLV-2 antibodies were analyzed. HIV-1 controllers and HTLV-2/HIV-1 chronic progressors were younger and with shorter time since HIV-1 diagnosis compared to HIV-1 chronic progressors. HIV-1 controllers and HTLV-2/HIV-1 chronic progressors had significantly higher CD8 T cell percentage (p = 0.002 and p = 0.016, respectively) and lower levels of HCV RNA loads (0.015 and 0.007, respectively) compared to that of HIV-1 chronic progressors. Multivariate analyses showed that gender and HTLV-2 infection were independently associated to HCV RNA load, while only HTLV-2 infection was independently associated to CD8 T cell percentage. The implication of HTLV-2 infection in the control of HIV-1 and HCV infections is worth being further analyze.

Proteomic Profile Associated With Loss of Spontaneous Human Immunodeficiency Virus Type 1 Elite Control.

BACKGROUND: Elite controllers (ECs) spontaneously control plasma human immunodeficiency virus type 1 (HIV-1) RNA without antiretroviral therapy. However, 25% lose virological control over time. The aim of this work was to study the proteomic profile that preceded this loss of virological control to identify potential biomarkers. METHODS: Plasma samples from ECs who spontaneously lost virological control (transient controllers [TCs]), at 2 years and 1 year before the loss of control, were compared with a control group of ECs who persistently maintained virological control during the same follow-up period (persistent controllers [PCs]). Comparative plasma shotgun proteomics was performed with tandem mass tag (TMT) isobaric tag labeling and nanoflow liquid chromatography coupled to Orbitrap mass spectrometry. RESULTS: Eighteen proteins exhibited differences comparing PC and preloss TC timepoints. These proteins were involved in proinflammatory mechanisms, and some of them play a role in HIV-1 replication and pathogenesis and interact with structural viral proteins. Coagulation factor XI, α-1-antichymotrypsin, ficolin-2, 14-3-3 protein, and galectin-3-binding protein were considered potential biomarkers. CONCLUSIONS: The proteomic signature associated with the spontaneous loss of virological control was characterized by higher levels of inflammation, transendothelial migration, and coagulation. Galectin-3 binding protein could be considered as potential biomarker for the prediction of virological progression and as therapeutic target in ECs.

Eradication of Hepatitis C Virus (HCV) Reduces Immune Activation, Microbial Translocation, and the HIV DNA Level in HIV/HCV-Coinfected Patients.

Background: There are contradictory data about the influence that hepatitis C virus (HCV) has on immune activation and inflammation in patients coinfected with human immunodeficiency virus (HIV) and HCV. Methods: HIV/HCV-coinfected patients receiving antiretroviral treatment who achieved a sustained virological response with interferon-free regimens were consecutively enrolled in a prospective study. The following factors were assessed before, immediately after the end of, and 1 month after the end of therapy: expression of HLA-DR/CD38, PD-1, and CD57 on CD4+ and CD8+ T-cells; measurement of the total HIV DNA load in peripheral blood mononuclear cells; and determination of plasma levels of soluble CD14 (sCD14), lipopolysaccharide (LPS), 16S ribosomal DNA (rDNA), interleukin 6 (IL-6), D-dimers, and high-sensitivity C-reactive protein (hsCRP). Results: Ninety-seven patients were consecutively included. At the end of therapy and 1 month later, there were significant reductions in the expression of HLA-DR and CD38 in CD4+ and CD8+ T cells, as well as levels of proviral HIV DNA, sCD14, LPS, 16S rDNA, and D-dimer (P < .001). By contrast, the expression of PD-1 and CD57 in CD4+ and CD8+ T cells and levels of IL-6 and hsCRP did not change. The improvement in levels of immune activation markers, proviral HIV DNA, and microbial translocation markers did not translate into an increased CD4+ T-cell count or increased ratio of the CD4+ T-cell count to the CD8+ T-cell count. Conclusions: HCV eradication in HIV/HCV-coinfected patients results in significant decreases in levels of immune activation markers, proviral HIV DNA load, microbial translocation markers, and D-dimers. These findings support the use of HCV treatment for all HIV/HCV-coinfected patients, even those with low-grade fibrosis.

Immune Correlates of Natural HIV Elite Control and Simultaneous HCV Clearance-Supercontrollers.

HIV-elite controllers are a minority group of HIV-infected patients with the ability to maintain undetectable HIV viremia for long time periods without antiretroviral treatment. A small group of HIV-controllers are also able to spontaneously clear the hepatitis C virus (HCV) whom we can refer to as "supercontrollers." There are no studies that explore immune correlates looking for the mechanisms implicated in this extraordinary phenomenon. Herein, we have analyzed HCV- and HIV-specific T-cell responses, as well as T, dendritic and NK cell phenotypes. The higher HCV-specific CD4 T-cell polyfunctionality, together with a low activation and exhaustion T-cell phenotype was found in supercontrollers. In addition, the frequency of CD8 CD161high T-cells was related with HIV- and HCV-specific T-cells polyfunctionality. Interesting features regarding NK and plasmacytoid dendritic cells (pDCs) were found. The study of the supercontroller's immune response, subjects that spontaneously controls both chronic viral infections, could provide further insights into virus-specific responses needed to develop immunotherapeutic strategies in the setting of HIV cure or HCV vaccination.

Factors Leading to the Loss of Natural Elite Control of HIV-1 Infection.

HIV-1 elite controllers (EC) maintain undetectable viral loads (VL) in the absence of antiretroviral treatment. However, these subjects have heterogeneous clinical outcomes, including a proportion that loses HIV-1 control over time. In this work, we compared, in a longitudinal design, transient EC, analyzed before and after the loss of virological control, with persistent EC. The aim was to identify factors leading to the loss of natural virological control of HIV-1 infection with a longitudinal retrospective study design. Gag-specific T-cell responses were assessed by in vitro intracellular polycytokine production quantified by flow cytometry. Viral diversity determinations and sequence dating were performed in proviral DNA by PCR amplification at limiting dilution of env and gag genes. The expression profile of 70 serum cytokines and chemokines was assessed by multiplex immunoassays. We identified transient EC as subjects with low Gag-specific T-cell polyfunctionality, high viral diversity, and high proinflammatory cytokine levels before the loss of control. Gag-specific T-cell polyfunctionality was inversely associated with viral diversity in transient controllers before the loss of control (r = -0.8; P = 0.02). RANTES was a potential biomarker of transient control. This study identified virological and immunological factors, including inflammatory biomarkers associated with two different phenotypes within EC. These results may allow a more accurate definition of EC, which could help in better clinical management of these individuals and in the development of future curative approaches.IMPORTANCE There is a rare group of HIV-infected patients who have the extraordinary capacity to maintain undetectable viral load levels in the absence of antiretroviral treatment, the so-called HIV-1 elite controllers (EC). However, there is a proportion within these subjects that eventually loses this capability. In this work, we found differences in virological and immune factors, including soluble inflammatory biomarkers, between subjects with persistent control of viral replication and EC that will lose virological control. The identification of these factors could be a key point for a right medical care of those EC who are going to lose natural control of viral replication and for the design of future immunotherapeutic strategies using as a model the natural persistent control of HIV infection.

Association of heterozygous CCR5Δ32 deletion with survival in HIV-infection: A cohort study.

The role of a 32 base pair deletion in the CCR5 gene (CCR5Δ32) in HIV-disease progression and response to combined antiretroviral therapy (cART) is well established. However, the impact of CCR5Δ32 in the long-term survival pre-cART and after cART introduction in a large cohort of HIV-infected patients is unknown. We analyzed the association of CCR5Δ32 deletion in the long-term survival of HIV-infected patients recruited between June 1981 and October 2016 (n = 1006). Clinical and epidemiological variables were recorded and CCR5Δ32 deletion was assessed by PCR and electrophoretic analysis. The association of CCR5Δ32 deletion with the time to death was analyzed by Log-Rank tests and Cox Regression models. The CCR5 WT/Δ32 prevalence was 13.4% (n = 135). We did not find any homozygous subject for CCR5Δ32 deletion. AIDS (n = 85, 41.5%) and non-AIDS (n = 87, 42.4%) events were the main causes of 205 deaths. CCR5Δ32 deletion was independently associated with survival (p = 0.022; hazard ratio (HR): 0.572, confidence interval (CI) [0.354-0.923]), after adjusting by HIV diagnosis before 1997, age at diagnosis, being on cART, risk of transmission, nadir CD4+ T-cell counts and CDC stage C. This result was reproduced when the analysis was restricted to patients on cART (p = 0.045; HR: 0.530 [0.286-0.985]). These results confirm the protective role of CCR5Δ32, and extend it to the long-term survival in a large cohort of HIV-infected patients. Beyond its antiviral effect, CCR5Δ32 enhanced the long-term survival of patients on cART.

CCR5+ CD8 T-cell levels and monocyte activation precede the onset of acute coronary syndrome in HIV-infected patients on antiretroviral therapy.

Acute coronary syndrome (ACS) is nowadays one of the leading causes of morbid-mortality in HIV-infected population, but innate and adaptive immune mechanisms preceding this event are unknown. In this work we comprehensively and longitudinally observed, by multiparametric flow cytometry and following a case-control design, increased CCR5+CD8+ T-cells levels and monocytes expressing activation and adhesion markers in HIV-infected patients who are going to suffer ACS. In addition, we found direct associations between activated CD8+ T-cells and myeloid cells that were only statistically significant in the group of patients with ACS and in the follow up time point just before the ACS. Our data highlight the important role of CCR5 in the onset of ACS and suggest this receptor as a marker of cardiovascular risk and potential therapeutic target to prevent the development of such non-AIDS-related event in HIV-infected patients.

Analysis of Non-AIDS-Defining Events in HIV Controllers.

BACKGROUND: Human immunodeficiency virus (HIV) controllers have the striking ability to maintain viremia at extremely low or undetectable levels without antiretroviral treatment. Even though these patients have been widely studied, information about clinical outcomes, especially concerning to non-AIDS-defining events (nADEs), is scarce. We have analyzed the frequency and rate of nADEs and their associated factors in a large multicenter HIV controller cohort. METHODS: Data on nADEs were recorded for 320 HIV controllers within the multicenter Spanish AIDS Research Network HIV Controllers Cohort (ECRIS). Percentages and crude incidence rates (CIRs) per 100 person-years of follow-up (PYFU) were calculated for the entire follow-up period and for 2 separate periods: the period under control and the period after loss of control. These rates were compared with those for 632 noncontrollers. Demographic and immunological data collected from the controllers were included in a multivariate model to assess factors that were independently associated with nADEs in HIV controllers. RESULTS: HIV controllers experience nADEs, albeit at lower rates than patients who do not spontaneously control the virus (1.252 [95% confidence interval {CI}, .974-1.586] per 100 PYFU and 2.481 [95% CI, 2.153-2.845] per 100 PYFU, respectively; P < .001). Hepatitis C virus (HCV) coinfection was the main factor associated with nADEs in all of the studied periods. Although hepatic events were the most prevalent, they represented only approximately 30% of the total events. CIRs of cardiovascular events increased in the post-loss-of-control period. CONCLUSIONS: HCV/HIV coinfection was the main factor associated with hepatic and extrahepatic nADEs in HIV controllers. The eradication of HCV infection may ameliorate the presence of comorbidities in these patients.

IFNγ⁻TNFα⁻IL2⁻MIP1α⁻CD107a⁺PRF1⁺ CD8 pp65-Specific T-Cell Response Is Independently Associated With Time to Death in Elderly Humans.

Persistent cytomegalovirus (CMV) infection has been suggested to be a major driving force in the immune deterioration and an underlying source of age-related diseases in the elderly. CMV antibody titers are associated with lower responses to vaccination, cardiovascular diseases, frailty, and mortality. CMV infection is also associated with shorter T-cell telomeres and replicative senescence. Although an age-related deregulation of CMV-specific T-cell responses could be an underlying cause of the relationship between CMV and immune defects, strong and polyfunctional responses are observed in elderly individuals, casting uncertainty on their direct role in age-related immune frailty. In this study, we longitudinally followed a cohort of healthy donors aged over 50 years, assessing their mortality rates and time to death during a 2-year period. Specific T-cell responses to the immunodominant antigen pp65 (IFNγ, TNFα, IL2, MIP1α, CD107a, and perforin production) were analyzed at the beginning of the 2-year observation period. A cytotoxic CD8 pp65-specific T-cell response, without cytokine or chemokine coexpression, was independently associated with all-cause mortality in these elderly individuals. This pp65-specific CD8 T-cell response could be a useful tool to identify individuals with depressed immune function and a higher risk of death.