Impacts of HIV Cure Interventions on Viral Reservoirs in Tissues

HIV reservoirs persist in infected individuals despite combination antiretroviral therapy and can be identified in secondary lymphoid tissues, in intestinal tissues, in the central nervous system as well as in blood. Clinical trials have begun to explore effects of small molecule interventions to perturb the latent viral infection, but only limited information is available regarding the impacts of HIV cure-related clinical interventions on viral reservoirs found in tissues. Of the 14 HIV cure-related clinical trials since 2012 that have evaluated the effects of small molecule interventions in vivo, four trials have examined the impacts of the interventions in peripheral blood as well as other tissues that harbor persistent HIV. The additional tissues examined include cerebral spinal fluid, intestines and lymph nodes. We provide a comparison contrast analyses of the data across anatomical compartments tested in these studies to reveal where peripheral blood analyses reflect outcomes in other tissues as well as where the data reveal differences between tissue outcomes. We also summarize the current knowledge on these topics and highlight key open questions that need to be addressed experimentally to move the HIV cure research field closer to the development of an intervention strategy capable of eliciting long-term antiretroviral free remission of HIV disease

Using animal models to overcome temporal, spatial and combinatorial challenges in HIV persistence research

Research challenges associated with understanding HIV persistence during antiretroviral therapy can be categorized as temporal, spatial and combinatorial. Temporal research challenges relate to the timing of events during establishment and maintenance of HIV persistence. Spatial research challenges regard the anatomical locations and cell subsets that harbor persistent HIV. Combinatorial research challenges pertain to the order of administration, timing of administration and specific combinations of compounds to be administered during HIV eradication therapy. Overcoming these challenges will improve our understanding of HIV persistence and move the field closer to achieving eradication of persistent HIV. Given that humanized mice and non-human primate HIV models permit rigorous control of experimental conditions, these models have been used extensively as in vivo research platforms for directly addressing these research challenges. The aim of this manuscript is to provide a comprehensive review of these recent translational advances made in animal models of HIV persistence

Worksite interventions for preventing physical deterioration among employees in job-groups with high physical work demands: Background, design and conceptual model of FINALE

<p>Abstract</p> <p>Background</p> <p>A mismatch between individual physical capacities and physical work demands enhance the risk for musculoskeletal disorders, poor work ability and sickness absence, termed physical deterioration. However, effective intervention strategies for preventing physical deterioration in job groups with high physical demands remains to be established. This paper describes the background, design and conceptual model of the FINALE programme, a framework for health promoting interventions at 4 Danish job groups (i.e. cleaners, health-care workers, construction workers and industrial workers) characterized by high physical work demands, musculoskeletal disorders, poor work ability and sickness absence.</p> <p>Methods/Design</p> <p>A novel approach of the FINALE programme is that the interventions, i.e. 3 randomized controlled trials (RCT) and 1 exploratory case-control study are tailored to the physical work demands, physical capacities and health profile of workers in each job-group. The RCT among cleaners, characterized by repetitive work tasks and musculoskeletal disorders, aims at making the cleaners less susceptible to musculoskeletal disorders by physical coordination training or cognitive behavioral theory based training (CBTr). Because health-care workers are reported to have high prevalence of overweight and heavy lifts, the aim of the RCT is long-term weight-loss by combined physical exercise training, CBTr and diet. Construction work, characterized by heavy lifting, pushing and pulling, the RCT aims at improving physical capacity and promoting musculoskeletal and cardiovascular health. At the industrial work-place characterized by repetitive work tasks, the intervention aims at reducing physical exertion and musculoskeletal disorders by combined physical exercise training, CBTr and participatory ergonomics. The overall aim of the FINALE programme is to improve the safety margin between individual resources (i.e. physical capacities, and cognitive and behavioral skills) and physical work demands, and thereby reduce the physical deterioration in a long term perspective by interventions tailored for each respective job-group.</p> <p>Discussion</p> <p>The FINALE programme has the potential to provide evidence-based knowledge of significant importance for public health policy and health promotion strategies for employees at high risk for physical deterioration.</p> <p>Trial registrations</p> <p>ISRCTN96241850, NCT01015716 and NCT01007669</p

Modest de novo Reactivation of Single HIV-1 Proviruses in Peripheral CD4+ T Cells by Romidepsin

A cure for human immunodeficiency virus (HIV-1) is restricted by the continued presence of a latent reservoir of memory CD4+ T cells with proviruses integrated into their DNA despite suppressive antiretroviral therapy (ART). A predominant strategy currently pursued in HIV-1 cure-related research is the “kick and kill” approach, where latency reversal agents (LRAs) are used to reactivate transcription from integrated proviruses. The premise of this approach is that “kicking” latent virus out of hiding allows the host immune system to recognize and kill infected cells. Clinical trials investigating the efficacy of LRAs, such as romidepsin, have shown that these interventions do induce transient spikes in viral RNA in HIV-1-infected individuals. However, since these trials failed to significantly reduce viral reservoir size or significantly delay time to viral rebound during analytical treatment interruptions, it is questioned how much each individual latent provirus is actually “kicked” to produce viral transcripts and/or proteins by the LRA. Here, we developed sensitive and specific digital droplet PCR-based assays with single-provirus level resolution. Combining these assays allowed us to interrogate the level of viral RNA transcripts from single proviruses in individuals on suppressive ART with or without concomitant romidepsin treatment. Small numbers of proviruses in peripheral blood memory CD4+ T cells were triggered to become marginally transcriptionally active upon romidepsin treatment. These novel assays can be applied retrospectively and prospectively in HIV-1 cure-related clinical trials to gain crucial insights into LRA efficacy at the single provirus level

Humanized NOG Mice for Intravaginal HIV Exposure and Treatment of HIV Infection

Humanized mice provide a sophisticated platform to study human immunodeficiency virus (HIV) virology and to test antiviral drugs. This protocol describes the establishment of a human immune system in adult NOG mice. Here, we explain all the practical steps from isolation of umbilical cord blood derived human CD34+ cells and their subsequent intravenous transplantation into the mice, to the manipulation of the model through HIV infection, combination antiretroviral therapy (cART), and blood sampling. Approximately 75,000 hCD34+ cells are injected intravenously into the mice and the level of human chimerism, also known as humanization, in the peripheral blood is estimated longitudinally for months by flow cytometry. A total of 75,000 hCD34+ cells yields 20%–50% human CD45+ cells in the peripheral blood. The mice are susceptible to intravaginal infection with HIV and blood can be sampled once weekly for analysis, and twice monthly for extended periods. This protocol describes an assay for quantification of plasma viral load using droplet digital PCR (ddPCR). We show how the mice can be effectively treated with a standard-of- care cART regimen in the diet. The delivery of cART in the form of regular mouse chow is a significant refinement of the experimental model. This model can be used for preclinical analysis of both systemic and topical pre-exposure prophylaxis compounds as well as for testing of novel treatments and HIV cure strategies

Characterization of Intact Proviruses in Blood and Lymph Node from HIV-Infected Individuals Undergoing Analytical Treatment Interruption.

The role of lymphoid tissue as a potential source of HIV-1 rebound following interruption of antiretroviral therapy (ART) is uncertain. To address this issue, we compared the latent viruses obtained from CD4+ T cells in peripheral blood and lymph nodes to viruses emerging during treatment interruption. Latent viruses were characterized by sequencing near-full-length (NFL) proviral DNA and env from viral outgrowth assays (VOAs). Five HIV-1-infected individuals on ART were studied, four of whom participated in a clinical trial of a TLR9 agonist that included an analytical treatment interruption. We found that 98% of intact or replication-competent clonal sequences overlapped between blood and lymph node. In contrast, there was no overlap between 205 latent reservoir and 125 rebound sequences in the four individuals who underwent treatment interruption. However, rebound viruses could be accounted for by recombination. The data suggest that CD4+ T cells carrying latent viruses circulate between blood and lymphoid tissues in individuals on ART and support the idea that recombination may play a role in the emergence of rebound viremia.IMPORTANCE HIV-1 persists as a latent infection in CD4+ T cells that can be found in lymphoid tissues in infected individuals during ART. However, the importance of this tissue reservoir and its contribution to viral rebound upon ART interruption are not clear. In this study, we sought to compare latent HIV-1 from blood and lymph node CD4+ T cells from five HIV-1-infected individuals. Further, we analyzed the contribution of lymph node viruses to viral rebound. We observed that the frequencies of intact proviruses were the same in blood and lymph node. Moreover, expanded clones of T cells bearing identical proviruses were found in blood and lymph node. These latent reservoir sequences did not appear to be the direct origin of rebound virus. Instead, latent proviruses were found to contribute to the rebound compartment by recombination

Administration of Panobinostat Is Associated with Increased IL-17A mRNA in the Intestinal Epithelium of HIV-1 Patients

Intestinal CD4+ T cell depletion is rapid and profound during early HIV-1 infection.This leads to a compromised mucosal barrier that prompts chronic systemic inflammation.The preferential loss of intestinal T helper 17 (Th17) cells in HIV-1 disease is a driver of the damage within the mucosal barrier and of disease progression.Thus, understanding the effects of new therapeutic strategies in the intestines has high priority. Histone deacetylase (HDAC) inhibitors (e.g., panobinostat) are actively under investigation as potential latency reversing agents in HIV eradication studies. These drugs have broad effects that go beyond reactivating virus, including modulation of immune pathways. We examined colonic biopsies from ART suppressed HIV-1 infected individuals (clinicaltrials.gov: NCT01680094) for the effects of panobinostat on intestinal T cell activation and on inflammatory cytokine production. We compared biopsy samples that were collected before and during oral panobinostat treatment and observed that panobinostat had a clear biological impact in this anatomical compartment. Specifically, we observed a decrease in CD69+ intestinal lamina propria T cell frequency and increased IL-17A mRNA expression in the intestinal epithelium. These results suggest that panobinostat therapy may influence the restoration of mucosal barrier function in these patients

Endotoxemia Is Associated with Altered Innate and Adaptive Immune Responses in Untreated HIV-1 Infected Individuals

BACKGROUND: Microbial translocation may contribute to the immunopathogenesis in HIV infection. We investigated if microbial translocation and inflammation were associated with innate and adaptive immune responses in adults with HIV. METHODOLOGY/PRINCIPAL FINDINGS: This was an observational cohort study. Sera from HIV-infected and HIV-uninfected individuals were analyzed for microbial translocation (soluble CD14, lipopolysaccharides [LPS], endotoxin core antibody, and anti-α-galactosyl antibodies) and inflammatory markers (high sensitivity C-reactive protein, IL-6, IL-1 receptor antagonist, soluble tumor necrosis factor receptor II, and IL-10) with enzyme-linked immunosorbent assays. Peripheral blood mononuclear cells (PBMC) from HIV-infected persons and healthy controls (primed with single-stranded HIV-1-derived RNA) were stimulated with LPS, and cytokine production was measured. Finally, HIV-infected patients were immunized with Prevnar 7vPnC±CpG 7909 followed by Pneumo Novum PPV-23. Effects of microbial translocation and inflammation on immunization were analyzed in a predictive regression model. We included 96 HIV-infected individuals, 76 on highly active antiretroviral therapy (HAART), 20 HAART-naive, and 50 healthy controls. Microbial translocation and inflammatory markers were higher among HIV-infected persons than controls. Cytokine levels following LPS stimulation were increased in PBMCs from HAART-naive compared to HAART-treated HIV-infected persons. Further, RNA-priming of PBMCs from controls acted synergistically with LPS to augment cytokine responses. Finally, high serum LPS levels predicted poor vaccine responses among HAART-naive, but not among HAART-treated HIV-infected individuals. CONCLUSIONS/SIGNIFICANCE: LPS acts synergistically with HIV RNA to stimulate innate immune responses in vitro and increasing serum LPS levels seem to predict poor antibody responses after vaccination among HAART-naive HIV-infected persons. Thus, our results suggest that microbial translocation may be associated with innate and adaptive immune dysfunction in untreated HIV infection