Intra-Host Evolution Analyses in an Immunosuppressed Patient Supports SARS-CoV-2 Viral Reservoir Hypothesis.

Throughout the SARS-CoV-2 pandemic, several variants of concern (VOCs) have been identified, many of which share recurrent mutations in the spike glycoprotein's receptor-binding domain (RBD). This region coincides with known epitopes and can therefore have an impact on immune escape. Protracted infections in immunosuppressed patients have been hypothesized to lead to an enrichment of such mutations and therefore drive evolution towards VOCs. Here, we present the case of an immunosuppressed patient that developed distinct populations with immune escape mutations throughout the course of their infection. Notably, by investigating the co-occurrence of substitutions on individual sequencing reads in the RBD, we found quasispecies harboring mutations that confer resistance to known monoclonal antibodies (mAbs) such as S:E484K and S:E484A. These mutations were acquired without the patient being treated with mAbs nor convalescent sera and without them developing a detectable immune response to the virus. We also provide additional evidence for a viral reservoir based on intra-host phylogenetics, which led to a viral substrain that evolved elsewhere in the patient's body, colonizing their upper respiratory tract (URT). The presence of SARS-CoV-2 viral reservoirs can shed light on protracted infections interspersed with periods where the virus is undetectable, and potential explanations for long-COVID cases.

Metformin Enhances Antibody-Mediated Recognition of HIV-Infected CD4+ T-Cells by Decreasing Viral Release.

The mechanistic target of rapamycin (mTOR) positively regulates multiple steps of the HIV-1 replication cycle. We previously reported that a 12-weeks supplementation of antiretroviral therapy (ART) with metformin, an indirect mTOR inhibitor used in type-2 diabetes treatment, reduced mTOR activation and HIV transcription in colon-infiltrating CD4+ T-cells, together with systemic inflammation in nondiabetic people with HIV-1 (PWH). Herein, we investigated the antiviral mechanisms of metformin. In a viral outgrowth assay performed with CD4+ T-cells from ART-treated PWH, and upon infection in vitro with replication-competent and VSV-G-pseudotyped HIV-1, metformin decreased virion release, but increased the frequency of productively infected CD4lowHIV-p24+ T-cells. These observations coincided with increased BST2/Tetherin (HIV release inhibitor) and Bcl-2 (pro-survival factor) expression, and improved recognition of productively infected T-cells by HIV-1 Envelope antibodies. Thus, metformin exerts pleiotropic effects on post-transcription/translation steps of the HIV-1 replication cycle and may be used to accelerate viral reservoir decay in ART-treated PWH.

Gaining Biological Insights through Supervised Data Visualization.

Dimensionality reduction-based data visualization is pivotal in comprehending complex biological data. The most common methods, such as PHATE, t-SNE, and UMAP, are unsupervised and therefore reflect the dominant structure in the data, which may be independent of expert-provided labels. Here we introduce a supervised data visualization method called RF-PHATE, which integrates expert knowledge for further exploration of the data. RF-PHATE leverages random forests to capture intricate featurelabel relationships. Extracting information from the forest, RF-PHATE generates low-dimensional visualizations that highlight relevant data relationships while disregarding extraneous features. This approach scales to large datasets and applies to classification and regression. We illustrate RF-PHATE's prowess through three case studies. In a multiple sclerosis study using longitudinal clinical and imaging data, RF-PHATE unveils a sub-group of patients with non-benign relapsingremitting Multiple Sclerosis, demonstrating its aptitude for time-series data. In the context of Raman spectral data, RF-PHATE effectively showcases the impact of antioxidants on diesel exhaust-exposed lung cells, highlighting its proficiency in noisy environments. Furthermore, RF-PHATE aligns established geometric structures with COVID-19 patient outcomes, enriching interpretability in a hierarchical manner. RF-PHATE bridges expert insights and visualizations, promising knowledge generation. Its adaptability, scalability, and noise tolerance underscore its potential for widespread adoption.

CD8 + T-cell priming is quantitatively but not qualitatively impaired in people with HIV-1 on antiretroviral therapy.

BACKGROUND: The induction of de novo CD8 + T-cell responses is essential for protective antiviral immunity, but this process is often impaired in people with HIV-1 (PWH). We investigated the extent to which the immune competence of naive CD8 + T cells, a key determinant of priming efficacy, could be preserved or restored in PWH via long-term antiretroviral therapy (ART). METHODS: We used flow cytometry, molecular analyses of gene transcription and telomere length, and a fully validated priming assay to characterize naive CD8 + T cells ex vivo and evaluate the induction of antigen-specific effector/memory CD8 + T cells in vitro , comparing age-matched healthy uninfected donors (HUDs), PWH on ART, and natural HIV-1 controllers (HICs). RESULTS: We found that naive CD8 + T cells were numerically reduced and exhibited a trend toward shorter telomere lengths in PWH on ART compared with HUDs and HICs. These features associated with impaired priming efficacy. However, we also found that naive CD8 + T cells were fully equipped proliferatively and transcriptionally in PWH on ART, enabling the generation of antigen-specific effector/memory CD8 + T cells with functional and phenotypic attributes comparable to those primed from HUDs. CONCLUSION: Our data suggest that naive CD8 + T cells in PWH on ART are intrinsically capable of generating functionally and phenotypically intact effector/memory CD8 + T cells in response to antigen, despite evidence of senescence and an overall numerical reduction that compromises priming efficacy relative to HUDs and HICs.

DAA-mediated HCV cure reduces HIV DNA levels in HCV/HIV coinfected people.

IMPORTANCE: Antiretroviral therapy (ART) for human immunodeficiency virus (HIV) can control virus replication and prolong the life of people living with HIV (PLWH). However, the virus remains dormant within immune cells in what is called the HIV reservoir. Furthermore, 2.3 million PLWH are also coinfected with hepatitis C virus (HCV) and are at risk of developing chronic liver disease and cancer. HCV treatment with direct acting antivirals (DAA) can completely cure the infection in more than 95% of treated individuals and improve their long-term health outcomes. In this study, we investigated how HCV treatment and cure affect the HIV reservoir. We demonstrate the beneficial impact of DAA treatment as it reduces the HIV reservoirs in particular in people infected with HCV before HIV. These results support the need for early ART and DAA treatment in HIV/HCV coinfections.

Is the Central Nervous System Reservoir a Hurdle for an HIV Cure?

There is currently no cure for HIV infection although adherence to effective antiretroviral therapy (ART) suppresses replication of the virus in blood, increases CD4+ T-cell counts, reverses immunodeficiency, and increases life expectancy. Despite these substantial advances, ART is a lifelong treatment for people with HIV (PWH) and upon cessation or interruption, the virus quickly rebounds in plasma and anatomic sites, including the central nervous system (CNS), resulting in disease progression. With recent advances in quantifying viral burden, detection of genetically intact viral genomes, and isolation of replication-competent virus from brain tissues of PWH receiving ART, it has become apparent that the CNS viral reservoir (largely comprised of macrophage type cells) poses a substantial challenge for HIV cure strategies. Other obstacles impacting the curing of HIV include ageing populations, substance use, comorbidities, limited antiretroviral drug efficacy in CNS cells, and ART-associated neurotoxicity. Herein, we review recent findings, including studies of the proviral integration sites, reservoir decay rates, and new treatment/prevention strategies in the context of the CNS, together with highlighting the next steps for investigations of the CNS as a viral reservoir.

Plasmatic HIV-1 soluble gp120 is associated with correlates of immune dysfunction and inflammation in ART-treated individuals with undetectable viremia.

BACKGROUND: Chronic inflammation persists in some people living with HIV (PLWH) during antiretroviral therapy and is associated with premature aging. The gp120 subunit of HIV-1 envelope sheds and can be detected in plasma, showing immunomodulatory properties even in the absence of detectable viremia. We evaluated whether plasmatic soluble gp120 (sgp120) and a family of gp120-specific anti-cluster A antibodies, linked to CD4 depletion in vitro, contribute to chronic inflammation, immune dysfunction, and sub-clinical cardiovascular disease in participants of the Canadian HIV and Aging cohort (CHACS) with undetectable viremia. METHODS: Cross-sectional assessment of sgp120 and anti-cluster A antibodies was performed in 386 individuals from CHACS. Their association with pro-inflammatory cytokines and subclinical coronary artery disease was assessed using linear regression models. RESULTS: High levels of sgp120 and anti-cluster A antibodies inversely correlated with CD4 count and CD4:CD8 ratio. The presence of sgp120 was associated with increased levels of IL-6. In participants with detectable atherosclerotic plaque and detectable sgp120, anti-cluster A antibodies and their combination with sgp120 levels correlated positively with the total volume of atherosclerotic plaques. CONCLUSIONS: sgp120 may act as a pan toxin causing immune dysfunction and sustained inflammation in a subset of PLWH, contributing to the development of premature comorbidities.

Tracking coreceptor switch of the transmitted/founder HIV-1 identifies co-evolution of HIV-1 antigenicity, coreceptor usage and CD4 subset targeting: the RV217 acute infection cohort study.

BACKGROUND: The CCR5 (R5) to CXCR4 (X4) coreceptor switch in natural HIV-1 infection is associated with faster progression to AIDS, but the mechanisms remain unclear. The difficulty in elucidating the evolutionary origin of the earliest X4 viruses limits our understanding of this phenomenon. METHODS: We tracked the evolution of the transmitted/founder (T/F) HIV-1 in RV217 participants identified in acute infection. The origin of the X4 viruses was elucidated by single genome amplification, deep sequencing and coreceptor assay. Mutations responsible for coreceptor switch were confirmed by mutagenesis. Viral susceptibility to neutralization was determined by neutralization assay. Virus CD4 subset preference was demonstrated by sequencing HIV-1 RNA in sorted CD4 subsets. FINDINGS: We demonstrated that the earliest X4 viruses evolved de novo from the T/F strains. Strong X4 usage can be conferred by a single mutation. The mutations responsible for coreceptor switch can confer escape to neutralization and drive the X4 variants to replicate mainly in the central memory (CM) and naïve CD4 subsets. Likely due to the smaller viral burst size of the CM and naïve subsets, the X4 variants existed at low frequency in plasma. The origin of the X4 viruses preceded accelerated CD4 decline. All except one X4 virus identified in the current study lost the conserved V3 N301 glycan site. INTERPRETATIONS: The findings demonstrate co-evolution of HIV-1 antigenicity, coreceptor usage and CD4 subset targeting which have implications for HIV-1 therapeutics and functional cure. The observations provide evidence that coreceptor switch can function as an evolutionary mechanism of immune evasion. FUNDING: Institute of Human Virology, National Institutes of Health, Henry M. Jackson Foundation for the Advancement of Military Medicine, Inc, Thai Red Cross AIDS Research Centre, Gilead Sciences, Merck, and ViiV Healthcare.

Plasma SARS-CoV-2 RNA elimination and RAGE kinetics distinguish COVID-19 severity.

Objectives: Identifying biomarkers causing differential SARS-CoV-2 infection kinetics associated with severe COVID-19 is fundamental for effective diagnostics and therapeutic planning. Methods: In this work, we applied mathematical modelling to investigate the relationships between patient characteristics, plasma SARS-CoV-2 RNA dynamics and COVID-19 severity. Using a straightforward mathematical model of within-host viral kinetics, we estimated key model parameters from serial plasma viral RNA (vRNA) samples from 256 hospitalised COVID-19+ patients. Results: Our model predicted that clearance rates distinguish key differences in plasma vRNA kinetics and severe COVID-19. Moreover, our analyses revealed a strong correlation between plasma vRNA kinetics and plasma receptor for advanced glycation end products (RAGE) concentrations (a plasma biomarker of lung damage), collected in parallel to plasma vRNA from patients in our cohort, suggesting that RAGE can substitute for viral plasma shedding dynamics to prospectively classify seriously ill patients. Conclusion: Overall, our study identifies factors of COVID-19 severity, supports interventions to accelerate viral clearance and underlines the importance of mathematical modelling to better understand COVID-19.

Transmission of highly virulent CXCR4 tropic HIV-1 through the mucosal route in an individual with a wild-type CCR5 genotype.

Nearly all transmitted/founder (T/F) HIV-1 are CCR5 (R5)-tropic. While previous evidence suggested that CXCR4 (X4)-tropic HIV-1 are transmissible, detection was not at the earliest stages of acute infection. Here, we identified an X4-tropic T/F HIV-1 in a participant in acute infection cohort. Coreceptor assays demonstrated that this T/F virus is strictly CXCR4 tropic. The participant experienced significantly faster CD4 depletion compared with R5 virus infected participants in the same cohort. Naïve and central memory CD4 subsets declined faster than effector and transitional memory subsets. All CD4 subsets, including naïve, were productively infected. Increased CD4+ T cell activation was observed over time. This X4-tropic T/F virus is resistant to broadly neutralizing antibodies (bNAbs) targeting V1/V2 and V3 regions. These findings demonstrate that X4-tropic HIV-1 is transmissible through the mucosal route in people with the wild-type CCR5 genotype and have implications for understanding the transmissibility and immunopathogenesis of X4-tropic HIV-1.

Measuring HIV reservoirs: Do we need to choose between quantity and quality?

The persistence of latent viral genomes in people receiving antiretroviral therapy (ART) is the main obstacle to a cure for HIV infection. Viral reservoirs can be defined as cells harboring HIV genomes that have the ability to produce infectious virions. Precise quantification of the cellular reservoirs of HIV is challenging because these cells are rare, heterogeneous and outnumbered by a larger number of cells carrying defective genomes. In addition, measuring the inducibility of these proviruses requires functional assays and remains technically difficult. The recent development of single cell and single viral genome approaches revealed additional layers of complexity: the cell subsets that harbor proviruses are heterogeneous and their ability to be induced is variable. A substantial fraction of intact HIV genomes may be permanently silenced after years of ART, revealing the underappreciated importance of induction assays. As such, a simple approach that would assess simultaneously the genetic intactness and the inducibility of the reservoir is still lacking. In here, we review recent advances in the development of methods to quantify and characterize persistently infected cells, and discuss how these findings can inform the design of future assays aimed at measuring the size of the intact and inducible HIV reservoir.

Donating One's Body to HIV Cure Research Through Canadian Medical Assistance in Dying: A Case Study.

BACKGROUND: Finding a cure for HIV is challenged by persisting reservoirs, the mapping of which necessitates invasive procedures. Inviting people with HIV (PWHIV) at the end of life to donate body specimens post-mortem through research autopsies is a novel approach, raising ethical concerns. OBJECTIVE: This case study aims to explore the motivations, barriers, and facilitators of a terminally-ill Canadian PWHIV who requested medical assistance in dying (MAID) and expressed interest in donating his body for HIV cure research. CASE PRESENTATION: An in-depth 3-hour and semi-structured interview was conducted with the participant. The interview transcription was thematically coded to identify motivations and perceived barriers and facilitators to participate in end-of-life HIV cure research. Our analysis identified six themes. Two themes expressed motivations: Collaboration in progress in health and science, seeing cure research as collaboration with professionals; and Opportunity to learn more, mostly about science and health. One theme expressed a barrier: Losing interest in or identification with long-term care research matters, especially those related to the management of long-term care. Three themes expressed by facilitators: Receiving information from professionals one trusts and knows, especially clinical and research teams; Perceiving research procedures as simple, useful, and embedded in care, perceiving clinical, educational, and interpersonal benefits that surpass costs of participation; and Perceiving research as one last way to contribute, that is, feeling useful or give back. CONCLUSION: Several circumstances facilitated the patient's participation: being a single man, having time to participate, having no strong religious belief, and valuing clear, direct communication. His motivations to participate in HIV cure research were altruistic, and also an experience of working with clinical and research teams. Finally, this perspective highlights HIV cure research participant candidates' need for education about research procedures.

Near full-length HIV sequencing in multiple tissues collected postmortem reveals shared clonal expansions across distinct reservoirs during ART.

HIV persists in tissues during antiretroviral therapy (ART), but the relative contribution of different anatomical compartments to the viral reservoir in humans remains unknown. We performed an extensive characterization of HIV reservoirs in two men who donated their bodies to HIV cure research and who had been on suppressive ART for years. HIV DNA is detected in all tissues, with large variations across anatomical compartments and between participants. Intact HIV genomes represent 2% and 25% of all proviruses in the two participants and are mainly detected in secondary lymphoid organs, with the spleen and mediastinal lymph nodes harboring intact viral genomes in both individuals. Multiple copies of identical HIV genomes are found in all tissues, indicating that clonal expansions are common in anatomical sites. The majority (>85%) of these expanded clones are shared across multiple tissues. These findings suggest that infected cells expand, migrate, and possibly circulate between anatomical sites.

An active HIV reservoir during ART is associated with maintenance of HIV-specific CD8+ T cell magnitude and short-lived differentiation status.

Before initiation of antiretroviral therapy (ART), HIV-specific CD8+ T cells are dysfunctional and short lived. To better understand the relationship between the HIV reservoir in CD4+ T cells and the magnitude and differentiation status of HIV-specific CD8+ T cells, we investigated these cells from acute and chronic HIV-infected individuals after 2 years of ART. Although both the HIV reservoir and the CD8+ T cell responses declined significantly after 2 years of ART, sustained HIV-specific CD8+ T cell responses correlated with a greater reduction of integrated HIV provirus. However, the magnitude of CD8+ T cells specific for HIV Gag, Pol, Nef, and Vif proteins positively associated with the active reservoir size during ART, measured as cell-associated RNA. Importantly, high HIV DNA levels strongly associate with maintenance of short-lived HIV-specific CD8+ T cells, regardless of ART initiation time. Our data suggest that the active reservoir maintains HIV-specific CD8+ T cell magnitude but prevents their differentiation into functional cells.

Spontaneous HIV expression during suppressive ART is associated with the magnitude and function of HIV-specific CD4+ and CD8+ T cells.

Spontaneous transcription and translation of HIV can persist during suppressive antiretroviral therapy (ART). The quantity, phenotype, and biological relevance of this spontaneously "active" reservoir remain unclear. Using multiplexed single-cell RNAflow-fluorescence in situ hybridization (FISH), we detect active HIV transcription in 14/18 people with HIV on suppressive ART, with a median of 28/million CD4+ T cells. While these cells predominantly exhibit abortive transcription, p24-expressing cells are evident in 39% of participants. Phenotypically diverse, active reservoirs are enriched in central memory T cells and CCR6- and activation-marker-expressing cells. The magnitude of the active reservoir positively correlates with total HIV-specific CD4+ and CD8+ T cell responses and with multiple HIV-specific T cell clusters identified by unsupervised analysis. These associations are particularly strong with p24-expressing active reservoir cells. Single-cell vDNA sequencing shows that active reservoirs are largely dominated by defective proviruses. Our data suggest that these reservoirs maintain HIV-specific CD4+ and CD8+ T responses during suppressive ART.

Transmission of highly virulent CXCR4 tropic HIV-1 through the mucosal route in an individual with a wild-type CCR5 genotype.

Nearly all transmitted/founder (T/F) HIV-1 are CCR5 (R5)-tropic. While previous evidence suggested that CXCR4 (X4)-tropic HIV-1 are transmissible, detection was not at the earliest stages of acute infection. Here, we identified an X4-tropic T/F HIV-1 in a participant in acute infection cohort. Coreceptor assays demonstrated that this T/F virus is strictly CXCR4 tropic. The participant experienced significantly faster CD4 depletion compared with R5 virus infected participants in the same cohort. Naïve and central memory CD4 subsets declined faster than effector and transitional memory subsets. All CD4 subsets, including naïve, were productively infected. Increased CD4 + T cell activation was observed over time. This X4-tropic T/F virus is resistant to broadly neutralizing antibodies (bNAbs) targeting V1/V2 and V3 regions. These findings demonstrate that X4-tropic HIV-1 is transmissible through the mucosal route in people with the wild-type CCR5 genotype and have implications for understanding the transmissibility and immunopathogenesis of X4-tropic HIV-1.

Plasmatic HIV-1 soluble gp120 is associated with immune dysfunction and inflammation in ART-treated individuals with undetectable viremia.

Background: Chronic inflammation persists in some people living with HIV (PLWH), even during antiretroviral therapy (ART) and is associated with premature aging. The gp120 subunit of the HIV-1 envelope glycoprotein can shed from viral and cellular membranes and can be detected in plasma and tissues, showing immunomodulatory properties even in the absence of detectable viremia. We evaluated whether plasmatic soluble gp120 (sgp120) and a family of gp120-specific anti-cluster A antibodies, which were previously linked to CD4 depletion in vitro , could contribute to chronic inflammation, immune dysfunction, and sub-clinical cardiovascular disease in participants of the Canadian HIV and Aging cohort (CHACS) with undetectable viremia. Methods: Cross-sectional assessment of plasmatic sgp120 and anti-cluster A antibodies was performed in 386 individuals from CHACS. Their association with pro-inflammatory cytokines, as well as subclinical coronary artery disease measured by computed tomography coronary angiography was assessed using linear regression models. Results: In individuals with high levels of sgp120, anti-cluster A antibodies inversely correlated with CD4 count (p=0.042) and CD4:CD8 ratio (p=0.004). The presence of sgp120 was associated with increased plasma levels of IL-6. In participants with detectable atherosclerotic plaque and detectable sgp120, sgp120 levels, anti-cluster A antibodies and their combination correlated positively with the total volume of atherosclerotic plaques (p=0.01, 0.018 and 0.006, respectively). Conclusion: Soluble gp120 may act as a pan toxin causing immune dysfunction and sustained inflammation in a subset of PLWH, contributing to the development of premature comorbidities. Whether drugs targeting sgp120 could mitigate HIV-associated comorbidities in PLWH with suppressed viremia warrants further studies. Key points: Soluble gp120 is detected in the plasma of people living with HIV-1 with undetectable viremia. The presence of soluble gp120 and anti-cluster A antibodies is associated with immune dysfunction, chronic inflammation, and sub-clinical cardiovascular disease.

Effects of Oral Cannabinoids on Systemic Inflammation and Viral Reservoir Markers in People with HIV on Antiretroviral Therapy: Results of the CTN PT028 Pilot Clinical Trial.

Chronic HIV infection is characterized by persistent inflammation despite antiretroviral therapy (ART). Cannabinoids may help reduce systemic inflammation in people with HIV (PWH). To assess the effects of oral cannabinoids during HIV, ten PWH on ART were randomized (n = 5/group) to increasing doses of oral Δ9-tetrahydrocannabinol (THC): cannabidiol (CBD) combination (2.5:2.5-15:15 mg/day) capsules or CBD-only (200-800 mg/day) capsules for 12 weeks. Blood specimens were collected prospectively 7-21 days prior to treatment initiation and at weeks 0 to 14. Plasma cytokine levels were determined via Luminex and ELISA. Immune cell subsets were characterized by flow cytometry. HIV DNA/RNA were measured in circulating CD4 T-cells and sperm by ultra-sensitive qPCR. Results from both arms were combined for statistical analysis. Plasma levels of IFN-γ, IL-1ß, sTNFRII, and REG-3α were significantly reduced at the end of treatment (p ˂ 0.05). A significant decrease in frequencies of PD1+ memory CD4 T-cells, CD73+ regulatory CD4 T-cells, and M-DC8+ intermediate monocytes was also observed (p ˂ 0.05), along with a transient decrease in CD28-CD57+ senescent CD4 and CD8 T-cells. Ki-67+ CD4 T-cells, CCR2+ non-classical monocytes, and myeloid dendritic cells increased over time (p ˂ 0.05). There were no significant changes in other inflammatory markers or HIV DNA/RNA levels. These findings can guide future large clinical trials investigating cannabinoid anti-inflammatory properties.

Soluble immune checkpoints as correlates for HIV persistence and T cell function in people with HIV on antiretroviral therapy.

Introduction: In people with HIV (PWH) both off and on antiretroviral therapy (ART), the expression of immune checkpoint (IC) proteins is elevated on the surface of total and HIV-specific T-cells, indicating T-cell exhaustion. Soluble IC proteins and their ligands can also be detected in plasma, but have not been systematically examined in PWH. Since T-cell exhaustion is associated with HIV persistence on ART, we aimed to determine if soluble IC proteins and their ligands also correlated with the size of the HIV reservoir and HIV-specific T-cell function. Methods: We used multiplex bead-based immunoassay to quantify soluble programmed cell death protein 1 (PD-1), cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), lymphocyte activation gene-3 (LAG-3), T cell immunoglobulin domain and mucin domain 3 (TIM-3), PD-1 Ligand 1 (PD-L1) and PD-1 Ligand 2 (PD-L2) in plasma from PWH off ART (n=20), on suppressive ART (n=75) and uninfected controls (n=20). We also quantified expression of membrane-bound IC and frequencies of functional T-cells to Gag and Nef peptide stimulation on CD4+ and CD8+ T-cells using flow cytometry. The HIV reservoir was quantified in circulating CD4+ T-cells using qPCR for total and integrated HIV DNA, cell-associated unspliced HIV RNA and 2LTR circles. Results: Soluble (s) PD-L2 level was higher in PWH off and on ART compared to uninfected controls. Higher levels of sPD-L2 correlated with lower levels of HIV total DNA and higher frequencies of gag-specific CD8+ T-cells expressing CD107a, IFNγ or TNFα. In contrast, the concentration of sLAG-3 was similar in uninfected individuals and PWH on ART, but was significantly elevated in PWH off ART. Higher levels of sLAG-3 correlated with higher levels of HIV total and integrated DNA, and lower frequency of gag-specific CD4+ T cells expressing CD107a. Similar to sLAG-3, levels of sPD-1 were elevated in PWH off ART and normalized in PWH on ART. sPD-1 was positively correlated with the frequency of gag-specific CD4+ T cells expressing TNF-a and the expression of membrane-bound PD-1 on total CD8+ T-cells in PWH on ART. Discussion: Plasma soluble IC proteins and their ligands correlate with markers of the HIV reservoir and HIV-specific T-cell function and should be investigated further in in large population-based studies of the HIV reservoir or cure interventions in PWH on ART.

Primary role of type I interferons for the induction of functionally optimal antigen-specific CD8+ T cells in HIV infection.

BACKGROUND: CD8+ T cells equipped with a full arsenal of antiviral effector functions are critical for effective immune control of HIV-1. It has nonetheless remained unclear how best to elicit such potent cellular immune responses in the context of immunotherapy or vaccination. HIV-2 has been associated with milder disease manifestations and more commonly elicits functionally replete virus-specific CD8+ T cell responses compared with HIV-1. We aimed to learn from this immunological dichotomy and to develop informed strategies that could enhance the induction of robust CD8+ T cell responses against HIV-1. METHODS: We developed an unbiased in vitro system to compare the de novo induction of antigen-specific CD8+ T cell responses after exposure to HIV-1 or HIV-2. The functional properties of primed CD8+ T cells were assessed using flow cytometry and molecular analyses of gene transcription. FINDINGS: HIV-2 primed functionally optimal antigen-specific CD8+ T cells with enhanced survival properties more effectively than HIV-1. This superior induction process was dependent on type I interferons (IFNs) and could be mimicked via the adjuvant delivery of cyclic GMP-AMP (cGAMP), a known agonist of the stimulator of interferon genes (STING). CD8+ T cells elicited in the presence of cGAMP were polyfunctional and highly sensitive to antigen stimulation, even after priming from people living with HIV-1. INTERPRETATION: HIV-2 primes CD8+ T cells with potent antiviral functionality by activating the cyclic GMP-AMP synthase (cGAS)/STING pathway, which results in the production of type I IFNs. This process may be amenable to therapeutic development via the use of cGAMP or other STING agonists to bolster CD8+ T cell-mediated immunity against HIV-1. FUNDING: This work was funded by INSERM, the Institut Curie, and the University of Bordeaux (Senior IdEx Chair) and by grants from Sidaction (17-1-AAE-11097, 17-1-FJC-11199, VIH2016126002, 20-2-AEQ-12822-2, and 22-2-AEQ-13411), the Agence Nationale de la Recherche sur le SIDA (ECTZ36691, ECTZ25472, ECTZ71745, and ECTZ118797), and the Fondation pour la Recherche Médicale (EQ U202103012774). D.A.P. was supported by a Wellcome Trust Senior Investigator Award (100326/Z/12/Z).