Biology subjects

Jones, R. B.

Publications and source records attributed to Jones, R. B..

At least 19 recordsLinked to original sources

bioRxiv2026

Potent and Broad HIV-1 Neutralization by a Bispecific CD4-CD4i Fusion Protein based on Single-Domain CD4-D1 and X5 CD4i antibody

Human immunodeficiency virus (HIV) infection remains a global health threat. Although antiretroviral therapy (ART) has significantly transformed HIV into a manageable chronic disease, emergence of drug resistance to current ART is a continuing concern. Broadly neutralizing antibodies, as well as reagents containing both a soluble CD4 mimetic and an HIV co-receptor inhibitor--such as CD4-CD4i antibodies--are promising strategies for the prevention and treatment of HIV infection. We previously developed a CD4 D1mimetic (mD1.22) with enhanced neutralization potency, surpassing that of the clinically validated sCD4-D1D2 mimetic. We also previously identified a novel CD4i antibody (X5) that targets the conserved coreceptor binding site on the gp120 core and recognizes an epitope partially overlapping with 17b monoclonal antibody binding site. X5 binding to gp120 was augmented by CD4 and modestly enhanced by CCR5. To leverage these favorable interactions, we designed and optimized sCD4-X5 bispecific antibodies by computational structure-aided modification of antibody size and fusion linkers between the two binding moieties. The bispecific D1X5, with a (G4S)7 long linker between D1 and X5 (IgG1-LL D1X5), exhibited broad neutralization against 11 diverse HIV subtypes across B, C, G clades and AC, BC recombinants. The TZM-bl cell neutralization assay showed IgG1-LL D1X5 neutralization geometric mean IC50 and IC80 are 0.6 g/mL and 3.4 g/mL respectively, which are within the range of potent bnAbs. This work has identified a novel single domain soluble CD4 based CD4-CD4i bispecific antibody with broad HIV-1 neutralization.

molecular biology↗

bioRxiv2026

Loss of Vpr-driven TRAIL-R2 expression protects HIV-infected cells from non-canonical NK cell TRAIL attack

HIV escapes sterilizing immunity through a variety of mechanisms, including the downregulation of MHC-I expression by HIV Nef and Vpu to counteract CD8+ T cell responses. While reduced MHC-I expression would be expected to support targeting by NK cells, a subpopulation of infected CD4+ T cells consistently resists multiple rounds of NK cell natural and antibody-dependent cytotoxicity. Studies further reveal that the HIV accessory protein Vpr induces expression of TNFRSF10B (TRAIL-R2) in CD4+ T cells, with survivors of NK cell targeting exhibiting relatively higher MHC-I and weaker expression of TRAIL-R2. In fact, reverse TRAIL signaling in NK cells leads to the release of perforin and granzymes, a pathway limited when TRAIL-R2 expression is diminished. Thus, independent of canonical death receptor signaling, TRAIL-R2 serves as an activating ligand that augments NK cell killing. These observations demonstrate that through Vpr, HIV can regulate the TRAIL/TRAIL-R2 axis to control NK cell functionality.

immunology↗

bioRxiv2026

HIV superinfection reveals sequential reservoir reactivation and immune-driven rebound dynamics

Superinfection, where a person with HIV acquires a second phylogenetically distinct strain, provides an opportunity to study reservoir evolutionary dynamics because the two strains can be tracked over time. We here characterize such a case that originally went undetected by clinical monitoring. The participant, who expressed the protective HLA-B*57:03 allele, initially controlled his subtype B infection but lost control after superinfection with a unique recombinant form (URF), which came to dominate in plasma without displacing the original B strain. Five years after initiating therapy, replication-competent proviruses from both strains persisted in blood, though URF proviruses dominated (80%). Despite their unequal reservoir representation, both strains rapidly rebounded upon treatment interruption, though most rebounding sequences belonged to a URF subclade that predated ART initiation. During the treatment interruption, genetically distinct viral lineages continually appeared in plasma, consistent with sequential "waves" of reactivation of diverse reservoir clones. Notably, immune escape variants emerged from the reservoir in a later "wave", displacing initial susceptible variants and underscoring the important role of immune responses in shaping rebound dynamics. Sequence analysis revealed that the URF transmitted/founder virus already harbored key HLA-B*57:03-associated escape mutations, likely explaining the loss of viral control after superinfection. Our results reveal that superinfection can remain undetected despite routine HIV clinical monitoring, that pre-adaptation of the superinfecting strain to host HLA can undermine immune control, that rebound virus can originate from reservoir clones predating ART initiation, and that immune responses can actively shape rebound dynamics by driving escape variant outgrowth during treatment interruption. IMPORTANCEThis case demonstrates that HIV superinfection can go undetected by routine clinical monitoring, underscoring the importance of assays that capture HIV diversity -- particularly those used to screen participants for cure trials. It also illustrates how pre-adaptation of a superinfecting strain to host HLA-restricted immune responses can evade existing immunity, with implications for understanding natural immune control, and underscoring the challenge of viral diversity for vaccine and immunotherapy design. The sequential emergence of genetically diverse HIV lineages during treatment interruption, including minority immune escape variants that emerged from the reservoir to displace the initial susceptible population, highlights the difficulty of predicting rebound virus composition and underscores the key role of immune responses in shaping rebound dynamics. Critically, escaped rebound variants can then reseed the reservoir, thereby enriching it in escaped forms, highlighting a specific vulnerability for cure approaches that harness natural immune responses to eliminate reservoir cells.

molecular biology↗

bioRxiv2026

Dynamic Antigen Expression and Intrinsic CTL Resistance in HIV Reservoir Clones

Reservoirs of clonally expanded CD4+ T-cells harboring rebound-competent HIV proviruses persist lifelong during ART. Latency is considered the principal barrier to viral eradication and has resisted pharmacological reversal, yet it appears that sustained immune pressure may still erode reservoirs. Recent advances have yielded glimpses into these exceptionally rare reservoir-harboring cells, implicating intrinsic pro-survival properties in their persistence. Here, we isolate and characterize populations of authentic reservoir clones (ARCs) that robustly proliferate and accumulate while producing infectious virus, without overtly succumbing to viral cytopathic effects. At any given moment, only small fractions of ARCs expressed HIV proteins, a state remarkably unperturbed by potent TCR or mitogenic stimulation. Nevertheless, sustained co-culture with cytotoxic T-lymphocytes (CTL) revealed extensive time-integrated antigenic vulnerability, culling clonal expansion of some ARCs by >90%. Notably, a regulatory T-cell ARC displayed pronounced cell-intrinsic resistance to CTL - a longstanding hypothesis we now directly demonstrate - linked to low oxidative stress and reversed with desferoxamine, a hypoxic stress inducer and FDA-approved therapeutic. Overall, we provide novel insights into the vulnerabilities of reservoir clones to potent, sustained CTL pressure and highlight intrinsic resistance pathways as actionable therapeutic targets, opening opportunities for advancing immune-based HIV cure strategies.

immunology↗

bioRxiv2025

HIV Nef amplifies mechanical heterogeneity to promote immune evasion

Intracellular pathogens must evade cytotoxic immunity to establish persistent infection. Although immune escape is typically viewed through a biochemical lens, the ability of certain pathogens to alter the mechanical properties of infected cells suggests that biophysical mechanisms may also contribute to the process. Here, we show that a subset of CD4+ T cells infected with the human immunodeficiency virus (HIV) resist elimination through a soft phenotype that inhibits killing by mechanosensitive cytotoxic T lymphocytes (CTLs). This phenotype arises from the combined effects of the HIV virulence factor Nef, which remodels the actin cytoskeleton, and intrinsic heterogeneity in the basal cytoskeletal properties of infected T cells. Pharmacological or genetic perturbations that reverse Nef signaling to the cytoskeleton or that stiffen the filamentous-actin cortex sensitize infected cells to CTL-mediated lysis. Taken together, these findings define a novel, biophysical paradigm of immune evasion with implications for HIV cure strategies.

immunology↗

bioRxiv2025

HIV-1 Integration Site Determines the Transcriptional Fate and Persistence of Integrated Proviruses

The mechanisms by which latent HIV-1 reservoirs persist during antiretroviral therapy is incompletely understood. Here, we derive a model system to measure clonal expansion and viral latency in which populations of human memory CD4+ T cells, each bearing a single transcriptionally active HIV-1 provirus are engrafted into immunodeficient mice. Over [~]2 months in vivo, clonal expansion and the establishment of latency occurred in subsets of engrafted infected cells. Clonal expansion in vivo was driven by T-cell receptor identity, but not by proviral insertional mutagenesis. The integration sites of proviruses that became latent in vivo were enriched on chromosome 19, in intergenic and centromeric satellite regions, and genes whose expression is atypically low. Pre-existing repressive epigenetic features were associated with latency for subsets of proviruses. Our findings suggest a confluency of genomic and epigenomic factors predispose certain genomic locations, including ZNF genes, to host proviruses that constitute the latent reservoir.

microbiology↗

bioRxiv2025

PRISM-Seq: An Ultra-sensitive Sequencing Approach For Mapping Lentiviral Integration Sites

Retroviral integration into host genomes is central to both HIV-1 persistence and the safety and function of lentiviral vectors used in gene and cell therapies. However, existing integration site assays remain limited by sensitivity, input requirements, and analytical complexity, and none have been validated at the single-molecule detection limit. Here, we introduce PRISM-seq, an ultra-sensitive workflow for genome-wide recovery of lentiviral-host junctions, paired with BulkIntSiteR, an open-source, fully automated pipeline for integration site annotation. We show that PRISM-seq accurately identifies proviral insertions across diverse genomic contexts, including euchromatin, heterochromatin, and repeat-rich centromeric regions, and detects high-confidence integration events down to a single input template molecule. By systematically characterizing assay- and amplification-associated noise, we developed a five-step quality control framework that removes PCR- and sequencing-derived artifacts. PRISM-seq also enables quantitative clonal tracking through replicate-based sampling and achieves performance comparable to or exceeding high-input assays at substantially reduced cost.

bioinformatics↗

bioRxiv2025

Molecular Plasticity of T Cells Informs Their Possible Adaptation in 4T1 Tumors

BackgroundThe triple-negative breast cancer (TNBC) microenvironment (TME) undergoes progressive reprogramming, transitioning from an early immune-active state to a late immune-suppressed state. While tumor cell plasticity has been extensively studied, the molecular plasticity of T cells in vivo remains poorly defined. ObjectivesTo characterize transcriptional changes in T cells during TNBC progression and identify stage-specific shifts in T cell function, polarization, and antigen-presenting cell (APC)-T cell interactions. ResultsTranscriptional analysis of T cells from BALB/c mice bearing 4T1 tumors at 1, 3, and 6 weeks revealed a decline in T cell-associated genes from 194 at 1 week to 156 at 6 weeks, with a significant late-stage loss of TCR diversity and contraction of natural killer T (NKT)- and {gamma}{delta} T cell-related transcripts. Cytokine and transcription factor dynamics reflected temporal T cell polarization: early (1 week) IL-12/{beta}-STAT4 signaling supports CD4+ type 1 T helper cell (Th1) and type 1 CD8+ cytotoxic T cell (Tc1) responses; intermediate (3 weeks) IL-21 and BCL6 expression suggest transient CD8+ cytotoxic follicular T cell (Tfc) skewing; and late (6 weeks) AhR and IL-1{beta} induction reflect interleukin 17/22 producing CD8+ T cell (Tc17/Tc22) transition. Pro-inflammatory cytokines and chemokines increased over time, while immunosuppressive mediators (e.g., IL-10) declined significantly. Antigen-presenting cell (APC)-T cell crosstalk deteriorated at 6 weeks, characterized by a reduction in the expression of co-stimulatory and APC genes. Despite an early dominance of M1-like macrophage signals (e.g., IL-12/{beta}), persistent expression of arginase 1 (ARG1) and other M2-associated genes indicated a stable tolerogenic niche. ConclusionsTNBC progression is characterized by progressive T cell functional decline, narrowing of TCR diversity, impaired APC-T cell interactions, and sustained macrophage-driven immunosuppression. These temporally coordinated immune shifts suggest tumor-driven adaptation toward immune evasion and identify potential windows for stage-specific immunotherapeutic intervention.

cancer biology↗

bioRxiv2025

Atorvastatin suppresses cardiac fibrosis and dysfunction induced by HIV and certain antiretroviral drugs in mice by blocking platelet TGFβ1

Cardiovascular disease (CVD) both atherosclerosis-related and heart failure with preserved ejection fraction (HFpEF) and linked to cardiac fibrosis, contributes to morbidity and mortality in people with HIV (PWH) receiving antiretroviral therapy (ART). In the REPRIEVE trial, pitavastatin reduced atherosclerotic CVD risk to a magnitude inconsistent with pitavastatins impact solely on LDL-cholesterol and inflammation. We hypothesized that HFpEF in PWH relates to HIV-induced fibrosis mediated by platelet TGF{beta}1, that it is accelerated by certain contemporary ART, and may also be inhibited by statins. ART drugs used in REPRIEVE, including a nucleoside/nucleotide, integrase inhibitor-based regimen (tenofovir (TDF), emtricitabine (FTC), and dolutegravir (DTG)), and the protease inhibitors ritonavir (RTV) and darunavir (DRV), and the impact of atorvastatin, were examined in two HIV mouse models: transgenic Tg26 mice and HIV-PDX mice engrafted with HIV-infected T cells. Tg26 and HIV-PDX mice had higher cardiac fibrosis than littermate controls without HIV (p<0.05). Administration of TDF-FTC-DTG or RTV, but not DRV, resulted in a further [~]2-fold increase in fibrosis (p<0.01). Higher cardiac fibrosis with intracardiac fat accumulation correlated with reduced diastolic function. Mice depleted of platelet TGF{beta}1 (TGF{beta}1Platelet-{Delta}Tg26), or treated with atorvastatin, were partially protected from HIV- and ART-induced cardiac fibrosis, steatosis, and diastolic dysfunction. Atorvastatin effects occurred independently of changes in inflammatory cytokines and total cholesterol. They correlated with reduced platelet activation and TGF{beta}1 signaling in cardiac endothelial cells, fibroblasts, and macrophages undergoing mesenchymal transition. These results indicate that certain ART regimens accelerate HIV-associated CVD characterized by HFpEF via platelet TGF{beta}1-dependent processes and mitigated by atorvastatin. They enhance understanding of the pleiotropic effects of statins in HIV/ART CVD and suggest a mechanism that might be targeted by antiplatelet agents or inhibition of TGF{beta} signaling. Key PointsO_LIContemporary ART regimens induce release of platelet TGF{beta}1 and are associated with cardiac fibrosis and diastolic dysfunction with ectopic fat deposition in HIV-infected mice. C_LIO_LIDepleting platelet TGF{beta}1 and/or treating with atorvastatin therapy suppresses HIV-ART-induced cardiac fibrosis, suggesting use of anti-platelet strategies to prevent heart failure among PWH. C_LI

biochemistry↗

bioRxiv2025

HIV-1 expression is heterogeneous among clones of CD4+ T cells carrying authentic intact latent proviruses

Antiretroviral therapy suppresses HIV-1 infection but is not curative because it fails to eliminate a reservoir of intact latent proviruses that reside primarily in CD4+ T cells. This compartment is composed of rare T cells that predominantly express memory and effector memory markers. The lack of precise understanding of the latent compartment has made it challenging to develop curative strategies for HIV-1 infection. Here we report on the properties of CD4+ T cells clones carrying intact latent proviruses, expanded in vitro from single cells obtained from the reservoir of people living with HIV-1. The latent proviruses in the clones were integrated into ZNF genes, non-genic satellite and centromeric regions, frequently associated with latency. Notably, the transcriptome of the cultured clones resembled their cells of origin. Despite their descent from single cells, only a fraction of the cells ranging from 0.4-14% expressed relatively low levels of HIV-1 that did not measurably alter host gene transcriptome. Latency reversing agents (LRAs) variably increased the number and amount of expression per cell, but the effects were modest and clone and LRA specific. The results suggest that pharmacologic and immunologic approaches to clear the reservoir should be optimized to accommodate intra- and inter-clonal diversity.

immunology↗

bioRxiv2025

Q4ddPCR (May the Fourth Be Precise): A Flexible, 4-Target Assay for High-Resolution HIV Reservoir Profiling

Precise and scalable quantification of the genetically intact HIV reservoir is critical for advancing curative strategies. However, current HIV reservoir assays such as the intact proviral DNA assay (IPDA) are limited by quantification failures or misclassification of defective proviral genomes due to HIV sequence heterogeneity. Q4ddPCR is a modular, droplet digital PCR assay that simultaneously targets four conserved regions in the HIV genome to improve specificity, reduce quantification gaps, and provide multi-layered readouts. We benchmarked Q4ddPCR against 3,650 near full-length proviral sequences from 13 virally suppressed people with HIV (PWH) generated by Q4PCR using the same primer/probe sets. Q4ddPCR enabled intact reservoir quantification in 95% of samples from three independent cohorts and closely matched sequence-confirmed Q4PCR reservoir measurements. In addition, multi-probe readouts revealed clonal intact reservoir dynamics that are not detectable by IPDA. In longitudinal samples from 42 participants over the first 4.5 years on antiretroviral therapy (ART), Q4ddPCR reported lower proviral frequencies and a steeper decline in intact proviral DNA compared to IPDA. Collectively, our findings confirm key predictions from mathematical modeling, demonstrating that multi-target assays provide greater specificity and more accurately capture the dynamics of the intact HIV reservoir.

immunology↗

bioRxiv2025

Development and validation of HIV SMRTcap for the characterization of HIV-1 reservoirs across tissues and subtypes

Human Immunodeficiency Virus type 1 (HIV-1) is responsible for the global HIV/AIDS epidemic and the establishment of an integrated HIV-1 reservoir remains the primary obstacle to cure. Upon therapy interruption, reactivation of the persistent HIV-1 reservoir propagates viral rebound and mediates continued immunological decline. While furthering understanding of the HIV-1 reservoir is essential for HIV-1 cure, commonly used sequencing strategies are often limited by the reliance on short-read sequencing across separate assays to determine integration sites and proviral integrity - something that does not always adequately resolve complex human genomic repeats or low complexity regions. Simultaneous identification of proviral integration sites and proviral integrity at the single molecule level would enable HIV-1 reservoir characterization with minimal imputation or bioinformatic reconstruction. Here we present HIV Single Molecule Real Time Capture (HIV SMRTcap), a novel molecular and computational pipeline that directly and simultaneously identifies HIV-1 integration sites, defines proviral integrity, and characterizes clonal expansion of HIV-1 provirus-containing cells with single molecule resolution. In combination with long-read, single-molecule, real-time (SMRT) sequencing and custom analytic pipelines, HIV SMRTcap enables a highly comprehensive characterization of HIV-1 reservoirs. Moreover, we demonstrate here that HIV SMRTcap performs robustly across the major global subtypes (HIV-1 subtype A, B, C, D and A/D recombinant viruses), and can use both cell- and tissue-derived inputs, including samples from antiretroviral therapy (ART) treated individuals with undetectable viral loads. Our results demonstrate that HIV SMRTcap serves as a comprehensive, robust method for unbiased HIV-1 reservoir characterization. Used alone, or in combination with single-cell based methods, HIV SMRTcap will enable novel exploration of viral reservoirs across subtypes and in tissue-specific compartments, providing critical information needed to inform HIV-1 cure.

genomics↗

bioRxiv2024

Multi-Omic Atlas reveals cytotoxic phenotype and ROS-linked metabolic quiescence as key features of CTL-resistant HIV-infected CD4+ T-cells

Cytotoxic T-lymphocytes (CTL) exert sustained pressure on reservoirs of HIV-infected cells that persist through years of antiretroviral therapy (ART). This selects for latently infected cells, but also potentially for cells that express HIV but possess intrinsic CTL resistance. We demonstrate that such resistance exists in HIV-infected CD4+ T-cells that survive rigorous CTL attack and map CTL susceptibility to cell identities and states defined by single-cell multi-omics and functional metabolic profiling. Cytotoxic CD4+ T-cells were prominently overrepresented amongst survivors, as were cells with quiescent metabolic profiles and low levels of reactive oxygen species (ROS) production. The induction of ROS production by treatment with deferoxamine sensitized these cells to CTL-mediated elimination. Reservoir-harboring cells from clinical samples share the above transcriptional features, being enriched for quiescent states. Our results provide an atlas for elucidating features of CTL resistance in HIV reservoirs, and identify oxidative stress as a therapeutic target to facilitate reservoir elimination.

immunology↗

bioRxiv2024

A Transcriptional Signature of Induced Neurons Differentiates Virologically Suppressed People Living With HIV from People Without HIV

Neurocognitive impairment is a prevalent and important co-morbidity in virologically suppressed people living with HIV (PLWH), yet the underlying mechanisms remain elusive and treatments lacking. Here, we explored for the first time, use of participant-derived directly induced neurons (iNs) to model neuronal biology and injury in PLWH. iNs retain age-and disease-related features of the donors, providing unique opportunities to reveal novel aspects of neurological disorders. We obtained primary dermal fibroblasts from six virologically suppressed PLWH (range: 27 - 64 years, median: 53); 83% Male; 50% White) and seven matched people without HIV (PWOH) (range: 27 - 66, median: 55); 71% Male; 57% White). iNs were generated using transcription factors NGN2 and ASCL1, and validated by immunocytochemistry and single-cell-RNAseq. Transcriptomic analysis using bulk-RNAseq identified 29 significantly differentially expressed genes between iNs from PLWH and PWOH. Of these, 16 genes were downregulated and 13 upregulated in PLWH iNs. Protein-protein interaction network mapping indicates that iNs from PLWH exhibit differences in extracellular matrix organization and synaptic transmission. IFI27 was upregulated in iNs from PLWH, which complements independent post-mortem studies demonstrating elevated IFI27 expression in PLWH-derived brain tissue, indicating that iN generation reconstitutes this pathway. Finally, we observed that expression of the FOXL2NB-FOXL2-LINC01391 genome locus is reduced in iNs from PLWH and negatively correlates with neurocognitive impairment. Thus, we have identified an iN gene signature of HIV through direct reprogramming of skin fibroblasts into neurons revealing novel mechanisms of neurocognitive impairment in PLWH. One sentence summaryDirect reprogramming of skin fibroblasts into neurons reveals unique gene signatures indicative of HIV infection in the context of viral suppression.

neuroscience↗

bioRxiv2024

The EZH2 inhibitor tazemetostat mitigates HIV immune evasion, reduces reservoir formation, and promotes durable CD8+ T-cell revitalization

Persistent HIV reservoirs in CD4 T-cells pose a barrier to curing HIV infection. We identified overexpression of enhancer of zeste homolog 2 (EZH2) in HIV-infected CD4 T- cells that survive cytotoxic T lymphocyte (CTL) exposure, suggesting a mechanism of CTL resistance. Inhibition of EZH2 with the FDA-approved drug tazemetostat increased surface expression of major histocompatibility complex class I (MHC-I) on CD4 T-cells, counterbalancing HIV Nef-mediated MHC-I downregulation. This improved CTL-mediated elimination of HIV-infected cells and suppressed viral replication in vitro. In a participant-derived xenograft mouse model, tazemetostat elevated MHC-I and the pro-apoptotic protein BIM in CD4 T-cells, facilitating CD8 T-cell-mediated reductions of HIV reservoir seeding. Additionally, tazemetostat promoted sustained skewing of CD8 T-cells toward less differentiated and exhausted phenotypes. Our findings reveal EZH2 overexpression as a novel mechanism of CTL resistance and support the clinical evaluation of tazemetostat to enhance clearance of HIV reservoirs and improve CD8+ T-cell function.

immunology↗

bioRxiv2024

Transcription of HIV-1 at sites of intact latent provirus integration

HIV-1 anti-retroviral therapy is highly effective but fails to eliminate a reservoir of latent proviruses leading to a requirement for life-long treatment. How the site of integration of authentic intact latent proviruses might impact their own or neighboring gene expression or reservoir dynamics is poorly understood. Here we report on proviral and neighboring gene transcription at sites of intact latent HIV-1 integration in cultured T cells obtained directly from people living with HIV, as well as engineered primary T cells and cell lines. Proviral gene expression was correlated to the level of endogenous gene expression under resting but not activated conditions. Notably, latent proviral promoters were 100-10,000X less active than in productively infected cells and had little or no measurable impact on neighboring gene expression under resting or activated conditions. Thus, the site of integration has a dominant effect on the transcriptional activity of intact HIV-1 proviruses in the latent reservoir thereby influencing cytopathic effects and proviral immune evasion.

immunology↗

bioRxiv2023

Immunogenicity of COVID-19 vaccines and their effect on the HIV reservoir in older people with HIV

Older individuals and people with HIV (PWH) were prioritized for COVID-19 vaccination, yet comprehensive studies of the immunogenicity of these vaccines and their effects on HIV reservoirs are not available. We followed 68 PWH aged 55 and older and 23 age-matched HIV-negative individuals for 48 weeks from the first vaccine dose, after the total of three doses. All PWH were on antiretroviral therapy (cART) and had different immune status, including immune responders (IR), immune non-responders (INR), and PWH with low-level viremia (LLV). We measured total and neutralizing Ab responses to SARS-CoV-2 spike and RBD in sera, total anti-spike Abs in saliva, frequency of anti-RBD/NTD B cells, changes in frequency of anti-spike, HIV gag/nef-specific T cells, and HIV reservoirs in peripheral CD4+ T cells. The resulting datasets were used to create a mathematical model for within-host immunization. Various regimens of BNT162b2, mRNA-1273, and ChAdOx1 vaccines elicited equally strong anti-spike IgG responses in PWH and HIV- participants in serum and saliva at all timepoints. These responses had similar kinetics in both cohorts and peaked at 4 weeks post-booster (third dose), while half-lives of plasma IgG also dramatically increased post-booster in both groups. Salivary spike IgA responses were low, especially in INRs. PWH had diminished live virus neutralizing titers after two vaccine doses which were rescued after a booster. Anti-spike T cell immunity was enhanced in IRs even in comparison to HIV- participants, suggesting Th1 imprinting from HIV, while in INRs it was the lowest. Increased frequency of viral blips in PWH were seen post-vaccination, but vaccines did not affect the size of the intact HIV reservoir in CD4+ T cells in most PWH, except in LLVs. Thus, older PWH require three doses of COVID-19 vaccine to maximize neutralizing responses against SARS-CoV-2, although vaccines may increase HIV reservoirs in PWH with persistent viremia.

immunology↗

bioRxiv2023

HIV reservoirs are dominated by genetically younger and clonally enriched proviruses

In order to cure HIV, we need to better understand the within-host evolutionary origins of the small reservoir of genome-intact proviruses that persists within infected cells during antiretroviral therapy (ART). Most prior studies on reservoir evolutionary dynamics however did not discriminate genome-intact proviruses from the vast background of defective ones. We reconstructed within-host pre-ART HIV evolutionary histories in six individuals and leveraged this information to infer the ages of intact and defective proviruses sampled after an average >9 years on ART, along with the ages of rebound and low-level/isolated viremia occurring during this time. We observed that the longest-lived proviruses persisting on ART were exclusively defective, usually due to large deletions. In contrast, intact proviruses and rebound HIV exclusively dated to the years immediately preceding ART. These observations are consistent with genome-intact proviruses having shorter lifespans, likely due to the cumulative risk of elimination following viral reactivation and protein production. Consistent with this, intact proviruses (and those with packaging signal defects) were three times more likely to be genetically identical compared to other proviral types, highlighting clonal expansion as particularly important in ensuring their survival. By contrast, low-level/isolated viremia sequences were genetically heterogeneous and sometimes ancestral, where viremia may have originated from defective proviruses. Results reveal that the HIV reservoir is dominated by clonally-enriched and genetically younger sequences that date to the untreated infection period when viral populations had been under within-host selection pressures for the longest duration. Knowledge of these qualities may help focus strategies for reservoir elimination. ImportanceCharacterizing the HIV reservoir that endures despite antiretroviral therapy (ART) is critical to cure efforts. Our observation that the oldest proviruses persisting during ART were exclusively defective, while intact proviruses (and rebound HIV) all dated to the years immediately pre- ART, explains why prior studies that sampled sub-genomic proviruses on-ART (which are largely defective) routinely found sequences dating to early infection, whereas those that sampled viral outgrowth sequences found essentially none. Together with our findings that intact proviruses were also more likely to be clonal, and that on-ART low-level/isolated viremia originated from proviruses of varying ages (including possibly defective ones), our observations indicate that: 1) on-ART and rebound viremia can have distinct within-host origins, 2) intact proviruses have shorter lifespans than grossly-defective ones, and therefore depend on clonal expansion for persistence, and 3) the HIV reservoir, being overall genetically younger, will be substantially adapted to within-host pressures, complicating immune-based cure strategies.

genetics↗