Biology subjects

Board, N. L.

Publications and source records attributed to Board, N. L..

7 recordsLinked to original sources

bioRxiv2026

Potential of HLA-E-targeting diabodies to induce lysis of HIV-1-infected cells by CD8+ T cells

A long-lived reservoir of cells harboring intact HIV-1 provirus persists throughout decades of antiretroviral therapy and can give rise to rapid viral rebound after treatment interruption. Some cure strategies employ cytotoxic T lymphocytes (CTL) to target this reservoir; however, the applicability and efficacy of immunotherapeutic strategies involving MHC class I-restricted CTL is limited by the polymorphic nature of MHC class I molecules and their downregulation by HIV-1 Nef. The non-polymorphic non-classical class I molecule HLA-E is stably expressed on HIV-1-infected CD4+ T cells and presents a potential universal target. We generated a single-chain diabody RLP-13 that redirects CTLs to target cells presenting a well-characterized peptide derived from Mycobacterium tuberculosis in the context of HLA-E. We verified the affinity and specificity of RLP-13. Through co-culture experiments, we confirmed that RLP-13 mediates polyfunctional, HLA-agnostic CTL responses. Using an HIV-1 reporter construct encoding the target peptide, we demonstrated robust and specific elimination of the HIV-1-expressing cell population. This proof-of-concept study shows that HLA-E antigens are promising immunotherapeutic targets that can bypass the limitations of classical MHC class I antigens - allelic variation and downregulation - and that such bispecific antibodies recognizing HIV-1-derived HLA-E binding epitopes could induce elimination of productively infected cells. SummarySengupta, Bachmann et al. utilize a novel HLA-E-restricted CD3-engaging single-chain diabody to induce antigen-specific polyfunctional CTL-responses that are HLA-type-independent. They further show that such biologics have potential to eliminate HIV-1-infected cells by targeting HLA-E-binding epitopes encoded in the HIV-1 provirus.

microbiology↗

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

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

A TCR-mimic bispecific antibody reduces HIV-1 provirus and delays viral rebound in HLA-matched humanized mice

Bispecific antibodies that reroute cytotoxic effectors toward infected cells are promising HIV-1 cure agents, yet existing formats bind Env and are limited by antigenic variation and Env down-regulation. We engineered a TCR-mimic single-chain diabody, HI12, that recognizes a conserved Pol-derived peptide presented by HLA-A*02:01 and evaluated its effect in HLA-matched, HIV-infected humanized mice. When administered during early antiretroviral therapy (ART), HI12 was well tolerated, activated HIV-specific CD8+ T cells and accelerated plasma virus decay. Treatment produced four-to six-fold reductions in intact and total proviral DNA within lymph-node and splenic CD4+ T cells, indicating substantive reservoir clearance. After ART interruption, HI12-treated animals showed a significant delay in viral rebound compared with controls, linking reservoir reduction to improved post-therapy control. These findings provide the first in-vivo evidence that a peptide-HLA-directed bispecific antibody can both shrink the intact HIV reservoir and defer viral recrudescence, supporting further development of TCR-mimic bispecific antibodies for cure strategies. Significance StatementAn HIV-1 cure strategy will require novel therapeutics to facilitate immune-mediated elimination of infected cells and reduction of blood and tissue reservoirs. Here, we demonstrate that an TCR-mimic bispecific antibody that recognizes a conserved Pol epitope can promote CD8+ T cell-mediated clearance of infected cells in HLA-matched, HIV-infected humanized mice. The bispecific antibody therapy reduced HIV-1 proviral DNA in lymph node and splenic tissues and improved post-therapy viral control. This study highlights the potential of developing novel TCR-mimic bispecific antibodies in HIV cure-directed strategies.

immunology↗

bioRxiv2025

Proviruses in CD4+ T cells reactive to autologous antigens contribute to nonsuppressible HIV-1 viremia

Antiretroviral therapy (ART) halts HIV-1 replication, reducing plasma virus levels to below the limit of detection, but it is not curative due to a reservoir of latently infected CD4+ T cells. In some people living with HIV-1 (PLWH), plasma HIV-1 RNA becomes persistently detectable despite optimal ART. This nonsuppressible viremia (NSV) is characterized by identical, non-evolving HIV-1 RNA variants expressed from infected CD4+ T cell clones. The mechanisms driving persistent virus production from a specific population of infected cells are poorly understood. We hypothesized that proviruses in cells responding to chronic immunologic stimuli, including self-associated antigens, may drive viral gene expression and NSV. Here, we demonstrate that stimulation of CD4+ T cells with autologous cell lysates induces virus production in an MHC-II-dependent manner. In 7 of 8 participants with NSV, we recovered viral RNA released ex vivo in response to autologous cell lysates that matched plasma virus. This process involves both defective and replication-competent proviruses residing in conventional T cells, and is also observed in PLWH with undetectable viremia. These findings suggest that recognition of self-associated antigens is an important cause of HIV-1 reservoir expression, which can contribute to persistent systemic inflammation and potential rebound upon ART interruption. One sentence summaryHIV-1 viremia not suppressed by effective ART can be caused by proviruses in CD4+ T cells reactive to autologous antigens.

immunology↗

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

Cognate Antigen Engagement Induces HIV-1 Expression In CD4+ T Cells From People On Long-Term ART

Despite antiretroviral therapy (ART), HIV-1 persists in latently-infected CD4+ T cells, preventing cure. Antigens drive the proliferation of infected cells, precluding latent reservoir decay. However, the relationship between antigen recognition and HIV-1 gene expression is poorly understood since most studies of latency reversal use agents that induce non-specific global T cell activation. Here, we isolated rare CD4+ T cells responding to cytomegalovirus (CMV) or HIV-1 Gag antigens from participants on long-term ART and assessed T cell activation and HIV-1 RNA expression upon co-culture with autologous dendritic cells (DCs) presenting cognate antigens. Physiological presentation of cognate antigens induced broad T cell activation (median 42-fold increase in CD154+CD69+ cells) and significantly increased HIV-1 transcription (median 4-fold), mostly through the induction of rare cells with higher viral expression. Thus, despite low proviral inducibility, physiologic antigen recognition can promote HIV-1 expression, potentially contributing to spontaneous reservoir activity on ART and viral rebound upon ART interruption.

immunology↗