Biology subjects

Dahal, L. N.

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

4 recordsLinked to original sources

bioRxiv2025

Innate immunocompetent iNSpheroids: A hiPSC-derived 3D model to study the central nervous system captures an early CNS response to rAAV

Gene therapies using adeno-associated viruses (AAVs) for central nervous system (CNS) disorders face challenges due to host immune responses not represented in classical preclinical models. Here, we present a human-induced pluripotent stem cell (hiPSC)-derived innate immunocompetent 3D CNS model that recapitulates neuroinflammatory hallmarks, serving as a platform for preclinical gene therapy development. Utilizing various scales of stirred-tank bioreactor systems, we generated (neurospheroids) iNSpheroids composed of neurons, astrocytes, and oligodendrocytes, alongside microglial cells (iMGL) to mimic the neuro-immune axis. These systems enabled large-scale production of iNSpheroids and subsequent miniaturization for co-culture experiments and screening of inflammatory stimuli, while maintaining a highly controlled environment. The iMGL-iNSpheroids demonstrated active neuron-microglia crosstalk and exhibited distinct inflammatory responses to a series of neuroinflammatory factors. iMGL-iNSpheroids mounted a mild and transient response to rAAV9, mediated by the activation of inflammatory pathways (e.g., TNF-via NF-{kappa}B activation) in glial cell populations. This model offers a valuable tool to dissect neuroinflammatory mechanisms, accelerating gene therapy development. TeaserImmune-competent 3D human CNS model recapitulates glial responses to rAAVs, enabling reliable preclinical gene therapy screening.

bioengineering↗

bioRxiv2025

LIMD1 Loss as an Early Driver of PD-L1 Upregulation and Immune Evasion in Lung Cancer

LIMD1 is a tumour suppressor gene frequently lost in non-small cell lung cancer (NSCLC), but its role in cancer-immune cell interactions remains unexplored. Here, we demonstrate that LIMD1 loss results in upregulation of the key immune checkpoint protein PD-L1. Using multi-region sequencing from the TRACERx dataset, we identify that LIMD1 loss is clonal in over 80% of squamous cell carcinoma (LUSC) and 40% of lung adenocarcinoma (LUAD) cases, correlating with increased PD-L1 expression. LIMD1 deficiency results in upregulation of basal and IFN{gamma}-induced PD-L1 expression in NSCLC cells and, consistent with its early loss during oncogenesis, in primary human small airway epithelial cells. Mechanistically, we demonstrate that LIMD1 interacts with the E3 ubiquitin ligase ARIH1 to mediate efficient PD-L1 ubiquitination and degradation, a process that is significantly impaired in LIMD1-deficient cells, resulting in increased PD-L1 stability. As a consequence, LIMD1-deficient tumour cells suppressed CD8+ T cell activation in vitro, and blockade of PD-L1 reversed this suppression. Clinically, we show that LIMD1 loss is associated with enhanced response to immune checkpoint inhibitors (ICIs) in NSCLC patient cohorts, revealing a novel cancer cell-intrinsic correlation of ICI efficacy. Our results uncover a tumour suppressor-mediated mechanism of PD-L1 expression and pave the way for stratified immunotherapy approaches in LIMD1-/- NSCLC.

cancer biology↗

bioRxiv2023

Soluble CTLA-4 raises the threshold for T-cell activation and modulates anti-tumour immunity

CTLA-4 is a crucial immune checkpoint receptor involved in the maintenance of immune homeostasis, tolerance, and tumour control. Antibodies targeting CTLA-4 have been promising treatment for numerous cancers, but the mechanistic basis of their anti-tumoral immune boosting effects are poorly understood. Although the ctla4 gene also encodes an alternatively-spliced soluble variant (sCTLA-4), preclinical/clinical evaluation of anti-CTLA-4-based immunotherapies have not considered the contribution of this isoform. Here, we explore the functional properties of sCTLA-4 and evaluate the efficacy of isoform-specific anti-sCTLA-4 antibody targeting in murine cancer model. We show that expression of sCTLA-4 in tumour cells suppresses CD8+ T-cells in vitro, and accelerates growth and experimental metastasis of murine tumours in vivo. These effects were accompanied by modification of the immune infiltrate, notably restraining CD8+ T-cells in a non-effector state. sCTLA-4 blockade with isoform-specific antibody reversed this restraint, enhancing intratumoural CD8+ T-cell activation and cytolytic potential, correlating with therapeutic efficacy and tumour control. This previously unappreciated role of sCTLA-4 suggests that better understanding of the biology and function of multi-gene products of immune checkpoint receptors needs to be fully elucidated for improved cancer immunotherapy.

cancer biology↗

bioRxiv2023

Synthetic STING agonists elicit powerful vaccine adjuvancy providing robust central memory and anti-tumour effects

Drugs that target the innate immune sensor STING are known to be effective in modulating the immune infiltrate of the tumour microenvironment. STING agonists have potential to enhance responses to checkpoint inhibitor therapy, however, their ability to influence and shape adaptive immune responses is poorly understood. Here, we investigated the impact of a range of synthetic STING agonists on antigen specific CD8+ T-cell responses to soluble antigen using the murine OT-1 adoptive transfer model with Ovalbumin as the antigen to monitor T cell responses. Our data demonstrate that synthetic STING agonists are able to stimulate antigen specific T-cell expansion in response to challenge in mice. This effect required expression of STING, an intact myeloid compartment and Type-I IFN and TNF signalling. Expanded T-cells post treatment differed from those induced by the established immune adjuvant, anti-CD40 antibody through lower induction of the immune checkpoint receptor PD-1. Furthermore, our data revealed a marked increase in the induction and persistence of CD8+ central memory cells after STING agonist and antigen challenge. Finally, we demonstrate that following rechallenge, STING agonism produced larger secondary responses that could be translated into enhanced tumour protection and survival. Therefore, synthetic STING agonists are capable of acting as potent immune adjuvants and can induce robust memory formation leading to better recall and tumour control. Critically, these benefits along with the lower expression of PD-1, have implications for their use as adjuvants for multiple immunotherapy and vaccine applications.

immunology↗