Hoang, R. E. | Search bioRxiv

bioRxiv2026

MAFA drives pancreatic islet maturation through coordination of cell growth, metabolic maturation, and resilience to stress.

Stem cell-derived islets (SC-islets) are a promising source for replacing the loss of insulin-secreting {beta} cells in insulin-dependent diabetes patients, but compared to their primary human counter parts, SC-islets are considered functionally immature and do not immediately regulate blood glucose after transplantation. To better understand the molecular underpinnings of {beta} cell maturation, we genetically engineered human pluripotent stem cells (hPSCs) to control the induction of the {beta} cell-maturing transcription factor MAFA or a MAFA S64F mutant variant associated with premature {beta} cell aging and senescence in SC-islets. We employ a combination of in vitro assays, multi-omics analyses, and murine in vivo transplantation studies to demonstrate that MAFA remodeling of the SC-islet transcriptional landscape promotes SC-islet biogenesis and metabolic maturation giving rise to greater SC-islet resilience and insulin secretory capacity. Parallel MAFA S64F studies highlight the importance for compensatory mechanisms for sustaining accelerated oxidative metabolism and enhanced insulin secretion without inducing intracellular stress responses and islet dysfunction. Overall, these results reveal MAFA-governed transcriptional changes that drive SC-islet functional maturation, which provides the foundation for maturing SC-islets in vitro, preserving {beta} cell integrity, and improving SC-islet cell replacement therapies.

cell biology↗

bioRxiv2026

A universal platform for simultaneous TCRα/β removal enables safer and more potent TCR therapies and autoimmune modeling

Adoptive T-cell therapies using tumour-specific T-cell receptors (TCRs) are limited by competition with endogenous receptors, which impairs efficacy and poses risks of off-target autoreactivity. Here we present a CRISPR-based platform that completely and selectively eliminates both endogenous TCR- and -{beta} chains without affecting introduced transgenic TCRs, irrespective of codon optimization. This approach achieves >90% deletion efficiency in Jurkat and primary human T cells, markedly enhancing the expression, pairing fidelity, and functional potency of transgenic receptors. Using a clinically relevant HLA-A*02:01-restricted DMF5 TCR, we show that dual TCR ablation boosts antigen-specific activation and cytotoxicity in vitro and significantly enhances tumor clearance in vivo in human immune system (HIS) mice, while preventing graft-versus-host disease (GVHD). Targeted locus amplification revealed that CRISPR-induced double-strand breaks did not alter lentiviral integration profiles, confirming genomic safety. Extending this approach to four insulin-reactive TCRs demonstrated that removal of endogenous receptors increased transduction efficiency and functional activity, with one (1E6) showing selective activation and infiltration of stem cell-derived islet grafts (SC-islets) in vivo. This study establishes a universal, safe, and scalable genome-editing platform for generating functionally precise human T cells. By integrating cancer immunotherapy and autoimmune disease modelling within a single framework, it provides a strong preclinical rationale for dual endogenous TCR removal as a route to improved specificity, safety, and therapeutic efficacy in TCR-based cell therapies.

immunology↗