Investigating the pathogenicity of the recessive HNF1A p.A251T variant in monogenic diabetes using iPSC-derived beta-like cells

Monogenic diabetes results from single-gene defects that impair pancreatic beta-cell function, with mutations in the HNF1A gene being a common cause. While most HNF1A mutations exhibit dominant inheritance, the p.A251T (c.751G>A) variant demonstrates recessive transmission. To define its pathogenic role, induced pluripotent stem cells (iPSCs) derived from a patient homozygous for HNF1A p.A251T, and CRISPR/Cas9-edited human embryonic stem cells carrying the hemizygous variant, were differentiated into pancreatic islet-like cells. Islet-like clusters carrying the p.A251T variant showed impaired differentiation into insulin-producing cells, while alpha-cell formation was increased. Correspondingly, transcriptomic analysis revealed decreases in beta- and increases in alpha cell-enriched genes. Functional studies in islet-like clusters demonstrated lowered glucose-stimulated insulin secretion and diminished intracellular Ca2+ responses, including altered beta-cell connectivity. Responses to sulphonylureas and to glucagon-like peptide-1 receptor agonist were retained. These results indicate that the HNF1A p.A251T variant impairs beta-cell lineage specification and function. Our findings establish patient-derived and genome-edited stem cell models as a translational platform to assess the pathogenicity of rare HNF1A variants and to guide precision therapy for monogenic diabetes. We also define intercellular connectivity as a partially-preserved feature of stem cell-derived human beta-like cells and show that this is affected in a model of monogenic diabetes. HighlightsO_LIPatient-derived and CRISPR/Cas9-edited human stem cells were used to model the rare recessive HNF1A p.A251T variant. C_LIO_LIThe HNF1A p.A251T variant impairs pancreatic islet differentiation, reducing mature beta cells and increasing the alpha-like cell population. C_LIO_LIThe variant compromises glucose-stimulated insulin secretion, which is partially rescued by sulphonylureas, and intercellular connectivity as assessed by Ca2+ imaging. C_LIO_LIThis study demonstrates that patient-derived stem cell models can assess HNF1A variant pathogenicity and supports precision therapy for monogenic diabetes. C_LI O_FIG O_LINKSMALLFIG WIDTH=171 HEIGHT=200 SRC="FIGDIR/small/24318788v2_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@3917d1org.highwire.dtl.DTLVardef@4a9e8eorg.highwire.dtl.DTLVardef@efabf8org.highwire.dtl.DTLVardef@1033ac0_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical abstract.C_FLOATNO Mechanistic impact of the HNF1A p.A251T variant on beta-cells identity and function. The HNF1A p.A251T mutation leads to multiple -cell defects, including reduced glucose-stimulated insulin secretion and diminished intracellular Ca{superscript 2} response to high glucose. Additionally, there is decreased expression of -cell commitment markers, suggesting impaired differentiation, and an increase in alpha-like cell populations. Together, these changes result in mildly disrupted -cell identity and function, indicating partial -cell functional insufficiency. C_FIG