ABSTRACT Aim of this research is to understand why endocrine b-cells in the pancreas of diabetic patients fail, with an eye toward identifying new genetic, biochemical, and cellular pathways that can be exploited as therapeutic targets to prevent and reverse this disease process. This grant has supported several original and widely reproduced discoveries, identifying a homeostatic loop orchestrated by Foxo transcription factors that integrates disparate hormonal and nutrient signals into a gene expression program intended to preserve b-cell function and identity. Signal achievements of this work have been the demonstration of three stages leading to b-cell failure: metabolic inflexibility, b-cell conversion into other pancreatic cell types, and dedifferentiation into an endocrine progenitor-like state. Since the last competing renewal, this grant has supported several major new findings: (i) identification of human diabetes-enriched islet cell types and master regulatory activities driving cell-state transitions; (ii) discovery of BACH2 as a driver of b/a-cell transition and b-cell dedifferentiation in humans; (iii) discovery of AFF3 as a driver of human b/a-cell transition; (iv) pathogenic role of aldehyde dehydrogenase 1 isoform A3 (ALHD1A3) in metabolic inflexibility leading to b-cell failure; (v) description of a “recovering b-cell” signature associated with reversal of metabolic inflexibility; (vi) identification of C2CD4A as a human diabetes susceptibility gene with a role in suppressing b-cell “disallowed” genes, thereby controlling metabolic flexibility; (vii) functional demonstration that regulation of mitochondrial complex III function through the oxidoreductase Cyb5r3 is critical in the progression of metabolic inflexibility into b-cell failure; (viii) identification of vitamin D-binding protein (Gc) as a marker of a-to-b- cell transition and potential functional mediator of b-cell dysfunction; (ix) demonstration of antagonistic epistasis between FoxO1 and Hnf4a in b-cells. The PI proposes to extend this work with the following specific aims: In Aim 1, to test pharmacologically and mechanistically whether BACH2 inhibition can reverse b-cell dedifferentiation and a/b-cell-state transitions. In Aim 2, to test the ability of newly synthesized ALDH1A3 inhibitors to reverse metabolic inflexibility, restoring to b-cell function. In Aim 3 to characterize C2CD4A, a putative corepressor encoded by a human diabetes susceptibility locus, as a potential link between type 2 diabetes genetics and metabolic inflexibility in b-cell dysfunction. This body of work offers immediate, tangible therapeutic applications with the potential to prevent or reverse b-cell failure, thus ensuring that we move from disease management to disease modification. It is envisioned that the successful completion of these aims will lead to new diabetes therapeutics.