Reprogramming insulin receptor activation with a de novo agonist to overcome severe insulin resistance

Abstract Computational protein engineering provides a powerful approach to address longstanding clinical challenges. Severe insulin resistance syndromes caused by mutations in the insulin receptor (IR) are life-threatening disorders for which effective long-term therapies remain lacking. Here, we define the in vivo activity and therapeutic potential of RF-409, a de novo–designed IR agonist that activates the receptor through a mechanism distinct from insulin. RF-409 exhibits markedly prolonged circulation compared to insulin and produces sustained improvements in glucose homeostasis without detectable adverse effects on body composition or liver function. In a patient-derived IR D707A mouse model of severe insulin resistance, RF-409—but not insulin—activates the mutant receptor, restoring glucose regulation and ameliorating hyperglycemia, hyperinsulinemia, lipoatrophy, and pancreatic atrophy. Mechanistically, RF-409 engages the IR through a noncanonical binding geometry while stabilizing an active conformation resembling that induced by insulin. Phosphoproteomic profiling shows that RF-409 elicits broadly insulin-like signaling with distinct temporal features in receptor-proximal regulation. Together, these findings establish a framework for reactivating dysfunctional receptors and suggest broader applications beyond rare receptoropathies, including diabetes and liver disease. Competing Interest Statement X.W., E.C., S.C., and D.B. are co-inventors on a provisional patent (IP: 50206.01US1) filed by the University of Washington covering RF-409 and its uses described in this manuscript. All other authors declare no competing interests.