Activatable prodrug strategies offer powerful means to control therapeutic presentation in space and time. Here, we report a single‐molecule prodrug design that enables glucose‐responsive activation of a glucagon analog for hypoglycemia protection. The system conjugates dasiglucagon with a synthetic pendant comprised of alternating arginine and phenylboronic acid (PBA) units, designed to couple peptide solubility to glucose concentration. The pendant modulates net charge through glucose‐dependent PBA–diol complexation, driving aggregation under normoglycemia and solubilization under hypoglycemia. The lead pendant contains five arginine–PBA repeats and exhibits optimal glucose‐responsive solubility and charge modulation, forming aggregates at high glucose and dissolving as glucose levels decline. Despite a modest reduction in receptor potency relative to native dasiglucagon, this approach provides significant prophylactic protection in a streptozotocin‐induced diabetic mouse model of insulin overdose, rescuing mice from hypoglycemia and eliminating mortality events. This work demonstrates a proof‐of‐concept for molecularly engineered, metabolite‐responsive glucagon prodrugs that function as on‐demand therapeutic depots. More broadly, it establishes a modular design paradigm for dynamic, self‐regulating peptide therapeutics based on charge modulation rather than external carrier systems.
DeWolf et al. (Sun,) studied this question.