Acute kidney injury (AKI) is a rapid loss of renal function that has become a global health concern, with incidence rates reaching up to 50% in critically ill patients. Despite its growing prevalence, no effective pharmacological therapy for AKI currently exists, underscoring an urgent need for new treatments. Here, we present a drug repositioning approach in which the diabetes drug ertiprotafib is formulated within a supramolecular nanomedicine platform to enable biomarker-guided AKI therapy. Covalent modification of ertiprotafib with a multifunctional peptide drives the drug to spontaneously self-assemble into supramolecular nano-assemblies (SENs) via intermolecular interactions. In an ischemia–reperfusion AKI mouse model, these SEN nanoparticles preferentially accumulate in inflamed kidneys by penetrating the interstitium through disrupted tight junctions, a disease-induced structural biomarker. Subsequently, biomarker-guided enzymatic activation of the assemblies by the upregulated cathepsin B in injured tubular cells triggers the release of ertiprotafib, resulting in protein tyrosine phosphatase 1B (PTP1B) inhibition and modulation of the Src/ERK signaling pathway to suppress inflammatory responses and ameliorate disease progression. In contrast, healthy tubular cells and other organs (with low cathepsin B expression) experience minimal prodrug activation, which mitigates ertiprotafib’s off-target toxicity. Collectively, this coordinated supramolecular nanotherapy demonstrates an effective and safer strategy for AKI treatment by repurposing an existing drug.