In vivo tracking of hepatic cathepsin B in diabetes using a sulfur-modified hemicyanine-based activatable imaging probe

We report the design and synthesis of CatP, a sulfur-substituted hemicyanine-based activatable probe, for the sensitive detection of cathepsin B (CatB) activity. This probe was applied to study the dynamic changes of hepatic CatB in diabetic mellitus (DM), a prevalent metabolic disease. The probe incorporates a Val-Cit linker specifically cleaved by CatB, triggering an intramolecular charge transfer (ICT) mechanism that generates strong near-infrared (750 nm) fluorescence and photoacoustic signals. Structural optimization through sulfur substitution significantly enhanced photoacoustic signal intensity compared to conventional oxygen-substituted analogs. In diabetic mouse models, CatP revealed significantly elevated hepatic CatB activity, which was normalized by metformin treatment as confirmed by both in vivo imaging and ex vivo immunofluorescence. Comprehensive biosafety evaluation demonstrated excellent biocompatibility with >90 % cell viability and no observable organ toxicity. These findings establish sulfur-modified hemicyanine as a promising scaffold for developing high-performance activatable probes, and demonstrate the value of CatP as a functional dye for studying protease activity in biomedical research.