Benzyl-Spacer-Engineered Prodrug: Enabling NIR Fluorescent Activation for Cancer Therapy in a Zebrafish Gallbladder Model

A novel near-infrared (NIR) prodrug, INM-Gem, was synthesized and characterized, which significantly improves the application of our previous molecular design for cancer therapy and diagnostics. Selective drug activation in tumor microenvironments via 1,6-elimination is enabled by the incorporation of a benzyl linker. Comparative studies reveal that our probe (i) is allowed to penetrate deeper into tissue (zebrafish), (ii) has less background signal interference, and (iii) involves greater specificity in biological systems than its predecessor. Chemical activation concurrently liberates gemcitabine and triggers NIR “turn-on” fluorescence at 673 nm (Stokes shift: 131 nm), facilitating sensitive, real-time drug-release monitoring. The dual-drug delivery and imaging functionality establish INM-Gem as a robust platform for precise bioimaging and targeted therapy. This work highlights the additional potential of a benzyl spacer in theranostics, wherein the cleavable linker helps enable this NIR-responsive modality for biomedical research.