Glucose‐Modified Viscosity‐Responsive Turn‐On Fluorescence Molecular Rotors for Wash‐Free Live‐Cell Imaging

The overexpression of glucose transporters (GLUTs) in cancer cells represents a promising diagnostic target. While 2‐NBDG , a fluorescent glucose analog, enables GLUT‐mediated imaging, it suffers from low brightness and requires high concentrations and washing. Herein, we report the development of turn‐on fluorescent probes for GLUTs based on the viscosity‐sensitive fluorophore in GFP for a wash‐free imaging of prostate cancer cell, PC‐3. A series of fluorescent molecular rotors (FMRs) were synthesized by modifying the 2‐methyl‐4‐( p ‐dimethylaminobenzylidene)‐5‐imidazolinone ( DMAB ) core. Structural modifications, such as cyclic amine substitution and julolidine incorporation, enhanced fluorescence quantum yields (Φ F up to 0.17 in glycerol) and viscosity sensitivity ( χ up to 0.87). Among them, the julolidine‐containing fluorophore Julo‐Ph showed the best performance. The DMAB ‐based fluorophores were conjugated to glucose or glucosamine to create GLUT‐targeted probes. A glucosamine‐bound conjugate, GlcN‐Julo‐Ph, was found to enable a wash‐free bright fluorescence imaging of PC‐3 cancer cells. Inhibition studies and docking simulations strongly suggested a GLUT‐mediated uptake of the fluorophore. GlcN‐Julo‐Ph outperformed 2‐NBDG in emitting a brighter intracellular fluorescence at a lower concentration, notably, under wash‐free conditions. Our findings shed light on the utility of viscosity‐sensitive FMRs for the design of turn‐on imaging probes and offer a promising platform for a rapid, low‐background cancer cell detection.