Activatable Multiplexed 19F NMR Probes for Dynamic Monitoring of Biomarkers Associated with Cellular Senescence

Simultaneous detection of multiple biomarkers is crucial to achieve specific and dynamic analysis of cellular senescence, given its intrinsic high heterogeneity. Current approaches for senescence detection largely rely on fluorescence imaging, but fluorescent probes inevitably suffer from issues including autofluorescence and spectral overlap when being applied for the simultaneous detection of multiple biomarkers. Herein, we report an alternative strategy and design activatable multiplexed senoprobes based on ¹⁹F NMR for dynamic monitoring of cellular senescence. Differing from previous approaches, our strategy has two unique advantages. First, this strategy utilizes the changes in the ¹⁹F chemical shift as the signal output, which features by its fingerprint and quantifiable characters, thereby significantly enhancing the detection throughput toward biomarkers with minimized spectral overlapping. Second, the background signal is minimized, benefiting from the extremely low abundance of F in biological samples, and the detection accuracy can thus be improved. As a proof of concept, two activatable ¹⁹F NMR molecular probes are synthesized that specially respond to two key senescence-associated biomarkers (β-gal and ROS) and have been successfully demonstrated for dynamical and quantitative assessment of the changes of these biomarkers in different cellular models of senescence, without causing obvious cytotoxicity. Owing to the flexible molecular design, this work may offer a useful platform to create diversified ¹⁹F NMR senoprobes for deep understanding of cellular senescence across a wide range of aging-related diseases.