Lin, Wenwei, et al. Bioconjugate chemistry 31.11 (2020): 2564-2575.
BODIPY FL Thalidomide is a highly sensitive fluorescent probe used for the development of a time-resolved fluorescence resonance energy transfer (TR-FRET) assay targeting the cereblon protein. As a component of an E3 ubiquitin ligase complex, cereblon is a critical target for molecular glues and PROTACs, which are gaining traction in drug discovery. However, prior assays lacked sufficient sensitivity for effective screening and characterization of cereblon ligands.
To overcome this, BODIPY FL Thalidomide was synthesized and demonstrated a high binding affinity to human cereblon (Kd = 3.6 nM). When applied in a TR-FRET assay, this probe significantly outperformed previously reported assays-showing 41- and 187-fold higher sensitivity than earlier fluorescence polarization and TR-FRET methods based on Cy5-tagged analogs. The assay was both selective (not detecting non-cereblon ligands) and robust, maintaining signal stability from 90 to 240 minutes across a range of thalidomide concentrations.
Importantly, BODIPY FL Thalidomide-mediated TR-FRET enables detection of minor changes in phthalimide activity due to thalidomide isomerization. This level of precision enhances drug development workflows involving cereblon-targeted degraders. Overall, BODIPY FL Thalidomide serves as a superior tool for high-throughput and precise quantification of cereblon ligand interactions, supporting the rational design of next-generation therapeutic agents.
