Affinity-Based Protein Profiling Reveals IDH2 as a Mitochondrial Target of Cannabinol in Receptor-Independent Neuroprotection.

Phytocannabinoids are attracting growing attention because of their potential for treatment of neurodegenerative diseases. Among them, the "minor" cannabinoid, cannabinol (CBN), has emerged as a promising neuroprotective agent, acting independently of classical cannabinoid receptors through as-yet unidentified mitochondrial targets. To uncover the molecular basis of its neuroprotective effects, we designed and synthesized a chemical probe based on CBN, incorporating a minimalist diazirine linker. Functional assays confirmed that the probe retains CBN's mitochondrial activity and exhibits strong mitochondrial enrichment, as demonstrated by fluorescence microscopy and click-correlative light and electron microscopy (click-CLEM). By affinity-based protein profiling (AfBPP), we identified isocitrate dehydrogenase 2 (IDH2) as a key mitochondrial target of CBN. This finding was further substantiated by siRNA knockdown studies, which revealed that the absence of IDH2 partially phenocopies CBN's effects, validating its role as a critical mediator of CBN's neuroprotective activity.