Abstract
Synthetic cannabinoid receptor agonists (SCRAs) represent a class of new psychoactive substances that pose great health risks attributed to their wide-ranging and severe adverse effects. Recent evidence has shown that SCRAs with key moieties can confer superagonism, yet this phenomenon is still not well understood. Here we report structure-activity relationships (SARs) of modular SCRAs contributing to superagonism by comparing eight compounds differing by their head moiety (l-valinate vs. l-tert-leucinate), core moiety (indole vs. indazole), and tail moiety (5-fluoropentyl vs. 4-fluorobenzyl) through different modes of bioluminescence resonance energy transfer (BRET) assays. We found that the l-tert-leucinate head moiety and indazole core moiety conferred superagonism across multiple Gα(i/o) proteins and β-arrestin-2. After generating the cannabinoid type 1 receptor (CB1R) mutant constructs, we found that transmembrane 2 (TM2) interactions to the head moiety of tested SCRAs at F170, F174, F177, and H178 are key to eliciting activity. Finally, we found that l-tert-leucinate SCRAs confer a high-efficacy response in ex vivo slice electrophysiology.