Mechanistic Basis for 5-HT2AR Over 5-HT2BR Activation

Selective 5-HT2AR agonism without 5-HT2BR activation is critical for safe psychedelic-inspired therapeutics. Here, we systematically probe the ligand-binding pockets of 5-HT2AR and 5-HT2BR and identify steric and conformational constraints at the side-extended pocket (SEP) and extended binding pocket (EBP) that dictate ligand orientation and receptor signaling. Guided by these insights, we design derivatives across tryptamine and phenethylamine scaffolds that integrate SEP and EBP engagement with scaffold modification to reinforce 5-HT2AR activation while limiting 5-HT2BR activity. Cryo-EM structures of 5-HT2AR and 5-HT2BR bound to IHCH-2330, a selective 5-HT2AR agonist and 5-HT2BR antagonist, confirm the design principle. These findings show differential activation mechanisms between 5-HT2AR and 5-HT2BR and provide a rational framework for engineering safer, functionally selective serotonergic compounds, mitigating 5-HT2BR–mediated side effects while preserving therapeutic efficacy. This study explores structural principles of receptor selectivity and establishes a strategy for designing selective 5-HT2AR agonists while reducing 5-HT2BR-associated valvulopathy risk.