The selective 5-HT receptor agonist 25CN-NBOH: Structure-activity relationship, in vivo pharmacology, and in vitro and ex vivo binding characteristics of [H]25CN-NBOH.

The remarkable effects exhibited by classical psychedelics in recent clinical trials have spawned considerable interest in 5-HT receptor (5-HTR) activation as a treatment strategy for several psychiatric/cognitive disorders. In this study we have continued our development of 25CN-NBOH, one of the most 5-HTR-selective agonists reported to date, as a pharmacological tool for exploration of 5-HTR expression and functions. The importance of the 2' and 3' positions in 25CN-NBOH as structural hotspots for its 5-HTR activity was investigated by synthesis and pharmacological characterization of six novel analogs at 5-HTR and 5-HTR in binding and functional assays. While the 5-HTR activity of 25CN-NBOH was retained in 3'-methyl, 2',3'-chroman, 2',3'-dihydrofuran and 2',3'-furan analogs, the 3'-methoxy and 3'-ethyl analogs displayed substantially lower binding affinities and agonist potencies than 25CN-NBOH. Interestingly, the 2',3'-substitution pattern was also a key determinant of agonist efficacy, as all six analogs exhibited low-efficacy partial agonism or de facto antagonism at the 5-HTR in the functional assays. Systemic administration of 25CN-NBOH and its close structural analog 25CN-NBMD induced robust head-twitch response in mice, a well-established behavioural effect of 5-HTR activation in vivo, and 25CN-NBOH mediated robust reductions in the activity of mice in an anxiety-related marble burying assay, which supports the proposed beneficial effects of 5-HTR activation on disorders characterized by cognitive rigidity. Finally, tritiated 25CN-NBOH exhibited high 5-HTR binding affinity (K ~1 nM) and selectivity against 5-HTR and 5-HTR in equilibrium and kinetic binding studies of the recombinant receptors, and in concordance [H]25CN-NBOH displayed substantial specific, ketanserin-sensitive binding to cortex and small levels of binding to choroid plexus in rat brain slices in autoradiography studies. In conclusion, this work delineates the subtle molecular determinants of the 5-HTR activity in 25CN-NBOH, substantiates the potential in this compound and its analogs as tools for in vivo studies of the 5-HTR, and introduces a novel selective agonist radioligand as another potentially valuable tool for future explorations of this receptor.