Structure-Based Design of a Chemical Probe Set for the 5-HT _5A Serotonin Receptor

Abstract

The 5-HT_5A receptor (5-HT_5AR), for which no selective agonists and a few antagonists exist, remains the least understood serotonin receptor. A single commercial antagonist, SB-699551, has been widely used to investigate the 5-HT_5AR function in neurological disorders, including pain, but this molecule has substantial liabilities as a chemical probe. Accordingly, we sought to develop an internally controlled probe set. Docking over 6 million molecules against a 5-HT_5AR homology model identified 5 mid-μM ligands, one of which was optimized to <b>UCSF678</b>, a 42 nM arrestin-biased partial agonist at the 5-HT_5AR with a more restricted off-target profile and decreased assay liabilities versus SB-699551. Site-directed mutagenesis supported the docked pose of <b>UCSF678</b>. Surprisingly, analogs of <b>UCSF678</b> that lost the 5-HT_5AR activity revealed that 5-HT_5AR engagement is nonessential for alleviating pain, contrary to studies with less-selective ligands. <b>UCSF678</b> and analogs constitute a selective probe set with which to study the function of the 5-HT_5AR.