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bioRxiv · 10.1101/2021.12.06.471449

Structure-based design of a chemical probe set for the 5-HT5A serotonin receptor

Kaplan, A. L.·Strachan, R. T.·Braz, J. M.·Craik, V.·Slocum, S.·Mangano, T.·Amabo, V.·O'Donnell, H. R.·Lak, P.·Basbaum, A.·Roth, B. L.·Shoichet, B. K.
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Abstract

The 5-HT5A receptor (5-HT5AR), for which no selective agonists and only a few antagonists exist, remains the least understood serotonin (5-HT) receptor. A single commercial antagonist (SB-699551) has been widely used to investigate central nervous system (CNS) 5-HT5AR function in neurological disorders, including pain. However, because SB-699551 has affinity for many 5-HTRs, lacks inactive property-matched controls, and has assay interference concerns, it has liabilities as a chemical probe. To better illuminate 5-HT5AR function, we developed a probe set through iterative rounds of molecular docking, pharmacological testing, and optimization. Docking over six million lead-like molecules against a 5-HT5AR homology model identified five mid-M ligand starting points with unique scaffolds. Over multiple rounds of structure-based design and testing, a new quinoline scaffold with high affinity and enhanced selectivity for the 5-HT5AR was developed, leading to UCSF678, a 42 nM arrestin-biased partial agonist at the 5-HT5AR with a much more restricted off-target profile and decreased assay liabilities vs. SB-699551. Site-directed mutagenesis supported the docked pose of UCSF678, which was also consistent with recent published 5-HTR structures. Surprisingly, property-matched analogs of UCSF678 that were either inactive across 5-HTRs or retained affinity for UCSF678s off-targets revealed that 5-HT5AR engagement is nonessential for alleviating pain in a mouse model, contrary to previous studies using less-selective ligands. Relative to SB-699551, these molecules constitute a well-characterized and more selective probe set with which to study the function of the 5-HT5A receptor. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=113 SRC="FIGDIR/small/471449v1_ufig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@8fb24dorg.highwire.dtl.DTLVardef@2dd84org.highwire.dtl.DTLVardef@1d3346org.highwire.dtl.DTLVardef@1117331_HPS_FORMAT_FIGEXP M_FIG Table of Contents Graphic C_FIG

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Kaplan, A. L., Strachan, R. T., Braz, J. M., Craik, V., Slocum, S., Mangano, T., Amabo, V., O'Donnell, H. R., Lak, P., Basbaum, A., Roth, B. L., Shoichet, B. K.. 2021-12-07. Structure-based design of a chemical probe set for the 5-HT5A serotonin receptor. https://doi.org/10.1101/2021.12.06.471449

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