Agonists and Antagonists of Protease-Activated Receptor 2 Discovered within a DNA-Encoded Chemical Library Using Mutational Stabilization of the Target

The discovery of ligands via affinity-mediated selection of DNA-encoded chemical libraries is driven by the quality and concentration of the protein target. G-protein-coupled receptors (GPCRs) and other membrane-bound targets can be difficult to isolate in their functional state and at high concentr...

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Bibliographic Details
Published in:SLAS discovery 2018-06, Vol.23 (5), p.429-436
Main Authors: Brown, Dean G., Brown, Giles A., Centrella, Paolo, Certel, Kaan, Cooke, Robert M., Cuozzo, John W., Dekker, Niek, Dumelin, Christoph E., Ferguson, Andrew, Fiez-Vandal, Cédric, Geschwindner, Stefan, Guié, Marie-Aude, Habeshian, Sevan, Keefe, Anthony D., Schlenker, Oliver, Sigel, Eric A., Snijder, Arjan, Soutter, Holly T., Sundström, Linda, Troast, Dawn M., Wiggin, Giselle, Zhang, Jing, Zhang, Ying, Clark, Matthew A.
Format: Article
Language:English
Subjects:
Allosteric Site - drug effects
Cell Line
DNA - genetics
HEK293 Cells
Humans
Ligands
Mutation - drug effects
Proteins - genetics
Receptors, G-Protein-Coupled - agonists
Receptors, G-Protein-Coupled - antagonists & inhibitors
Receptors, G-Protein-Coupled - genetics
Small Molecule Libraries - pharmacology
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Summary:The discovery of ligands via affinity-mediated selection of DNA-encoded chemical libraries is driven by the quality and concentration of the protein target. G-protein-coupled receptors (GPCRs) and other membrane-bound targets can be difficult to isolate in their functional state and at high concentrations, and therefore have been challenging for affinity-mediated selection. Here, we report a successful selection campaign against protease-activated receptor 2 (PAR2). Using a thermo-stabilized mutant of PAR2, we conducted affinity selection using our >100-billion-compound DNA-encoded library. We observed a number of putative ligands enriched upon selection, and subsequent cellular profiling revealed these ligands to comprise both agonists and antagonists. The agonist series shared structural similarity with known agonists. The antagonists were shown to bind in a novel allosteric binding site on the PAR2 protein. This report serves to demonstrate that cell-free affinity selection against GPCRs can be achieved with mutant stabilized protein targets.
ISSN:2472-5552
2472-5560
DOI:10.1177/2472555217749847