Discovery of Protease-Activated Receptor 4 (PAR4)-Tethered Ligand Antagonists Using Ultralarge Virtual Screening
Shannon T. Smith, Jackson B. Cassada, Lukas von Bredow, Kevin Erreger, Emma M. Webb, Trevor A. Trombley, Jacob J. Kalbfleisch, Brian J. Bender, Irène Zagol-Ikapitte, Valerie M. Kramlinger, Jacob L. Bouchard, Sidnee G. Mitchell, Maik Tretbar, Brian K. Shoichet, Craig W. Lindsley, Jens Meiler, Heidi E. Hamm
Abstract
Here, we demonstrate a structure-based small molecule virtual screening and lead optimization pipeline using a homology model of a difficult-to-drug G-protein-coupled receptor (GPCR) target. Protease-activated receptor 4 (PAR4) is activated by thrombin cleavage, revealing a tethered ligand that activates the receptor, making PAR4 a challenging target. A virtual screen of a make-on-demand chemical library yielded a one-hit compound. From the single-hit compound, we developed a novel series of PAR4 antagonists. Subsequent lead optimization via simultaneous virtual library searches and structure-based rational design efforts led to potent antagonists of thrombin-induced activation. Interestingly, this series of antagonists was active against PAR4 activation by the native protease thrombin cleavage but not the synthetic PAR4 agonist peptide AYPGKF.