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Selective Agonists and Antagonists of Formylpeptide Receptors: Duplex Flow Cytometry and Mixture-Based Positional Scanning Libraries

Selective Agonists and Antagonists of Formylpeptide Receptors: Duplex Flow Cytometry and Mixture-Based Positional Scanning Libraries

Authors :
Bruce S. Edwards
José L. Medina-Franco
Jon R. Appel
Radleigh G. Santos
Marc A. Giulianotti
Susan M. Young
Clemencia Pinilla
Richard A. Houghten
Larry A. Sklar
Tina Yates-Gibbins
Publication Year :
2013
Publisher :
The American Society for Pharmacology and Experimental Therapeutics, 2013.

Abstract

The formylpeptide receptor (FPR1) and formylpeptide-like 1 receptor (FPR2) are G protein-coupled receptors that are linked to acute inflammatory responses, malignant glioma stem cell metastasis, and chronic inflammation. Although several N-formyl peptides are known to bind to these receptors, more selective small-molecule, high-affinity ligands are needed for a better understanding of the physiologic roles played by these receptors. High-throughput assays using mixture-based combinatorial libraries represent a unique, highly efficient approach for rapid data acquisition and ligand identification. We report the superiority of this approach in the context of the simultaneous screening of a diverse set of mixture-based small-molecule libraries. We used a single cross-reactive peptide ligand for a duplex flow cytometric screen of FPR1 and FPR2 in color-coded cell lines. Screening 37 different mixture-based combinatorial libraries totaling more than five million small molecules (contained in 5,261 mixture samples) resulted in seven libraries that significantly inhibited activity at the receptors. Using positional scanning deconvolution, selective high-affinity (low nM K(i)) individual compounds were identified from two separate libraries, namely, pyrrolidine bis-diketopiperazine and polyphenyl urea. The most active individual compounds were characterized for their functional activities as agonists or antagonists with the most potent FPR1 agonist and FPR2 antagonist identified to date with an EC₅₀ of 131 nM (4 nM K(i)) and an IC₅₀ of 81 nM (1 nM K(i)), respectively, in intracellular Ca²⁺ response determinations. Comparative analyses of other previous screening approaches clearly illustrate the efficiency of identifying receptor selective, individual compounds from mixture-based combinatorial libraries.

Details

Language :
English
Database :
OpenAIRE
Accession number :
edsair.doi.dedup.....f6fd3bea86cee3f9b35a23291c7c92d8