Received for publication July 7, 2004.
Revised August 10, 2004.
Accepted for publication August 12, 2004.

A novel nonpeptide ligand for formyl peptide receptor-like 1

Abstract

Formyl peptide receptor-like 1 (FPRL1) is a G protein-coupled receptor that binds natural and synthetic peptides as well as lipoxin A4 and mediates important biological functions. To facilitate its pharmacological characterization, we screened a compound library and identified a substituted quinazolinone (Quin-C1, 4-butoxy-N-[2-(4-methoxy-phenyl)-4-oxo-1,4-dihydro-2H-quinazolin-3-yl]-benzamide) as a ligand for FPRL1. Quin-C1 induces chemotaxis and secretion of beta-glucuronidase in peripheral blood neutrophils with a potency of approximately 1/1,000 of that of the peptide agonist WKYMVm. In studies using transfected rat basophilic leukemia (RBL) cell lines expressing either FPR or FPRL1, Quin-C1 induced enzyme release from RBL-FPRL1 but not RBL-FPR cells. Likewise, Quin-C1 selectively stimulates calcium mobilization in RBL-FPRL1 cells, a response that was markedly inhibited by pertussis toxin. Quin-C1 also stimulates phosphorylation of extracellular signal-regulated protein kinases 1 and 2, and induces internalization of an FPRL1 fused to green fluorescent protein. In degranulation assays, both the FPRL1-selective peptide agonist MMK1 and Quin-C1 exhibited lower efficacy and potency than WKYMVm, with EC₅₀ values of 7.17 x 10^-8 M and 1.88 x 10^-6 M, respectively, as compared to the EC₅₀ value for WKYMVm (2.29 x 10^-8 M). However, Quin-C1 did not induce neutrophil superoxide generation at up to 100 micromolar concentration. Based on these results, we conclude that Quin-C1 is a novel nonpeptide ligand that binds to FPRL1 and selectively stimulates FPRL1-mediated functions. Quin-C1 is a prototype of substituted quinazolinones based on which further structural modifications may be made to improve its efficacy and potency for FPRL1.

Key words: Chemotactic peptides, Gi family, Combinatorial chemistry, Receptor binding studies, Receptor-mediated, Leukocytes/Mast cells