Investigation into ligand selectivity and bias at the formyl peptide receptor family

Formyl peptide receptors (FPRs) mediate both pro-inflammatory and resolution phases of the inflammatory response involved in many disease states. Harnessing their potential for pharmaceutical development requires an accurate picture of their signalling and regulation to the many test compounds developed. This study compares distinct responses of mouse and human FPR subtypes to several ligands in an attempt to clarify the dual nature of FPR signalling. Here we expressed human and mouse variants of FPR1 and FPR2 in HEK293 cells, and assessed competition binding, BRET assays to measure the interaction between receptors and either Arrestin 3 or mini-Gsi, internalization and ERK1/2 phosphorylation. Concentration response curves for 11 distinct ligands at each subtype were generated, then analysed to determine EC50s, Emax values and ligand bias. All compounds were less potent than WKYMVm across receptor subtype, with strength of signalling correlating with affinity estimates. Rank order of potency was maintained across the signalling pathways. Notably, MMK1 was specific for human FPR2, and BMS-986235 selective for FPR2 over FPR1 in both species. Little evidence of pathway bias was detectable, with the notable exception of the recently described pepducin F2Pal10. The majority of tested ligands exhibit efficacy at each subtype meaning conclusions of physiological receptor function based on these compounds should be treated circumspectly. It is not possible to determine distinct signalling profiles that would explain pro-resolution versus inflammatory physiology, and the most likely explanation for these data would be a combination of FPR1 and FPR2 responses.