Integration of virtual screening with high-throughput flow cytometry to identify novel small molecule formylpeptide receptor antagonists.
The formylpeptide receptor (FPR) family of G-protein-coupled receptors contributes to the localization and activation of tissue-damaging leukocytes at sites of chronic inflammation. We developed a FPR homology model and pharmacophore (based on the bovine rhodopsin crystal structure and known FPR ligands, respectively) for in silico screening of approximately 480,000 drug-like small molecules. A subset of 4324 compounds that matched the pharmacophore was then physically screened with the HyperCyt flow cytometry platform in high-throughput, no-wash assays that directly measure human FPR binding, with samples (each approximately 2500 cells in 2 microl) analyzed at 40/min. From 52 confirmed hits (1.2% hit rate), we identified 30 potential lead compounds (inhibition constant, Ki= 1-32 microM) representing nine distinct chemical families. Four compounds in one family were weak partial agonists. All others were antagonists. This virtual screening approach improved the physical screening hit rate by 12-fold (versus 0.1% hit-rate in a random compound collection), providing an efficient process for identifying small molecule antagonists.
Edwards
Bruce S
BS
Department of Cytometry, University of New Mexico, Albuquerque, NM 87131-0001, USA.
Bologa
Cristian
C
Young
Susan M
SM
Balakin
Konstantin V
KV
Prossnitz
Eric R
ER
Savchuck
Nikolay P
NP
Sklar
Larry A
LA
Oprea
Tudor I
TI
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D004353:Drug Evaluation, Preclinical; D005434:Flow Cytometry / Q000379:methods*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D044042:Receptors, Formyl Peptide / Q000819:agonists / Q000037:antagonists & inhibitors* / Q000737:chemistry
D044042:Receptors, Formyl Peptide