title: Identification of a tetrahydroquinoline analog as a pharmacological inhibitor of the cAMP-binding protein Epac.
abstract: The cAMP-binding protein Epac is a therapeutic target for the treatment of various diseases such as cardiac hypertrophy and tumor invasion. This points out the importance to develop Epac inhibitors to better understand the involvement of these cAMP sensors in physiology and pathophysiology. Here, we have developed a functional fluorescence-based high-throughput assay with a Z' value around 0.7 for screening Epac-specific antagonists. We identified an Epac1 inhibitor compound named CE3F4 that blocked Epac1 guanine nucleotide exchange activity toward its effector Rap1 both in cell-free systems and in intact cells. CE3F4 is a tetrahydroquinoline analog that fails to influence protein kinase A holoenzyme activity. CE3F4 inhibited neither the interaction of Rap1 with Epac1 nor directly the GDP exchange on Rap1. The kinetics of inhibition by CE3F4 indicated that this compound did not compete for binding of agonists to Epac1 and suggested an uncompetitive inhibition mechanism with respect to Epac1 agonists. A structure-activity study showed that the formyl group on position 1 and the bromine atom on position 5 of the tetrahydroquinoline skeleton were important for CE3F4 to exert its inhibitory activity. Finally, CE3F4 inhibited Rap1 activation in living cultured cells, following Epac activation by either 8-(4-chlorophenylthio)-2'-O-methyl-cAMP, an Epac-selective agonist, or isoprenaline, a non-selective β-adrenergic receptor agonist. Our study shows that CE3F4 and related compounds may serve as a basis for the development of new therapeutic drugs.
authors.affiliation: Université Paris-Sud, CIBLOT-IFR 141, Faculté de Pharmacie, 92296 Châtenay-Malabry Cedex, France.
authors.forename: Delphine
authors.initials: D
authors.lastname: Courilleau
authors.forename: Malik
authors.initials: M
authors.lastname: Bisserier
authors.forename: Jean-Christophe
authors.initials: JC
authors.lastname: Jullian
authors.forename: Alexandre
authors.initials: A
authors.lastname: Lucas
authors.forename: Pascal
authors.initials: P
authors.lastname: Bouyssou
authors.forename: Rodolphe
authors.initials: R
authors.lastname: Fischmeister
authors.forename: Jean-Paul
authors.initials: JP
authors.lastname: Blondeau
authors.forename: Frank
authors.initials: F
authors.lastname: Lezoualc'h
mesh_terms: D002352:Carrier Proteins; D000242:Cyclic AMP / Q000737:chemistry / Q000378:metabolism; D004353:Drug Evaluation, Preclinical; D020662:Guanine Nucleotide Exchange Factors / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D007700:Kinetics; D011485:Protein Binding / Q000187:drug effects; D011804:Quinolines / Q000737:chemistry / Q000494:pharmacology*; D020827:rap1 GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D002352:Carrier Proteins; D020662:Guanine Nucleotide Exchange Factors; D011804:Quinolines; C116122:RAPGEF3 protein, human; C542964:1,2,3,4-tetrahydroquinoline; D000242:Cyclic AMP; D020827:rap1 GTP-Binding Proteins