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CELLULAR AND MOLECULAR

Discovery of -Aminoazaheterocycle-Methylglyoxal Adducts as a New Class of High-Affinity Inhibitors of Cystic Fibrosis Transmembrane Conductance Regulator Chloride Channels

Département de Pharmacochimie Moléculaire, Centre National de la Recherche Scientifique/Université de Grenoble-Joseph Fourier, Grenoble, France (C.R., M.-C.M., B.B., J.-L.D.); Institut de Physiologie et Biologie Cellulaires, Université de Poitiers, Centre National de la Recherche Scientifique, Poitiers, France (C.N., P.M., S.N., R.R., Fr.B.); and Institut du thorax, Institut National de la Santé et de la Recherche Médicale Unité 533, Nantes, France (Fl.B., C.G.)

The cystic fibrosis transmembrane conductance regulator (CFTR) represents the main Cl^– channel in the apical membrane of epithelial cells for cAMP-dependent Cl^– secretion. Here we report on the synthesis and screening of a small library of nontoxic -aminoazaheterocycle-methylglyoxal adducts, inhibitors of wild-type (WT) CFTR and G551D-, G1349D-, and F508del-CFTR Cl^– channels. In whole-cell patch-clamp experiments of Chinese hamster ovary (CHO) cells expressing WT-CFTR, we recorded rapid and reversible inhibition of forskolin-activated CFTR currents in the presence of the adducts 5a and 8a,b at 10 pM concentrations. Using iodide efflux experiments, we compared concentration-dependent inhibition of CFTR with glibenclamide (IC₅₀ = 14.7 µM), 3-[(3-trifluoromethyl)phenyl]-5-[(4-carboxyphenyl-)methylene]-2-thioxo-4-thiazolidinone (CFTR_inh-172) (IC₅₀ = 1.2 µM), and -aminoazaheterocycle-methylglyoxal adducts and identified compounds 5a (IC₅₀ = 71 pM), 8a,b (IC₅₀ = 2.5 nM), and 7a,b (IC₅₀ = 3.4 nM) as the most potent inhibitors of WT-CFTR channels. Similar ranges of inhibition were also found when these compounds were evaluated on CFTR channels with the cystic fibrosis mutations F508del (in temperature-corrected human airway epithelial F508del/F508del CF15 cells)-, G551D-, and G1349D-CFTR (expressed in CHO and COS-7 cells). No effect of compound 5a was detected on the volume-regulated or calcium-regulated iodide efflux. Picomolar inhibition of WT-CFTR with adduct 5a was also found using a 6-methoxy-N-(3-sulfopropyl)-quinolinium fluorescent probe applied to the human tracheobronchial epithelial cell line 16HBE14o–. Finally, we found comparable inhibition by 5a or by CFTR_inh-172 of forskolin-dependent short-circuit currents in mouse colon. To the best of our knowledge, these new nontoxic -aminoazaheterocycle-methylglyoxal adducts represent the most potent compounds reported to inhibit CFTR chloride channels.

Address correspondence to: Dr. Jean-Luc Décout, Département de Pharmacochimie Moléculaire, Université de Grenoble, Centre National de la Recherche Scientifique, Bât. E, rue de la Chimie, BP 53, 38041 Grenoble Cedex 9, France. E-mail: jean-lucdecout{at}ujf-grenoble.fr

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