Deletion of Phe-508 (Delta F508) is the most common mutation in the cystic fibrosis transmembrane conductance regulator (CFTR) causing cystic fibrosis. Delta F508-CFTR has defects in both channel gating and endoplasmic reticulum-to-plasma membrane processing. We identified six novel classes of high affinity potentiators of defective Delta F508-CFTR Cl- channel gating by screening 100,000 diverse small molecules. Compounds were added 15 min prior to assay of iodide uptake in epithelial cells co-expressing Delta F508-CFTR and a high sensitivity halide indicator (YFP-H148Q/I152L) in which Delta F508-CFTR was targeted to the plasma membrane by culture at 27 degrees C for 24 h. Thirty-two compounds with submicromolar activating potency were identified; most had tetrahydrobenzothiophene, benzofuran, pyramidinetrione, dihydropyridine, and anthraquinone core structures (360-480 daltons). Further screening of >1000 structural analogs revealed tetrahydrobenzothiophenes that activated DeltaF508-CFTR Cl- conductance reversibly with Kd < 100 nm. Single-cell voltage clamp analysis showed characteristic CFTR currents after Delta F508-CFTR activation. Activation required low concentrations of a cAMP agonist, thus mimicking the normal physiological response. A Bayesian computational model was developed using tetrahydrobenzothiophene structure-activity data, yielding insight into the physical character and structural features of active and inactive potentiators and successfully predicting the activity of structural analogs. Efficient potentiation of defective Delta F508-CFTR gating was also demonstrated in human bronchial epithelial cells from a Delta F508 cystic fibrosis subject after 27 degrees C temperature rescue. In conjunction with correctors of defective Delta F508-CFTR processing, small molecule potentiators of defective Delta F508-CFTR gating may be useful for therapy of cystic fibrosis caused by the Delta F508 mutation.

Nanomolar affinity small molecule correctors of defective ΔF508-CFTR chloride channel gating / Yang, Hong; Shelat, Anang A.; Guy, R. Kiplin; Gopinath, Vadiraj S.; Ma, Tonghui; Du, Kai; Lukacs, Gergely L.; Taddei, Alessandro; Folli, Chiara; Pedemonte, Nicoletta; Galietta, Luis J. V.; Verkman, A. S.. - In: THE JOURNAL OF BIOLOGICAL CHEMISTRY. - ISSN 0021-9258. - 278:37(2003), pp. 35079-35085. [10.1074/jbc.M303098200]

Nanomolar affinity small molecule correctors of defective ΔF508-CFTR chloride channel gating

Galietta, Luis J. V.;
2003

Abstract

Deletion of Phe-508 (Delta F508) is the most common mutation in the cystic fibrosis transmembrane conductance regulator (CFTR) causing cystic fibrosis. Delta F508-CFTR has defects in both channel gating and endoplasmic reticulum-to-plasma membrane processing. We identified six novel classes of high affinity potentiators of defective Delta F508-CFTR Cl- channel gating by screening 100,000 diverse small molecules. Compounds were added 15 min prior to assay of iodide uptake in epithelial cells co-expressing Delta F508-CFTR and a high sensitivity halide indicator (YFP-H148Q/I152L) in which Delta F508-CFTR was targeted to the plasma membrane by culture at 27 degrees C for 24 h. Thirty-two compounds with submicromolar activating potency were identified; most had tetrahydrobenzothiophene, benzofuran, pyramidinetrione, dihydropyridine, and anthraquinone core structures (360-480 daltons). Further screening of >1000 structural analogs revealed tetrahydrobenzothiophenes that activated DeltaF508-CFTR Cl- conductance reversibly with Kd < 100 nm. Single-cell voltage clamp analysis showed characteristic CFTR currents after Delta F508-CFTR activation. Activation required low concentrations of a cAMP agonist, thus mimicking the normal physiological response. A Bayesian computational model was developed using tetrahydrobenzothiophene structure-activity data, yielding insight into the physical character and structural features of active and inactive potentiators and successfully predicting the activity of structural analogs. Efficient potentiation of defective Delta F508-CFTR gating was also demonstrated in human bronchial epithelial cells from a Delta F508 cystic fibrosis subject after 27 degrees C temperature rescue. In conjunction with correctors of defective Delta F508-CFTR processing, small molecule potentiators of defective Delta F508-CFTR gating may be useful for therapy of cystic fibrosis caused by the Delta F508 mutation.
2003
Nanomolar affinity small molecule correctors of defective ΔF508-CFTR chloride channel gating / Yang, Hong; Shelat, Anang A.; Guy, R. Kiplin; Gopinath, Vadiraj S.; Ma, Tonghui; Du, Kai; Lukacs, Gergely L.; Taddei, Alessandro; Folli, Chiara; Pedemonte, Nicoletta; Galietta, Luis J. V.; Verkman, A. S.. - In: THE JOURNAL OF BIOLOGICAL CHEMISTRY. - ISSN 0021-9258. - 278:37(2003), pp. 35079-35085. [10.1074/jbc.M303098200]
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11588/739374
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