The Screening And Molecular Pharmacology Study Of Natural Flavonoid Cystic Fibrosis Transmembrane Conductance Regulator(CFTR) Chloride Channel Activators

Cystic fibrosis transmembrane conductance regulator(CFTR)protein is a kind of Cl- channel, which belong to the the super family of ATP-binding cassette (ABC) transporters.Its main function is to mediate cAMP-dependent Cl- secretion in various epithelia and it also interacts with other ion channels.The mutations of CFTR cause cystic fibrosis (CF)which is the most prevalent lethal autosomal recessive genetic disease in Caucasians . Until now there are more than 1300 mutations known in CFTR gene .Among them over 66% mutations are caused by the deletion of the Phe-508 (ΔF508-CFTR) which is retarded at the endoplasmic reticulum and fail to be located on cell plasma membrane, due to misfolding and /or defective interactions with molecular chaperons and leads to channel function defective. Even presence at the apical membrane, the channel function can restore just after stimulated by the CFTR activators. Utilizing the combinatorial chemistry in the past few years, we got some small molecule activators, which could stimulateΔF508-CFTR .But these activators had low affinity and their security called in question. So recent years the emphasis become to find some new activators that have little side effects .A stably transfected epithelial cell line co-expressingΔF508-CFTR or Wild type- CFTR(Wt-CFTR) and a green fluorescent protein mutant with ultra-high halide sensitivity (YFP-H148Q/I152L) was generated as assay for the drug screening. Using this cell model, 15 flavonoid compounds including Baicalein,Farrerol and so on, were screened for promoting CFTR-mediated halide transport. We identified two compounds (Baicalein and Farrerol) with strong activity and two compounds (Silymarin and Lysionotin) with relatively weak activity in activatingΔF508-CFTR and wt-CFTR Cl- channel. Pharmacological properties of these compounds were characterized systematically in terms of the dose-response, reversibility, activition time-course and so on. The four compounds could activate defectiveΔF508-CFTR Cl- channel as well as Wt-CFTR Cl- channel in a dose-dependent manner. Half of the maximum activations (Vmax) of the compouds could be reached within 10 minutes and the activation were reversible. Preliminary structure-activity analysis indicated that the existence of hydroxyl at both 6 and 8 position are imperative for the activity of flavonoid compounds. The four compounds are fast-acting and reversibleΔF508-CFTR potentiators that worth further in vivo pharmacological studies as candidate CF drugs.