Cystic fibrosis (CF) is the most common lethal genetic disorder in Caucasians, affecting approximately 1 in 2500 newborns and is due to mutations in the gene for the cystic fibrosis transmembrane conductance regulator (CFTR). Despite gradual improvements in treatment over the past twenty years, most individuals with CF currently do not survive past age 30, and new approaches to treatment are urgently needed. This proposal will investigate the basic pathophysiologic defect produced by a range of CFTR mutations in epithelial cells, with the long range goal of developing specific pharmacological reagents which will activate the mutant protein product. We have already shown that the most common mutation (deltaF508) and a number of other missinse mutations produce CFTR in Xenopus oocytes which can be activated by high levels of cyclic AMP. This observation will now be extended to a variety of mammalian epithelial cell lines, studying the effect of expression of wild-type and mutant forms of CFTR on chloride channel conductance. The intracellular processing of wild-type and mutant CFTR will be simultaneously studied using a specific antibody. An explanation for the severity and organ specificity of particular CF mutations will be sought by comparison of the in vitro properties of CFTR, expressed in different epithelial cells, with the in vivo phenotype. Perhaps most importantly, pharmacologic agents will be sought which have the potential to activate deltaF508 and other mutant CFTR proteins in the airway; from preliminary data on Xenopus oocytes, phosphodiesterase inhibitors, agents which stimulate adenylate cyclase, and phosphatase inhibitors are all worth investigating.
