Homeostasis of the airway surface liquid (ASL) in lung is critically dependent on chloride ion (Cl-) transport, mediated by the apical channel, CFTR (cystic fibrosis transmembrane conductance regulator). CFTR gene mutations associated with the most common autosomal recessive disease in Caucasians, cystic fibrosis (CF) result in loss of CFTR function and severe impairment of the ASL volume regulation. Deletion of F508 (F508del) in the CFTR gene is present in 90% of CF patients; it blocks CFTR biosynthetic processing, reduces CFTR Cl- channel function, and decreases CFTR mRNA stability and translation. It is estimated that the classic severe CF phenotype develops when the CFTR channel function is less than 1% of normal and at least 10% function is needed to alleviate the severe phenotype. Therapies targeting the basic molecular defects in CF demonstrate potential but are insufficient for most patients. While the FDA-approved drug VX-770 potentiates CFTR channel ?open? probabilities and improves lung function for less than 10% of patients with the rare mutation G551D, the combined use of VX-770 and a corrector VX-809 that rescues the folding and processing defect was only marginally effective for F508del patients. CF patients with more severe lung disease were entirely resistant to such therapy. Transforming growth factor (TGF)-?1 contributes to resistance of corrector therapy by blocking CFTR translation and represses ancillary ion channels critical to residual ASL homeostasis in CF, namely BK (a Ca+2-activated K+ channel) and ANO1 (a Ca+2-activated Cl- channel). High TGF-?1 levels are seen in 40% of F508del homozygous patients due to polymorphisms in the TGF-?1 gene. Air pollutants, including cigarette smoke also increase TGF-?1 levels. The concomitant upregulation of TGF-?1 in turn increases the severity of CF lung disease in F508del homozygous patients and presents a major block to therapies aimed at F508del-CFTR rescue. We propose that targeting the pathogenic TGF-?1 activity would improve the residual ASL volume homeostasis in CF by restoring the function of BK and ANO1 channels. It would also increase the efficacy of therapy for F508del CF patients by restoring the diminished 508del-CFTR translation and allowing increased activity of the mutant CFTR channel function. Thus, defining TGF-?1 mediators in lung tissue presents an opportunity to design novel drugs eliminating the pathogenic TGF-?1 activity in CF patients. Our central hypothesis is that the targeted reduction of TGF-?1 activity will ameliorate negative effects on CFTR, BK, and ANO1 channels and ASL homeostasis, and provide a novel approach to treat F508del CF patients.
Our specific aims are to test the hypothesis: (1) that Dab2 is a TGF-?1 adaptor that inhibits the residual ASL homeostasis in CF bronchial epithelium by directing nuclear transport of Smad3 to repress the ancillary channels, BK and ANO1; (2) that the Dab2-Smad3 interactions upregulates microRNAs that repress the ancillary channels and block corrector-mediated rescue of F508del-CFTR; and (3) that a Dab2-Smad3 interaction is required for the pathogenic activity of TGF-?1 in CF bronchial epithelium.
A molecule called CFTR controls the movement of salt and water to maintain normal lung function. Severe lung disease results from the absent CFTR function in a disease called cystic fibrosis (CF). Ancillary molecules provide small compensation for the absent CFTR function in CF but TGF-beta, a protein produced by patients, inhibits the molecules, and destroys the ability of medications to restore the absent CFTR function. Learning how to block TGF-beta will help to treat CF lung disease and reduce the suffering and healthcare cost associated with CF and with many other common forms of lung disease exacerbated by TGF-beta, which we propose in this application.