PROJECT SUMMARY/ABSTRACT Recently developed and approved therapies for cystic fibrosis (CF) consist of small-molecule modulators that aid in folding, trafficking, and channel gating of mutant CF transmembrane conductance regulator (CFTR). However, these advances still only partially restore function of the most common mutant of CFTR. The goal of this Fellowship is to explore the efficacy and mechanism of action of a new class of CFTR modulators―prostones―that could serve as CFTR activators to be used in combination with currently available structure-correcting CFTR modulators to restore mutant CFTR function to levels comparable with wildtype CFTR. The guiding hypothesis is that the prostaglandin derivative lubiprostone (an FDA-approved drug developed to treat chronic constipation) acts through endogenous prostaglandin signaling pathways to specifically activate wildtype and modulator-rescued mutant CFTR in airway epithelia. In preliminary studies, it was observed that lubiprostone increases CFTR-mediated short-circuit current in non-CF and modulator-rescued CF primary- derived nasal epithelial cultures. Furthermore, it was observed that pharmacological blockade of prostaglandin receptors abolished the action of lubiprostone on CFTR activity. Specific Aim 1 will more thoroughly quantitatively evaluate the effect of lubiprostone on CFTR function in non-CF and CF airway epithelia. In this Aim, the efficacy of lubiprostone in affecting CFTR activity and transepithelial fluid transport will be explored in common and rare mutations of CFTR that lead to CF and in primary-cultured epithelia derived from both upper (nasal) and lower (bronchial) regions of the airway. Specific Aim 2 seeks to elucidate the mechanism of action of prostones in the airway by identifying the roles of prostaglandin signaling pathways in mediating endogenous CFTR activity, as well as CFTR activation by lubiprostone. In this Aim, the signaling pathway of the endogenous prostaglandin, prostaglandin E2, will be explored. The extent and mechanism-of-action by which prostaglandin E2 is responsible for regulating CFTR activity on the airway will be investigated. Lastly, in preparation for drug development, rigorous ligand-receptor binding studies will be performed to quantify and compare lubiprostone-receptor binding properties to functional properties of lubiprostone activation of CFTR in airway epithelia. The current project will provide invaluable technical and professional training in support of the applicant’s career goals. This work has the promise to establish an FDA-approved drug, lubiprostone, as an additional pharmacological tool in restoring function to mutant CFTR for the treatment of CF. Furthermore, by elucidating the mechanism of action of CFTR activation by lubiprostone, this work will provide a better understanding of the cellular pathways regulating endogenous CFTR activity in airway epithelia and shed light on how these natural mechanisms of CFTR activation can be ph...