Impact of CFTR Correctors on the Channel Activity of Rescued F508del CFTR in Airway Epithelial Cells

Abstract

Cystic fibrosis (CF) is caused by the functional deficiency of a cAMP-activated anion channel known as the cystic fibrosis transmembrane conductance regulator (CFTR). Over 90% of CF patients carry at least one copy of the F508del CFTR gene. The F508del mutation causes a global conformational change that prevents the nascent protein from exiting the endoplasmic reticulum, leading to loss of function at the cell surface. CFTR correctors have been developed to bind to the nascent F508del CFTR, change its conformation, and mobilize it to the plasma membrane. Rescued F508del CFTR (rF508del) does not fully regain channel activity. Lumacaftor, the first CFTR corrector approved by the FDA, acutely inhibits rF508del channel opening (Ambrosetti et al. Pediatric Pulmonology 54(S2): 190-191). Our objective is to evaluate the impact of the other two FDA-approved CFTR correctors, tezacaftor and elexacaftor, on rF508del channel activity. We hypothesize that different CFTR correctors impact rF508del channel activity differently. We utilized an electrode-based, real-time iodide efflux assay to evaluate the acute impact of CFTR correctors on the cAMP-dependent anion channel activity of rF508del in a CF bronchial epithelial cell line stably expressing F508del CFTR (CFBE-DF) at 37°C. CFBE-DF cells were incubated with a corrector for over 24 hours before iodide efflux assay was performed in the acute presence or absence of the same corrector. Genistein was used as a potentiator. F508del processing was assessed by quantitative immunoblotting. The lack of acute exposure to 5μM lumacaftor results in a 2% increase in the mean F508del processing (n=3, p=0.94) and a 48% increase in channel activity (n=3, p=0.04). F508del processing peaks at 1μM lumacaftor but the channel activity peaks at a much lower 0.05μM lumacaftor. In contrast, the lack of acute exposure to 5μM tezacaftor results in an 8% reduction in the mean channel activity (n>3, p=0.79) and a 17% reduction in the mean F508del processing (n>3, p=0.43). Both F508del processing and channel activity peak at 5μM tezacaftor. The lack of acute exposure to 2μM elexacaftor results in an 11% decrease in the mean F508del channel activity (n=4. p=0.54). F508del channel activity peaks at 2μM elexacaftor. We conclude that, unlike lumacaftor, tezacaftor and elexacaftor do not inhibit the channel activity of rescued F508del CFTR in CFBE-DF cells. A better understanding of the impact of CFTR correctors on the channel activity of rescued CFTR processing mutants will inform the development of highly effcacious CFTR modulator combinations for the treatment of CF patients. This work is supported by NIH (5P30DK072482), Samford University, and Cystic Fibrosis Foundation. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.