P170 Trikafta® modulates release of extracellular vesicles in cystic fibrosis

Abstract

Dysregulation of CFTR leads to chronic inflammation in Cystic Fibrosis (CF) airways. Extracellular vesicles (EVs) have emerged as an important biomarkers of disease. We previously reported elevated levels of EVs in CF with distinct age-related protein profiles. The levels of EVs and its associated proteins in CF in the presence of CF modulators Trikafta (elexacaftor, tezacaftor, ivacaftor), however remains unexplored. The study aims to characterise and study the effect of Trikafta on EVs and its associated protein targets in CF airway models and serum samples from patients with cystic fibrosis (PWCF). Bronchial brushings from CF patients were passaged and differentiated by air liquid interface (ALI) cultures. After 21 days, TEER was recorded and differentiation of mucociliary layer was confirmed with MUC5AC staining. Further the cultures were treated with Trikafta. EVs released into the apical and basal compartments before and after treatment were extracted by differential centrifugation and characterised by NTA and mass spectrometry (MS). The therapeutic restoration of CFTR was examined using immunoblotting. Serum samples from PWCF on Trikafta were also examined for EV release and content before and after 12 months treatment. After 21 days, of ALI culture. EVs were isolated before and after treatment with Trikafta showed a significant 1.5-fold decrease in EVs in both apical and basal compartments respectively (PWCF (Apical) Control: 4.2E+07 ± 3E+06; Treated 3.2E+07 ± 2E+06; (Basal) Control: 1.3E +08 ± 1.6E+07; Treated 8.1E+07 ± 5E+06) demonstrating that Trikafta treatment diminishes epithelial EV releases. MS based proteomics revealed unique EV protein fingerprints post Trikafta treatment for both epithelial EVs and serum EVs with changes in expression of immune proteins were observed. The study demonstrates that Trikafta can modulate EV release and content in PWCF demonstrating novel mechanisms by which CFTR modulators can regulate the inflammatory landscape in CF.