Abstract

Trikafta (elexacaftor/tezacaftor/ivacaftor, ETI) is approved for cystic fibrosis (CF) patients with at least one F508del mutation in the CFTR gene or another responsive mutation based on in vitro data. However, the pharmacological effects of ETI on F508del-CFTR remain incompletely defined in vitro. To explore the mechanisms underlying Trikafta’s clinical efficacy, we used primary bronchial epithelial cells from F508del homozygous patients and CFBE41o- cells expressing F508del-CFTR. We assessed CFTR maturation, turnover, chloride transport, and thermal stability under various ETI concentrations and treatment durations at physiological temperature using electrophysiology (Ussing chamber, patch-clamp) and biochemical assays. We found that ETI efficacy on F508del-CFTR is strongly influenced by both treatment duration and concentration. Reducing ETI from standard doses, i.e. E (3 µM), T (18 µM), I (1 µM), to 33%, 11%, 3.3%, and 1.1% decreased function and maturation, but 33% retained most of the corrective effect. After 2 hours of treatment, around 50% of the CFTR-dependent current was preserved, unlike in untreated cells. Notably, replacing elexacaftor with bamocaftor further improved F508del-CFTR maturation and function compared to ETI, though it did not affect the rate of current decline over time. These findings highlight the importance of optimizing ETI dose and exposure duration, as both significantly affect F508del-CFTR stability and function. The retained efficacy at reduced 44
concentrations suggests possible individualized dosing strategies, particularly for patients experiencing adverse effects with full-dose ETI.