PROJECT SUMMARY Cystic fibrosis (CF) is a progressive disease affecting around 30,000 people in the US and is caused by a mutation in a gene affecting the CFTR protein that regulates mucus composition. In airways in the lung this leads to mucus stasis, infection, and remodeling that result in mucus plugs, poor ventilation, and progressive airway destruction. Highly-effective CFTR modulators, now available to >90% of patients, have revolutionized clinical care, with increases in pulmonary function via more effective mucociliary clearance. Burdensome maintenance therapies like airway clearance treatment (ACT) require around 2 dedicated hours per day and have been questioned by patients, families, and medical providers. In a recent CF-community survey, airway clearance was ranked as the most burdensome therapy. Traditional studies of therapy withdrawal pose some patient risk, since measurement is via relatively insensitive pulmonary function testing (PFT) and lung-function reductions can have permanent consequences. Breakthroughs in structural and hyperpolarized-gas MRI demonstrate exquisite sensitivity to CF lung disease and can be used to monitor regional and subtle changes over time, much more precisely than PFTs, and with regional specificity. MRI provides a unique opportunity to safely evaluate ACT. The overarching goal of our proposal is to determine effectiveness of ACT in the era of highly effective CFTR modulators by studying structure-function relationships via MRI in patients of varying severity who have stopped and restarted ACT. We will achieve this goal via three separate, hypothesis-driven Aims in this clinical trial: Hypothesis 1: Patients who have relatively low structural defects will have fewer ventilation defects and higher pulmonary function, and these defects will relate to frequency of ACT usage. Specific Aim 1: To perform UTE and hyperpolarized Xe MRI in 30 CF patients aged 12-21, approximately 15 of whom have self-withdrawn ACT, to regionally characterize obstructive severity and correlate regional structural lung abnormalities (via UTE FLORET MRI) to functional deficits (via Xe MRI) Hypothesis 2: Patients who have self-withdrawn ACT after initiation of effective modulators will demonstrate increases in regional ventilation after reinitiating treatment, with greater ventilation increases in patients with a higher level of lung abnormalities. Specific Aim 2: To perform a stepwise ACT re-initiation trial in fifteen 12-21 y.o. patients who have self-withdrawn airway clearance treatment (defined as ≤ 3x/week). UTE and Xe MRI, spirometry, and multiple-breath washout will be performed at baseline, after increasing treatment to 7x/week for 1 week and then 14x/week for 2 weeks, with daily logging to aid compliance and study engagement. Hypothesis 3: Patients who currently use 2x daily ACT and have few lung abnormalities on MRI can reduce to 1x daily or less, with no significant functional changes in the lung. Specific Aim 3: To per...