C70-20 Persistent Inflammatory Airway Epithelial Remodeling After Long Term Cftr Modulator Therapy

Abstract Introduction The cystic fibrosis (CF) airway epithelium is characterized by detrimental structural and functional changes (termed remodeling) associated with chronic inflammation and infection secondary to CFTR ion channel dysfunction. Elexacaftor+tezacaftor+ivacaftor (ETI) therapy improves CFTR function and clinical outcomes for people with CF (pwCF) but long term impact on airway epithelial remodeling is still unclear. By studying an adult cohort of pwCF over two years, we recently demonstrated that ETI decreases inflammation and improves remodeling. Here we compare pwCF on ETI to healthy donors to investigate the extent to which remodeling is mitigated by long-term ETI treatment. Methods Single cell RNA sequencing (scRNAseq) was used to compare cellular compositional and molecular programming changes between adult pwCF after ∼6 months of ETI and age matched healthy donors using nasal brush biopsies. Spatial transcriptomics was used to probe interactions between epithelial, immune, and structural cell compartments in CF (with and without ETI) and healthy donor lungs. Outcomes were validated using airway tissue immunofluorescence, primary airway epithelial air-liquid interface cultures (ALIs), and airway stem cell self-renewal assays. Results scRNAseq demonstrated prominent differences between pwCF on ETI and healthy controls consistent with persistent inflammation, ciliated cell dysfunction, and mucous and squamous metaplasia in CF. Data indicated that the CF airways remain infiltrated by more neutrophils, which have activated antimicrobial sensing pathways and are primed for NETosis and cytolytic activity despite ETI. Mononuclear phagocyte populations in CF were dominated by monocytes and monocyte derived macrophages with aberrant programming that were molecularly similar to those identified in non-ETI treated CF sputum by a prior study. CF epithelial cells expressed significantly higher levels of interferon signaling related genes, but largely failed to upregulate class I and II major histocompatibility (MHC) family gene expression, indicating aberrant inflammatory signaling. The airway basal stem cell pool remained compositionally and functionally altered in CF with lower self-renewal and differentiation capacity. Spatial transcriptomics identified focally altered basal stem cell pool heterogeneity underlying areas of mucous vs. squamous remodeling, which may receive differential input from nearby immune and structural cells. Conclusion ETI leads to improvements in CF epithelial inflammation and dysfunction, which are not fully rectified when ETI is initiated in adults with existing airway disease and may rebound during the course of long-term therapy. The airway stem cell pool remains focally disrupted. Unraveling the molecular mechanisms driving these responses will have implications for long-term ETI use and potentially future gene and cell therapies. This abstract is funded by: CFF, NIH