DEVELOPING NOVEL MODEL SYSTEMS AND TREATMENTS FOR CYSTIC FIBROSIS

Cystic fibrosis (CF) is caused by loss-of-function mutations in the CF transmembrane regulator (CFTR) gene. Pharmacotherapies directed at CFTR have been developed but are not effective for mutations that reduce mRNA or protein expression. Replacing airway epithelial cells carrying mutant CFTR with cells that express a normal copy of the gene (i.e., cell therapy) has the potential to serve as a one-time cure for CF lung disease regardless of the causative CFTR mutation. This dissertation describes novel model systems and advancements in the development of CF cell therapy. In chapter 1, I contextualize the major challenges facing translation of cell therapy for CF by summarizing primary literature and describing the current state of the field. Chapter 2 compares the proliferative and competitive properties of airway epithelial cell progenitor populations with the goal of finding the optimal cell candidate for CF cell therapy. In chapter 3, I develop an in vitro model of cell engraftment using the air-liquid interface (ALI) culture system in which well-differentiated CF ALI cultures are injured and non-CF cells are delivered at the peak of injury. Using this model, we found that long-term cell engraftment requires severe injury with partial epithelial denudation. In chapter 4, I develop methods to culture airway tissue explants for at least 14 days ex vivo. Airway tissue explants can be created from human or animal tissue, can be cryopreserved without loss of viability, and could serve as a translational steppingstone between in vitro and in vivo studies. In chapter 5, we develop airway epithelial cell lines from cell donors carrying rare CFTR variants. These cell lines recapitulated primary cell morphology and mirrored the parent primary cell and cell donors’ clinical response to CFTR modulators. I then used cell lines carrying premature termination codon (PTC) CFTR variants to assess eukaryotic release factor 3a (eRF3a) degraders for their ability to promote ribosomal readthrough and rescue CFTR function. Finally, in chapter 6, I summarize major findings, acknowledge remaining gaps in knowledge, and propose future studies. In summary, this work establishes and employs novel model systems to develop therapies for CF.