Impaired intestinal free fatty acid transport followed by chylomicron malformation, not pancreatic insufficiency, cause metabolic defects in cystic fibrosis.

Intestinal disease is one of the earliest manifestations of cystic fibrosis (CF) in children and is closely tied to deficits in growth and nutrition, both of which are directly linked to future mortality. Patients are treated aggressively with pancreatic enzyme replacement therapy and a high-fat diet to circumvent fat malabsorption, but this does not reverse growth and nutritional defects. We hypothesized that defects in chylomicron production could explain why CF body weights and nutrition are so resistant to clinical treatments. We used gold standard intestinal lipid absorption and metabolism approaches, including mouse mesenteric lymph cannulation, in vivo chylomicron secretion kinetics, transmission electron microscopy, small intestinal organoids, and chylomicron metabolism assays to test this hypothesis. In mice expressing the G542X mutation in cystic fibrosis transmembrane conductance regulator (CFTR-/- mice), we find that defective FFA trafficking across the epithelium into enterocytes drives a chylomicron formation defect. Furthermore, G542X mice secrete small, triglyceride-poor chylomicrons into the lymph and blood. These defective chylomicrons are cleared into extraintestinal tissues at ∼10-fold faster than WT chylomicrons. This defect in FFA absorption resulting in dysfunctional chylomicrons cannot be explained by steatorrhea or pancreatic insufficiency and is maintained in primary small intestinal organoids treated with micellar lipids. These studies suggest that the ultrahigh-fat diet that most people with CF are counselled to follow may instead make steatorrhea and malabsorption defects worse by overloading the absorptive capacity of the CF small intestine.

In vivo editing of lung stem cells for durable gene correction in mice.

In vivo genome correction holds promise for generating durable disease cures; yet, effective stem cell editing remains challenging. In this work, we demonstrate that optimized lung-targeting lipid nanoparticles (LNPs) enable high levels of genome editing in stem cells, yielding durable responses. Intravenously administered gene-editing LNPs in activatable tdTomato mice achieved >70% lung stem cell editing, sustaining tdTomato expression in >80% of lung epithelial cells for 660 days. Addressing cystic fibrosis (CF), NG-ABE8e messenger RNA (mRNA)-sgR553X LNPs mediated >95% cystic fibrosis transmembrane conductance regulator (CFTR) DNA correction, restored CFTR function in primary patient-derived bronchial epithelial cells equivalent to Trikafta for F508del, corrected intestinal organoids and corrected R553X nonsense mutations in 50% of lung stem cells in CF mice. These findings introduce LNP-enabled tissue stem cell editing for disease-modifying genome correction.