rAAV-CFTRAR Rescues the Cystic Fibrosis Phenotype in Human Intestinal Organoids and Cystic Fibrosis Mice

Rationale: Gene therapy holds promise for a curative mutation independent treatment applicable to all patients with cystic fibrosis (CF). The various viral vector-based clinical trials conducted in the past have demonstrated safety and tolerance of different vectors, but none have led to a clear and persistent clinical benefit. Recent clinical breakthroughs in recombinant adeno-associated viral vector (rAAV)-based gene therapy encouraged us to reexplore an rAAV approach for CF. Objectives: We evaluated the preclinical potential of rAAV gene therapy for CF to restore chloride and fluid secretion in two complementary models: intestinal organoids derived from subjects with CF and a CF mouse model, an important milestone toward the development of a clinical rAAV candidate for CF gene therapy. Methods: We engineered an rAAV vector containing a truncated CF transmembrane conductance regulator (CFTR Delta R) combined with a short promoter (CMV173) to ensure optimal gene expression. A rescue in chloride and fluid secretion after rAAV-CFTR Delta R treatment was assessed by forskolin-induced swelling in CF transmembrane conductance regulator (CFTR)-deficient organoids and by nasal potential differences in Delta F508 mice. Measurements and Main Results: rAAV-CFTR Delta R transduction of human CFTR-deficient organoids resulted in forskolin-induced swelling, indicating a restoration of CFTR function. Nasal potential differences demonstrated a clear response to low chloride and forskolin perfusion in most rAAV-CFTR Delta R-treated CF mice. Conclusions: Our study provides robust evidence that rAAV-mediated gene transfer of a truncated CFTR functionally rescues the CF phenotype across the nasal mucosa of CF mice and in patient derived organoids. These results underscore the clinical potential of rAAV-CFTR Delta R in offering a cure for all patients with CF in the future.