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Abstract
Recombinant adeno-associated virus (AAV) vectors expressing the cystic fibrosis transmembrane
conductance regulator (CFTR) gene have been used to deliver CFTR to the airway epithelium
of cystic fibrosis (CF) patients. However, no significant CFTR function has been demonstrated
likely due to low transduction efficiencies of the AAV vectors. To improve AAV transduction
efficiency for human airway epithelium (HAE), we generated a chimeric AAV library
and performed directed evolution of AAV on an in vitro model of human ciliated airway
epithelium. Two independent and novel AAV variants were identified that contained
capsid components from AAV-1, AAV-6, and/or AAV-9. The transduction efficiencies of
the two novel AAV variants for human ciliated airway epithelium were three times higher
than that for AAV-6. The novel variants were then used to deliver CFTR to ciliated
airway epithelium from CF patients. Here we show that our novel AAV variants, but
not the parental, AAV provide sufficient CFTR delivery to correct the chloride ion
transport defect to ~25% levels measured in non-CF cells. These results suggest that
directed evolution of AAV on relevant in vitro models will enable further improvements
in CFTR gene transfer efficiency and the development of an efficacious and safe gene
transfer vector for CF lung disease.