Increased Contact Time Improves Adenovirus-Mediated CFTR Gene Transfer to Nasal Epithelium of CF Mice

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Abstract
Multiple dosing with recombinant adenoviral vectors containing the cystic fibrosis transmembrane conductance regulator (CFTR) cDNA to the nasal mucosa of cystic fibrosis (CF) transgenic mice reportedly results in only partial correction of the CF defect in chloride (Cl¯) secretion without normalizing sodium (Na+) hyperabsorption, perhaps indicating inefficient gene transfer into the nasal airway epithelium in vivo. In this study, we have examined whether optimizing vector administration such as contact time could improve gene transfer efficiency. Changes in basal nasal potential difference (PD), and in PD (ΔPD) following addition of amiloride and subsequent removal of Cl¯ from the luminal perfusate were assayed. As reported previously, the basal nasal PD was significantly more negative in CF mice (–24.9 ± 2.1 mV) than in normal mice (–6.3 ± 1.2 mV). Normal mouse nasal mucosa exhibited a large hyperpolarization in response to low Cl¯ substitution (ΔPD of 8.5 ± 1.9 mV), whereas the nasal mucosa of the CF mouse depolarized in response to this treatment. No correction of either the Cl¯ or Na+ transport defects were observed when 5 × 109 IU of Ad2/CFTR-5 were administered to the nasal passage of CF mice over a period of 5–20 min. However, when CF mice were perfused over a period of 60 min with the same dose of vector, a significant response (ΔPD of 5.9 ± 1.1 mV) to low Cl¯ substitution was detected 2 days later. In these mice, the basal nasal PD (–10.5 ± 1.4 mV) and the response to amiloride were also reduced, indicating a partial correction of the Na+ transport defect. Expression of functional CFTR activity was transient with no measurable ΔPD signals observed by day 7 post-treatment. These results suggest that prolonging the contact between an adenoviral vector and the respiratory epithelium enhances the efficiency of gene transfer and can result in improved correction of the CF Na+ and Cl¯ ion transport defects. Therefore, strategies that improve internalization of viral vectors and that prolong their contact time with target cells may result in the improved clinical efficacy of such vectors. Gene transfer using recombinant adenoviral vectors containing the cystic fibrosis transmembrane conductance regulator (CFTR) cDNA is being evaluated as a potential treatment for patients with cystic fibrosis (CF). Early clinical studies indicate that the efficiency of adenovirus-mediated gene transfer to the nasal epithelium of CF patients may be low. This study focuses on efforts to improve the efficiency of gene transduction using the nasal mucosa of transgenic CF mice as a model. We show that prolonging the incubation time of the vector with the airway epithelium enhanced the extent of adenovirus-mediated gene transfer. Levels of functional CFTR expression were detected that were dose-dependent and persisted for up to 4 days post-infection. These studies indicate that interventions that facilitate a greater residence time of the vector with the airway epithelium may enhance the efficiency of gene transduction to these cells.

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