Optimization of adeno-associated virus vector-mediated gene transfer to the respiratory tract

Abstract

An efficient adeno-associated virus (AAV) vector was constructed for the treatment of respiratory diseases. AAV serotypes, promoters and routes of administration potentially influencing the efficiency of gene transfer to airway cells were examined in the present study. Among the nine AAV serotypes (AAV1-9) screened in vitro and four serotypes (AAV1, 2, 6, 9) evaluated in vivo, AAV6 showed the strongest transgene expression. As for promoters, the cytomegalovirus (CMV) early enhancer/chicken β-actin (CAG) promoter resulted in more robust transduction than the CMV promoter. Regarding delivery routes, intratracheal administration resulted in strong transgene expression in the lung, whereas the intravenous and intranasal administration routes yielded negligible expression. The combination of the AAV6 capsid and CAG promoter resulted in sustained expression, and the intratracheally administered AAV6-CAG vector transduced bronchial cells and pericytes in the lung. These results suggest that AAV6-CAG vectors are more promising than the previously preferred AAV2 vectors for airway transduction, particularly when administered into the trachea. The present study offers an optimized strategy for AAV-mediated gene therapy for lung diseases, such as cystic fibrosis and pulmonary fibrosis.

Mosaic Recombinant Adeno-associated Virus Vector rAAV/DJ/CAG for Targeted Gene Delivery to Melanoma Cells Metastasized to the Lung

“…In our research, we focused on the use of CMV and CAG promoters. The choice of a promoter is extremely important in targeted gene therapy and, thus, in the individualization of treatment (90)(91)(92). We noticed that improved transduction efficiency in melanoma cells in in vitro and in vivo conditions was obtained by the CAG promoter (Figures 3, 7, and 8).…”

“…The expression of CAG promoter in the lungs lasted up to 12 months while that of CMV expression up to 8 months. However, the CMV promoter has an immediate effect that provides strong but short-term expression (90,91). However, it is very susceptible to silencing in some cell types (89).…”

Effect of promoter, promoter mutation and enhancer on transgene expression mediated by episomal vectors in transfected HEK293, Chang liver and primary cells

“…Previous studies indicated that the endogenous mammalian promoters can provide higher expression than viral promoters [26,27]. The EF-1α promoter has been shown to be one of the strongest promoters in various cell lines [28,29]. Indeed, the EF-1α promoter is often active in cells where viral promoters fail to drive downstream gene expression and are gradually silenced [30,31].…”

Genetic therapies for cystic fibrosis lung disease

“…These studies exemplify the challenge of developing efficient AAV vectors for evaluation in relevant CF animal models, including pigs [4*], sheep [7], ferrets [8] and mice [6], but also provide efficient transduction of human epithelia for clinical translation. For example, nine naturally occurring AAV serotypes tested in human cells and murine lungs identified AAV6 as the strongest candidate in both human and mouse epithelium [5]. However, other serotypes, including AAV2/5, i.e.…”