Development of Novel Formulations That Enhance Adenoviral-Mediated Gene Expression in the Lung in Vitro and in Vivo

Abstract

Despite remarkable progress in the development of both viral and non-viral gene delivery vectors for cystic fibrosis therapy, low efficiency of gene transfer to the airway epithelium is a major obstacle to clinical application. Here we develop formulations that enhance cellular absorption of adenoviral vectors. We selected excipients from a panel of pharmaceutically acceptable com-pounds known to enhance drug absorption. Transduction efficiency of the virus in the presence of each ingredient was assessed in vitro and in vivo. Mannitol and chitosan substantially enhanced transduction efficiency in vitro and augmented expression in vivo by 4 and 8 log units, respectively. The most successful formulation (a blend of sucrose, mannitol, and Pluronic F68) transduced 100% of an A549 cell population in vitro and produced areas of intense gene expression in both large and small airways in vivo with minimal toxicity. Dose response studies also indicate that when placed in this formulation, the viral dose can be lowered by 1/2 log while maintaining superior levels of transgene expression. This formulation also enhanced the physical stability of the virus. No significant loss in titer was detected from a lyophilized formulation after storage at 25 degrees C for 30 days.

An optimized protocol for detection of E. coli β-galactosidase in lung tissue following gene transfer

“…Although X-gal staining has been successfully used for transgene detection in lungs transduced with adeno-and lentiviral vectors (e.g., Croyle et al 2001;Kobinger et al 2001;Medina et al 2003;Wang et al 2004), the visualization of LacZ expression from AAV vectors remains more challenging. In this study, we systematically tested parameters to optimize the detection of b-gal activity in lungs.…”

Microencapsulation of PEGylated Adenovirus within PLGA Microspheres for Enhanced Stability and Gene Transfection Efficiency

“…Viruses have coat proteins on the surface that are very sensitive to denaturation against environmental pH, temperature, redox potential, and shear-stress (21)(22)(23)(24). This has lead to the expectation that the microencapsulation of ADV within PLGA microspheres would result in a significant loss of viral infectivity due to the harsh physical conditions imposed by the double emulsion and solvent evaporation method.…”

Natural Polymers as Components of Blends for Biomedical Applications

“…When chitosan is blended with egg phosphatidylcholine, advanced injectable drug delivery systems for pharmaceutical applications can be obtained [157]. The stability of such blends is due to the specific interactions between the molecules.…”