Mechanism by which calcium phosphate coprecipitation enhances adenovirus-mediated gene transfer

Delivery of a normal copy of CFTR cDNA to airway epithelia may provide a no vel treatment for cystic fibrosis lung disease. Unfortunately, current vect ors are inefficient because of limited binding to the apical surface of air way epithelia. We recently reported that incorporation of adenovirus in a c alcium phosphate coprecipitate (Ad:CaPi) improves adenovirus-mediated gene transfer to airway epithelia in vitro and in vivo. To understand better how coprecipitation improves gene transfer we tested the hypothesis that incor poration in a CaPi coprecipitate increases the binding of adenovirus to the apical surface of differentiated human airway epithelia. When a Cy3-labell ed adenovirus was delivered in a coprecipitate, binding increased 54-fold a s compared with adenovirus alone. Moreover, infection by Ad:CaPi was indepe ndent of fiber knob-CAR and penton base-integrin interactions. After bindin g to the cell surface, the virus must enter the cell in order to infect We hypothesized that Ad:CaPi may stimulate fluid phase endocytosis, thereby fa cilitating entry. However, we found that neither adenovirus nor Ad:CaPi cop recipitates altered fluid phase endocytosis. Nevertheless, Ad:CaPi preferen tially infected cells showing endocytosis. Thus, CaPi coprecipitation impro ves adenovirus-mediated gene transfer by coating the epithelial surface wit h a layer of virus which enters cells during the normal process of endocyto sis.