319. Improving the Odds of Adenovirus-Mediated Gene Therapy By Upregulation of the Coxsackievirus and Adenovirus Receptor

Adenovirus-mediated gene therapy has been limited by the fact that the primary receptor for most adenovirus serotypes, the Coxsackievirus and adenovirus receptor (CAR), is inaccessible or not expressed on many cell types of interest. Few mechanisms have been discovered that regulate CAR expression and tissue specific localization. In the airway, CAR is mostly considered a cell-cell adhesion protein localized at the basolateral surface of polarized epithelia. Recently, an alternate isoform of CAR, CAREx8, has been identified at the apical surface of polarized airway epithelia and is implicated in viral infection from the apical surface. We hypothesized that upregulation of cellular mechanisms that facilitate endogenous CAREx8 protein expression at the apical surface would enhance adenovirus gene transfer. Using polarized model epithelial cell lines and primary human airway epithelia, we found that IL-8, a proinflammatory cytokine and a neutrophil chemoattractant, stimulates the protein expression and apical localization of CAREx8 via activation of AKT/S6K and inhibition of GSK3β. IL-8- mediated upregulation of CAREx8 increased AdV5-β-Gal entry and transduction by approximately 5-fold. Moreover, we found that infiltrating neutrophils bind CAREx8 at the apical surface of a polarized epithelium and, surprisingly, neutrophils enhance AdV5-β-Gal entry into the epithelium by 2-3 fold. The effect of IL-8 and neutrophils on AdV infection could be blocked by fiber-knob from AdV5 but not AdV3, a non-CAR binding serotype, indicating the importance of CAR. These findings suggest that acute inflammation may enhance adenovirus infection. Moreover, therapeutics that stimulate the AKT/S6K pathway or inhibit GSK3β may be able to augment adenovirus-mediated gene therapy.