Role of Integrin Expression in Adenovirus-Mediated Gene Delivery to the Intestinal Epithelium

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Abstract
Adenoviral vectors are being developed for oral delivery of therapeutic genes to the intestine. Initial studies in the rat using mucolytics and direct application of adenovirus encoded with the interleukin-1 receptor antagonist gene to the jejunum produced limited gene expression. The goal of this study was to determine the role of integrins in adenovirus-mediated gene delivery to the intestinal epithelium. Integrins are involved in cellular differentiation and tight junction formation and mediate adenoviral internalization. Results from Caco-2 and IEC-18 cells suggest that, as enterocytes differentiate, cell-surface integrin expression decreases. Pretreatment of Caco-2 cells with RGD peptides reduced adenoviral transduction efficiency by 80% in undifferentiated cells and 20% in differentiated cells. Both differentiated and undifferentiated IEC-18 cells showed a 70% drop in transduction when pretreated with the peptide. Infection inhibition studies with monoclonal antibodies further suggest that αvβ3 and α6β1 integrins play significant roles in adenoviral internalization in the intestine. Expression of integrins in cell culture models of the intestine correlated with in vivo expression in intestinal segments. These results indicate that the ileum is a prime target for efficient adenovirus-mediated gene transfer in the rat. To enhance transduction in differentiated enterocytes (probable targets for oral gene delivery), Caco-2 cells were treated with interleukin-1β (a cytokine known to increase integrin expression) prior to administration of the virus. Transduction efficiency increased four-fold. Previous studies of adenovirus-mediated gene transfer of the interleukin-1 receptor antagonist protein to rat jejunum have produced limited gene expression. In the present study, we have shown that integrins play a significant role in efficient adenoviral infection of the intestinal epithelium. As enterocytes differentiate, integrin expression decreases. This coincides with significant reduction in adenoviral transduction efficiency. Results from two in vitro models of the intestinal epithelium (Caco-2 and IEC-18 cells) show that αvβ3 integrins have the most influence on adenoviral internalization. Results from screening of rat intestinal segments for expression of the αvβ5 integrin suggest that, based on integrin expression, the ileum is a prime target for efficient adenovirus-mediated gene transfer. We have also found that administration of immunomodulating agents and inflammatory disease states provide a favorable environment for efficient internalization of adenoviral vectors due to up-regulation of integrin receptors.