MECHANISM OF ADENOVIRUS IMPROVEMENT OF CATIONIC LIPOSOME-MEDIATED GENE-TRANSFER

Substantial effort has been focused on the development of highly effic ient gene transfer strategies. Although viral and non-viral methods ha ve been elaborated, mechanisms of gene delivery are still poorly under stood. We exploited our recent observation that replication-deficient type 5 adenovirus dramatically enhances lipofectAMINE-mediated gene tr ansfer (lipoadenofection) in differentiated cells to elucidate the mec hanism of adenovirus action in this process. Heat-induced denaturation of viral capsid abolishes adenovirus action whereas inactivation of v iral genome by short treatment with UV has no effect. Electron microsc opic observations reveal the formation of a complex containing adenovi rus and lipofectAMINE which probably carries DNA into cells via endocy tosis. Anti-adenovirus antiserum or monoclonal anti-alpha(v) beta(3) i ntegrin antibody inhibits lipoadenofection, at least partially. Neutra lization of endosomal compartments with chloroquine, ammonium chloride or monensin does not prevent adenovirus improvement of gene transfer. Hence, adenovirus-lipofectAMINE-DNA complexes in which viral particle s are each encompassed by three lipid layers. penetrate cells via an e ndocytic pathway involving probably the adenovirus receptor and alpha( v) beta(3) integrin. The resulting efficient transfer and expression o f plasmid DNA proceeds from a mechanism in which adenoviral endosomoly tic activity appears to be required while viral genome is not essentia l. (C) 1997 Elsevier Science B.V.