Abstract

The adenovirus capsid proteins cooperate to mediate a highly efficient gene transduction event in permissive tissue culture cell lines. Cellular entry is initiated by the fiber protein, which binds with high affinity to the coxsackie-and-adenovirus receptor. Subsequent to fiber binding, penton base facilitates virus internalization by binding αv integrins through an RGD motif. The highly efficient nature of adenovirus infectivity into a wide variety of cell types has led to the development of adenovirus vectors for therapeutic purposes. While adenovirus vectors (AdV) have considerable potential in a number of clinical applications, they are severely limited due a promiscuous tropism in vivo, a lack of cell-specific targeting, and a potent activation of the host immune response. We propose that the limitations associated with current AdV are intimately connected to capsid protein-host cell interactions. The primary goals of this study are: (1) to understand how capsid proteins influence AdV transduction and immune activation in vitro and in vivo, (2) to modify capsid proteins in a manner that blunts AdV infectivity and immunoreactivity, and (3) to develop highly targeted vectors whose cellular distribution can be controlled through fiber protein manipulations. Our experimental strategy takes advantage of the well-characterized molecular genetics of adenovirus. By manipulating viral late gene expression, we generated a panel of AdV expressing fiber proteins with differential binding functions. These fiber-modified vectors were further altered by deleting the integrin binding capacity of penton base. Each of the viruses was tested for transduction and immune activation in a mouse model of systemic administration. The results demonstrate that capsid-modified AdV are severely compromised in their ability to mediate transduction and host immune stimulation. These modified viruses were then used as platforms to generate vectors with targeting potential to mouse melanoma cells. Development of the targeted vectors uncovered a potentially novel role for the fiber protein in virion packaging and growth. The adenovirus vectors generated in this study contribute to our understanding of virus-host cell interactions in vivo and have attributes that may be beneficial in therapeutic applications such as gene therapy, vaccine development, and cancer targeting.