136. Vector Host Interactions of Group B Adenoviruses in CD46 Transgenic Mice
Authors: Anuj Gaggar, Dmitry M. Shayakhmetov, Sonia Marwah, Shaoheng Ni, Andre M. Lieber
Journal: Molecular Therapy (2004)

Adenovirus (Ad) is a useful tool for gene delivery into a wide variety of cell types. Most adenoviral vectors used for gene therapy are based on serotype 5, which binds to cells via interactions between the viral fiber protein and its receptor, called Coxsackie and Adenovirus Receptor (CAR). Despite the widespread use of Ad5-based vectors for gene delivery, many therapeutic targets, including dendritic cells, hematopoietic stem cells, and malignant cancer cells, have very low levels of CAR and therefore are not efficiently infected by Ad5-based vectors. There is a growing body of evidence that Ads from subgroup B can circumvent limitations of Ad5 based-vectors and efficiently deliver genes to these cell types. We, and others, have shown that group B Ads use CD46, a complement-regulatory molecule, for initial cellular attachment. Although many studies have used group B Ads, little is known about their interactions with the host. In order to better understand the impacts of group B Ads on the host, we used transgenic mice expressing human CD46 in a pattern closely mimicking that of humans. We used a vector based on an Ad5 capsid with the fiber and shaft domain derived from serotype 35, a representative group B Ad (Ad5/35). We analyzed CD46 transgenic mice in three aspects: 1) biodistribution of Ad5/35 applied either intravenously (IV) or intraperitoneally (IP) 2) Kupffer cell uptake of Ad immediately following tail vein injection in normal mice versus transgenic mice and 3) levels of innate immune responses following systemic viral application of Ad5/35 or Ad5 in normal and transgenic mice after IP or IV injection. For biodistribution studies, mice were injected with 5 × 109 pfu/mouse either through the tail vein or within the peritoneum. Immunohistochemistry analysis of organs 72 hours post virus injection revealed reporter gene expression in the liver, lymph nodes, and spleen following IV injection. Interestingly, the spleen showed reporter gene expression in the marginal zone. Mice injected IP showed significantly lower levels of gene expression in all tissues analyzed with only lymph nodes and the spleen showing appreciable transduction. Initial analyses reveal that IV injection of adenovirus results in substantial increase in Ad DNA accumulation over IP injections in all analyzed tissues. To determine Kupffer cell uptake of virus immediately after application, vector conjugated to fluorophore Cy-3 was injected into the tail vein of normal and transgenic mice, and 30 min. after injection, the mouse livers were harvested and stained with Kupffer cell specific markers to assess viral uptake. An increase in virus in the liver was seen in transgenic mice compared to normal mice and much of this virus was associated with Kupffer cells. Finally, to assess levels of innate toxicity elicited by either Ad5 or Ad5/35, we will examine inflammatory cytokines (such as TNFa, IFN-γ, IL-6, IL-10, IL-12, and MCP-1) six hours after systemic application in CD46 transgenic mice as compared to normal mice. Together, these studies will allow for a better understanding of how group B Ads interact with a model host and will provide information on the potential future uses of group B Ads for gene therapy in human patients.