Objectives: Given the propensity of human Adenoviruses (Ads) to more readily infect cells of epithelial rather than mesenchymal origin, we hypothesized that non-human Ad knob proteins might enhance gene transfer to odontoblasts (mesenchymal) relative to ameloblasts (epithelial). Methods: Chimeric Ads comprised of E1/E3 deleted serotype 5 human Ads in which the knob or entire knob/shaft region of the fiber protein has been replaced by non-human serotype Adenoviruses were obtained. All viruses contained a Luciferase reporter gene driven by the CMV promoter. Viruses used are as follows: Ad5 (control), Ad5-MK (murine knob), Ad5-OvF (complete ovine fiber), Ad5-PK (porcine knob), Ad5-PF (complete porcine fiber), Ad5-CK2 (canine serotype 2 knob) and Ad5-CK2.pK7 (canine serotype 2 knob w/ 7-lysine motif). These vectors were tested on the human ameloblast and odontoblast cell lines AI-W (amelo) and JP-D (odonto). Gene transfer was measured by plating 30,000 cells/well in 24-well culture plates and 24 hours later incubating cell cultures with 100 viral particles per cell of the appropriate virus for 2 hours at 37C. Luciferase gene activity was assayed with a Promega Luciferase Assay System. Results: Results of gene transfer experiments are presented in Figure 1, with each bar representing the average of 5 culture wells, with standard deviation represented by error bars. In accordance with prior findings, Ad5 accomplished negligible gene transfer to both ameloblasts and odontoblasts. All non-human knobs substantially enhanced gene delivery to ameloblasts (p<0.05). Canine knobs (CAV2 and CAV2pK7) substantially enhanced gene delivery to odontoblasts (p<0.05), with the CAV2 knob chimera reaching >20% of the gene transfer levels seen in ameloblasts. Conclusions: In light of the documented difficulty associated with Ad-mediated gene transfer to cells of mesenchymal origin, we have identified a non-human Ad knob capable of robust gene transfer to odontoblasts, with an appealing infectivity profile relative to ameloblasts.

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