Abstract 714: Direct Comparison of AAV Serotypes 1–9; Onset of Gene Expression, Time to Peak Expression, and Fate of Viral Genomes in Mice after Skeletal Muscle, and Tail Vein Injection
Background- Viral delivery vectors founded on the adeno-associated virus (AAV) serotype 2 platform are currently being evaluated for human gene transfer. All viral genes can be removed, resulting in only the transgene of interest and the viral terminal repeats. These small non-enveloped single-stranded DNA viruses have been associated with long duration expression in experimental animals, and these qualities make AAV serotypes ideal systems when considering viral gene delivery. However, while scores of serotypes have been described, few have been compared directly. In this study we have directly compared AAV serotypes 1–9 by two routes of injection, skeletal muscle and tail vein.
Methods and Results- Previous studies of AAV serotypes 1–5 have demonstrated uniformity in production and purification. Those results guided this comparison of luciferase gene expression-efficiency between AAV serotypes 1–9 after single injection into mouse muscle, or tail vein, of 1 x1011 viral particles into the gastrocnemius (100μL total volume) or tail vein (250μL total volume). Kinetic information including rate of onset, peak expression, and level of peak expression were determined for each animal studied. Results in muscle demonstrate that AAV1 and 6 have the highest level of gene expression, reach peak expression faster, and retained higher transgene copy numbers than other serotypes. Expression levels of AAV5, 8 and 9 were similar, but ~10 fold lower than AAV1 and 6, and AAV2, 3, 4 and 7 were ~50 times lower than AAV1 and 6. The kinetic data confirms previous observations of delayed onset for AAV2 as well as demonstrates a similar lag for AAV3, 4, and 7 in muscle. Tail vein injection of AAV3 and AAV8 resulted in whole body luminescence with AAV8 having higher overall levels of expression. While the expression was similar in AAV1 and 6 animals, AAV6 expression was more pronounced in the thoracic cavity and AAV1 had higher levels of hepatosplenic expression.
Conclusion- AAV serotypes 1 and 6 show the best overall muscle, with AAV6 showing a propensity for thoracic expression. Combining imaging and end point quantitative data can be broadly applied to evaluating AAV serotypes by various routes. By maintaining certain standard conditions new serotypes can be similarly evaluated.