Abstract 400: AAV2.9 Vectors Attain Robust and Sustained Gene Expression in Two Models of Vascular Gene Transfer
Background: Effective gene therapy of vascular disorders remains an elusive target. The main reasons for the relatively low efficacy of vascular gene therapy are poor and transient transduction of arterial tissue. AAV2, pseudotyped with serotype 9 capsid (AAV2.9), has been previously demonstrated to achieve effective transduction of heart, lung and skeletal muscle; however, no study to date has investigated AAV2.9 efficacy for vascular gene transfer. We hypothesized that the AAV2.9 may possess significant transduction potency for arterial tissue in vivo.
Methods in vitro: Rat aortic smooth muscle (A10) and bovine aortic endothelial (BAEC) cells were transduced with AAV2.9GFP and Ad5GFP at GC/cell and particle/cell ratios of 1x 104.
Methods in vivo: In a model of local dwelling arterial delivery, 5x1010 GC of AAV2.9Luc or 5x1010 particles of Ad5 were intralumenally delivered to denuded rat carotid arteries. In a model of stent-based gene delivery, using our previously published bisphosphonate-bare metal vector attachment ( PNAS 103:159–164, 2006), stents configured with AdLuc or AAV2.9Luc tethered via a hydrolysable linker were deployed in rat carotids. In both models arterial reporter expression was studied by serial intravital bioluminescence imaging.
Results: In vitro, peak GFP expression achieved with AAV2.9 in A10 and BAEC was 18- and 110-fold lower than with Ad vector. In rat carotid studies with intralumenally delivered AAV2.9Luc vector, progressively increasing levels of site-specific luciferase activity over time were shown (0.9±0.3x106, 8.2±3x106, 13.3±4.9x106, 26.3±5.4x106 and 16.7 ± 3.75x106 ph/sr/min at 4, 15, 31, 60 and 150 days post-delivery). Luc signal at day 4 following Ad5Luc delivery was 10-fold lower than with AAV2.9Luc, and dropped to background levels 15 days post-administration. Sustained Luc expression was also observed with AAV2.9 vectors tethered on stents per intravital bioluminescence imaging.
Conclusions: Despite lower in vitro levels of transgene expression of AAV2.9 compared to Ad5, the hybrid AAV2.9 serotype demonstrates significantly greater arterial gene transfer in vivo, resulting in higher and sustained levels of transgene expression.