Abstract 268: Targeting Adeno-associated Viral Vectors to Atherosclerotic Plaques
Introduction: Targeted delivery of biological agents selective for atherosclerotic plaques could provide a useful agent for treatment and/or imaging of atherosclerosis. Adeno-associated virus 2 (AAV2) is a commonly used gene therapy vector, however it has a broad tropism and following systemic administration mediates inefficient delivery of transgenes to vascular cells. AAV2 tropism can be altered using small targeting peptides identified by phage display. The aim of this study was to insert atherosclerotic plaque targeting peptides into the receptor binding site of the virus capsid to create vectors that selectively target atherosclerotic plaques.
Methods: Two putative atherosclerotic plaque targeting peptides (CAPGPSPSKC and CNH-RYMQMC) were engineered into the receptor binding domain of the AAV2 capsid to produce AAV2-CAP and AAV2-CNH. Viruses expressing the LacZ and eGFP reporter genes were produced and their ability to mediate vascular-specific gene delivery was tested.
Results: In a non-vascular cell line (HeLa) neither of the peptide-modified viruses produced significantly higher levels of transduction than unmodified AAV2. AAV2-CNH transduced murine, rat and human endothelial cell lines 10–100- fold higher than AAV2 (p<0.05). AAV2-CNH transduction was mediated through a proteasome-insensitive pathway, suggesting that peptide alters both receptor binding and trafficking of the virus. In addition, AAV2-CNH transduced cells over-expressing the putative receptor membrane type 1 matrix metalloproteinase (MT1-MMP) at significantly higher levels than control cells. Surface plasmon resonance confirmed that the vector binds to MT1-MMP. Modified AAV2 vectors were further characterized in vivo in ApoE−/− mouse model of atherosclerosis. Following intravenous injection of the vectors real-time PCR was used to detect virus in tissues. Compared to control AAV2, substantially higher levels (10–100 fold) of AAV2-CAP and AAV2-CNH were detected in the brachiocephalic artery and aorta (sites of atherosclerotic plaque formation), with reduced levels in all other organs examined, including liver and spleen.
Conclusions: AAV2-CAP and AAV2-CNH may provide useful tools to selectively deliver genes to atherosclerotic vasculature.