P238Efficient gene transfer to human vascular cells in vitro and ex vivo using adenovirus serotype 49

Abstract

Introduction: Neointima formation and vascular remodelling through vascular smooth muscle cell (VSMC) migration and proliferation can limit the long term success of coronary interventions, for example in coronary artery bypass grafting (CABG). Gene therapy may provide a novel treatment strategy to improve long-term patency rates and reduce the need for repeat surgery. CABG surgery is potentially suitable for gene therapy as it allows ex vivo treatment of the vein before grafting. Therapeutic gene delivery to endothelial cells (ECs) and vascular smooth muscle cells (VSMCs) using recombinant adenovirus (Ad) 5 can successfully reduce pathological remodelling in experimental animal models. However, the Ad5 receptor coxsackie adenovirus receptor (CAR) is only expressed at low levels on vascular cells. Additionally, the common prevalence of neutralising antibodies against Ad5 in humans may limit the efficacy of Ad5 based gene therapy, although this requires testing in the clinical setting. Thus, alternative Ad serotypes may provide more efficient vascular gene transfer with lower neutralising antibody titer, thereby improving the prospect of effective gene therapy. Here we investigate the potential of Ad49, a novel species D adenovirus, as a vector for vascular gene delivery. Methods and Results: Transduction of primary human VSMCs and ECs with Ad49 was 4 fold higher than Ad5 (p<0.001) and 30 fold higher than Ad35 (p<0.0001) when infected with 1000 viral particles per cell. Ad49 could also transduce HVSMCs more efficiently than Ad5 following a short exposure time <60 minutes. Ex vivo incubation of whole mouse aorta with Ad49GFP resulted in transduction of endothelial cells seen by co-localisation of GFP and CD31 immunohistochemistry. Similarly, luminal infusion of Ad49GFP into intact human saphenous vein ex vivo also resulted in endothelial cell transduction. The Ad49 receptor is unidentified however, blocking CAR, CD46 and removal of sialic acid, classical candidate adenovirus receptors, did not alter Ad49 transduction. Importantly serum neutralisation studies in 103 patient serum samples from Glasgow cardiovascular clinics found Ad5 neutralisation rates were high (40/103) but no pre-existing immunity to Ad49 was observed. In conclusion, we demonstrate that Ad49 efficiently transduces vascular cells in vitro and ex vivo in intact tissue and may provide a new vector for vascular gene therapy.