394. Effective Focal, Homogeneous Gene Transfer after Coronary Infusion of Adenoviruses

Abstract

Myocardial gene therapy has been inhibited by the lack of effective in vivo delivery. Previously published models utlize aortic occlusion and long delivery times at low body temperatures. These reqirements are not feasible for eventual human usage. We previously investigated several parameters relevant to gene transfer efficiency in ex vivo and in vitro models. Here we extend those findings to an in vivo model. In a porcine intracoronary infusion model, we investigated the folllowing parameters hypothesized to affected gene efficacy efficiency: (1) vascular permeability agents (2) virus contact time; (3) virus concentration; (4) vasodilatory agents (5) coronary flow rate; (6) participation of the coronary sinus (CS) in delivery. Methods: Domestic piglets (5–7kg) underwent coronary catheterization using conventional techniques. The left anterior descending artery (LAD) was subselected for pretreatment and virus solutions infusion through a 2.7Fr balloon catheter. The level of infusion was consistently just distal to the second diagonal branch. For CS delivery, the CS was cannulated and the great cardiac vein was selected using a balloon catheter. Animals were sacrificed 3–6 days after the procedure, and hearts were stained with X-gal using conventional methods. The transgene efficacy was estimated by percentage of stained cells in the focal area. For all studies, we used Adbgal, a recombinant adenovirus encoding the E. coli b-galactosidase gene. Results: Baseline parameters included sildenafil 25mg PO, infusion Ca concentration 1mM, pretreatment over 3 minutes with 5μg/μl VEGF, 5mg/ml adenosine, 250mg/ml nitroglycerin, and 5 × 109 pfu/ml Adbgal infusion with the same concentration of adenosine and nitroglycerin at 6ml/min flow rate and 2 min contact time. The baseline gene transfer efficiency was 40% (Figure 1). Vascular permeability agents (adenosine, nitroglycerin, VEGF, 50 mM Ca2+, 8-Br-cGMP) increased transgene efficacy compared to the baseline regimen. Adenosine also appeared to compensate for lower flow rates. Increasing flow rate and contact time also improved gene transfer efficiency. Notably, simultaneous infusion through LAD and CS showed the highest transgene efficacy of 80% of myocytes in the target area. Conclusion: Adenovirus gene transfer efficiency can be improved by manipulation of vascular permeability agents, coronary flow characteristics, virus concentration and contact time.