A systematic evaluation of cardiac gene transfer protocol in a large animal model

M. T. Vo (Heidelberg)¹, V. Horchemer (Heidelberg)¹, D. Kehr (Heidelberg)¹, E. Meinhardt (Heidelberg)¹, M. Pott (Heidelberg)¹, P. Raake (Augsburg)², N. Frey (Heidelberg)¹, P. Schlegel (Heidelberg)^1
1Universitätsklinikum Heidelberg Klinik für Innere Med. III, Kardiologie, Angiologie u. Pneumologie Heidelberg, Deutschland; ²Universitätsklinikum Augsburg I. Medizinische Klinik Augsburg, Deutschland

Adeno-associated virus (AAV)-based gene therapy (GT) represents a promising treatment option for genetic cardiomyopathies and heart failure. One major obstacle hampering translation of potent GT  is robust myocardial transgene expression. While vector optimization addresses tissue specificity, addition of loco-regional delivery strategies allows for a reduction in vector dose while maintaining a high vector concentration in the target region. Only this combination will allow for cost-effective therapies.  Therefore, assessing an effective and clinically applicable delivery strategy is key to achieving successful transgene expression. Coronary artery and sinus balloon occlusions are common procedures in interventional cardiology. We hypothesize that occlusion during delivery will increase cardiac AAV uptake and gene expression by extending the local dwell time.

Pre-screened animals with low neutralizing antibody titers (nAb, <1:8) were randomized into three groups and received 1×10^13 viral genome copies per animal of a recombinant AAV6 expressing firefly luciferase (rAAV6.fLuc) using three different application protocols.We compared a) antegrade delivery (cAD) with b) pressure-controlled infusion and antegrade occlusion (pcAO) and c) pressure-controlled infusion with antegrade/retrograde double occlusion (pcARO). Protocols pcAO and pcARO was achieved through three 30-second occlusion cycles with two-minute recovery periods. The animals were continuously monitored using electrocardiogram and blood sampling to determine the safety of the procedures. 28 days after gene-therapy, organs were harvested and biodistribution and transgene expression was measured.

Coronary artery occlusion (pcAO and pcARO) generally resulted in higher transgene expression in the target area when compared to antegrade gene delivery without occlusion. Simultaneous balloon occlusion of the coronary artery and the sinus (pcARO) resulted in the highest transgene expression amongst the three tested protocols. The vector DNA in heart to liver ratio was highest in the pcARO group. All procedures were well tolerated. 

These data suggest that the combination of coronary artery and sinus balloon occlusions optimally supports antegrade delivery of AAV gene therapy vectors, which has important implication for both pre-clinical and clinical applications of gene therapy products.