Ex vivo gene therapy prevents chronic graft vascular disease in cardiac allografts

Objective: We hypothesized that ex vivo hyperbaric transfection of antisense oligodeoxynucleotides for blockade of intercellular adhesion molecule-1, an important mediator of cell adhesion and T-cell costimulation, would reduce chronic graft vascular disease in cardiac allografts. Methods: PVG hearts underwent ex vivo transfection with antisense, reverse antisense intercellular adhesion molecule-1 oligodeoxynucleotide (80 μmol/L), or saline solution at 3 atm pressure for 45 minutes at 4°C and were transplanted heterotopically into ACI recipients with or without treatment with intercellular adhesion molecule-1 (1A29) or leukocyte function associated antigen-1 (WT.1) monoclonal antibodies. Transfection efficiency was confirmed with fluorescein isothiocyanate-labeled oligodeoxynucleotides and fluorescent microscopy. Efficacy of intracellular adhesion molecule-1 blockade was assessed with the use of immunohistochemistry. Graft reperfusion injury was evaluated at 6 to 24 hours by neutrophil infiltration (myeloperoxidase [MPO]), cardiac edema (%wt/wt), and histologic injury (percent contraction band necrosis). Grafts from recipients treated with cyclosporine A (5 mg/kg per day, days 0 to 9) were scored for chronic graft vascular disease on postoperative day 90 ranging from 0 (no involvement) to 4 (>50% vascular occlusion). Results: Transfection was highly efficient (fluorescein isothiocyanate-labeled oligodeoxynucleotides in 48% ± 5% of total myocardial nuclei) and effective at blocking intracellular adhesion molecule-1 expression (positive area in allografts taken on postoperative day 3 was reduced from 100% ± 0% to 52% ± 14%, n = 4). Blockade with antisense oligodeoxynucleotides versus monoclonal antibodies was less effective at preventing reperfusion injury while more effective at reducing chronic graft vascular disease (score 0.98 ± 0.48, p < 0.05). Reverse antisense oligodeoxynucleotides and vector control (antisense oligodeoxynucleotide infusion without pressure) groups failed to demonstrate this beneficial effect. Conclusion: Hyperbaric transfection of antisense oligodeoxynucleotides proved highly efficient, effective at blockade of intracellular adhesion molecule-1, and demonstrated a sequence-specific reduction in chronic graft vascular disease. This highly targeted alteration of donor organ immunogenicity may have an important future role in clinical immunosuppressive strategies.