A normothermic ex vivo organ perfusion delivery method for cardiac transplantation gene therapy

• Published in: Scientific reports 2019-05, Vol.9 (1), p.8029-8029, Article 8029
• Main Authors: Bishawi, Muath, Roan, Jun-Neng, Milano, Carmelo A., Daneshmand, Mani A, Schroder, Jacob N., Chiang, Yuting, Lee, Franklin H., Brown, Zachary D., Nevo, Adam, Watson, Michael J., Rowell, Trevelyn, Paul, Sally, Lezberg, Paul, Walczak, Richard, Bowles, Dawn E.
• Format: Article
• Language: English
• Subjects: 13; 42; 42/44; 631/443/592; 692/4019/2773; Adenoviridae - genetics; Allografts; Allografts - chemistry; Animals; Autografts; Background levels; Blood; Copy number; Coronary artery; Coronary vessels; Cytomegalovirus; Deoxyribonucleic acid; DNA; Expression vectors; Feasibility Studies; Female; Gene therapy; Gene transfer; Gene Transfer Techniques; Genes, Reporter - genetics; Genetic Therapy - methods; Genetic Vectors - genetics; Heart; Heart Failure - genetics; Heart Failure - therapy; Heart transplantation; Heart Transplantation - methods; Heart transplants; Humanities and Social Sciences; Humans; Liver - chemistry; Luciferases - analysis; Luciferases - genetics; Models, Animal; multidisciplinary; Myocardium; Myocardium - chemistry; Organ Preservation - methods; Organ Preservation Solutions - chemistry; Perfusion; Perfusion - methods; Proteins; Psoas muscle; Science; Science (multidisciplinary); Spleen; Sus scrofa; Transplantation, Homologous - methods; Vectors (Biology)
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/s41598-019-43737-y

Clinically, both percutaneous and surgical approaches to deliver viral vectors to the heart either have resulted in therapeutically inadequate levels of transgene expression or have raised safety concerns associated with extra-cardiac delivery. Recent developments in the field of normothermic ex vivo cardiac perfusion storage have now created opportunities to overcome these limitations and safety concerns of cardiac gene therapy. This study examined the feasibility of ex vivo perfusion as an approach to deliver a viral vector to a donor heart during storage and the resulting bio distribution and expression levels of the transgene in the recipient post-transplant. The influence of components (proprietary solution, donor blood, and ex vivo circuitry tubing and oxygenators) of the Organ Care System (OC) (TransMedics, Inc., Andover MA) on viral vector transduction was examined using a cell-based luciferase assay. Our ex vivo perfusion strategy, optimized for efficient Adenoviral vector transduction, was utilized to deliver 5 × 10 13 total viral particles of an Adenoviral firefly luciferase vector with a cytomegalovirus (CMV) promotor to porcine donor hearts prior to heterotopic implantation. We have evaluated the overall levels of expression, protein activity, as well as the bio distribution of the firefly luciferase protein in a series of three heart transplants at a five-day post-transplant endpoint. The perfusion solution and the ex vivo circuitry did not influence viral vector transduction, but the serum or plasma fractions of the donor blood significantly inhibited viral vector transduction. Thus, subsequent gene delivery experiments to the explanted porcine heart utilized an autologous blood recovery approach to remove undesired plasma or serum components of the donor blood prior to its placement into the circuit. Enzymatic assessment of luciferase activity in tissues (native heart, allograft, liver etc.) obtained post-transplant day five revealed wide-spread and robust luciferase activity in all regions of the allograft (right and left atria, right and left ventricles, coronary arteries) compared to the native recipient heart. Importantly, luciferase activity in recipient heart, liver, lung, spleen, or psoas muscle was within background levels. Similar to luciferase activity, the luciferase protein expression in the allograft appeared uniform and robust across all areas of the myocardium as well as in the coronary arteries. Importantly, despite high cop