The broad long-term objectives of this proposal are to achieve targeted, efficient and durable gene transfer to the transplanted heart and, using biologically relevant genes for isoforms of nitric oxide synthase (NOS), to normalize coronary arterial vasoreactivity and reduce cardiac allograft vasculopathy (CAV) after allotransplantation in both a small and large animal model. The central hypotheses of this proposal are that, using the unique transplant setting, targeted, efficient and durable gene transduction of blood vessels of the heart or the myocardium can be achieved and that abnormalities in coronary vascular relaxation and the development of CAV after heart transplantation can be reduced by efficient cardiac transduction with endothelial or inducible NOS (eNOS or iNOS). CAV is the greatest obstacle to long-term patient survival after cardiac transplantation. Gene therapy may be particularly applicable in the setting of transplantation as the donor organ is uniquely available for genetic modification ex vivo prior to implantation into a recipient. Major challenges to successful clinical gene genetic modification ex vivo prior to implantation into a recipient. Major challenges to successful clinical gene therapy include the need for optimal vectors and delivery systems to transfer genetic material to tissues in vivo. Recently, for the first time, physical and chemical conditions of gene delivery to the heart have been shown to allow selective targeting of the vector to the media of the coronary arteries of the myocardium. Using a normothermic perfusion system, the usual period of storage of the donor heart before transplantation can be transformed into a period for optimal gene delivery without affecting the viability of the graft.
The first aim of transformed into a period for optimal gene delivery without affecting the viability of the graft.
The first aim of the current proposal is to identify the most suitable vector (adenovirus or lentivirus) and delivery system using chemical modification of a normothermic perfusion system for efficient, targeted and durable gene delivery in the heterotopic rat heart transplant model.
The second aim i s to study, using the most suitable vector and same delivery system from Specific Aim 1, the effects of eNOS or iNOS gene transfer to either the coronary vasculature or myocardium of the donor heart on coronary arterial vascular reactivity and on CAV after heterotopic pig heart allotransplantation. NOS will be studied as the gene of choice as reduced nitric oxide bioavailability is characteristic of vessels undergoing CAV. Abnormal endothelium dependent relaxation precedes CAV, so that successful NOS gene transfer might be expected to impact on the early stages of the disease. Before gene transfer technology to the transplanted heart can progress to the clinic, satisfactory experience in a large animal model will be necessary. The porcine model is particular suitable from a physiological and pathological viewpoint and is also the animal of choice for future clinical xenotransplantation. Success in achieving the specific aims of this proposal opens up the potential for clinical gene therapy to avoid CAV after heart transplantation. Implications also exist for the application of these techniques for the modification of non-allograft arteriosclerosis.
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