Adenoviral/lentiviral vectors for genetic modification

用于基因修饰的腺病毒/慢病毒载体

• 批准号: 6661537
• 负责人: CHRISTOPHER G.A. MCGREGOR
• 金额: $ 30.7万
• 依托单位: MAYO CLINIC ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-09-01 至 2003-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6661537
• 关键词: angiogenesis; blood vessel disorder; coronary vessels; enzyme therapy; gene therapy; heart transplantation; homologous transplantation; isolation perfusion; laboratory rat; nitric oxide; nitric oxide synthase; nonhuman therapy evaluation; postoperative complications; swine; transfection /expression vector

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 is 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.

该提案的广泛长期目标是实现有针对性、高效和持久的基因转移到移植心脏，并利用一氧化氮合酶（NOS）亚型的生物学相关基因，在小型和大型动物模型中使冠状动脉血管反应性正常化并减少同种异体移植后的心脏同种异体移植血管病变（CAV）。该提案的中心假设是，利用独特的移植环境，可以实现心脏或心肌血管的靶向、高效和持久的基因转导，并且可以通过内皮型或诱导型NOS（eNOS或iNOS）进行有效的心脏转导来减少心脏移植后冠状血管舒张异常和CAV的发展。 CAV是心脏移植后患者长期生存的最大障碍。基因治疗可能特别适用于移植，因为供体器官在植入受体之前可以进行离体基因改造。在植入受体之前成功进行离体临床基因遗传修饰的主要挑战。成功的临床基因治疗的主要挑战包括需要最佳的载体和递送系统来将遗传物质转移到体内组织。最近，首次显示基因递送至心脏的物理和化学条件允许载体选择性靶向心肌冠状动脉中层。使用常温灌注系统，移植前供体心脏的通常储存期可以转变为最佳基因递送期，而不影响移植物的活力。第一个目标是在不影响移植物活力的情况下转化为最佳基因传递时期。当前提案的第一个目标是通过对常温灌注系统进行化学修饰来确定最合适的载体（腺病毒或慢病毒）和递送系统，以便在异位大鼠心脏移植模型中实现高效、有针对性和持久的基因递送。第二个目标是使用Specific Aim 1中最合适的载体和相同的递送系统，研究eNOS或iNOS基因转移到供体心脏的冠状脉管系统或心肌对异位猪心脏同种异体移植后冠状动脉血管反应性和CAV的影响。 NOS 将作为首选基因进行研究，因为一氧化氮生物利用度降低是接受 CAV 的血管的特征。异常的内皮依赖性松弛先于 CAV，因此成功的 NOS 基因转移可能会对该疾病的早期阶段产生影响。在移植心脏的基因转移技术能够进入临床之前，需要在大型动物模型中获得令人满意的经验。从生理和病理角度来看，猪模型特别适合，也是未来临床异种移植的首选动物。成功实现该提案的具体目标开启了临床基因治疗避免心脏移植后 CAV 的潜力。这些技术对于改善非同种异体移植动脉硬化的应用也存在意义。