GENETIC ENGINEERING OF A FAILURE RESISTANT VEIN GRAFT

静脉旁路移植术是治疗冠状动脉和外周循环动脉粥样硬化闭塞性疾病的主要方法。静脉移植失败，最终导致血栓性闭塞，会导致显著的发病率和死亡率，以及需要昂贵的再干预。基因转移技术的发展为在分子水平上改变静脉移植物生物学的预防策略提供了可能。该提案旨在制定一项转化研究计划，以解决目前限制基因工程应用于静脉移植失败问题的关键障碍。将采用多学科方法，采用最先进的基于病毒的载体系统，结合体内基因表达的分子和功能分析。该计划将支持申请人的发展，在遗传方法培训的血管外科医生，成为一个独立的研究者能够进行临床前和临床评价的遗传干预血管疾病。研究和培训将在基因转移技术领域一位公认的领导者的指导下进行，其环境旨在通过提供关键的核心技术和专门知识来简化这种方法的发展。该方法分为两个不同的组成部分。第一个涉及严格评估当前和不断发展的病毒载体系统，其应用于静脉移植。将检查新型腺病毒、腺相关病毒、假型逆转录病毒和慢病毒载体系统的递送效率、基因表达的稳定性和宿主反应。静脉移植物内皮将构成这些策略的主要目标。将研究组织特异性（即内皮）启动子与病毒启动子对转基因表达的影响。第二阶段将集中于产生抗血栓性静脉移植物表面的特定治疗目标。设计用于增加静脉内皮的天然抗凝剂（硫酸乙酰肝素合成、ADP水解、蛋白C活化）或血栓溶解（纤溶酶活化）功能的基因构建体将用于静脉移植的动物模型中。长期目标是开发一种安全、高效和有效的术中基因治疗方案，该方案将设计抗闭塞的静脉旁路移植物。

Vein bypass grafting is a mainstay of therapy for patients with atherosclerotic occlusive diseases of the coronary and peripheral circulations. Failure of vein grafts, ultimately resulting in thrombotic occlusion, incurs significant morbidity and mortality as well as the need for costly reinterventions. The development of gene transfer technology offers the potential for preventive strategies designed to modify vein graft biology at the molecular level. This proposal seeks to develop a translational research program for addressing the critical obstacles currently limiting the application of genetic engineering to the problem of vein graft failure. A multidisciplinary approach, employing state of the art viral-based vector systems combined with molecular and functional analysis of gene expression in-vivo, will be employed. The program will support the development of the applicant, a vascular surgeon trained in genetic approaches, into an independent investigator capable of conducting preclinical and clinical evaluations of genetic interventions for vascular disease. The research and training will take place under the mentorship of a recognized leader in gene transfer technology, in an environment designed to streamline the development of such approaches by providing critical core technologies and expertise. The approach is divided into two distinct components. The first involves the rigorous evaluation of current and evolving viral vector systems for their application to vein grafting. Novel adenovirus, adeno-associated virus, pseudotyped retrovirus, and lentivirus vector systems will be examined for delivery efficiency, stability of gene expression, and host responses. The vein graft endothelium will constitute the primary target of these strategies. The effects of tissue specific (i.e. endothelial) versus viral promoters on transgene expression will be studied. The second phase will focus on the specific therapeutic goal of generating a thromboresistant vein graft surface. Gene constructs designed to increase natural anticoagulant (heparan sulfate synthesis, ADP hydrolysis, protein C activation) or thrombolytic (plasmin activation) functions of venous endothelium will be employed in animal models of vein grafting. The long term objective is to develop a safe, efficient, and effective protocol for intraoperative gene therapy that will engineer vein bypass grafts that are resistant to occlusion.