Polymer Carrier for Ischemia-Inducible Gene Therapy

描述(由申请人提供)： 尽管缺血性心脏病(IHD)的治疗取得了进展，但每年仍有超过50万美国人进展到IHD末期和充血性心力衰竭。基因治疗已经成为治疗IHD的一种很有前途的选择。在最初的人类试验中，使用血管内皮生长因子(VEGF)的基因治疗最近被证明有助于保护动物冠状动脉结扎后的心肌功能，并有助于缓解心肌缺血症状。然而，基因疗法的广泛适用性仍然存在重大限制。目前将基因输送到心肌的方法受到患者发病率和死亡率的潜在限制，或者是因为低的基因转染率。此外，同时影响缺血和非缺血心肌的IHD的基因治疗与血管瘤的形成和动脉粥样硬化斑块的进展有关。在基因治疗能够广泛应用于IHD之前，还需要开发安全、高效的基因转染和调控基因表达的技术。我们建议：1)开发两种新型的多功能阳离子水溶性脂聚合物，以最小的毒性实现高水平的靶向基因转移；2)开发治疗IHD的基因结构，其表达受心肌内氧分压的调节。我们的脂聚合物含有阳离子DNA缩合剂、亲水间隔物和疏水分子。这些聚合物具有良好的溶解性、有效的溶酶体逃逸和有效的摄取。促红细胞生成素EPO增强子、EPO 3‘-非翻译区(3’-UTR)和氧依赖降解结构域将被整合到血管内皮生长因子基因中，以促进心肌缺血区域的稳定蛋白表达。这些基因构建物将使用水溶性脂聚合物载体输送到心肌。在体外证明了高转染率和缺血诱导的基因表达后，我们将在手术模型中检测体内血管内皮生长因子的调节表达。这些研究将是1)以最小毒性有效地转导心肌的临床有用载体的发展，以及2)仅限于心肌缺血区表达的治疗性基因的开发，有助于使基因治疗在IHD中的安全应用成为临床现实。

DESCRIPTION (provided by applicant): Despite advances in the treatment of ischemic heart disease (IHD), more than 500,000 Americans progress to end-stage IHD and congestive heart failure each year. Gene therapy has emerged as a promising option for the treatment of IHD. Gene therapy using Vascular Endothelial Growth Factor (VEGF) has recently been demonstrated to help preserve myocardial function following coronary artery ligation in animals and to help relieve symptoms of myocardial ischemia in initial human trials. Significant limitations remain, however, to the broad applicability of gene therapy. Current methods of gene delivery to the myocardium are limited by the potential for patient morbidity and mortality or, alternatively, by low transfection efficiency. Further, gene therapy for IHD that affects both ischemic and non-ischemic myocardium is associated with angioma formation and atherosclerotic plaque progression. Before gene therapy can be widely applied to IHD, techniques for safe, efficient gene transfection and regulated gene expression will need to be developed. We propose to: 1) develop two novel multifunctional cationic water-soluble lipopolymers to achieve high levels of targeted gene transfection with minimal toxicity and 2) develop therapeutic gene constructs for IHD whose expression is regulated by oxygen tension within the myocardium. Our lipopolymers contain a cationic DNA condensing agent, a hydrophilic spacer, and a hydrophobic molecule. These polymers offer good solubility, effective lysosomal escape and efficient uptake. The erythropoietin EPO enhancer, the EPO 3'- untranslated region (3'-UTR) and an oxygen dependent degradation domain will be incorporated into the VEGF gene to promote stable protein expression in only ischemic regions of the myocardium. These gene constructs will be delivered to the myocardium using water-soluble lipopolymer carriers. After demonstrating high transfection efficiency and ischemia-inducible gene expression in vitro, we will test the regulated expression of VEGF in vivo in surgical models of IHD. These studies will be integral to the development of 1) a clinically useful vector for efficient transfection of the myocardium with minimal toxicity and 2) therapeutic genes whose expression is limited to ischemic regions of the myocardium, helping to make the safe application of gene therapy for IHD a clinical reality.