Heme Oxygenas Gene Therapy of Heart Ischemia-reperfusion

心脏移植是各种终末期心脏病患者的首选治疗方法。在过去的二十年里，免疫抑制药物的发展、技术创新和术后护理的改进显著改善了心脏移植后的结果。然而，心脏移植物缺血再灌注(I-R)损伤仍然是导致早期移植物功能障碍和长期致病的主要发病率和死亡率来源。相当多的证据支持一个重要的 血红素加氧酶(HO-1)在脑缺血再灌注保护中的作用这种酶将血红素分解为胆绿素、游离铁和一氧化碳(CO)，后者起到减少炎症和细胞死亡的作用。这项建议的目的是开发合适的HSV-HO-1基因载体，特异性地靶向心脏血管内皮细胞的感染，并在足够的水平和持续时间内表达HO，以改善这种类型的缺血性疾病。在四个特定的目标中，将设计一个重定向的HSV载体(HSV-HOT)，其中内皮特异性受体VEGF-R2(血管内皮生长因子受体)[和/或Tie-2(血管生成素受体)]将代替天然的HSV受体HveA和HveC用于病毒的附着和穿透。如果需要， 我们还将探索一种替代的重定向策略，该策略利用靶向可溶性适配器(Aim 1)。一种高度工程的、复制缺陷的载体主干将用于基因传递。该载体无细胞毒性，高度稳定，能够高效表达转基因，适合于高效价生产和纯化，使用的细胞系是为补充反式病毒功能缺陷而设计的。临床前研究将在体外使用人内皮细胞(AIM 2)进行，随后将在啮齿动物(AIM 3)、猪和灵长类动物(AIM 4)心脏移植模型中进行体内研究。病媒应用研究将包括开发对心脏血管进行病媒管理的程序、病媒分析 剂量和分布，HO-1表达和副产物合成(如CO)的特征，以及I-R保护的评估。我们的长期目标是开发在心脏移植受者身上进行早期I期试验所需的临床前有效性和安全性数据。

Heart transplantation is the preferred therapy for patients with a variety of end-stage heart diseases. Over the last two decades, developments in immunosuppressive medications, technical innovations, and improvements in postoperative care have significantly improved outcomes following heart transplantation. Cardiac allograft ischemia-reperfusion (I-R) injury, however, remains a major source of morbidity and mortality leading to both early allograft dysfunction as well as long-term morbidity. Considerable evidence supports an important role for heme oxygenase (HO-1) in protection against I-R. This enzyme catabolizes heme into biliverdin, free iron and carbon monoxide (CO), which act to reduce inflammation and cell death. The goal of this proposal is to develop suitable HSV-HO-1 gene vectors that specifically target infection of heart blood vessel endothelium and express HO in sufficient levels and duration to ameliorate this type of ischemic disease. In four specific aims, a retargeted HSV vector (HSV-HOT) will be engineered in which the endothelial-specific receptors VEGF-R2 (vascular endothelial growth factor receptor) [and/or Tie-2 (angiopoietin receptor)] will be used for virus attachment and penetration in lieu of the natural HSV receptors HveA and HveC. If needed, we will also explore an alternative retargeting strategy that utilize a targeting soluble adapter (Aim 1). A highly engineered, replication defective vector backbone will be used for gene delivery. This vector is noncytotoxic, highly stable, capable of vigorous transgene expression and suitable for high titer manufacture and purification using cell lines that are engineered to complement the defective viral functions in trans. Preclinical studies will be carried out in vitro using human endothelial cells (Aim 2) followed by in vivo studies in rodent (Aim 3), pig and primate (Aim 4) heart transplantation models. Vector application studies will include the development of procedures for vector administration to heart blood vessels, analyses of vector dosing and distribution, characterization of HO-1 expression and byproduct synthesis (e.g. CO), and evaluation of protection from I-R. It is our long-term goal to develop the preclinical efficacy and safety data needed to perform early phase I trials in heart transplant recipients.