Vector Identification and Gene Delivery Approach in Pigs

猪的载体鉴定和基因传递方法

• 批准号: 6738715
• 负责人: Judith K Gwathmey
• 金额: $ 18.88万
• 依托单位: GWATHMEY, INC.
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-09-10 至 2005-08-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6738715
• 关键词: adeno associated virus group; bioenergetics; biotechnology; calcium flux; calcium transporting ATPase; cardiac myocytes; disease /disorder model; gene delivery system; gene expression; gene therapy; genetic promoter element; genetic transduction; heart failure; nerve /myelin protein; recombinant virus; sarcoplasmic reticulum; swine; therapy design /development; transfection /expression vector

DESCRIPTION (provided by applicant): The goal of this proposal is to develop a novel therapy for molecular inotropy, specifically, viral-mediated myocardial delivery of the calcium regulatory protein SERCA 2a (sarcoplasmic reticulum Ca 2¿ATPase) to large animals exhibiting heart failure in an attempt to restore function and improve survival without worsening energetic parameters. Congestive heart failure (CHF) represents an enormous clinical problem demanding effective therapeutic approaches. Despite advances in traditional approaches to its treatment, including pharmacologic management, myocardial revascularization, mechanical assist devices, and transplantation, CHF remains a leading cause of death worldwide. Therefore, a novel therapy aimed at decreasing the morbidity and mortality of CHF and improving the quality of life for millions of patients is particularly attractive. Cardiac gene therapy has shown promise in early animal studies and lends itself to the treatment of heart failure. A defect in intracellular calcium handling is known to be a key abnormality in both human and experimental CHF. Deficient Ca 2¿uptake by the sarcoplasmic reticulum (SR) during relaxation in failing hearts from humans and animal models has been associated with a decrease in the expression and activity of SR Ca2+ATPase (SERCA2a). Our preliminary work over the last five years have shown that 1) Gene transfer is an effective means of introducing the SERCA2a gene into myocytes in vitro and in vivo and 2) that increasing the expression of SERCA2a restores contractility and normalizes intracellular calcium cycling in a rodent model of CHF. We are now extending our experiments from rodents to porcine models. We aim to 1) determine the efficiency of transduction of viral vectors in cardiomyocytes isolated from pig hearts; 2) determine the efficiency of transduction of the AAV vectors and El-E4 deleted recombinant adenovirus following gene transfer in vivo in pigs; 3) test the proximal human brain natriuretic (hBNP) promoter (-408 to +100 relative to transcription start site) for the ability to be induced by pressure-overload versus ischemia.

描述（由申请人提供）： 该提案的目标是开发一种分子正性肌力的新疗法，特别是病毒介导的心肌递送钙调节蛋白SERCA 2a（肌浆网Ca 2¿ATP酶）到表现出心力衰竭的大型动物，以试图恢复功能和尽管充血性心力衰竭（CHF）的传统治疗方法（包括药物管理）取得了进展，但在不恶化能量参数的情况下提高生存率仍然是一个巨大的临床问题。血运重建、机械辅助装置和移植，CHF 仍然是全球死亡的主要原因，因此，旨在降低 CHF 发病率和死亡率并改善数百万患者生活质量的新疗法特别有吸引力。细胞内钙处理缺陷被认为是人类和实验性 CHF 缺钙的一个关键异常。人类和动物模型衰竭心脏舒张期间肌浆网 (SR) 的摄取与 SR Ca2+ ATP 酶 (SERCA2a) 表达和活性的降低有关。我们过去五年的初步工作表明 1。 ) 基因转移是在体外和体内将 SERCA2a 基因引入肌细胞的有效方法，2) 增加 SERCA2a 的表达可以恢复收缩性并使细胞内钙循环正常化。我们现在将我们的实验从啮齿动物扩展到猪模型，我们的目标是 1) 确定从猪心脏分离的病毒载体的转导效率；2) 确定 AAV 载体和 El 的转导效率。 -E4在猪体内基因转移后删除重组腺病毒3)测试近端人脑钠尿(hBNP)启动子(相对于转录起始-408至+100)位点），以了解压力过载与缺血诱导的能力。