Targeted gene delivery and activation by ultrasound

描述（由申请人提供）：基因治疗的成功主要依赖于新技术的发展，这些新技术可用于产生位点特异性基因传递，而不会引起全身毒性和免疫原性反应，以及调节体内转基因表达的能力。尽管过去进行了大量的努力，但基因治疗的这些主要障碍仍然存在。本研究旨在开发新型超声系统，通过提供新的技术来克服这些障碍：1)靶向基因传递到内脏器官；2)体内转基因表达的空间和时间调控。在初步研究中，我们已经证明了超声介导的体外基因转移的可行性，利用培养的HeLa细胞和碎石冲击波（LSW）介导的猪肾脏体内位点特异性基因传递。我们也证明了超声可以作为一种简单方便的物理手段来激活热休克蛋白（hsp） 70B启动子控制下的转基因表达。本提案中概述的工作将是这些初步研究的重要扩展，并将包括多学科和全面的研究，将创新的工程方法与细胞和分子生物学的现代技术以及动物实验相结合。我们的具体目标是：1。用于靶向基因传递和激活的新型超声系统的发展。2. 新型超声系统在水和组织幻象中产生的声场和相关空化泡动力学的物理特性。3. 超声介导基因体外传递的优化。4. 超声靶向基因在猪肾中的传递；超声调控骨骼肌基因传递和激活——慢性肾衰竭大鼠模型中促红细胞生成素（Epo）基因治疗的新方法由于超声波可以穿透组织深处并聚焦于身体的特定器官，因此超声波靶向基因在体内的传递和激活的前景非常吸引人。这种基因传递和激活的多用途物理方法对于多种疾病的分子治疗具有重要价值，包括癌症、心血管疾病和各种肾脏疾病（多囊肾、肾癌、急性肾小球肾炎和慢性间质疾病）。

DESCRIPTION (provided by applicant): The success of gene therapy relies critically on the development of new technologies that can be used to produce site-specific gene delivery without causing systemic toxicity and immunogenic response, as well as the ability to regulate transgene expression in vivo. Despite extensive efforts in the past, these primary obstacles in gene therapy still exist. This proposal aims to develop novel ultrasound systems that have the potential to overcome these obstacles by providing new techniques for 1) targeted gene delivery to internal organs and 2) spatial and temporal regulation of transgene expression in vivo. In preliminary studies, we have already demonstrated the feasibility of ultrasound-mediated gene transfer in vitro using cultured HeLa cells and lithotripter shock wave (LSW)-mediated site-specific gene delivery in vivo in porcine kidney. We have also shown that ultrasound can be used as a simple and convenient physical means to activate transgene expression under the control of heat shock protein (hsp) 70B promoter. The work outlined in this proposal will be a significant expansion of these preliminary studies and will encompass a multidisciplinary and comprehensive investigation combining innovative engineering approaches with modem techniques in cell and molecular biology, and animal experimentations. Our specific aims are: 1. Development of novel ultrasound systems for targeted gene delivery and activation. 2. Physical characterization of the acoustic fields and associated cavitation bubble dynamics produced by the novel ultrasound systems both in water and in tissue phantoms. 3. Optimization of ultrasound-mediated gene delivery in vitro. 4. Ultrasound-targeted gene delivery in porcine kidney, and 5. Ultrasound-regulated gene delivery and activation in skeletal muscle -a novel approach for erythropoietin (Epo) gene therapy in a rat model with chronic renal failure. Because ultrasound can penetrate deep into tissue and be focused on specific organs of the body, the prospect for ultrasound-targeted gene delivery and activation in vivo is very appealing. Such a versatile physical method for gene delivery and activation could be of great value for molecular therapies of a wide range of diseases, including cancer, cardiovascular disease, and various renal disorders (polycystic kidney, renal cancer, acute glomerulonephritis, and chronic interstitial disease).