Vascular-targeted gene therapy to block proliferation of smooth muscle cells using a novel adenovirus vector

已经出现了许多策略来改善急性损伤后血管壁的功能，包括靶向内皮细胞功能、血管平滑肌细胞增殖和迁移以及外膜祖细胞。microRNA是小的非编码RNA，其具有通过靶向3 'UTR中的种子序列靶向数百个基因的能力。我们已经证明miRNA在血管平滑肌细胞功能和血管重塑的发展中是必不可少的（Deng 2015/MacDonald 2013）。我们已经鉴定出具有阻断血管平滑肌细胞增殖能力的新型miRNA。一个主要的进步将是开发新的方法，通过基于病毒的递送方法将治疗性miRNA递送到血管壁中。与Havenga博士合作多年，促进了用于基因治疗的新型腺病毒载体的开发（Waddington Cell，2008）。最近，我们已经鉴定、纯化并测试了一种称为Ad 20/42/42的新型人腺病毒载体，其显示出向内皮细胞有效递送基因，但向肝（腺病毒载体积累的主要部位）的递送非常有限。此外，该病毒在一般人群中显示出非常低水平的预先存在的中和作用（约10% vs 60%，人腺病毒5），使其成为应用于人基因治疗的非常有吸引力的病毒。因此，在本项目中，我们将在血管损伤后的血管基因治疗环境中测试该载体（以及在项目过程中可用的其他新的有前途的载体）。因此，该项目将在以下方面具有突出的培训潜力：病毒学、载体构建和繁殖血管细胞的体外细胞培养急性血管损伤和病毒递送的体内模型治疗性miRNA的体内评估该项目还受益于与意大利的里雅斯特Giacca实验室合作开发治疗性miRNA。

Many strategies have emerged to improve the function of the blood vessel wall following acute injury, including targeting endothelial cell function, vascular smooth muscle cell proliferation and migration and adventitial progenitor cells. microRNA are small non-coding RNA that have the capacity to target many hundreds of genes through targeting seed sequences in the 3'UTR. We have shown the miRNA are essential in vascular smooth muscle cell function and development of vascular remodelling (Deng 2015/MacDonald2013). We have identified novel miRNA that have the capacity to block vascular smooth muscle cell proliferation. A major step forward would be the development of novel approaches to deliver therapeutic miRNA into the vessel wall by virus-based delivery approaches. Working with the Dr Havenga over many years has facilitated the development of novel adenovirus-based vectors for gene therapy (Waddington Cell, 2008). Recently, we have identified, purified and tested a novel human adenovirus vector called Ad20/42/42 that displays effective delivery of genes to endothelial cells but very restricted delivery to the liver - the main site of adenovirus vector accumulation. Further, this virus shows very low levels of pre-existing neutralisation in the general population (approximately 10% vs 60% for human adenovirus 5) making this a very attractive virus for application to human gene therapy. In this project we will therefore test this vector (and other novel promising vectors that become available during the course of the project) in the setting of vascular gene therapy post injury of the vasculature. The project will therefore have outstanding training potential in:Virology, vector construction and propagationIn vitro cell culture of vascular cells In vivo models of acute vascular injury and delivery of virusEvaluation of therapeutic miRNA in vivo.The project additionally benefits from a collaboration on the development of therapeutic miRNA with the Giacca lab in Trieste, Italy.