Enhanced delivery method for distribution of lentiviral and AAV vectors

广泛和可控的病毒载体分布是实现人类基因治疗帕金森病的关键。鉴于人类纹状体的体积，需要开发方法来充分覆盖病媒。对流增强递送（CED）是最近开发的用于将小分子和大分子递送到实体组织中的靶向部位的方法，其利用整体流动机制将大分子递送和分配到临床上显著体积的组织。该技术是基于一个压力输注系统，“推动”和替代周围细胞的实质细胞外液。携带AAV-2颗粒的流体的递送导致载体在CNS内的显著扩散。该递送系统 通过改善分布体积、提供靶组织内药物浓度的均匀分布以及将治疗因子完全递送至靶部位而提供显著的优点。AAV-2与细胞表面上的硫酸肝素蛋白聚糖结合并增强AAV-2与细胞的相互作用。至于其他肝素结合蛋白，AAV-2与肝素的结合亲和力低，细胞转导需要高亲和力受体。初步数据表明，将肝素添加到AAV-2颗粒中允许AAV-2在解剖结构内的稳健的基于对流的递送。该项目将系统地检查CED与肝素组合沿着对AAV在整个猴纹状体中分布的影响。将测试的相同方法也将应用于VSV-g假型慢病毒载体。

Widespread and controllable distribution of viral vectors is essential to achieve human gene therapy for Parkinson's disease. Given the volume of the human striatum methods development is required for adequate vector coverage. Convection-enhanced delivery (CED), a recently developed approach for delivery of small and large molecules to targeted sites in solid tissues, utilizes a bulk flow mechanism to deliver and distribute macromolecules to clinically significant volumes of tissue. The technique is based on a pressure infusion system that "pushes" and replaces extracellular fluid in surrounding cells of the parenchyma. Delivery of fluid carrying the AAV-2 particles results in significant spread of the vector within CNS. This delivery system offers significant advantages by improving the volume of distribution, providing uniform distribution of drug concentration within the targeted tissue, and complete delivery of the therapeutic factor to the target site. AAV-2 binds to heparin-sulfate proteoglycans on the cell surface and enhances the interaction of AAV-2 with cells. As for other heparin-binding proteins the binding affinity of AAV-2 to heparin is low and a high-affinity receptor is required for cell transduction. Preliminary data indicate that heparin addition to AAV-2 particles allows for robust convection based delivery of AAV-2 within anatomical structures. This project will systematically examine the impact of combined CED along with heparin to distribute AAV throughout the monkey striatum. The same methodologies will be tested will also be applied to VSV-g pseudotyped lentivirus vectors.