Delivery of Sleeping Beauty Transposons to Dog Liver for Gene Therapy

将睡美人转座子输送到狗肝脏进行基因治疗

基本信息

批准号：
7537339
负责人：
PERRY B. HACKETT
金额：
$ 29.09万
依托单位：
DISCOVERY GENOMICS, INC.
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-08-15 至 2011-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7537339
关键词：
Aftercare Animal Model Animals Beta-glucuronidase Biodistribution Biological Biological Assay Blood Blood flow Blood specimen Canis familiaris Catheters Cells Communities Condition DNA DNA delivery Development Disease Dyes Effectiveness Enzymes Erythropoietin Evaluation Factor IX Factor VIII Gene Expression Gene Transfer Genes Genome Genomics Goals Harvest Hematopoietic Hemophilia A Hepatic artery Hepatocyte Hereditary Disease Human Infusion procedures Injection of therapeutic agent Invasive Liver Localized Lysosomal Storage Diseases Mammals Measures Mediating Methods Modeling Monitor Mus Operative Surgical Procedures Organ Placement Plasmids Polymerase Chain Reaction Procedures Public Health Recovery Reporter Genes Reporting Sleeping Beauty Small Business Funding Mechanisms Small Business Innovation Research Grant Solutions System Tail Techniques Testing Therapeutic Thoracic Surgical Procedures Tissues Transgenic Organisms Transposase Validation Veins Viral base cell type clinically relevant design gene therapy hepatic vein human tissue interest non-viral gene therapy plasmid DNA pressure research study response scale up therapeutic gene transgene expression uptake

项目摘要

DESCRIPTION (provided by applicant): The overall goal of Discovery Genomics, Inc. is to develop the Sleeping Beauty transposon system for human gene therapy. To achieve this to treat hemophilia, we must be able to direct uptake and long-term expression of therapeutic genes in targeted tissues such as the liver. Only one method of plasmid delivery has been effective for high-level gene expression in the liver in mice - the rapid, high-pressure delivery known as hydrodynamic injection. In the mouse, hydrodynamic delivery of DNA requires injection of a large volume (10% vol/wt) of a DNA solution through the tail vein in less than 10 seconds. For larger animals, this procedure is conjectured to be unacceptable on a whole animal basis. In this highly focused project we propose to develop catheter-based, local hydrodynamic delivery to the liver of dogs that will require minimal surgery and amounts of therapeutic DNA. These features are especially important for the delivery of Factor IX and Factor VIII-expressing transposons for treatment of hemophilia. Recognizing that delivery to only one cell type in the liver (and other organs) is not possible, we will to couple physical and biological controls over gene expression to our transgenic constructs that will reduce their expression in hematopoietic cells to reduce undesirable immunological responses. This project will dovetail with our ongoing SBIR project to deliver Factor VIII and IX genes in dogs via open-chest surgery for treatment of hemophilia. The Specific Aims of the project are to 1) develop appropriate transposons with reporter genes and recovery cassettes that can be used to evaluate the efficacy of hydrodynamic delivery in dogs, 2) determine an optimal, catheter-based delivery procedure that can be used to inject DNA with minimal surgical intervention into canine livers, and 3) demonstrate that long-term gene expression will result from transposition of Sleeping Beauty transposons into canine hepatocytes using the procedure(s) developed in Aim 2. Public Health Relevance: Delivery of Sleeping Beauty Transposons to Dog Liver for Gene Therapy. There currently is no efficient and effective method for delivery of Sleeping Beauty transposons to liver in humans for gene therapy. The goal of this project is to develop a transposon delivery system that will allow hydrodynamic delivery of SB transposons in dogs as a model large animal for humans.

描述（由申请人提供）：Discovery Genomics, Inc.的总体目标是开发用于人类基因治疗的睡美人转座子系统。为了实现血友病的治疗，我们必须能够在肝脏等目标组织中指导治疗基因的摄取和长期表达。只有一种质粒递送方法对小鼠肝脏中的高水平基因表达有效——快速、高压递送，称为流体动力注射。在小鼠中，DNA 的流体动力传递需要在 10 秒内通过尾静脉注射大量（10% vol/wt）的 DNA 溶液。对于较大的动物，推测该程序在整个动物基础上是不可接受的。在这个高度集中的项目中，我们建议开发基于导管的局部流体动力学输送到狗的肝脏，这将需要最少的手术和大量的治疗性 DNA。这些特征对于递送表达因子 IX 和因子 VIII 的转座子以治疗血友病尤其重要。认识到仅递送至肝脏（和其他器官）中的一种细胞类型是不可能的，我们将对基因表达的物理和生物控制与我们的转基因构建体结合起来，这将减少它们在造血细胞中的表达，从而减少不良的免疫反应。该项目将与我们正在进行的 SBIR 项目相吻合，通过开胸手术向狗提供因子 VIII 和 IX 基因以治疗血友病。该项目的具体目标是 1) 开发带有报告基因和恢复盒的适当转座子，可用于评估狗中流体动力学递送的功效，2) 确定可用于注射的最佳基于导管的递送程序DNA 以最少的手术干预进入犬肝脏，3) 证明使用目标 2 中开发的程序将睡美人转座子转座到犬肝细胞中将产生长期基因表达。公共卫生相关性：将睡美人转座子输送到狗肝脏进行基因治疗。目前尚无高效且有效的方法将睡美人转座子递送至人类肝脏进行基因治疗。该项目的目标是开发一种转座子传递系统，允许在作为人类大型动物模型的狗体内进行水动力传递 SB 转座子。