SB Transposons for Gene Therapy

用于基因治疗的 SB 转座子

• 批准号: 7886257
• 负责人: PERRY B. HACKETT
• 金额: $ 36.72万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-01 至 2014-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7886257
• 关键词: Adult; Aftercare; Alkaline Phosphatase; Animal Model; Animals; Behavioral; Beta-glucuronidase; Biodistribution; Biological Assay; Biological Models; Brain; Canis familiaris; Catheters; Chromosomes; DNA; DNA delivery; Development; Disease; Dyes; Effectiveness; Enzymes; Equipment and supply inventories; Evaluation; Future; Gene Delivery; Gene Expression; Genes; Genome; Glycosaminoglycans; Goals; Grant; Harvest; Hemophilia A; Hepatic; Hepatocyte; Human; Immune response; Individual; Infusion procedures; Injection of therapeutic agent; Insertional Mutagenesis; L-Iduronidase; Liver; Maintenance; Mammals; Measures; Mediating; Medical; Mental Retardation; Methods; Modeling; Monitor; Mucopolysaccharidoses; Mucopolysaccharidosis I; Mucopolysaccharidosis VII; Mus; Neurologic; Organ; Physiological; Plasma; Plasmids; Procedures; Protocols documentation; Recombinants; Recovery; Reporter; Reporter Genes; Risk; Sampling; Site; Sleeping Beauty; System; Tissues; Transgenes; Treatment Cost; Treatment Efficacy; Vacuole; Validation; Viral Genes; design; dog genome; effective therapy; enzyme replacement therapy; gene delivery system; gene therapy; gene transfer vector; hepatic vein; in vivo; minimally invasive; mouse model; non-viral gene delivery; novel; plasmid DNA; preference; pressure; promoter; public health relevance; relating to nervous system; research study; scale up; therapeutic gene; therapeutic transposons; uptake; vector

DESCRIPTION (provided by applicant): New non-viral gene transfer procedures are needed for human gene therapy in order to achieve long-term maintenance and expression of newly introduced genes. The purpose of this project is to continue evaluation and validation of the Sleeping Beauty (SB) transposon system for its efficacy to catalyze integration of transgenes to cure mucopolysaccharidosis diseases in murine and canine model systems. In the past few years we have demonstrated our ability to deliver both 2-glucuronidase (GUSB) and a-L-iduronidase (IDUA) encoding genes to mouse liver and thereby correct accumulation of storage vacuoles in non-neural tissues of MPS I mice. Here, we propose to extend these studies in order to determine the efficacy of the SB transposon system as a gene transfer vector in the mouse brain and in the liver of a larger animal as a prelude to scale-up for human gene therapy. Our results indicate that hydrodynamic injection of therapeutic transposons is the best method for non-viral gene delivery to livers of adult mice for long-term expression. We hypothesize that this procedure also can be efficacious in humans if applied to individual organs or tissues in a relatively non-invasive manner using catheter delivery. Accordingly, the project has three sub-goals. The first will employ the mouse model system: 1) Examine the duration of expression of IDUA following delivery of SB transposon vectors in the presence of immune responses (Aim1). The second is to develop an effective delivery of SB transposons into dog liver (Aims 2 and 3). The third sub-goal is to examine the subsequent duration of expression of GUSB and the effects in the MPS VII dog model (Aim 4). We will examine the physiological, neurological, and behavioral effects of expression of GUSB in enzyme-deficient animals, as well as inventory the integration site preferences of the transposon vector following hydrodynamic delivery into chromosomes of dog liver to help evaluate potential risks from insertional mutagenesis. The experiments in this project will provide an assessment of the capacity of SB as a gene-transfer vector to target therapeutically important organs in an in vivo gene therapy protocol. The results will lay the groundwork for its future application to human gene therapy of a variety of diseases, e.g., several MPS diseases and hemophilia, which can be treated by gene delivery to the liver. PUBLIC HEALTH RELEVANCE: Mucopolysaccharidoses (MPS) are medical conditions caused by deficiencies in any of several enzymes that result in physiological abnormalities and mental retardation. Recombinant enzyme replacement therapy is available but treatment costs can range from $100,000 to $1,000,000 per year. The goal of this grant is to determine whether transposons containing therapeutic genes can cure MPS disease in dogs as a model for human gene therapy.

描述（由申请人提供）：人类基因治疗需要新的非病毒基因转移程序，以实现新引入基因的长期维持和表达。本项目的目的是继续评价和验证睡美人（SB）转座子系统在小鼠和犬模型系统中催化转基因整合以治愈粘多糖样变性疾病的功效。在过去的几年中，我们已经证明了我们能够将2-葡萄糖醛酸酶（GUSB）和α-L-艾杜糖醛酸酶（IDUA）编码基因递送至小鼠肝脏，从而纠正MPS I小鼠非神经组织中储存空泡的积累。在这里，我们建议扩展这些研究，以确定SB转座子系统作为基因转移载体在小鼠大脑和更大动物的肝脏中作为人类基因治疗规模扩大的前奏的功效。我们的研究结果表明，治疗性转座子的流体动力学注射是非病毒基因递送到成年小鼠肝脏进行长期表达的最佳方法。我们假设，如果使用导管递送以相对非侵入性的方式应用于个体器官或组织，则该程序在人体中也可以有效。因此，该项目有三个次级目标。第一个将采用小鼠模型系统：1）在存在免疫应答的情况下，检查SB转座子载体递送后IDUA表达的持续时间（Aim 1）。第二个是开发SB转座子到狗肝脏中的有效递送（目的2和3）。第三个子目标是检查GUSB表达的后续持续时间和MPS VII犬模型中的作用（目标4）。我们将研究GUSB在酶缺陷动物中表达的生理、神经和行为效应，以及在流体动力学递送到狗肝脏染色体中后清点转座子载体的整合位点偏好，以帮助评估插入诱变的潜在风险。在这个项目中的实验将提供一个评估的能力，SB作为一个基因转移载体，在体内基因治疗方案的目标治疗重要的器官。这一结果将为其未来应用于人类多种疾病的基因治疗奠定基础，几种MPS疾病和血友病，这可以通过基因递送到肝脏来治疗。 公共卫生关系：粘多糖贮积症（MPS）是由导致生理异常和智力迟钝的几种酶中的任何一种缺乏引起的医学病症。重组酶替代疗法是可用的，但治疗费用可能从每年10万美元到100万美元不等。这项资助的目的是确定含有治疗基因的转座子是否可以治愈狗的MPS疾病，作为人类基因治疗的模型。