Towards gene therapy of mitochondrial disease

线粒体疾病的基因治疗

基本信息

批准号：
7845571
负责人：
MICHAEL D KOOB
金额：
$ 18.75万
依托单位：
UNIVERSITY OF MINNESOTA
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-05-15 至 2011-10-30
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): A wide array of inherited human diseases are caused by mutations in our mitochondrial DNA (mtDNA) genome and in mitochondrial genes encoded in the nuclear genome, but there are currently no effective therapies for these clinically devastating diseases. We are able to engineer yeast mtDNA and, moreover, we have developed technology that allows us for the first time to engineer mammalian mitochondrial genomes and reintroduce these genomes into mouse embryos. We propose to use this technology to develop a gene therapy for Friedreich's ataxia (FRDA), an autosomal recessive neurodegenerative disease caused by defects in frataxin, a nucleus-encoded mitochondrial protein. We will focus our initial efforts on correcting the molecular deficits associated with the complete loss of frataxin in yeast. This well characterized model of FRDA will enable us to systematically assess both general and protein-specific features required to efficiently express a fully functional form of frataxin from the mitochondrial genome. We will use the information and reagents developed in this initial phase of the project to engineer a set of mouse mtDNA genomes suitable for correcting deficits in a mouse model of FRDA. We will evaluate the efficiency with which these genomes compensate for the loss of the mouse nuclear frataxin gene by packaging them in mitochondria and injecting them into single cell embryos of FRDA knockout mice. Because this gene knockout mutation leads to loss of mitochondrial function and so is embryonic lethal, we can readily assay the functionality of the mitochondrial frataxin genes by their ability to either partially rescue (i.e., generate viable embryonic cells) or fully rescue this phenotype (i.e., generate viable mice). This experimental system will therefore allow us to optimize both our mitochondrial transfer technology and our mitochondrial frataxin gene constructs. Once we have completed the work described in this application we will be in an excellent position to develop a gene therapy approach for mouse models of FRDA and to work towards adapting these therapies to treating FRDA and other mitochondrial diseases in humans. PUBLIC HEALTH RELEVANCE: A wide array of inherited human diseases are caused by mutations in our mitochondrial DNA (mtDNA) genome and in mitochondrial genes encoded in the nuclear genome, but there are currently no effective therapies for these clinically devastating diseases. The experiments described in this proposal will give us both greater molecular insights into one of these diseases (FRDA) and will allow us to develop mtDNA engineering and transfer technologies that will serve as indispensable tools for developing therapies to treat these diseases.

描述（由申请人提供）：多种遗传性人类疾病是由我们的线粒体DNA（mtDNA）基因组和核基因组中编码的线粒体基因的突变引起的，但目前还没有针对这些临床毁灭性疾病的有效疗法。我们能够改造酵母线粒体DNA，此外，我们还开发了技术，使我们能够首次改造哺乳动物线粒体基因组并将这些基因组重新引入小鼠胚胎中。我们建议利用这项技术开发一种针对弗里德赖希共济失调 (FRDA) 的基因疗法，FRDA 是一种由 frataxin（一种核编码线粒体蛋白）缺陷引起的常染色体隐性神经退行性疾病。我们最初的努力将集中于纠正与酵母中 frataxin 完全丧失相关的分子缺陷。这种经过充分表征的 FRDA 模型将使我们能够系统地评估从线粒体基因组中有效表达全功能形式的 frataxin 所需的一般特征和蛋白质特异性特征。我们将利用该项目初始阶段开发的信息和试剂来设计一组适合纠正 FRDA 小鼠模型缺陷的小鼠 mtDNA 基因组。我们将通过将这些基因组包装在线粒体中并将其注射到 FRDA 敲除小鼠的单细胞胚胎中来评估这些基因组补偿小鼠核 frataxin 基因缺失的效率。由于这种基因敲除突变会导致线粒体功能丧失，因此是胚胎致死性的，因此我们可以通过线粒体 frataxin 基因部分拯救（即产生可行的胚胎细胞）或完全拯救这种表型（即产生可存活的胚胎细胞）的能力来轻松测定线粒体 frataxin 基因的功能。，产生可存活的小鼠）。因此，该实验系统将使我们能够优化线粒体转移技术和线粒体 frataxin 基因构建体。一旦我们完成了本申请中描述的工作，我们将处于有利地位，为 FRDA 小鼠模型开发基因治疗方法，并努力使这些疗法适应人类 FRDA 和其他线粒体疾病的治疗。公共卫生相关性：多种人类遗传性疾病是由线粒体 DNA (mtDNA) 基因组和核基因组中编码的线粒体基因突变引起的，但目前还没有针对这些临床破坏性疾病的有效疗法。该提案中描述的实验将使我们对这些疾病之一（FRDA）有更深入的分子了解，并使我们能够开发 mtDNA 工程和转移技术，这些技术将成为开发治疗这些疾病的疗法不可或缺的工具。