Towards gene therapy of mitochondrial disease

点击翻译按钮获取中文摘要

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.