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TAT-Mediated Delivery of Frataxin for Friedreichs Ataxia

TAT 介导的 Frataxin 递送治疗弗里德赖希共济失调

基本信息

批准号：
7230193
负责人：
Ronald Mark Payne
金额：
$ 15.91万
依托单位：
INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
依托单位国家：
美国
项目类别：
财政年份：
2006
资助国家：
美国
起止时间：
2006-05-02 至 2009-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7230193
关键词：
9q13 Affect Animal Model Animals Ataxia Biology Brain Cardiomyopathies Cell membrane Cells Cessation of life Characteristics Chimeric Proteins Chromosomes Cleaved cell Clinical Control Animal Cultured Cells Data Defect Degenerative Disorder Development Disease Dose Evaluation Exhibits Fetal Tissues Fibroblasts Frequencies Friedreich Ataxia Funding Mechanisms Genes Genetic Transcription Genome Goals Green Fluorescent Proteins HIV HIV-1 Hand Heart Human Hypertrophic Cardiomyopathy Introns Iron Iron-Binding Proteins Knockout Mice Left Localized Malate Dehydrogenase Mediating Membrane Potentials Messenger RNA Methods Mitochondria Mitochondrial Diseases Mitochondrial Matrix Mitochondrial Proteins Mus Neurologic Nuclear Numbers Oxidants Patients Peptides Phenotype Placenta Postdoctoral Fellow Process Production Proteins Reaction Reporting Research Ethics Committees Research Personnel Research Project Grants Skeletal Muscle Stress System Technology Tertiary Protein Structure Testing Therapeutic Tissues Trans-Activators Transcription Process Transgenic Organisms Translating Translational Research Trinucleotide Repeats Triplet Multiple Birth United States National Institutes of Health Viral Viral Vector Yeasts antioxidant therapy base cellular transduction concept cost frataxin gene therapy human TYRP1 protein intraperitoneal iron chelation therapy mitochondrial membrane motor control multidisciplinary new technology novel pre-clinical programs research study therapy development

项目摘要

DESCRIPTION (provided by applicant): The goal of this translational research project is the preclinical development of a novel therapy for Friedreich's Ataxia (FA). FA is the most common human ataxia and results from inadequate production of the Frataxin protein most often due to a triplet expansion in the nuclear FRDA gene. The gene cannot be translated. Frataxin is an iron-binding protein targeted to the mitochondrial matrix. In its absence, iron accumulation in mitochondria causes oxidant stress destroying mitochondrial and nuclear function. Progressive ataxia and cardiomyopathy are prominent clinical findings with early death from hypertrophic cardiomyopathy. Experiments in cell culture have shown that replacement of the FRDA gene can restore mitochondrial function indicating that if Frataxin can be produced, the cell will be rescued. To overcome the limitations of viral vectors for delivering gene products to mitochondria inside of cells, we will use protein transduction domains (PTD) that cross cell membranes. We have recently shown that the Transactivator of Transcription (TAT) peptide from the human immunodeficiency virus can deliver proteins to mitochondria. We have further developed methods to localize these proteins to mitochondria by including a mitochondrial targeting sequence (MTS) in the fusion protein construct. Our preliminary data shows that a TAT-Frataxin fusion protein crosses both cell and mitochondrial membranes and is localized because the MTS is recognized and cleaved leaving the fusion protein trapped in the mitochondria. This 2 year project will: 1) Develop and test TAT-Frataxin fusion proteins using both cells in culture, and normal mice, for correct localization, processing, and assembly. The most efficient fusion construct for the Frataxin protein will be determined (year 1). 2) Show that TAT-Frataxin can rescue the phenotype of fibroblast cells in culture from FRDA patients (year 1). 3) Show that TAT-Frataxin can rescue the phenotype of an animal model transgenic for conditional loss of the FRDA gene in heart and brain (year 2). These mice are now in hand. The long term goal of this project is the development of a novel technology supporting therapy development for FA. The identification of this candidate therapeutic will allow preclinical evaluation of the strategy in an animal model of FA and will be used to establish a larger multidisciplinary team approach for treatment of this disease through use of the U01, or other, funding mechanism (PAR-02-139).

描述（由申请人提供）：这个转化研究项目的目标是一种治疗弗里德赖希共济失调（FA）的新疗法的临床前开发。FA是最常见的人类共济失调，是由Frataxin蛋白产生不足引起的，最常见的原因是核FRDA基因的三重扩增。基因不能被翻译。Frataxin是一种针对线粒体基质的铁结合蛋白。在缺乏铁的情况下，线粒体中的铁积累会引起氧化应激，破坏线粒体和细胞核功能。进行性共济失调和心肌病是肥厚性心肌病早期死亡的突出临床表现。细胞培养实验表明，替换FRDA基因可以恢复线粒体功能，这表明如果Frataxin能够产生，细胞将得到拯救。为了克服病毒载体将基因产物传递到细胞内线粒体的局限性，我们将使用跨细胞膜的蛋白质转导结构域（PTD）。我们最近发现，来自人类免疫缺陷病毒的转录反激活因子（TAT）肽可以将蛋白质传递到线粒体。通过在融合蛋白构建中加入线粒体靶向序列（MTS），我们进一步开发了将这些蛋白定位到线粒体的方法。我们的初步数据表明，TAT-Frataxin融合蛋白穿过细胞和线粒体膜，并被定位，因为MTS被识别和切割，使融合蛋白被困在线粒体中。这个为期2年的项目将：1)使用培养细胞和正常小鼠开发和测试TAT-Frataxin融合蛋白，以正确定位，加工和组装。将确定最有效的Frataxin蛋白融合构建体（第1年）。2)证明TAT-Frataxin可以挽救FRDA患者培养的成纤维细胞的表型（第1年）。3)证明TAT-Frataxin可以挽救心脏和大脑中FRDA基因条件缺失转基因动物模型的表型（第2年）。这些老鼠现在已经到手了。该项目的长期目标是开发一种支持FA治疗发展的新技术。这种候选治疗方法的确定将允许在FA动物模型中对该策略进行临床前评估，并将用于通过使用U01或其他资助机制（PAR-02-139）建立更大的多学科团队治疗该疾病的方法。

项目成果

期刊论文数量（3）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Cardiomyopathy of Friedreich's ataxia: use of mouse models to understand human disease and guide therapeutic development.

DOI：
10.1007/s00246-011-9943-6
发表时间：
2011-03
期刊：
PEDIATRIC CARDIOLOGY
影响因子：
1.6
作者：
Payne, R. Mark;Pride, P. Melanie;Babbey, Clifford M.
通讯作者：
Babbey, Clifford M.

TAT fusion protein transduction into isolated mitochondria is accelerated by sodium channel inhibitors.

DOI：
10.1021/bi101057v
发表时间：
2010-11-09
期刊：
BIOCHEMISTRY
影响因子：
2.9
作者：
Rayapureddi, Jayanagendra P.;Tomamichel, Wendy J.;Walton, Sonia T.;Payne, R. Mark
通讯作者：
Payne, R. Mark

The Heart in Friedreich's Ataxia: Basic Findings and Clinical Implications.