Gene Transfer and NMR Studies in Alpha-Mannosidosis Brain

α-甘露糖苷沉积症脑中的基因转移和核磁共振研究

• 批准号: 8068082
• 负责人: JOHN H WOLFE
• 金额: $ 9.96万
• 依托单位: CHILDREN'S HOSP OF PHILADELPHIA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-06-01 至 2010-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8068082
• 关键词: Affect; Aftercare; Age; Animals; Area; Biological Assay; Brain; Brain Diseases; Cells; Cellular Morphology; Child; Clinical; Clinical Trials; Complementary DNA; Development; Diffusion weighted imaging; Disease; Distal; Early treatment; Enzymes; Felis catus; Gene Delivery; Gene Expression; Gene Proteins; Gene Transfer; Genes; Grant; Gray unit of radiation dose; Histology; Human; Image; Inborn Genetic Diseases; Individual; Inherited; Injection of therapeutic agent; Lesion; Life; Ligands; Longitudinal Studies; Lysosomal Storage Diseases; Magnetic Resonance; Magnetic Resonance Imaging; Maps; Measures; Mediating; Mental Retardation; Messenger RNA; Methods; Modality; Modeling; Molecular Biology; Monitor; Mutation; Nature; Neurologic; Neurology; Normal Statistical Distribution; Nuclear Magnetic Resonance; Operative Surgical Procedures; Pathology; Pathway interactions; Physiological; Positron-Emission Tomography; Protons; Reporter Genes; Resolution; Rodent; Route; Serotyping; Site; Spectrum Analysis; Syndrome; Testing; Time; Tissues; Treatment Efficacy; Work; alpha Mannosidase B; alpha-Mannosidosis; brain cell; brain volume; cell type; clinically relevant; enzyme deficiency; gene therapy; gray matter; improved; neurogenetics; neurophysiology; public health relevance; research study; restoration; therapeutic gene; vector; white matter

DESCRIPTION (provided by applicant): Lysosomal storage diseases (LSDs) are a large group of inherited enzyme deficiencies that produce fatal degenerative syndromes in children, with most having severe brain disease manifest as mental retardation. Neurogenetic diseases have lesions throughout the brain, thus it is thought that global correction will be needed. Vector mediated transfer of a normal cDNA of the gene can correct defective cells, but current vectors mediate relatively limited gene delivery in the brain. Nevertheless, CNS gene therapy may work in most LSDs because the enzyme is secreted from genetically corrected cells and taken up by neighboring cells; and the enzyme may be transported via axonal pathways to distal sites. To study approaches to gene therapy in a brain that is significantly larger than a rodent brain, we will use a cat model of human alpha- mannosidosis (AMD), caused by a mutation in the lysosomal alpha-mannosidase (LAMAN) gene. In the current grant period we found: 1) An AAV1 serotype vector increased the amount of total gene transfer and it expanded transduction to both white and gray matter areas. 2) A single surgical procedure injecting the vector into several tracks spaced around the brain produced enough normal LAMAN to reverse large areas of storage lesions. Clinically, the treated AMD cats showed promising improvements in the neurological syndrome, but not complete resolution. 3) Several magnetic resonance (MR) modalities were evaluated to non-invasively monitor disease pathology in the living animal brain. Magnetization transfer imaging (MTI) showed that significant improvements in the white matter could be measured in AMD cat brains treated by gene therapy. Diffusion-weighted imaging (Dw-MRI) and spectroscopy (1H-MRS) were able to detect disease in the gray matter areas, where most of the storage is seen. 4) Pathology studies demonstrated significant restoration of normal brain cell morphology in treated cats, in both gray and white matter, but some regions of uncorrected disease remained. To improve on these advances, we propose in the continuation grant to: 1) Evaluate a method to monitor vector gene expression in the living brain by PET. 2) Investigate alternate vector delivery approaches to achieve more complete correction of the CNS. 3) Study the ability of Dw-MRI and 1H-MRS to measure changes in the gray matter after treatment, using a 3T clinical magnet. 4) Study long-term correction (>1 year), the effects of starting treatment earlier in life, and evaluate functional correlates of disease correction in neuro-physiological studies. Together, these experiments in a mammalian brain that is significantly larger than a rodent brain provide a robust translational model to study the potential of AAV-mediated gene therapy to correct the global brain lesions in neurogenetic LSDs. The translational nature of these studies involves a team of experts from several fields, including neurology, MRI and PET imaging, gene vector development, molecular biology, neurophysiology, and pathology. PUBLIC HEALTH RELEVANCE: The lysosomal storage diseases are a large group of inherited disorders that have severe disease in the brain. We are studying gene therapy methods to correct the brain disease. The proposed studies are directed towards improving gene and protein delivery sufficiently to make them feasible for human clinical trials.

描述（由申请人提供）：溶酶体贮积病（LSD）是一大类遗传性酶缺乏症，可在儿童中产生致命的退行性综合征，其中大多数患有严重的脑部疾病，表现为智力迟钝。神经遗传性疾病在整个大脑中都有病变，因此认为需要进行整体纠正。载体介导的基因的正常cDNA的转移可以纠正缺陷细胞，但是目前的载体介导的脑中的基因递送相对有限。然而，CNS基因治疗可能在大多数LSD中起作用，因为该酶从遗传校正的细胞分泌并被邻近细胞吸收;并且该酶可以通过轴突途径转运到远端部位。为了研究在显著大于啮齿动物脑的脑中进行基因治疗的方法，我们将使用由溶酶体α-甘露糖苷酶（LAMAN）基因突变引起的人α-甘露糖苷沉积症（AMD）的猫模型。在本研究期间，我们发现：1）AAV 1血清型载体增加了总基因转移量，并将转导扩展到白色和灰质区域。2)一个单一的外科手术将载体注射到大脑周围的几个轨道上，产生了足够的正常LAMAN来逆转大面积的储存病变。临床上，治疗的AMD猫显示出神经系统综合征的有希望的改善，但没有完全解决。3)几个磁共振（MR）模式进行了评估，以非侵入性监测疾病病理在活体动物大脑。磁化传递成像（MTI）显示，在基因治疗的AMD猫脑中，可以测量到白色物质的显着改善。弥散加权成像（DW-MRI）和波谱（1H-MRS）能够检测到灰质区域的疾病，其中大部分存储被发现。4)病理学研究表明，给药猫的正常脑细胞形态（灰质和白色物质）显著恢复，但仍存在一些未纠正的疾病区域。为了改进这些进展，我们在继续资助中提出：1）评估一种通过PET监测活脑中载体基因表达的方法。2)研究替代载体递送方法以实现CNS的更完全矫正。3)使用3 T临床磁体研究DW-MRI和1H-MRS测量治疗后灰质变化的能力。4)研究长期矫正（>1年），在生命早期开始治疗的影响，并在神经生理学研究中评估疾病矫正的功能相关性。总之，这些实验中的哺乳动物的大脑是显着大于啮齿动物的大脑提供了一个强大的翻译模型，研究的潜力，AAV介导的基因治疗，以纠正全球脑病变的神经遗传性LSD。这些研究的转化性质涉及来自多个领域的专家团队，包括神经学，MRI和PET成像，基因载体开发，分子生物学，神经生理学和病理学。 公共卫生相关性：溶酶体贮积病是一大组遗传性疾病，在大脑中有严重的疾病。我们正在研究基因治疗方法来纠正脑部疾病。拟议的研究旨在充分改善基因和蛋白质的递送，使其适用于人体临床试验。