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，大多数患有严重脑部疾病的患者表现为智力低下。神经遗传疾病的病变遍及整个大脑，因此认为需要进行整体校正。载体介导的正常基因 cDNA 转移可以纠正有缺陷的细胞，但目前的载体在大脑中介导的基因传递相对有限。尽管如此，中枢神经系统基因疗法可能对大多数 LSD 有效，因为这种酶是由经过基因校正的细胞分泌并被邻近细胞吸收的。并且酶可以通过轴突途径转运至远端部位。为了研究比啮齿动物大脑大得多的大脑中的基因治疗方法，我们将使用人类α-甘露糖苷中毒（AMD）的猫模型，该模型是由溶酶体α-甘露糖苷酶（LAMAN）基因突变引起的。在当前资助期内，我们发现：1) AAV1 血清型载体增加了总基因转移量，并将转导范围扩大到白质和灰质区域。 2）通过一次外科手术将载体注射到大脑周围间隔的几个轨道中，产生了足够的正常 LAMAN 来逆转大面积的存储损伤。临床上，接受治疗的 AMD 猫的神经系统综合症显示出有希望的改善，但并未完全解决。 3) 对几种磁共振（MR）模式进行了评估，以非侵入性地监测活体动物大脑中的疾病病理学。磁化转移成像 (MTI) 表明，经过基因治疗的 AMD 猫大脑中白质的显着改善。弥散加权成像 (Dw-MRI) 和光谱学 (1H-MRS) 能够检测灰质区域的疾病，大部分存储都在灰质区域。 4) 病理学研究表明，接受治疗的猫的灰质和白质正常脑细胞形态显着恢复，但一些未纠正的疾病区域仍然存在。为了改进这些进展，我们在继续拨款中建议：1）评估一种通过 PET 监测活体大脑中载体基因表达的方法。 2) 研究替代载体传递方法以实现更完整的中枢神经系统校正。 3) 使用 3T 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）、早期开始治疗的效果，并评估神经生理学研究中疾病矫正的功能相关性。总之，这些在比啮齿动物大脑大得多的哺乳动物大脑中进行的实验提供了一个强大的转化模型，用于研究 AAV 介导的基因治疗纠正神经遗传性 LSD 中的整体大脑损伤的潜力。这些研究的转化性质涉及来自多个领域的专家团队，包括神经病学、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类遗传性疾病，会导致严重的脑部疾病。我们正在研究纠正脑部疾病的基因治疗方法。拟议的研究旨在充分改善基因和蛋白质的传递，使其可用于人体临床试验。