Advancing Gene Therapy for Late Infantile Neuronal Ceroid Lipofuscinosis

推进晚期婴儿神经元蜡质脂褐质沉着症的基因治疗

• 批准号: 8382982
• 负责人: Beverly L. Davidson
• 金额: $ 22.65万
• 依托单位: UNIVERSITY OF IOWA
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-07-01 至 2014-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8382982
• 关键词: 15 year old; 2 year old; Affinity; Affinity Chromatography; Antibodies; Bacteriophages; Behavior; Binding; Biological Assay; Blood - brain barrier anatomy; Blood Vessels; Brain; Bypass; CLN2 gene; Capsid; Cell Surface Receptors; Cell membrane; Cell surface; Central Nervous System Diseases; Cessation of life; Child; Childhood; Complex; Cultured Cells; Defect; Dependovirus; Detection; Disease; Endothelial Cells; Endothelium; Engineering; Enzymes; Epilepsy; Functional disorder; Gene Delivery; Glycoproteins; Goals; Green Fluorescent Proteins; Home environment; Homing; Human; Hydrolase; Impaired cognition; In Vitro; Infantile neuronal ceroid lipofuscinosis; Label; Liquid Chromatography; Lysosomal Storage Diseases; Mass Spectrum Analysis; Measures; Mediating; Membrane Proteins; Methods; Molecular; Motor; Mus; Mutation; Nerve Degeneration; Neuraxis; Neurodegenerative Disorders; Neuropil; Pathology; Patients; Peptide Receptor; Peptides; Phage Display; Phage Receptors; Polysaccharides; Protein Binding; Proteins; Proteoglycan; Recombinants; Relative (related person); Reporter; Research; Rodent; Rodent Model; Specificity; Structure; Surface; Technology; Testing; Therapeutic; Therapeutic Agents; Translations; Travel; Tropism; United States National Institutes of Health; Vascular Endothelial Cell; Vascular Endothelium; Viral; Virus; Virus Receptors; Vision; Work; base; disease-causing mutation; effective therapy; experience; gene therapy; improved; in vivo; knock-down; mind control; mouse model; neuropathology; novel; novel strategies; overexpression; prevent; receptor; research study; restoration; tandem mass spectrometry; therapeutic development; transduction efficiency; tripeptidyl-peptidase I; vector; virus tropism

DESCRIPTION (provided by applicant): Late Infantile Neuronal Ceroid Lipofuscinosis (LINCL) is a childhood-onset autosomal recessive neurodegenerative disease. This disorder, a lysosomal storage disease (LSD), is caused by mutations in CLN2, the gene that encodes the lysosomal hydrolase tripeptidyl peptidase I (TPP1). TPP1 deficiency causes epileptic seizures, vision impairment, cognitive and motor dysfunction, and is invariably fatal by late childhood. Currently, there is no treatment or cure for this devastating disease. The major barrier to treatment is the delivery of bioactive TPP1 enzyme to the central nervous system, a goal hindered by the selectivity of the blood-brain-barrier (BBB). In recent studies, our lab has engineered a brain-homing adeno-associated virus (AAV) gene delivery vector. Brain tropism of the virus is conferred by a peptide, GMNAFRA, which we identified by in vivo phage display panning in a mouse mode of LINCL. The GMN peptide has affinity for the luminal surface of brain vascular endothelial cells, which are the primary component of the BBB. When delivered peripherally, this peptide-modified virus (GMN-AAV) travels to, and transduces brain endothelia. Significantly, delivery of GMN-AAV encoding CLN2 in LINCL mice restored TPP1 expression in the brain and corrected neuropathological defects. Thus, this novel gene therapy approach bypasses the BBB by engineering endothelial cells to secrete recombinant enzyme directly into the underlying neuropil. In order to advance this exciting and promising gene therapy approach to LINCL patients, elucidation of the identity of the brain endothelial receptor molecule that interacts with the GMN peptide is required. The overall goal of this proposal is to perform experiments to discover this molecule, which mediates GMN-AAV brain targeting and transduction. Our hypothesis is that the receptor is a plasma membrane-associated protein or glycan moiety expressed on the surface of brain vascular endothelial cells in TPP1 deficient brain. In aim one, we will use affinity purification and quantitative mass spectrometry methods to isolate and identify the GMN-AAV receptor. In aim two, we will use a mammalian glycan array developed by the NIH-operated Consortium for Functional Glycomics to determine whether GMN-AAV binds to specific glycans or classes of glycans that may be present on the surface of brain endothelial cells. Completion of these aims will enable us to advance the translation of this novel gene therapy to children with LINCL. In addition, this therapeutic strategy could be broadly applied to other LSDs that cause central nervous system dysfunction and neurodegeneration. PUBLIC HEALTH RELEVANCE: The proposed research will identify the cell-surface receptor that mediates binding and transduction of brain vascular endothelia by an AAV gene delivery vector. This vector can restore TPP1 enzyme expression and prevent neuropathology in a rodent model of Late Infantile Neuronal Ceroid Lipofuscinosis (LINCL), a currently untreatable fatal neurodegenerative disorder. Results of these studies are anticipated to advance this highly promising gene therapy to treatment of LINCL patients.

描述（由申请人提供）：晚期婴儿神经元蜡质脂褐质沉着症（LINCL）是一种儿童期发病的常染色体隐性神经退行性疾病。这种疾病是一种溶酶体贮积病 (LSD)，是由 CLN2 突变引起的，CLN2 是编码溶酶体水解酶三肽基肽酶 I (TPP1) 的基因。 TPP1 缺乏会导致癫痫发作、视力障碍、认知和运动功能障碍，并且在儿童晚期总是致命的。目前，这种毁灭性疾病尚无治疗方法或治愈方法。治疗的主要障碍是将生物活性 TPP1 酶输送到中枢神经系统，而血脑屏障 (BBB) 的选择性阻碍了这一目标的实现。在最近的研究中，我们的实验室设计了一种脑归巢腺相关病毒（AAV）基因传递载体。该病毒的脑向性是由一种肽 GMNAFRA 赋予的，我们通过在 LINCL 小鼠模式中进行体内噬菌体展示淘选来鉴定该肽。 GMN 肽对脑血管内皮细胞的管腔表面具有亲和力，而脑血管内皮细胞是 BBB 的主要成分。当外周递送时，这种肽修饰病毒 (GMN-AAV) 传播并转导脑内皮细胞。值得注意的是，在 LINCL 小鼠体内注射编码 CLN2 的 GMN-AAV 可恢复大脑中 TPP1 的表达并纠正神经病理学缺陷。因此，这种新颖的基因治疗方法通过改造内皮细胞将重组酶直接分泌到下面的神经纤维中来绕过血脑屏障。为了向 LINCL 患者推进这种令人兴奋且有前途的基因治疗方法，需要阐明与 GMN 肽相互作用的脑内皮受体分子的身份。该提案的总体目标是进行实验来发现这种介导 GMN-AAV 大脑靶向和转导的分子。我们的假设是，该受体是 TPP1 缺陷大脑中脑血管内皮细胞表面表达的质膜相关蛋白或聚糖部分。在目标一中，我们将使用亲和纯化和定量质谱方法来分离和鉴定 GMN-AAV 受体。在目标二中，我们将使用由 NIH 运营的功能糖组学联盟开发的哺乳动物聚糖阵列来确定 GMN-AAV 是否​​与可能存在于脑内皮细胞表面的特定聚糖或一类聚糖结合。完成这些目标将使我们能够推进本文件的翻译 针对 LINCL 儿童的新型基因疗法。此外，这种治疗策略可以广泛应用于其他导致中枢神经系统功能障碍和神经退行性变的LSD。 公共健康相关性：拟议的研究将鉴定通过 AAV 基因传递载体介导脑血管内皮结合和转导的细胞表面受体。该载体可以恢复 TPP1 酶的表达并预防晚期婴儿神经元蜡质脂褐质沉积症 (LINCL) 啮齿动物模型中的神经病理学，LINCL 是一种目前无法治疗的致命性神经退行性疾病。这些研究的结果预计将推动这种极具前景的基因疗法治疗 LINCL 患者。