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Recombinant Enzyme Fusion Protein for Lysosomal Storage Disorders

用于治疗溶酶体贮积症的重组酶融合蛋白

基本信息

批准号：
7413300
负责人：
YUN ZHANG
金额：
$ 45.66万
依托单位：
ARMAGEN TECHNOLOGIES, INC.
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-03-01 至 2010-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7413300
关键词：
Adult Affect Affinity Affinity Chromatography Animal Model Anions Antibodies Beta-glucuronidase Binding Biological Assay Bioreactors Blood - brain barrier anatomy Blood Vessels Blood capillaries Brain Brain Part Bypass COS Cells Canis familiaris Carbohydrates Cations Cells Cephalic Cessation of life Child Chimeric Proteins Chinese Hamster Ovary Cell Chromatography Clinical Trials Cloning Complementary DNA Condition Conditioned Culture Media Contracts DNA Restriction Enzymes DNA Sequence Analysis Dermatan Sulfate Development Dihydrofolate Reductase Disease Dose Drug Kinetics Electroporation Engineering Enzyme-Linked Immunosorbent Assay Enzymes Equus caballus Filtration Future Genes Genetic Engineering Glucuronides Goals Grant Guanosine Monophosphate Heparitin Sulfate Hereditary Disease Histamine Human Hydrolysis Immunoglobulin G Inborn Errors of Metabolism Injection of therapeutic agent Institution Insulin Receptor Isoelectric Focusing Laboratories Legal patent Light Liver Macaca mulatta Mass Spectrum Analysis Measures Mediating Methods Methotrexate Modeling Molecular Sieve Chromatography Molecular Weight Monoclonal Antibodies Mucopolysaccharidosis VII Mus Mutate Neuraxis Organ Orphan Drugs Outsourcing Patients Pharmaceutical Preparations Pharmacologic Substance Pharmacology and Toxicology Phase Phase II Clinical Trials Phase III Clinical Trials Plasmids Polyacrylamide Gel Electrophoresis Polymerase Chain Reaction Preparation Primates Process Production Protein Engineering Proteins Radiolabeled Recombinant Antibody Recombinant Fusion Proteins Recombinant Proteins Recombinants Research Rodent Running Serum Small Business Funding Mechanisms Small Business Innovation Research Grant Sodium Dodecyl Sulfate Solutions Spinal Cord Staging Structure System Technology Testing Therapeutic Therapeutic Effect Transfection United States Food and Drug Administration Western Blotting Work abstracting brain cell capillary cell bank chimeric antibody commercialization dalton design enzyme activity enzyme replacement therapy expression vector fusion gene glucuronide human INSR protein in vivo intravenous administration intravenous injection lysosomal glycosyl hydrolase milligram molecular trojan horse novel novel strategies peptide permease preclinical study progressive neurodegeneration prototype radiotracer receptor receptor binding size small molecule therapeutic enzyme transcytosis uptake vector

项目摘要

DESCRIPTION (provided by applicant): Zhang, Yun Abstract There are over 40 lysosomal storage disorders, and most of these diseases affect adversely the central nervous system (CNS). The mainstay of treatment is enzyme replacement therapy (ERT). However, ERT is not effective for the brain, because the enzymes do not cross the brain capillary wall, which forms the blood-brain barrier (BBB) in vivo. Without treatment of the CNS, the young patients are destined to progressive neurodegeneration and death. The limiting factor in the future treatment of these diseases is the transport of the enzyme across the BBB. Bypass of the BBB with direct injection into the brain is not effective, because only a small part of the brain is treated with a trans-cranial delivery system. Conversely, virtually all cells of the brain can be treated with a trans-vascular delivery system that enables the enzyme to cross the BBB following intravenous administration. A new approach to the BBB delivery of large molecules such as enzymes is the molecular Trojan horse technology. A bi-functional fusion protein is produced with genetic engineering, wherein the missing recombinant enzyme is fused to a BBB molecular Trojan horse. The latter is a genetically engineered protein that is able to cross the human BBB by receptor- mediated transcytosis on endogenous BBB peptide transport systems. Pre-clinical studies show that a large enzyme with a molecular weight >100,000 Daltons, can be delivered to brain via transport across the BBB, following attachment to a BBB receptor-specific Trojan horse. The present work will produce a novel fusion gene encoding a lysosomal enzyme and a genetically engineered molecular Trojan horse, which will allow the production of the corresponding fusion protein, AGT-180. The fusion gene will be incorporated in a eukaryotic expression vector followed by permanent transfection of cells. These phase II studies will enable production of a master cell bank and development of the purification and downstream processing of the fusion protein. Fusion proteins comprised of BBB targeting antibodies and recombinant enzymes could be therapeutic in the treatment of the brain in human lysosomal storage disorders. 1 Zhang, Yun Project Narrative Lysosomal storage disorders are serious inborn errors of metabolism, and about 75% of the ~40 lysosomal storage disorders affect the brain. The mainstay of treatment is Enzyme Replacement Therapy (ERT). However, ERT is ineffective in the brain, because the enzymes do not cross the blood-brain barrier (BBB). The present work will produce a novel recombinant fusion protein that is able to both (a) bind a human BBB receptor to trigger transport into the brain, and (b) retain high lysosomal enzyme activity. This novel drug, designated AGT-180, will be a model for the treatment of the brain for multiple genetic diseases. 1

描述（申请人提供）：张云摘要溶酶体贮积症有 40 多种，其中大多数疾病对中枢神经系统（CNS）产生不利影响。主要治疗方法是酶替代疗法（ERT）。然而，ERT 对大脑无效，因为酶不能穿过脑毛细血管壁，而脑毛细血管壁在体内形成血脑屏障 (BBB)。如果不治疗中枢神经系统，年轻患者注定会出现进行性神经退行性病变和死亡。未来治疗这些疾病的限制因素是酶穿过血脑屏障的运输。绕过血脑屏障直接注射到大脑中的方法并不有效，因为只有一小部分大脑接受了经颅给药系统的治疗。相反，几乎所有大脑细胞都可以通过跨血管递送系统进行治疗，该系统使酶能够在静脉注射后穿过血脑屏障。一种通过血脑屏障传递酶等大分子的新方法是分子特洛伊木马技术。通过基因工程产生双功能融合蛋白，其中缺失的重组酶与BBB分子特洛伊木马融合。后者是一种基因工程蛋白，能够通过内源性 BBB 肽转运系统上受体介导的转胞吞作用穿过人类 BBB。临床前研究表明，分子量 >100,000 道尔顿的大酶在附着于 BBB 受体特异性特洛伊木马后，可以通过 BBB 运输输送到大脑。目前的工作将产生一种编码溶酶体酶的新型融合基因和一个基因工程分子特洛伊木马，这将允许产生相应的融合蛋白AGT-180。融合基因将被整合到真核表达载体中，然后永久转染细胞。这些第二阶段研究将能够生产主细胞库并开发融合蛋白的纯化和下游加工。由 BBB 靶向抗体和重组酶组成的融合蛋白可用于治疗人类溶酶体贮积症的大脑。 1 张云项目叙述溶酶体贮积症是严重的先天性代谢错误，约 40 种溶酶体贮积症中约 75% 影响大脑。主要治疗方法是酶替代疗法（ERT）。然而，ERT 在大脑中无效，因为酶不能穿过血脑屏障 (BBB)。目前的工作将产生一种新型重组融合蛋白，它能够（a）结合人类 BBB 受体以触发转运到大脑中，并且（b）保留高溶酶体酶活性。这种新药被命名为 AGT-180，将成为治疗大脑多种遗传性疾病的模型。 1