Identifying targeting antibodies in an in vitro model of the human blood-brain barrier during ischemic stroke

在缺血性中风期间人血脑屏障的体外模型中识别靶向抗体

• 批准号: 10796765
• 负责人: Moriah Katt
• 金额: $ 15.64万
• 依托单位: WEST VIRGINIA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-08 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10796765
• 关键词: 3-Dimensional; ABCB1 gene; Acclimatization; Acute; Address; Alteplase; Analysis of Variance; Animal Model; Animals; Antibodies; Apical; Apoptosis; Architecture; Area; Astrocytes; Autopsy; Basal lamina; Benchmarking; Binding; Biodistribution; Biological Assay; Blood; Blood - brain barrier anatomy; Blood brain barrier dysfunction; Brain; Brain Ischemia; Brain region; Brain-Derived Neurotrophic Factor; CASP3 gene; CRISPR interference; Cadaver; Carbon Dioxide; Cell Line; Cell Surface Proteins; Cell surface; Cells; Cellular Morphology; Centers of Research Excellence; Central Nervous System Diseases; Cerebrovascular system; Cessation of life; Chimeric Proteins; Chinese Hamster Ovary Cell; Chronic; Chronic Phase; Clinic; Clinical; Clinical Trials; Collagen; Collagen Type IV; Communities; Confidence Intervals; Contralateral; Control Groups; Coupled; Data; Data Analyses; Detection; Development; Devices; Dextrans; Diameter; Disease; Disease Progression; Dose; Drug Delivery Systems; Drug Transport; Electrical Resistance; Endothelial Cells; Endothelium; Environment; Enzyme-Linked Immunosorbent Assay; Excision; Exposure to; Extracellular Matrix; FDA approved; Failure; Fc Immunoglobulins; Fibronectins; Flow Cytometry; Gel; Generations; Glass; Glial Fibrillary Acidic Protein; Glucose; Growth; Growth Factor; Guide RNA; Health; Hour; Housing; Human; Hyaluronic Acid; Hydrogels; Hypoxia; Immunoglobulin G; Immunoprecipitation; In Situ Nick-End Labeling; In Vitro; Incubated; Individual; Infarction; Injections; Injury; Insulin Receptor; Intravenous; Ipsilateral; Ischemia; Ischemic Stroke; Label; Laminin; Lead; Length; Leukocytes; Libraries; Ligands; Liposomes; Lymphocyte; Maintenance; Mass Spectrum Analysis; Measurement; Measures; Mediating; Methods; Microfluidic Microchips; Microscopy; Modeling; Monitor; Morphology; Motor; Mus; Mutagens; Names; Nature; Neurites; Neuroglia; Neurons; Neuroprotective Agents; Nutrient; Nutrient availability; Organ; Outcome; Oxygen; PC12 Cells; Patients; Penetration; Perfusion; Permeability; Persons; Phage Display; Pharmaceutical Preparations; Phenotype; Phosphorylation; Physiological; Plasmids; Production; Proteins; Pump; Quality of life; Recovery; Rehabilitation therapy; Reperfusion Therapy; Reporting; Resected; Rhodamine; Rhodamine 123; Risk; Rod; Rodent; Running; Scheme; Slide; Source; Specificity; Stains; Stroke; Supporting Cell; Surface; System; TFRC gene; Tariquidar; Techniques; Testing; Therapeutic; Tight Junctions; Time; Tissue Sample; Tissues; Tracer; Transfection; Translating; Traumatic Brain Injury; Tubulin; United States National Institutes of Health; Universities; Validation; Vascular Endothelium; Visualization; West Virginia; Western Blotting; Work; Yeasts; antibody libraries; antibody test; aquaporin 4; blood-brain barrier crossing; blood-brain barrier function; candidate identification; cell immortalization; cell motility; cell type; chronic stroke; cross reactivity; density; deprivation; design; design and construction; disability; disulfide bond; efficacy testing; experimental study; fabrication; field study; human disease; human model; human tissue; improved; in vitro Model; induced pluripotent stem cell; inflammatory marker; insight; lead candidate; live cell microscopy; lucifer yellow; monolayer; mouse model; neuroinflammation; neuronal growth; neuroprotection; non-invasive imaging; normoxia; occludin; overexpression; placebo group; polydimethylsiloxane; power analysis; preclinical study; prevent; protein folding; receptor; recruit; response; screening; shear stress; side effect; small molecule therapeutics; stem cell model; stem cells; stroke model; stroke recovery; stroke therapy; success; symptom management; targeted treatment; trafficking; transcytosis; translational therapeutics; transport inhibitor; uptake; vector

Stroke is one of the leading causes of disability and death in the US. One of the contributing factors to this is the lack of FDA approved treatments for chronic stroke. Aside from tissue plasminogen activator (tPA), there are no other approved therapeutics with disease modifying effects; current therapy is entirely focused on symptom management and rehabilitation. This is in part due to the lack of understanding of the underlying mechanisms involved, as well as the difficulty of delivering therapeutics through the blood-brain barrier (BBB) and into the brain. This project aims to address both of these difficulties through the development of a human induced pluripotent stem cell (hiPSC) derived model of the BBB incorporating hiPSC derived brain microvascular endothelial like cells (BMECs), astrocytes, and neurons. Using a 3D microvessel platform ischemia will be modeled using perturbations in shear flow, oxygen concentration, and nutrient availability. The resultant phenotype of the cells will be investigated using live cell microscopy to measure permeability of fluorescent tracer molecules, as well as morphological changes in the cells. Using this model, as well as a simplified 2D model of ischemia, a library of antibodies will be screened for improved targeting and delivery to the human ischemic brain. Finally, the antibodies identified in the screen will be tested for efficacy in delivering known non-brain penetrant neuroprotectant growth factors to the ischemic brain both in the in vitro model and in a mouse model of stroke. In summary, this project will establish a human specific model of the BBB during ischemic stroke and identify antibodies that are able to specifically target and deliver therapeutics to the ischemic region of the brain.

中风是美国残疾和死亡的主要原因之一。其中一个促成因素是 缺乏FDA批准的慢性中风治疗方法。除了组织纤溶酶原激活物（tPA）， 没有其他批准的具有疾病改善作用的治疗方法;目前的治疗完全集中在 症状管理和康复这部分是由于缺乏对 涉及的潜在机制，以及通过血脑递送治疗剂的困难， 血脑屏障（BBB），进入脑内。本项目旨在通过以下方式解决这两个难题： 人诱导多能干细胞（hiPSC）衍生的BBB掺入模型的开发 hiPSC衍生的脑微血管内皮样细胞（BMEC）、星形胶质细胞和神经元。使用3D 微血管平台缺血将使用剪切流，氧浓度， 和养分供应。将使用活细胞显微镜研究细胞的所得表型 以测量荧光示踪分子的渗透性以及细胞中的形态变化。 使用该模型以及简化的缺血2D模型，将筛选抗体库， 改善了对人缺血性脑的靶向和递送。最后，筛选出的抗体 将测试在将已知的非脑渗透性神经保护生长因子递送至脑组织中的功效。 在体外模型和中风的小鼠模型中均观察到缺血性脑。总之，该项目将 建立缺血性中风期间BBB的人类特异性模型，并鉴定能够 特异性地靶向并递送治疗剂至脑的缺血区域。