Monoclonal Antibody Drug Development for Alzheimer?s Disease

阿尔茨海默病单克隆抗体药物开发

基本信息

批准号：
7498752
负责人：
William M Pardridge
金额：
$ 31.45万
依托单位：
UNIVERSITY OF CALIFORNIA LOS ANGELES
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-08-15 至 2011-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7498752
关键词：
A Mouse Affinity Affinity Chromatography Age-Months Alzheimer&apos s Disease Alzheimer&apos s disease model Amino Acids Amyloid Animals Antibodies Binding Binding Sites Biochemical Biological Assay Bioreactors Blood Blood - brain barrier anatomy Blood capillaries Brain Brain hemorrhage Breeding COS Cells Cations Cerebrum Chimeric Proteins Chinese Hamster Chinese Hamster Ovary Cell Chromatography Clinical Trials Cloning Complementary DNA Condition DNA Sequence Data Dementia Deposition Depth Disease Disruption Dose Drug Delivery Systems Drug Kinetics Electroporation Engineering Fc Receptor Filtration G-substrate Genes Genetic Engineering Genotype Goals Harvest Hemorrhage Histocytochemistry Human Hybridomas IgG1 Immune Immunoglobulin G Immunoglobulin Variable Region Immunoradiometric Assays Immunotherapy Inbred BALB C Mice Injection of therapeutic agent Insulin Receptor Label Laboratories Light Measurement Mediating Mediation Medicine Methods Monoclonal Antibodies Mus Ovary Peptides Peripheral Peritoneal Pharmaceutical Preparations Pharmacology Plasma Plasmids Polymerase Chain Reaction Proteins Prussian blue Public Health Publishing Purpose RNA Splicing Radio Rattus Reaction Research Rodent Saline Senile Plaques Series Serum-Free Culture Media Staging Structure Testing Therapeutic Transfection Transferrin Receptor Transgenic Mice Transgenic Organisms Validation Western Blotting Work amyloid peptide antibiotic G 418 capillary drug development drug discovery human INSR protein in vivo interest lipofection molecular trojan horse mouse model nervous system disorder neurobehavior novel therapeutics pre-clinical receptor receptor binding uptake variable region gene

项目摘要

DESCRIPTION (provided by applicant): The dementia of Alzheimer's disease (AD) is caused by the deposition of Ab amyloid in brain over many years. Once the amyloid plaque forms in brain, it is permanent in the absence of plaque disaggregation therapy. The most potent form of plaque diasaggregation therapy is a monoclonal antibody (MAb) against the Ab amyloid peptide of AD, and passive immune therapy of AD is currently being tested in clinical trials. However, the anti-Ab MAb does not cross the blood-brain barrier (BBB). Consequently, very high doses of MAb must be administered to lower plaque in brain of AD transgenic mice, and these high doses causes brain microhemorrhage. The present research will genetically engineer a new form of anti-Ab MAb that is enabled to cross the BBB via receptor-mediated transport in both the blood-to-brain and brain-to-blood directions. The PI has previously genetically engineered a chimeric MAb against the mouse transferrin receptor (TfR) that crosses the BBB in the blood-to-brain direction via receptor-mediated transport on the mouse BBB TfR, and also crosses the BBB in the brain-to-blood direction on the BBB Fc receptor (FcR). In addition, the PI has genetically engineered, and expressed a single chain Fv (ScFv) antibody against the amino terminal portion of the Ab peptide. The present research will produce a new fusion protein, wherein the anti-Ab ScFv is fused to the carboxyl terminus of the chimeric MAb against the mouse TfR, and this new fusion protein is designated the TfRMAb-A2ScFv bi-specific antibody (BSA). Following the genetic engineering of the new BSA, the protein will be transiently expressed in COS cells, and the bi-functionality of the BSA will be demonstrated with a mouse TfR binding assay and an Ab binding assay. The BSA will then be permanently expressed in Chinese hamster ovary (CHO) cells, followed by selection, and dilutional cloning, and purification with affinity and cation exchange chromatography. The purified BSA will then be used to treat APPswe/PSEN1(dE9) double transgenic mice over 3 month period. Control mice will be treated with either saline or with the conventional high dose anti-Ab MAb that does not cross the BBB. The mice will be evaluated for neurobehavior, plasma Ab levels, brain Ab plaque content, and brain micro-hemorrhage using Prussian blue histochemistry. This research will provide the necessary pre-clinical pharmacology to support an IND filing for the treatment of humans with AD using genetically engineered fusion antibodies that both cross the BBB via receptor-mediation and bind and disaggregate Ab amyloid plaque of AD. PUBLIC HEALTH RELEVANCE: The dementia of Alzheimer's disease (AD) is caused by the deposition of Ab amyloid in brain over many years, and once the amyloid plaque forms in brain, it is permanent in the absence of plaque disaggregation therapy. The most potent form of plaque diasaggregation therapy is a monoclonal antibody (MAb) against the Ab amyloid peptide of AD, and passive immune therapy of AD is currently being tested in clinical trials; however, the anti-Ab MAb does not cross the blood-brain barrier (BBB). The present research will test in AD transgenic mice the efficacy of a new genetically engineered fusion antibody that both crosses the BBB via receptor-mediation and binds and disaggregates Ab amyloid plaque of AD.

描述（由申请人提供）：阿尔茨海默氏病的痴呆（AD）是由多年来AB淀粉样蛋白在大脑中沉积的。一旦在大脑中形成淀粉样菌斑，它就会永久存在，在没有斑块分解疗法的情况下。牙菌斑二聚体疗法的最有效形式是针对AD的AB淀粉样蛋白肽的单克隆抗体（MAB），而AD的被动免疫治疗目前正在临床试验中进行测试。但是，抗AB MAB不会越过血脑屏障（BBB）。因此，必须将非常高剂量的mAb施用到AD转基因小鼠大脑的较低斑块上，并且这些高剂量会引起脑部微视频诊断。本研究将从遗传上设计一种新形式的抗AB mAB，该形式可以通过受体介导的运输在血液到脑和脑对血液方向上越过BBB。 PI先前已经针对小鼠转铁蛋白受体（TFR）进行了基因设计的嵌合MAB，该mAb通过受体介导的小鼠BBB TFR在血液到脑方向上跨BBB跨BBB，也可以在BBB FC受体（FCR）上的脑对血方向上的BBB越过BBB。此外，PI具有基因设计，并表达了针对AB肽的氨基末端部分的单链FV（SCFV）抗体。本研究将产生一种新的融合蛋白，其中抗AB SCFV与小鼠TFR融合到嵌合MAB的羧基末端，并将这种新的融合蛋白指定为TFRMAB-A2SCFV BI特异性抗体（BSA）。遵循新BSA的基因工程，将在COS细胞中瞬时表达蛋白质，并使用小鼠TFR结合测定法和AB结合测定法证明BSA的双功能。然后，BSA将在中国仓鼠卵巢（CHO）细胞中永久表达，然后选择和稀释克隆，并使用亲和力和阳离子交换色谱法进行纯化。然后，纯化的BSA将在3个月内用于治疗Appswe/psen1（DE9）双转基因小鼠。对照小鼠将用盐水或不穿越BBB的常规高剂量抗ab mAB处理。使用普鲁士蓝色的组织化学，将评估小鼠的神经行为，血浆AB水平，脑AB斑块含量和脑微毛。这项研究将提供必要的临床前药理学，以支持使用基因工程融合抗体对AD治疗AD的IND归档，这些抗体既通过受体中的受体介绍，又结合AD的AB淀粉样蛋白。公共卫生相关性：阿尔茨海默氏病的痴呆（AD）是由多年来AB淀粉样蛋白在大脑中的沉积引起的，一旦大脑中的淀粉样蛋白斑块形式，它在没有斑块分解疗法的情况下是永久性的。牙菌斑二聚体疗法的最有效形式是针对AD的AB淀粉样蛋白肽的单克隆抗体（MAB），而AD的被动免疫治疗目前正在临床试验中进行测试。但是，抗AB MAB不会越过血脑屏障（BBB）。本研究将在AD转基因小鼠中测试一种新的基因工程融合抗体的功效，该抗体既通过受体中的受体进行跨BBB，并结合AD的AB淀粉样蛋白斑块。