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：
7674768
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/7674768
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 Pharmacology Clinical Trials Cloning Complementary DNA DNA Sequence Data Dementia Deposition Disease 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 Plasma Plasmids Proteins Prussian blue Publishing RNA Splicing Radio Rattus Reaction Research Rodent Saline Senile Plaques Series Serum-Free Culture Media Staging Structure Testing Therapeutic Transfection Transferrin Receptor Transgenic Mice Validation Western Blotting Work amyloid peptide animal efficacy 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 public health relevance 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（申请人提供）：阿尔茨海默病（AD）痴呆是由于Ab淀粉样蛋白在大脑中长年沉积引起的。一旦淀粉样蛋白斑块在大脑中形成，在没有斑块解聚治疗的情况下，它是永久性的。最有效的斑块解聚疗法是针对 AD 的 Ab 淀粉样肽的单克隆抗体 (MAb)，而 AD 的被动免疫疗法目前正在临床试验中进行测试。然而，抗Ab MAb 不能穿过血脑屏障(BBB)。因此，必须施用非常高剂量的单克隆抗体来降低 AD 转基因小鼠脑中的斑块，而这些高剂量会导致脑微出血。目前的研究将通过基因工程设计出一种新型抗抗体单克隆抗体，该单克隆抗体能够通过受体介导的血液至大脑和大脑至血液方向的运输穿过血脑屏障。 PI 此前已对小鼠转铁蛋白受体 (TfR) 进行了基因工程改造，该单克隆抗体通过小鼠 BBB TfR 上受体介导的转运以血液至大脑方向穿过 BBB，并且还穿过大脑至大脑方向的 BBB。 - BBB Fc 受体 (FcR) 上的血液方向。此外，PI还经过基因工程改造，表达针对Ab肽氨基末端部分的单链Fv (ScFv)抗体。目前的研究将产生一种新的融合蛋白，其中抗Ab ScFv融合到针对小鼠TfR的嵌合MAb的羧基末端，这种新的融合蛋白被命名为TfRMAb-A2ScFv双特异性抗体（BSA）。新 BSA 的基因工程之后，该蛋白将在 COS 细胞中瞬时表达，并且 BSA 的双功能将通过小鼠 TfR 结合测定和 Ab 结合测定来证明。然后 BSA 将在中国仓鼠卵巢 (CHO) 细胞中永久表达，然后进行选择、稀释克隆，并通过亲和层析和阳离子交换层析进行纯化。然后，纯化的 BSA 将用于治疗 APPswe/PSEN1(dE9) 双转基因小鼠 3 个月。对照小鼠将用盐水或不穿过 BBB 的常规高剂量抗抗体 MAb 进行治疗。将使用普鲁士蓝组织化学评估小鼠的神经行为、血浆抗体水平、脑抗体斑块含量和脑微出血。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治疗人类 AD 的 IND 申请，该融合抗体通过受体介导穿过 BBB，并结合和分解 AD 的 Ab 淀粉样斑块。公共健康相关性：阿尔茨海默病 (AD) 痴呆是由 Ab 淀粉样蛋白在大脑中多年沉积引起的，一旦淀粉样蛋白斑块在大脑中形成，在没有斑块解聚治疗的情况下，它是永久性的。最有效的斑块解聚疗法是针对 AD 的 Ab 淀粉样肽的单克隆抗体 (MAb)，AD 的被动免疫疗法目前正在临床试验中进行测试；然而，抗Ab MAb 不能穿过血脑屏障(BBB)。目前的研究将在 AD 转基因小鼠中测试一种新的基因工程融合抗体的功效，该抗体既通过受体介导穿过 BBB，又结合并分解 AD 的 Ab 淀粉样斑块。