Activated protein C mechanisms of brain white matter protection and new therapies for brain white matter ischemic injury

激活蛋白C脑白质保护机制及脑白质缺血性损伤新疗法

基本信息

批准号：
10391557
负责人：
William J Mack
金额：
$ 84.37万
依托单位：
UNIVERSITY OF SOUTHERN CALIFORNIA
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-07-15 至 2026-04-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10391557
关键词：
Address Agonist Amino Acid Sequence Anti-Inflammatory Agents Anticoagulants Apoptotic Arteries Astrocytes Axon Biological Products Blood - brain barrier anatomy Blood Vessels Blood flow Brain Cells Clinic Clinical Clinical Trials Collaborations Data Dementia Endothelium Engineering Exhibits Extravasation Factor VIIIa Factor Va Fiber Functional disorder GTP-Binding Proteins Gene Expression Generations Goals In Vitro Inflammatory Injury Ischemic Stroke Knowledge Lesion Ligands Magnetic Resonance Imaging Microglia Modeling Mus Mutation N-terminal Nerve Degeneration Neurons Oligodendroglia PAR-1 Receptor Pathway interactions Peptides Platelet Activation Point Mutation Prevention therapy Proteolysis Role Signal Transduction Site Stroke Testing Therapeutic Thrombin Toxic effect Translating Traumatic Brain Injury activated Protein C activated protein C receptor analog arrestin 2 base behavior test cell type disability improved in vivo in vivo Model insight ischemic injury mimetics mortality mutant neuropathology novel novel therapeutics peptidomimetics phase II trial prevent protective effect response stroke model stroke patient stroke therapy success vascular cognitive impairment and dementia vasoconstriction white matter white matter injury

项目摘要

ABSTRACT Stroke in small brain vessels in subcortical white matter (WM) regions account for 25% of all strokes. It leads to vascular cognitive impairment and dementia (VCID), and is the second leading cause of dementia overall. Despite such clinical importance, the pathophysiology of ischemic WM injury (WMI) and VCID is still poorly understood. Moreover, there is no yet an approved therapy for prevention and/or treatment of WM strokes and VCID. Here, we propose collaborative studies between the Zlokovic and Griffin labs on activated protein C (APC) pathways in the WM, and to evaluate therapeutic potential of APC-based therapies for ischemic WMI using a model of vasoconstriction of small brain vessels in the WM. Our previous studies using models of large artery infracts, brain trauma and neurodegeneration led to discovery of vasculoprotective, blood-brain barrier (BBB)-stabilizing, neuroprotective, and anti-inflammatory activities of APC and its cytoprotective-selective mutants. In 2019, these findings have been translated into successfully completed phase 2 trial for ischemic stroke of 3K3A-APC, a 2nd generation cytoprotective-selective APC analog with >90% loss of anticoagulant activity. However, whether activation of APC pathways in the WM is beneficial or not during ischemic WMI, remains unknown. Our goals include: 1) providing proof of concept for hypothesized mechanisms for protective activities of APC in the WM; and 2) characterizing novel protease activated receptor 1 (PAR1)-related P1-47 and PAR3-related P3-42 APC-mimetic peptides, and 3) testing improved 3rd generation APC R-46-selective biologics for treating and preventing ischemic WMI and WM stroke. Our pilot data support our hypotheses that: i) APC will be beneficial for ischemic WMI via PAR1 cleavage at Arg46 to protect WM fiber tracts, oligodendrocytes and BBB from ischemic WMI (AIM 1); ii) APC-mimetic peptides derived from PAR1 and PAR3 sequences (e.g,, P1-47 and P3-42, (i.e., the tethered PAR agonists created by APC cleavages) exhibit synergistic biased agonism, and will elicit -arrestin 2-dependent cytoprotective signaling in brain endothelium and oligodendrocytes in vitro and in vivo after WM stroke (AIM 2); and iii) E56K-APC and D180E-APC newly engineered APC mutants have enhanced ability to cleave PAR1 at Arg46 and will provide improved APC biologics for WM stroke therapy (AIM 3). To address our hypotheses, we will use i) WM model of stroke; ii) new mouse lines carrying R41Q-PAR1 and R46Q-PAR1 point mutations, and -arrestin 2-/- and G12-/- mice; iii) new APC-mimetic PAR1- and PAR3-related peptides with the respective PAR1 and PAR3 tethered-ligand amino acid sequences; iv) new APC R46-cleavage site selective biologics; v) in vivo mutiparametric longitudinal MRI of WM lesion volume, BBB integrity, blood flow, structural and connectivity changes, and tract-tracing based connectomics for circuit level analysis; vi) behavior tests; vii) immunohistology, neuropathology; and viii) oligodendrocyte cultures and in vitro BBB model. If successful, new knowledge generated from this project could translate to the clinic as new therapies for WM stroke and VCID.

抽象的皮质下白质（WM）区域的小脑血管中的中风占所有中风的25％。它导致血管认知障碍和痴呆（VCID），是痴呆症总体上的第二大原因。尽管如此临床重要性，但缺血性WM损伤（WMI）和VCID的病理生理仍然知之甚少。此外，尚无批准的预防和/或治疗WM中风和VCID的疗法。在这里，我们提出了Zlokovic和Griffin Labs之间的合作研究，以使用WM的缺血性WMI评估基于APC的疗法的活化蛋白C（APC）途径，并使用WM中小脑容器的血管收缩模型来评估缺血性WMI的治疗潜力。我们先前使用大型动脉违规，脑外伤和神经退行性的模型的研究导致了APC及其细胞保护性突变体的血管保护性，血脑屏障（BBB）稳定，神经保护性和抗炎活性。在2019年，这些发现已转化为成功完成2阶段试验的3K3A-APC缺血性中风，这是第二代细胞保护性选择APC类似物，抗凝活性损失> 90％。但是，在缺血性WMI期间，WM中APC途径的激活是否有益。我们的目标包括：1）为保护WM中APC活动的假设机制提供概念验证；和2）表征新型保护性活化受体1（PAR1）相关的P1-47和PAR3相关的P3-42 APC模拟辣椒，以及3）测试改善了第三代APC R-46选择性生物学，用于治疗和预防缺血性WMI和WMI stroke。我们的飞行员数据支持我们的假设：i）APC通过ARG46处的PAR1裂解对缺血性WMI有益，以保护WM纤维区域，少突胶质细胞和BBB免受缺血性WMI的影响（AIM 1）； ii）源自PAR1和PAR3序列（例如，P1-47和P3-42，（即，由APC裂解产生的束缚的PAR激动剂）暴露于协同偏见的激动剂，将引起braimodent的braimodectim和Oligy In In In In In In In In In In In In In In In In In In In In In In In In In In In In In In In In In In In In In In In In In In In In In In In In Inim Inim Inim Inim Inim Inim Inium和Oligim Introcy In In Inim Inim In Inium和Oligy Introcy Introcy， WM中风（AIM 2）和III）E56K-APC和D180E-APC新设计的APC突变体具有增强的ARG46清除PAR1的能力，并将为WM中风疗法提供改进的APC生物学（AIM 3）。为了解决我们的假设，我们将使用I）中风模型； ii）携带R41Q-PAR1和R46Q-PAR1点突变的新小鼠系，以及-arrestin 2 - / - 和G12 - / - 小鼠； iii）与各个PAR1和PAR3系在一起的氨基酸序列的新型APC模拟PAR1和PAR3相关的Petides； iv）新的APC R46裂解位点选择性生物制剂； v）WM病变体积，BBB完整性，血流，结构和连通性变化以及基于路追踪的连接组学用于电路水平分析； vi）行为测试； vii）免疫组织学，神经病理学；和VIII）少突胶质细胞培养物和体外BBB模型。如果成功，该项目产生的新知识可能会转化为WM中风和VCID的新疗法。

项目成果

期刊论文数量（1）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Protection of ischemic white matter and oligodendrocytes in mice by 3K3A-activated protein C.

DOI：
10.1084/jem.20211372
发表时间：
2022-01-03
期刊：
The Journal of experimental medicine
影响因子：
0
作者：
Huuskonen MT;Wang Y;Nikolakopoulou AM;Montagne A;Dai Z;Lazic D;Sagare AP;Zhao Z;Fernandez JA;Griffin JH;Zlokovic BV
通讯作者：
Zlokovic BV