Induced Therapeutic Overexpression of Acetylcholinesterase in Vivo

体内诱导乙酰胆碱酯酶的治疗性过度表达

基本信息

批准号：
7633244
负责人：
Richard Lee Rotundo
金额：
$ 32.2万
依托单位：
UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2006
资助国家：
美国
起止时间：
2006-09-30 至 2011-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7633244
关键词：
Acetylcholinesterase Affect Amino Acids Appearance Blood Circulation Catalytic Domain Cell Line Cell surface Cells Cessation of life Chemicals Collaborations Dose Endoplasmic Reticulum Enzyme Stability Enzymes Event Exhibits Exposure to Extramural Activities Future Half-Life Human Isoflurophate Label Lead Length Lethal Dose 50 Measures Medical Research Methods Modification Molecular Mus Muscle Muscle Cells Nervous system structure Neuromuscular Junction Organophosphates Peptides Peripheral Nerves Pesticides Physiologic pulse Principal Investigator Process Proteins Recovery Research Research Design Research Institute Sarin Serum Signal Transduction Site Skeletal Muscle Solubility Soman System Tail Testing Therapeutic Time Tissues Translations Work base cholinergic design fluorophore improved in vivo mimetics mouse model nerve agent novel overexpression programs research study synthetic peptide tissue culture tissue/cell culture transmission process uptake

项目摘要

DESCRIPTION (provided by applicant): Acetylcholinesterase (AChE), the enzyme that terminates cholinergic transmission in the nervous system, is the target of nerve agents and organophosphate pesticides. Inactivation of AChE can result in rapid debilitation or death, thus the use of easily synthesized AChE inactivating nerve agents such as soman and sarin by terrorists looms as a potential catastrophic event. We have previously shown that about 80% of the newly-synthesized AChE molecules are catalytically inactive and subsequently degraded in the endoplasmic reticulum. In addition, while studying the assembly of the multimeric enzyme, we discovered that the noncatalytic subunit appears to "rescue" the catalytic subunits from degradation and, moreover, stabilizes the tetramers for subsequent transport to the muscle cell surface and secretion. Based on these observations we have designed and tested peptides that can mimic the ability of the non-catalytic subunits to rescue the enzyme and increase its expression. Our specific aims on this project are; 1) test the hypothesis that the non-catalytic subunit of AChE increases expression of the enzyme by rescuing the catalytic subunits from degradation. The objective is to determine the molecular mechanisms underlying the observed increase in AChE expression to develop more effective inducing peptides; 2) Optimize the synthetic peptides designed to rescue AChE for their cellular uptake and retention as well as ability to induce tetramerization and increased enzyme stability; 3) Test the ability of the non-catalytic AChE subunit mimetic peptides to increase AChE expression in vivo using a mouse model; and 4) Determine the ability of the synthetic peptides to protect mice from exposure to organophosphate nerve agents and/or to improve survival when administered after exposure. Together these studies could lead to a novel method for increasing AChE expression in vivo for protection against organophosphate compounds as well as enhancing the rate of recovery following exposure.

描述（由申请人提供）：乙酰胆碱酯酶（AChE）是一种终止神经系统中胆碱能传递的酶，是神经毒剂和有机磷农药的目标。乙酰胆碱酯酶失活可导致快速衰弱或死亡，因此恐怖分子使用容易合成的乙酰胆碱酯酶失活神经毒剂（例如梭曼和沙林）可能会成为潜在的灾难性事件。我们之前已经证明，大约 80% 新合成的 AChE 分子没有催化活性，随后在内质网中降解。此外，在研究多聚体酶的组装时，我们发现非催化亚基似乎“拯救”了催化亚基免于降解，而且稳定了四聚体，以便随后转运到肌肉细胞表面和分泌。基于这些观察，我们设计并测试了可以模仿非催化亚基拯救酶并增加其表达的能力的肽。我们该项目的具体目标是： 1) 检验 AChE 的非催化亚基通过拯救催化亚基免于降解来增加酶表达的假设。目的是确定观察到的 AChE 表达增加的分子机制，以开发更有效的诱导肽； 2) 优化旨在拯救AChE的合成肽的细胞摄取和保留以及诱导四聚化和增加酶稳定性的能力； 3) 利用小鼠模型测试非催化AChE亚基模拟肽体内增加AChE表达的能力； 4)确定合成肽保护小鼠免于暴露于有机磷酸酯神经毒剂和/或在暴露后施用时提高存活率的能力。这些研究共同可能产生一种增加体内 AChE 表达的新方法，以防止有机磷酸酯化合物的侵害，并提高暴露后的恢复率。