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：
7900676
负责人：
Richard Lee Rotundo
金额：
$ 17万
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/7900676
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）是终止神经系统中胆碱能传播的酶，是神经剂和有机磷酸盐农药的靶标。酸痛的失活可能导致衰弱或死亡，从而使用易于合成的aChE aChE灭活神经剂，例如索曼人和恐怖分子的沙林，可能是一种潜在的灾难性事件。我们以前已经表明，大约80％的新合成的ACHE分子是催化活性的，随后在内质网中降解。此外，在研究多聚酶的组装时，我们发现非催化亚基似乎“营救”催化亚基免于降解，此外，还可以使四聚体稳定，以便随后运输到肌肉细胞表面和分泌。基于这些观察结果，我们设计和测试了肽，可以模仿非催化亚基营救酶并增加其表达的能力。我们对该项目的具体目标是； 1）检验以下假设：ACHE的非催化亚基通过从降解中营救催化亚基来增加酶的表达。目的是确定观察到的ACHE表达增加的分子机制，以开发出更有效的诱导肽。 2）优化旨在挽救其细胞摄取和保留率的酸痛的合成肽，以及诱导四聚体和提高酶稳定性的能力； 3）测试非催化性ACHE亚基模拟肽使用小鼠模型在体内增加体内表达的能力； 4）确定合成肽保护小鼠免于暴露于有机磷酸盐神经剂的能力和/或在暴露后施用时提高生存率。这些研究共同导致一种新的方法来增加体内ACHE表达，以防止有机磷酸化合物，并提高暴露后的恢复速率。