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Epilepsy in Focal Cortical Malformations

局灶性皮质畸形中的癫痫

基本信息

批准号：
10598453
负责人：
Angelique Bordey
金额：
$ 43.59万
依托单位：
YALE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-04-01 至 2024-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10598453
关键词：
Address Adult Affect Age Aging Binding Biochemical Biology Birth CCI-779 Cells Cortical Dysplasia Cortical Malformation Cyclic AMP Cyclic Nucleotides Data Development Disease Dose Electroporation Epilepsy Excision FRAP1 gene Generations Genes Genetic Diseases Heart Human Hyperactivity Individual Lead Light Medial Messenger RNA Modeling Mus Mutation Neurobiology Neurons Operative Surgical Procedures Pathway interactions Patients Peripheral Phosphorylation Physiological Post-Translational Protein Processing Prefrontal Cortex Protein Isoforms Proteins Refractory SDZ RAD Science Seizures Serious Adverse Event Sirolimus TSC1 gene Testing Time Tissues Translations Treatment Protocols Tuberous Sclerosis beta-adrenergic receptor conventional therapy genetic variant hippocampal pyramidal neuron improved in utero in vivo mTOR inhibition mouse model mutant neuronal excitability new therapeutic target novel patient population personalized medicine prevent receptor side effect therapeutic target

项目摘要

Abstract Tuberous sclerosis complex (TSC) and focal cortical dysplasia type II (FCDII) are caused by mutations in mTOR pathway genes leading to mTOR hyperactivity, focal malformations of cortical development (fMCD), and seizures in 80-90% of the patients. The current definitive treatments for epilepsy are surgical resection or treatment with everolimus, which inhibits mTOR activity (only approved for TSC). Because both options have severe limitations, there is a major need to better understand the mechanisms leading to seizures to improve life-long epilepsy treatment in TSC and FCDII. To investigate such mechanisms, we recently developed a murine model of fMCD-associated epilepsy that recapitulates the human TSC and FCDII disorders. fMCD are defined by the presence of misplaced, dysmorphic cortical neurons expressing hyperactive mTOR – for simplicity we will refer to these as “mutant” neurons. In our model and in human TSC tissue, we made a surprising finding that mutant neurons express HCN4 channels, which are not normally functionally expressed in cortical neurons. These data led us to ask several important questions based on the known biology of HCN4 channels: (1) As HCN4 channels are responsible for the pacemaking activity of the heart, can HCN4 channel expression lead to repetitive firing of mutant neurons resulting in seizures? (2) HCN4 is the most cAMP-sensitive of the four HCN isoforms. Do coincident increases in cAMP (e.g., β-adrenergic receptors) and hyperpolarization or depolarizations drive HCN4 channel opening and neuronal firing? (3) HCN4 channel mRNA is expressed in cortical neurons. Is the abnormal HCN4 expression in mutant neurons due to increased translation via mTOR? (4) Seizures can start at any age in patients that have been seizure-free for decades, but we do not know why. Can this be explained by worsening of mTOR hyperactivity with age leading to a progressive increase in HCN4 expression until there is enough HCN4 channels to depolarize cells and reach firing threshold upon activation? (5) There is no selective blocker of the HCN4 channel and blocking other HCN channels would have serious central and peripheral side-effects. Identifying the mechanisms responsible for functional HCN4 expression may therefore provide alternative therapeutic targets. Do binding partners and/or post-translational modifications contribute to HCN4 abnormal expression in mutant neurons? We will address these questions in three aims testing our central hypothesis that abnormal mTOR- and translation- dependent expression of HCN4 channels leads to repetitive neuronal firing and seizures in TSC and FCDII. Aim 1: Test the hypothesis that abnormal HCN4 channel expression in murine TSC/FCDII mutant neurons contribute to neuron excitability and seizure activity. Aim2: Test the hypothesis that abnormal HCN4 expression is mTOR- and translation-dependent and increases with age and seizures. Aim 3: Test the hypothesis that HCN4 binding partners and posttranslational modifications are necessary for its functional expression and function. The proposed studies will be performed through a collaborative effort between the Bordey and Calderwood labs that together combine unique and extensive expertise in in vivo neurobiology, and biochemical and protein science.

摘要多发性硬化症（TSC）和局灶性皮质发育不良II型（FCDII）是由基因突变引起的。在导致mTOR过度活跃的mTOR通路基因中，皮质发育的局灶性畸形（fMCD），80-90%的患者出现癫痫发作。目前癫痫的确切治疗方法是手术切除或用抑制mTOR活性的依维莫司治疗（仅批准用于TSC）。由于这两种选择都有严重的局限性，因此非常需要更好地了解导致癫痫发作的机制，以改善TSC和FCDII的终身癫痫治疗。到为了研究这些机制，我们最近开发了一种fMCD相关癫痫的小鼠模型它概括了人类TSC和FCDII疾病。fMCD的定义是表达过度活跃的mTOR的错位的畸形皮质神经元-为了简单起见，我们将参考这些是“突变的”神经元。在我们的模型和人类TSC组织中，我们有一个令人惊讶的发现，突变的神经元表达HCN 4通道，其在皮质神经元中通常不功能性表达。神经元这些数据让我们基于HCN 4的已知生物学提出了几个重要问题（1）由于HCN 4通道负责心脏的起搏活动，HCN 4通道是否可以通道表达导致突变神经元重复放电导致癫痫发作？(2)HCN 4是四种HCN同种型中最cAMP敏感的。cAMP的同步增加（例如，β-肾上腺素能的受体）和超极化或去极化驱动HCN 4通道开放和神经元放电？ (3)HCN 4通道mRNA在皮质神经元中表达。突变型HCN 4表达异常神经元由于增加翻译通过mTOR？(4)癫痫发作可以在患者的任何年龄开始，几十年来一直没有被发现，但我们不知道为什么。这是否可以解释为随着年龄的增长，mTOR过度活跃导致HCN 4表达的进行性增加，直到出现足够的HCN 4通道去激活细胞并达到激活阈值？(5)没有 HCN 4通道的选择性阻断剂和阻断其他HCN通道将具有严重的中枢神经系统损伤。和外周副作用确定负责功能性HCN 4表达的机制因此可以提供替代的治疗靶点。结合伴侣和/或翻译后修饰有助于突变神经元中HCN 4的异常表达？我们将解决这些问题三个目标的问题测试我们的中心假设，异常mTOR-和翻译- HCN 4通道的依赖性表达导致TSC中重复的神经元放电和癫痫发作， FCDII。目的1：验证小鼠TSC/FCDII中HCN 4通道表达异常的假设，突变神经元有助于神经元兴奋性和癫痫发作活动。目标2：检验假设， HCN 4的异常表达是mTOR依赖性的，并且随着年龄的增长而增加，癫痫发作目的3：检验HCN 4结合配偶体和翻译后修饰是其功能表达和功能所必需的。将开展拟定研究通过Bordey和Calderwood实验室之间的合作努力，它们共同联合收割机结合了独特的以及在体内神经生物学、生物化学和蛋白质科学方面的广泛专业知识。