NMDAR Mutations & Neurodevelopmental Disorder: from Mechanism to Targeted Therapy

描述(申请人提供)：神经发育障碍与大脑功能障碍有关，影响儿童的行为、记忆或学习能力。这种残疾给个人、他们的家庭以及社会带来了毁灭性的精神、情感和经济后果。一些残疾的分子基础涉及各种离子通道家族的致病突变，包括NMDAR(NMDAR)。由两个甘氨酸结合的GluN1亚基和两个谷氨酸结合的GluN2亚基组装形成的阳离子选择性NMDAR通道调节兴奋性突触电流中缓慢的、钙离子渗透的成分，这可以触发突触强度的变化，突触强度是学习的细胞关联。NMDAR在正常的大脑发育中也扮演着重要的角色。仅在过去的三年里，就有大量突变(&gt；140)的报道，这导致了一种观点，即这些突变存在于神经疾病患者的子集中，特别是早期发作的难治性癫痫。令人惊讶的是，在有神经问题的儿童患者中发现的NMDAR突变的发生率为5.7%，与Na+、K+、Ca2+通道和GABA受体的突变相似或更高。NMDAR亚单位的突变在患有广泛神经发育问题的儿童中被发现，包括注意缺陷多动障碍(ADHD)、自闭症谱系障碍、发育迟缓、精神发育迟缓、精神分裂症、智力残疾和难治性癫痫。不幸的是，几乎没有对这些突变的功能分析，这使得在临床表型背景下评估突变的影响是不可能的。我们提出了4条实验路线，解决神经疾病潜在的分子机制，建议源于NMDAR亚单位的突变。我们将研究跨膜结构域(TM)、连接子和配体结合结构域(LBD)突变的功能效应，并测试FDA批准的药物纠正突变诱导的功能获得的能力。所有实验都将利用包含0、1或2个突变NMDAR亚单位的受体，从而能够评估杂合子患者的功能。目的1.人类TM连接区的NMDAR突变对功能有何影响？我们将分析跨膜结构域或相关连接子中的26个突变。我们将合作努力，以获得开放渠道配置的晶体。目的2.人类NMDAR配体结合区突变对功能有何影响？我们将评估36个配体结合域突变的功能效应，并合作获得结晶学数据。目的3.人类NMDAR突变如何影响神经元的运输和功能？我们将分析NMDAR介导的突触电流在转染突变NMDAR亚单位的切片培养中的特性。目的4.NMDAR通道病可以治疗吗？我们将评估FDA批准的NMDAR拮抗剂在功能获得NMDAR突变时的效力(IC50)，并评估NMDAR突变的神经毒性潜力。

DESCRIPTION (provided by applicant): Neurodevelopmental disorders are associated with disabilities in brain function that affect a child's behavior, memory or ability to learn. Such disabilities carry devastating mental, emotional, and economic consequences for the individuals, their families, as well as society. The molecular bases for a subset of disabilities involve disease-causing mutations in various ion channel families, including NMDA receptors (NMDARs). The cation-selective NMDAR channels formed from assembly of two glycine-binding GluN1 subunits and two glutamate-binding GluN2 subunits mediate a slow, Ca2+-permeable component of excitatory synaptic currents that can trigger changes in synaptic strength, a cellular correlate of learning. NMDARs also play an important role in normal brain development. A large number of mutations (>140) have been reported in just the last three years, leading to the view that these mutations are present in a subset of patients with neurological disorders, particularly early onset intractable seizures. Surprisingly, the incidence of NMDAR mutations found in pediatric patients presenting with neurological problems is 5.7%, similar to or higher than that for Na+, K+ , Ca2+ channels and GABA receptors. Mutations in NMDAR subunits have been identified in children with a broad range of neurodevelopmental problems, including attention deficit hyperactivity disorder (ADHD), autism spectrum disorders, developmental delay, mental retardation, schizophrenia, intellectual disability, and intractable seizures. Unfortunately, virtually no functional analysis of these mutations exists, making it impossible to evaluate effects of mutations in the context of clinical phenotype. We proposed 4 lines of experimentation addressing the molecular mechanism underlying neurological diseases suggested to arise from mutations in NMDAR subunits. We will study the functional effects of mutations in the transmembrane domain (TM), linkers, and ligand binding domains (LBD) and test the ability of FDA-approved drugs to rectify the mutation-induced gain-of-function. All experiments will utilize receptors that contain 0, 1, or 2 mutant NMDAR subunits, enabling an assessment of function in heterozygous patients. Aim 1. How do human NMDAR mutations in the TM- linker regions impact function? We will analyze 26 mutations in the transmembrane domain or associated linkers. We will collaborate on efforts to obtain crystals of the open channel configuration. Aim 2. How do human NMDAR mutations in the ligand binding domains impact function? We will evaluate the functional effects of 36 mutations in the ligand binding domain, and collaborate to obtain crystallographic data. Aim 3. How do human NMDAR mutations influence neuronal trafficking and function? We will analyze the properties of NMDAR-mediated synaptic current in slice cultures transfected with mutant NMDAR subunits. Aim 4. Are NMDAR channelopathies treatable? We will evaluate the potency (IC50) of FDA-approved NMDAR antagonists at gain-of-function NMDAR mutations and evaluate the neurotoxic potential of NMDAR mutations.