Multi-scale disease modeling of SCN2A-related epilepsy due to gain-of-function variants

癫痫影响着全球1%的人口，在美国有300万人。 一个人越来越多的儿童癫痫与离子通道基因的致病变异有关，包括 电压门控钠通道基因SCN 2A。NINDS的2020年癫痫研究基准优先 确定遗传变异如何导致癫痫和相关的神经发育障碍。SCN 2A变体， 表现为功能丧失与严重的神经发育障碍和迟发性癫痫有关。 另一方面，功能获得性SCN 2A变体主要具有早发性癫痫的表型。 编码的钠通道（NaV1.2）在兴奋性多巴胺能神经元中的早期高度表达， 发展，提供了一个独特的机会，研究兴奋性神经元功能障碍如何导致早发性 癫痫动物和人类组织来源的神经元模型为Dravet如何 综合征导致中间神经元功能障碍和癫痫。在SCN 2A相关疾病中， 阐明功能丧失如何导致自闭症谱系障碍伴迟发性癫痫。由于缺乏现成的 虽然现有的疾病模型，但对兴奋性神经元功能障碍如何在早期 会导致早发性癫痫这项提案将利用两种早发性癫痫的变种， SCN 2A具有趋同的临床表型，但具有不同的生物物理机制。患者源性神经元 模型和鼠标模型提供了定义多个机械收敛点的机会， 量表：从单个神经元到影响癫痫表型的神经回路。目标1将决定如何获得两个- 编码错义突变M1879 T和E430 A的功能缺失的SCN 2A变体通过以下方式赋予增加的兴奋性： 不同的机制。在隔离和基本电路中对iPSC衍生神经元的功能分析将 定义不同的变异如何影响兴奋性，从而向癫痫表型收敛。目标2将 在癫痫小鼠中定义海马高水平回路扰动， 概括了SCN 2A-E430 A人类癫痫性脑病。兴奋性变化的离体分析， 突触信号和海马体中的网络输出将导致对功能获得如何产生新的理解。 SCN 2A变体影响神经元网络。EEG和深度电极将提供与 体内癫痫表型。这项建议将推动获奖者作为一个独立的物理学家，科学家， 结合遗传性癫痫多尺度建模的新专业知识，重点相关的教学法，以及多样化的 专业从事神经发育和遗传疾病的职业发展团队，所有这些都是高度协作的 环境促进初级教师的发展。该奖项将提供一个平台，以1）定义特定于变体的 自限性和难治性癫痫中癫痫表型的贡献，2）研究如何靶向 癫痫回路功能障碍影响未来R 01资助的独立回路输出和癫痫表型 research.

Project Summary Epilepsy affects up to 1% of the population worldwide, and 3 million in the United States alone. A growing proportion of pediatric epilepsies are tied to causative variants in ion channel genes, including the voltage-gated sodium channel gene SCN2A. The 2020 Epilepsy Research Benchmarks of NINDS prioritize identifying how genetic variants cause epilepsy and related neurodevelopmental disorders. SCN2A variants that manifest with loss-of-function are associated with severe neurodevelopmental disorders and late-onset epilepsy. On the other hand, gain-of-function SCN2A variants predominantly have a phenotype of early-onset epilepsy. The encoded sodium channel (NaV1.2) is highly expressed in excitatory glutamatergic neurons early in development, presenting a unique opportunity to examine how excitatory neuron dysfunction leads to early-onset epilepsy. Animal and human tissue-derived neuron models have brought mechanistic insight to how Dravet syndrome results in interneuron dysfunction and epilepsy. Among SCN2A-related diseases, animal models illuminate how loss-of-function leads to autism spectrum disorder with late-onset epilepsy. Due to lack of readily available disease models, there is sparse mechanistic understanding of how excitatory neuron dysfunction early in development leads to early-onset epilepsy. This proposal will exploit two early-onset epilepsy variants of SCN2A that have a convergent clinical phenotype yet divergent biophysical mechanisms. Patient-derived neuron models and mouse models provide the opportunity to define the point of mechanistic convergence at multiple scales: from single neurons to neural circuits influencing epilepsy phenotype. Aim 1 will determine how two gain- of-function SCN2A variants, encoding missense mutations M1879T and E430A, confer increased excitability by distinct mechanisms. Functional analysis of iPSC-derived neurons in isolation and in elementary circuits will define how the different variants impact excitability and thus converge toward an epileptic phenotype. Aim 2 will define hippocampal higher-level circuit perturbations in epileptic mice designed with genome editing to recapitulate the SCN2A-E430A human epileptic encephalopathy. Ex vivo analysis of changes in excitability, synaptic signaling, and network output in the hippocampus will lead to new understanding of how gain-of-function SCN2A variants affect neuronal networks. EEG and depth electrodes will provide spatiotemporal correlate to the in vivo epilepsy phenotype. This proposal will propel the awardee to independence as a physician-scientist by incorporating new expertise in multi-scale modeling of genetic epilepsy, focused relevant didactics, and a diverse career development team specializing in neurodevelopmental and genetic disorders, all in a highly collaborative environment fostering junior faculty development. This award will provide a platform to 1) define variant-specific contributions to epilepsy phenotype in self-limited and intractable epilepsies and 2) investigate how targeted epileptic circuit dysfunction influences circuit output and epilepsy phenotype in future R01-funded independent research.