Epigenetics-Based Autism Treatment with Animal Models and Human Stem Cells

摘要 该项目旨在发现针对自闭症核心症状的新的药物干预措施，包括社交 缺陷和重复的行为。自闭症的致病因素之一是Shank3基因的缺失。 编码谷氨酸能突触的支架蛋白。我们将使用Shank3缺陷小鼠模型和 人类干细胞衍生的神经元参与了这一药物发现的努力。遗传学研究发现，许多 自闭症中被破坏的基因是介导组蛋白甲基化/去甲基化的组蛋白修饰酶， 它们在转录调控中起着关键作用。我们的初步研究发现，组蛋白赖氨酸4 二甲基化(H3K4me2，与基因激活有关)在儿童的前额叶皮质(PFC)中显著减少 自闭症患者和Shank3基因缺陷小鼠。H3K4me2被赖氨酸专一性的组蛋白去甲基化 1(LSD1，KDM1A)，发现该基因在Shank3基因缺陷小鼠的PFC神经元中增加。我们假设 抑制LSD1上调H3K4me2并恢复基因表达可能能够改善孤独症样症状 因此，为自闭症提供了一种新的治疗策略。结合了行为，生化， 电生理学、基因组和干细胞方法将被用来检验这一假设。目标1，我们将 表观遗传剂在自闭症小鼠模型中的表观遗传学变化和治疗效果的特征。这个 组蛋白甲基化标记和组蛋白去甲基酶的改变也将在Shank3-PFC中进行检测。 缺陷小鼠和自闭症患者的死后组织。目的2，揭示分子机制。 自闭症模型的潜在表观遗传学治疗。突触反应和神经元兴奋性将是 在用LSD1抑制剂治疗的Shank3缺陷小鼠中记录。全基因组范围的基因表达改变 组蛋白甲基化将使用RNAseq和ChIPseq进行检测。确定的关键字的因果作用 LSD1抑制剂的治疗效果中的分子也将被确定。在目标3中，我们将研究 Shank3基因致ASD患者神经细胞的分子改变及治疗策略 单倍体功能不全。为了找出在Shank3小鼠模型中发现的表观遗传治疗策略是否可能 同样在自闭症患者身上工作，我们将使用创新的干细胞技术来检测LSD1的能力 逆转ASD患者诱导后神经元突触缺陷和分子畸变的抑制剂 多能干细胞。这项研究的结果不仅揭示了重要的自闭症之间的机制联系 风险因素，但也揭示了一种基于机制的自闭症治疗策略。

Summary This project aims to discover novel pharmacological intervention for core symptoms of autism, including social deficits and repetitive behaviors. One of the causal factors of autism is the loss of Shank3 gene, which encodes a scaffolding protein at glutamatergic synapses. We will use Shank3-deficient mouse models and human stem cell-derived neurons in this drug discovery endeavor. Genetics studies have found that many of genes disrupted in autism are histone-modifying enzymes that mediate histone methylation/demethylation, which play a key role in transcriptional regulation. Our preliminary studies have found that histone lysine 4 dimethylation (H3K4me2, linked to gene activation) is significantly decreased in the prefrontal cortex (PFC) of autistic humans and Shank3-deficient mice. H3K4me2 is demethylated by lysine-specific histone demethylase 1 (LSD1, KDM1A), which is found to be increased in PFC neurons of Shank3-deficient mice. We hypothesize that inhibiting LSD1 to elevate H3K4me2 and restore gene expression may be able to ameliorate autism-like phenotypes, therefore providing a novel therapeutic strategy for autism. Combined behavioral, biochemical, electrophysiological, genomic and stem cell approaches will be used to test this hypothesis. Aim 1, we will characterize epigenetic changes and therapeutic effects of epigenetic agents in mouse models of autism. The alteration of histone methylation marks and histone demethylases will also be examined in PFC of Shank3- deficient mice and autism human postmortem tissues. Aim 2, we will reveal the molecular mechanisms underlying epigenetic treatment of autism models. Synaptic responses and neuronal excitability will be recorded in Shank3-deficient mice treated with LSD1 inhibitors. Genome-wide alteration of gene expression and histone methylation will be examined using RNAseq and ChIPseq. The causal role of identified key molecules in the therapeutic effects of LSD1 inhibitors will also be determined. In Aim 3, we will examine the molecular alteration and treatment strategy in human neurons from ASD patient with Shank3 haploinsufficiency. To find out whether the epigenetic treatment strategy found in Shank3 mouse models might also work in autism patients, we will use the innovative stem-cell technology to examine the capability of LSD1 inhibitors to reverse synaptic deficits and molecular aberrations in ASD patient’s neurons derived from induced pluripotent stem cells. Results from this study will not only reveal the mechanistic link among important autism risk factors, but also uncover a mechanism-based treatment strategy for autism.