Modeling Down Syndrome Neural Phenotypes with Chromosomal Silencing

项目总结： 唐氏综合症(DS)，或21三体，是导致儿童智力障碍的主要遗传原因， 大约每700名活产婴儿中就有1名携带额外的21号染色体副本。与不太常见的情况相比 单基因疾病中，DS的发病机制尚不清楚。DS的治疗需要以下两种之一 识别传统疗法或潜在的染色体疗法靶向的分子途径 使三体染色体上的许多可能的破坏性基因沉默。一扇治疗之窗 干预在于阿尔茨海默病的病理，几乎所有的DS患者都在中年遭受这种疾病的折磨。 最近，通过定向插入转基因，额外的染色体已经以一种诱导的方式沉默了。 将XIST基因导入人诱导多能干细胞(IPSC)。Xist通常使一个X静音 雌性的染色体，提供了一种自然的剂量补偿机制。染色体沉默 DS细胞为研究DS的病理机制提供了强大的等基因和等表观遗传学模型，标志着世界上第一个 迈向DS患者染色体治疗的目标。拟议的工作将调查 沉默DS iPSC来源的神经细胞上的额外染色体，研究DS和阿尔茨海默病- 特定的表型。 目的1：探讨染色体沉默对小鼠DS神经表型的影响。 在体外，IPSCs将通过传统的二维神经元培养分化为神经元 技术和三维有机体。脑有机体是一种最近开发的工具，它已经被 被证明是一个有用的人脑发育模型，并已被用于研究大脑的紊乱 发展。将比较来自具有两个和三个Cl.21功能副本的IPSCs的神经元 DS神经细胞被认为具有的表型。其中包括胶质细胞与神经元比率的增加，改变 树突棘形态改变，线粒体形态改变。立体文化将被用来 研究较少探索的病理，如皮质板层的改变。这一目标还将解决 有丝分裂后细胞是否支持染色体沉默是一个重要的治疗问题。 目的2：相同的染色体沉默系统将被用于研究阿尔茨海默氏症相关 神经元表型。包括细胞内和细胞外淀粉样蛋白沉积以及细胞内淀粉样蛋白沉积。 过度磷酸化的tau沉积。由于与一般智力残疾相比，其发病时间相对较晚， 阿尔茨海默病的DS成分最适合进行治疗干预。研究的效果 阿尔茨海默病表型的染色体沉默为阿尔茨海默病的发病机制提供了一个强有力的模型 这也使这一新颖的策略更接近于治疗学。 这项提议试图利用一种新的染色体沉默技术来更好地模拟人类神经 DS和相关AD的表型。

Project Summary: Down syndrome (DS), or trisomy 21, is the leading genetic cause of intellectual disability in children, with approximately 1 in 700 live births carrying an extra copy of chromosome 21. Compared to less common single gene disorders, DS pathogenesis is still poorly understood. Treatment for DS would require either identification of molecular pathways to target with conventional therapies or, potentially, chromosomal therapy to silence the many possibly disruptive genes on the trisomic chromosome. One window of therapeutic intervention lies in the Alzheimer’s disease pathology that almost all DS patients suffer from in middle age. Recently, the extra chromosome has been silenced in an inducible manner by targeted insertion of a transgene for the XIST gene into human induced pluripotent stem cells (iPSC). XIST normally silences one X chromosome in females, providing a natural mechanism of dosage compensation. Chromosomal silencing in DS cells provides a powerful isogenic and isoepigenetic model for studying DS pathology and marks the first step towards the goal of chromosomal therapy for DS patients. The proposed work will investigate the effect of silencing the extra chromosome on DS iPSC-derived neuronal cells, investigating both DS and Alzheimer- specific phenotypes. Aim 1: In order to investigate the effect that chromosomal silencing has on DS neural phenotypes in vitro, iPSCs will be differentiated into neurons using conventional two-dimensional neuronal culturing techniques and three-dimensional organoids. Cerebral organoids are a recently-developed tool that have been shown to be a useful model for human brain development, and have been used to study disorders of brain development. Neurons derived from iPSCs with two and three functional copies of Chr.21 will be compared for phenotypes that DS neural cells are thought to possess. These include an increased glia:neuron ratio, altered dendritic spine morphology, and altered mitochondrial morphology. Three-dimensional cultures will be used to investigate less well-explored pathologies such as alterations in cortical lamination. This aim will also address the important therapeutic question of whether post-mitotic cells can support chromosomal silencing. Aim 2: The same chromosomal silencing system will be used to investigate Alzheimer’s-associated neuronal phenotypes. These include intra and extracellular amyloid deposition as well as intracellular hyperphosphylated tau deposition. Due to its relatively late onset compared to general intellectual disability, the Alzheimer’s disease component of DS is most suitable for therapeutic intervention. Studying the effect of chromosomal silencing on Alzheimer’s phenotypes provides a strong model for Alzheimer’s pathogenesis while also bringing this novel strategy one step closer to therapeutics. This proposal seeks to utilize a novel chromosomal silencing technique to better model human neural phenotypes in DS and associated AD.