Preserving cellular aspects of aging in patient-specific models of ALS

在ALS患者特异性模型中保留衰老的细胞方面 项目摘要 诱导多能干细胞（iPSC）生物学为人类体外神经退行性疾病提供了巨大的希望 因为这些细胞可以产生人脑中的任何细胞，一个活的“虚拟大脑”， 实验操作，具有与个体患者完全相同的基因组成， 先前在尸检和临床样本中鉴定的神经退行性表型。这些 “患者特异性”体外测试系统使靶点发现、药物筛选和治疗证据成为可能。 在患者细胞中的概念研究比目前可能的翻译过程早得多。 尽管有这些独特的优势，年龄作为一个关键的致病风险因素的保护目前是一个主要的问题。 这些制度的局限性。这部分是由于1）细胞中与年龄相关的特征的丧失， 再生到胚胎状态，和2）分化方案中的缺陷， 从胚胎细胞中产生成熟的神经元。直接的转录因子介导的重编程， 被称为“谱系转换”的患者成纤维细胞向诱导运动神经元的转化代表了一种替代方法， 在体外产生人类神经元的方法。来自我们实验室、合作者和其他人的新数据显示， 通过谱系转换产生的神经元更好地保留了年龄相关和疾病相关的缺陷。 在这项为期1年的可行性研究中，我们将比较由谱系转换产生的运动神经元， 成纤维细胞（BMP-MN），以及通过从iPSC定向分化产生的那些，所述iPSC从成纤维细胞（BMP-MN）重编程。 相同的成纤维细胞样品（iPSC-MN）。成纤维细胞样品来自患有GGGGCC的患者。 C9 ORF 72基因中的六核苷酸重复扩增突变，已知其引起一种形式的 肌萎缩侧索硬化症（ALS）和额颞叶痴呆，以及匹配的对照。第一个假设是 这意味着，EST-MN转录组将与死后组织的转录组和来自 ALS患者我们的合作者Verge Genomics创建了一个创新的，大数据驱动的ALS基因 使用来自39项ALS相关研究的公共和专有基因表达数据， 将在项目中使用。 核质转运缺陷已成为一种表型，其中年龄和C9- ALS相关 差异已经确定。 因此，我们的第二个假设是，M-MN将具有显著更多的 与iPSC-MN相比，年龄相关的核质转运缺陷明显， 免疫荧光共聚焦显微镜。综合起来，这两项研究将证明这一原则， ADM-MN创造了上级ALS体外测试系统，保留了衰老的重要疾病相关方面 对iPSC银行和疾病建模领域产生了巨大的影响。AcuraStem公司发展人类 用于临床前人体验证其自身CNS治疗方法的细胞模型和测定，以及其 发展伙伴。

Preserving cellular aspects of aging in patient-specific models of ALS Project Summary Induced pluripotent stem cell (iPSC) biology holds great promise for human ​in vitro ​neurodegenerative disease modeling because these cells can give rise to any cell in the human brain, a living “virtual brain” amenable to experimental manipulation, having the exact same genetic makeup as individual patients and displaying neurodegenerative phenotypes previously identified in postmortem and clinical samples. These “patient-specific” ​in vitro testing systems enable target discovery, drug screening and therapeutic proof of concept studies in patient cells much earlier in the translational process than is currently possible. Despite these unique advantages, the preservation of age as a key pathogenic risk factor is presently a major limitation of these systems. This is in part due to 1) the loss of age-related characteristics in cells that are rejuvenated to an embryonic state, and 2) to deficiencies in the differentiation protocols that are unable to produce mature neurons from embryonic cells. The direct, transcription factor mediated reprogramming, also known as “lineage conversion”, of patient fibroblasts into induced motor neurons represents an alternative approach for generating human neurons ​in vitro. New data from our lab, our collaborators’ and others’ show that neurons generated through lineage conversion better retain age-related and disease-associated deficits. In this 1-year feasibility study we will compare motor neurons generated by lineage conversion from fibroblasts (fib-MNs), with those generated by directed differentiation from iPSCs reprogrammed from the same fibroblast samples (iPSC-MNs). The fibroblast samples are from patients having the GGGGCC hexanucleotide repeat expansion mutation in the ​C9ORF72 ​gene, which is known to cause a form of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia, and matched controls. Our 1st hypothesis is that fib-MN transcriptomes will be significantly more similar to those of post-mortem tissue and those from ALS patients. Our collaborator Verge Genomics has created an innovative, big-data-driven ALS gene expression signature using ​public and proprietary gene expression data from 39 ALS-relevant studies that we will use in the project. Nucleocytoplasmic transport defects have emerged as one phenotype where both age and ​C9-​ALS related differences have been identified. ​Therefore, our ​2nd hypothesis is that fib-MNs will have significantly more pronounced age-related nucleocytoplasmic transport defects than iPSC-MNs as measured by immunofluorescence confocal microscopy. Taken together, these two studies would prove the principle that fib-MNs create superior ALS in-vitro testing systems retaining important, disease-relevant aspects of aging having tremendous impact ​on the iPSC banking and disease modeling fields. AcuraStem Inc. develops human cell models and assays for preclinical human validation of its own CNS therapeutics, as well as those of its development partners.