Pathobiology of Neurodegeneration in C9ORF72 Repeat Expansion

在这份名为“C9ORF72重复扩增神经退行性变的病理生物学”的P01再申请中，我们试图评估C9ORF72重复扩增的机制，C9ORF72重复扩增是肌萎缩性侧索硬化症（ALS）和额颞叶痴呆（FTD）的最常见原因，以改善c9FTD/ALS患者的诊断和预后。我们组建了一个世界级的团队，汇集了神经学、遗传学、神经病理学和细胞生物学方面的专业知识，他们紧密合作，并拥有所有资源。我们在阐明扩增的重复RNA转录物和表观遗传变化如何分别驱动c9FTD/ALS的毒性和单倍功能不全方面取得了重大进展，发现重复扩增的大小确实影响疾病的严重程度，并鉴定了c9FTD/ALS患者血液中可检测到的潜在生物标志物。我们现在提供的证据表明，在C9ORF72突变携带者的脑组织、成纤维细胞和血液中可以检测到赖氨酸9处组蛋白3的异常甲基化。我们还发现TMEM106B是C9FTD/ALS疾病表型的第一个遗传修饰因子。利用梅奥诊所神经内科的优势，我们开始对44个C9ALS谱系进行纵向研究，以确定扩展大小、组蛋白赖氨酸残基三甲基化、mRNA表达水平和TMEM106B基因型是否与c9FTD/ALS的表型变异相关。此外，我们已经生产并鉴定了在细胞和动物模型以及人体组织中检测五种重复相关的非atg （RAN）翻译肽[poly(GP), poly(GA), poly(GR), poly（PA）和poly(PR)]的关键抗体。我们还提供证据表明，泛素-2、tau和p62/sequestosome存在于不同脑区和脊髓的神经元包裹体中，这表明除了TDP-43之外，这些蛋白可能在伴有TDP-43病理的额颞叶变性（FTLD-TDP）的发病机制中发挥作用。我们提供的初步数据表明，tdp -43阴性/ c9rant阳性的神经元包涵体也可以用二甲基精氨酸抗体检测到，这表明一种与非组蛋白甲基化有关的新的疾病机制。简而言之，我们的多学科研究将提高对c9orf72相关神经变性的理解，识别潜在的生物标志物和治疗靶点，并开发令人信服的脑、生物流体和活检资源，以帮助未来的药物发现。

DESCRIPTION (provided by applicant): In this P01 resubmission proposal entitled "Pathobiology of Neurodegeneration in C9ORF72 Repeat Expansion," we seek to evaluate the mechanisms of C9ORF72 expanded repeats, the most common cause of amyotrophic lateral sclerosis (ALS) and front temporal dementia (FTD), to improve the diagnosis of and prognosis for patients suffering from c9FTD/ALS. We have assembled a world-class team combining expertise in neurology, genetics, neuropathology, and cell biology that has worked closely together and has all resources in place. Our significant progress to elucidate how expanded repeat RNA transcripts and epigenetic changes may respectively drive toxicity and haploinsufficiency in c9FTD/ALS has led to the discovery that repeat expansion size does affect disease severity, and the identification of a potential biomarker detectable in blood of c9FTD/ALS patients. We now present evidence that aberrant methylation of histone 3 at lysine 9 is detectable in brain tissue, fibroblasts and blood of C9ORF72 mutation carriers. We also identified TMEM106B as the first genetic modifier of disease phenotype in C9FTD/ALS. In drawing upon the strengths of the Mayo Clinic Neurology Department, we have begun longitudinal studies of 44 C9ALS pedigrees to determine whether expansion size, tri-methylation of histone lysine residues, mRNA expression levels and TMEM106B genotypes, correlate with phenotypic variability in c9FTD/ALS. In addition, we have since produced and characterized antibodies critical for detecting each of the five repeat-associated non-ATG (RAN) translation peptides [poly(GP), poly(GA), poly(GR), poly(PA) and poly(PR)] in cell and animal models as well as human tissue. We also provide evidence that ubiquilin-2, tau and p62/sequestosome are present in neuronal inclusions in various brain regions and spinal cord, indicating that these proteins, in addition to TDP-43, may play a role in pathogenesis of front temporal lobar degeneration with TDP-43 pathology (FTLD-TDP). We provide preliminary data that TDP-43-negative/C9RANT-positive neuronal inclusions can also be detected with antibodies to dimethylarginine, suggesting a new disease mechanism involving non-histone protein methylation. Simply put, our multi-disciplinary studies will improve understanding of C9ORF72-related neurodegeneration, identify potential biomarkers and therapeutic targets, and develop a compelling brain, biofluid and biopsy resource to aid future drug discovery.