Pathobiology of Neurodegeneration in C9ORF72 repeat expansion

在这篇题为“C9orf72重复扩张神经退行性变的病理生物学”的P01提案更新中，我们寻求改善C9orf72相关肌萎缩性侧索硬化症（ALS）和额颞叶痴呆（FTD）患者的诊断和预后。我们的综合项目和核心将通过确定“c9ALS/FTD”的治疗靶点和生物标志物来实现这一目标，并继续努力阐明C9orf72基因中G4C2重复扩增导致神经退行性变的机制。我们已经组建了一支世界级的研究团队，他们在神经学、遗传学、神经病理学、“组学”、疾病建模、生物标志物和生物信息学方面具有专业知识，他们紧密合作，并拥有所有资源。我们在确定c9ALS/FTD的病理机制方面取得了重大进展，发现从扩展的G4C2重复序列非典型翻译的poly（GR）蛋白与c9ALS/FTD患者的神经退行性相关，并导致小鼠神经元丢失，poly（GR）诱导应激颗粒的形成并损害其分解。后者可以解释c9ALS/FTD中观察到的核胞质转运缺陷，因为我们发现核胞质转运因子异常地被招募到应激颗粒中。在拟议的项目中，我们将以这些令人兴奋的发现为基础，解决需要解答的新问题，以开发有效的治疗c9ALS/FTD的方法。事实上，分别使用下一代质谱法和单核RNA测序对c9ALS患者和（G4C2）149小鼠的脑组织进行深度蛋白质组学和转录组学分析，将揭示代表c9ALS靶通路的蛋白质和基因网络。此外，通过研究重复靶向反义寡核苷酸（ASO）处理的（G4C2）149只小鼠和c9ALS患者来源的神经元培养物的蛋白质组和转录组，我们将确定对治疗有反应的蛋白质共表达模块和枢纽蛋白，从而代表潜在的治疗靶点和候选生物标志物。由于缺乏有助于ALS早期诊断、预测疾病进展和确认靶标的生物标志物，潜在ALS治疗方法的开发和测试受到了严重阻碍，因此后者是一种迫切需要。为了使这三个研究项目能够有效地实现其目标，研究核心将收集c9ALS患者的生物体液和脑组织，这也将有助于分析和整合数据，以及验证研究结果。总的来说，我们的多学科协同研究将提高我们对c9orf72相关神经变性的理解，确定潜在的生物标志物和治疗靶点，并为c9ALS领域开发全面的脑和生物流体资源。

In this P01 proposal renewal entitled “Pathobiology of Neurodegeneration in C9orf72 Repeat Expansion”, we seek to improve the diagnosis of and prognosis for patients suffering from C9orf72-associated amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Our integrated Projects and Cores will accomplish this by identifying therapeutic targets and biomarkers for “c9ALS/FTD” and continuing with our productive endeavors to elucidate the mechanisms by which G4C2 repeat expansions in the C9orf72 gene cause neurodegeneration. We have assembled a world-class team of investigators with expertise in neurology, genetics, neuropathology, “-omics”, disease modeling, biomarkers and bioinformatics that have worked closely together and have all resources in place. Our significant progress to determine the pathomechanisms underpinning c9ALS/FTD has led to the discoveries that poly(GR) proteins atypically translated from expanded G4C2 repeats correlate with neurodegeneration in c9ALS/FTD patients and cause neuronal loss in mice, and that poly(GR) induces the formation of stress granules and impairs their disassembly. The latter may account for the observed nucleocytoplasmic transport defects in c9ALS/FTD given our finding that nucleocytoplasmic transport factors are aberrantly recruited to stress granules. In the proposed projects, we will build upon these exciting findings and tackle new questions in need of answers in order to develop effective treatments for c9ALS/FTD. Indeed, deep proteomic and transcriptomic profiling, respectively, using next-generation mass spectrometry and single-nucleus RNA sequencing of brain tissues from c9ALS patients and (G4C2)149 mice will uncover protein and gene networks that represent target pathways for c9ALS. Furthermore, by interrogating the proteome and transcriptome of (G4C2)149 mice and c9ALS patient-derived neuronal cultures treated with repeat-targeting antisense oligonucleotides (ASO), we will identify protein co-expression modules and hub proteins that respond to treatment and thus represent potential therapeutic targets and candidate biomarkers. The latter represent a critical need given that the development and testing of potential ALS treatments have been seriously hampered by the lack of biomarkers to aid in the early diagnosis of ALS, to forecast disease progression, and to confirm target engagement. To enable all three research projects to effectively achieve their goals, biofluids and brain tissues from c9ALS patients will be collected by the research cores, which will also assist with analyzing and integrating data, as well as validating findings. Overall, our multi-disciplinary, synergistic studies will improve our understanding of C9orf72-related neurodegeneration, identify potential biomarkers and therapeutic targets, and develop a comprehensive brain and biofluid resource for the c9ALS field.