Probing Post-Translational Modification in Neurodegenerative Protein Aggregation with a Novel Antibody-Based Technology

利用基于抗体的新型技术探索神经退行性蛋白质聚集的翻译后修饰

• 批准号: MR/S033947/1
• 负责人: Francesco Aprile
• 金额: $ 153.37万
• 依托单位: Imperial College London
• 依托单位国家: 英国
• 项目类别: Fellowship
• 财政年份: 2020
• 资助国家: 英国
• 起止时间: 2020 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=MR%2FS033947%2F1
• 关键词: Probing; Post; Translational; Modification; Neurodegenerative

Dementia, including Alzheimer's disease, is among the most common causes of death in the U.K. Worldwide, it is estimated that 50 million people suffer from dementia, and that this number will rise to 150 million by 2050 unless effective treatments are found. The formation of amyloid aggregates is a hallmark of many of these pathologies. In the last decade, research efforts have focused on understanding the aggregation process and the associated toxicity caused by monitoring amyloid formation of a single protein in isolation under highly controlled conditions. This approach helped unveil the physical phenomena that regulate amyloid formation; nevertheless, often it provides overly simplistic descriptions of the process, as it fails to account for the contributions of many coordinated events that occur in the human body. The most striking evidence of this fact is the observation that aggregates formed in vitro are significantly different from those observed in patients, which, for example, show multiple chemical modifications (also called post-translational modifications or PTMs), which likely arise as a result of the interactions of these proteins within the complex environment of the brain.In theory, antibodies are ideal tools to accurately investigate amyloid processes, as they can be used for both highly quantitative in vitro studies and imaging of complex biological samples. As such, antibodies could, in principle, enable a high level of interdisciplinarity, which is necessary to extrapolate complex information intricate biological systems. Nevertheless, despite their potential, the use of antibodies in the amyloid field is currently hindered by several challenges linked to their production. In particular, current antibody-discovery strategies do not always allow one to target specific protein regions (i.e. epitopes) a priori or to target specific chemical features (such as modified epitopes) or conformations (such as aggregated).As a UKRI Future Leader Fellow at the Department of Chemistry of Imperial College London, my goal will be to develop an innovative antibody-discovery platform, which combines molecular and chemical biology with protein design. I will exploit this platform to characterise the mechanisms of formation and nature of amyloids formed in vivo. In particular, I will use this platform to generate antibodies to understand the role of PTMs in the amyloid aggregation process.A vast body of literature has identified PTMs of amyloid proteins in people affected by dementia. As the exact role of PTMs in amyloid formation and toxicity has yet to be determined, current drug discovery approaches generally disregard PTMs and are based on unmodified proteins. Nevertheless, such drug discovery approaches have yet to lead to a cure. PTMs represent an unexplored opportunity for therapeutic intervention and diagnosis of dementia. Despite their clinical promise, PTMs are difficult to characterise in biological contexts due to the complex makeup of in vivo samples. The high specificity and affinity afforded by antibodies make them perfect probes for identifying and localising PTMs in heterogeneous contexts.To understand the relevance of PTMs in dementia, I propose to generate antibodies that target specific PTM-amyloids, which is currently not possible with state-of-the-art methodology. I will then use these antibodies to study aggregates from biological samples and to determine which modifications are responsible for the formation and toxicity of these self-assemblies. To do so, I will combine biophysical methods (such as protein aggregation studies), imaging on biological samples, and C. elegans studies.The results of this study will enable the identification of new pathological mechanisms and biomarkers towards novel diagnostic and therapeutic approaches against dementia.

痴呆症，包括阿尔茨海默病，是英国最常见的死亡原因之一。据估计，全球有5000万人患有痴呆症，到2050年，这一数字将上升到1.5亿，除非找到有效的治疗方法。淀粉样蛋白聚集体的形成是许多这些病理的标志。在过去的十年中，研究工作集中在了解聚集过程和相关的毒性，通过监测在高度控制的条件下分离的单个蛋白质的淀粉样蛋白形成。这种方法有助于揭示调节淀粉样蛋白形成的物理现象；然而，它往往对这一过程提供了过于简单化的描述，因为它没有考虑到发生在人体中的许多协调事件的贡献。这一事实最令人震惊的证据是，在体外形成的聚集体与在患者中观察到的聚集体明显不同，例如，在患者中观察到的聚集体表现出多种化学修饰（也称为翻译后修饰或PTMs），这可能是这些蛋白质在复杂的大脑环境中相互作用的结果。理论上，抗体是准确研究淀粉样蛋白过程的理想工具，因为它们既可以用于高定量的体外研究，也可以用于复杂生物样品的成像。因此，原则上，抗体可以实现高水平的跨学科，这对于推断复杂的信息和复杂的生物系统是必要的。然而，尽管它们具有潜力，抗体在淀粉样蛋白领域的应用目前受到与它们的生产相关的几个挑战的阻碍。特别是，目前的抗体发现策略并不总是允许一个人先验地靶向特定的蛋白质区域（即表位）或靶向特定的化学特征（如修饰的表位）或构象（如聚集）。作为伦敦帝国理工学院化学系UKRI未来领导者研究员，我的目标是开发一个创新的抗体发现平台，将分子和化学生物学与蛋白质设计相结合。我将利用这个平台来描述体内形成的淀粉样蛋白的形成机制和性质。特别是，我将利用这个平台来生成抗体，以了解PTMs在淀粉样蛋白聚集过程中的作用。大量文献已经在痴呆症患者身上发现了淀粉样蛋白的ptm。由于PTMs在淀粉样蛋白形成和毒性中的确切作用尚未确定，目前的药物发现方法通常不考虑PTMs，而是基于未修饰的蛋白质。然而，这种药物发现方法尚未导致治愈。PTMs为痴呆症的治疗干预和诊断提供了一个尚未开发的机会。尽管具有临床前景，但由于体内样品的复杂组成，PTMs很难在生物学背景下进行表征。抗体的高特异性和亲和力使其成为在异质环境中识别和定位PTMs的完美探针。为了了解ptm与痴呆的相关性，我建议产生针对特定ptm淀粉样蛋白的抗体，这是目前最先进的方法无法实现的。然后，我将使用这些抗体来研究来自生物样品的聚集体，并确定哪些修饰对这些自组装的形成和毒性负责。为此，我将结合生物物理方法（如蛋白质聚集研究）、生物样品成像和秀丽隐杆线虫研究。这项研究的结果将有助于识别新的病理机制和生物标志物，为新的诊断和治疗痴呆症的方法。

项目成果

A Chemical Mutagenesis Approach to Insert Post-translational Modifications in Aggregation-Prone Proteins.

• DOI: 10.1021/acschemneuro.2c00077
• 发表时间: 2022-06-15
• 期刊: ACS CHEMICAL NEUROSCIENCE
• 影响因子: 5
• 作者: Ge, Ying;Masoura, Athina;Yang, Jingzhou;Aprile, Francesco A.
• 通讯作者: Aprile, Francesco A.