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

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，除非找到有效的治疗方法，否则到2050年，这一数字将上升到1.5亿。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，研究工作的重点是了解聚集过程和相关的毒性，这些过程是在高度可控的条件下分离监测单一蛋白质的淀粉样蛋白形成所引起的。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，它通常对这一过程提供了过于简单化的描述，因为它未能解释发生在人体内的许多协调事件的贡献。这一事实最显著的证据是观察到，体外形成的聚集体与患者观察到的聚集体显著不同，例如，患者体内观察到的聚集体显示出多种化学修饰(也称为翻译后修饰或PTM)，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，因为它们可以用于高定量的体外研究和复杂生物样本的成像。因此，抗体在原则上可以实现高度的跨学科，这是推断复杂信息错综复杂的生物系统所必需的。然而，尽管有潜力，抗体在淀粉样蛋白领域的使用目前受到与其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人们事先针对特定的蛋白质区域(即表位)，或者针对特定的化学特征(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的UKRI未来领导者研究员，我的目标将是开发一个创新的抗体发现平台，它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在体内形成的淀粉样蛋白的形成机制和性质。特别是，我将利用这个平台产生抗体，以了解PTM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中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忽略PTMS，而是基于未经修饰的蛋白质。然而，这种药物发现方法尚未带来治愈方法。PTM为痴呆症的治疗干预和诊断提供了一个尚未探索的机会。尽管PTM在临床上很有希望，但由于体内样本的复杂组成，很难在生物学背景下描述它们的特征。抗体提供的高特异性和亲和力使它们成为在不同环境中识别和定位PTM的完美探针。为了了解PTM在痴呆症中的相关性，我建议生成针对特定PTM-淀粉样蛋白的抗体，这在目前最先进的方法学中是不可能的。然后，我将使用这些抗体来研究生物样本中的聚集体，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)、生物样本成像和线虫研究。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，为治疗痴呆症提供新的诊断和治疗方法。

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.