Centre for Misfolding Diseases, Department of Chemistry, University of Cambridge, CB2 1EW Cambridge, United Kingdom;
Centre for Misfolding Diseases, Department of Chemistry, University of Cambridge, CB2 1EW Cambridge, United Kingdom.
Proc Natl Acad Sci U S A. 2020 Jun 16;117(24):13509-13518. doi: 10.1073/pnas.1919464117. Epub 2020 Jun 3.
Protein misfolding and aggregation is the hallmark of numerous human disorders, including Alzheimer's disease. This process involves the formation of transient and heterogeneous soluble oligomers, some of which are highly cytotoxic. A major challenge for the development of effective diagnostic and therapeutic tools is thus the detection and quantification of these elusive oligomers. Here, to address this problem, we develop a two-step rational design method for the discovery of oligomer-specific antibodies. The first step consists of an "antigen scanning" phase in which an initial panel of antibodies is designed to bind different epitopes covering the entire sequence of a target protein. This procedure enables the determination through in vitro assays of the regions exposed in the oligomers but not in the fibrillar deposits. The second step involves an "epitope mining" phase, in which a second panel of antibodies is designed to specifically target the regions identified during the scanning step. We illustrate this method in the case of the amyloid β (Aβ) peptide, whose oligomers are associated with Alzheimer's disease. Our results show that this approach enables the accurate detection and quantification of Aβ oligomers in vitro, and in and mouse hippocampal tissues.
蛋白质错误折叠和聚集是许多人类疾病的标志，包括阿尔茨海默病。这个过程涉及到瞬态和异质可溶性寡聚物的形成，其中一些具有高度细胞毒性。因此，开发有效诊断和治疗工具的主要挑战是检测和定量这些难以捉摸的寡聚物。在这里，为了解决这个问题，我们开发了一种两步理性设计方法，用于发现寡聚体特异性抗体。第一步包括一个“抗原扫描”阶段，在这个阶段中，设计了一组初始抗体来结合覆盖目标蛋白整个序列的不同表位。通过体外实验，可以确定寡聚物中暴露的区域，但不包括纤维状沉积物中的区域。第二步涉及“表位挖掘”阶段，在这个阶段中，设计了第二组抗体来专门针对扫描阶段中确定的区域。我们在淀粉样β (Aβ) 肽的情况下说明了这种方法，其寡聚物与阿尔茨海默病有关。我们的结果表明，这种方法能够准确检测和定量体外和小鼠海马组织中的 Aβ 寡聚物。
