Moghbel Marc, Roth Aline, Baptista Daniela, Miteva Kapka, Burger Fabienne, Montecucco Fabrizio, Vuilleumier Nicolas, Mach François, Brandt Karim J
Endotelix Diagnostics Sàrl Geneva Switzerland.
Division of Cardiology, Department of Medicine, Faculty of Medicine, Foundation for Medical Research University of Geneva Geneva Switzerland.
Res Pract Thromb Haemost. 2022 Oct 24;6(7):e12828. doi: 10.1002/rth2.12828. eCollection 2022 Oct.
Antiphospholipid antibody (aPL) syndrome (APS) is an autoimmune disease characterized by the presence of antiphospholipid antibodies and thromboembolic or pregnancy complications. Although cryptic epitope R39-R43 belonging to beta-2-glycoprotein 1 (β2GP1) has been identified as the main antigenic determinant for aPLs, we have recently demonstrated that the epitope is a motif determined by the polarity, rather than by the sequence or charge of amino acids.
In the present study, we wanted to identify the association of residues needed to obtain the highest aPL affinity.
Based on the epitope R39-R43 and our identified motif, we generated a printed peptide microarray of 676 different peptides. These peptides have been then screened for their ability to interact with the plasmas from 11 well-characterized APS patients and confirmed by surface plasma resonance assay.
We identified a peptide that selectively bound immunoglobulin G (IgG) derived from APS patients with 100 times more affinity than β2GP1, Domain I, or epitope R39-R43. This peptide is able to inhibit the activity of IgG derived from APS patients in vitro. We have also generated a monoclonal IgG antibody against this peptide. Using both peptide and monoclonal antibody, we have been able to develop a fully standardized indirect colorimetric immunoassay with highly sensitivity. The identification of the optimized peptide offers a new standardized and accurate tool for diagnostics of APS. Furthermore, having increased affinity for aPL, this peptide could represent a useful tool as prevention strategy for APS and an alternative to the use of anticoagulants.
抗磷脂抗体（aPL）综合征（APS）是一种自身免疫性疾病，其特征为存在抗磷脂抗体以及血栓栓塞或妊娠并发症。尽管已确定属于β2糖蛋白1（β2GP1）的隐蔽表位R39 - R43是aPL的主要抗原决定簇，但我们最近证明该表位是由极性决定的基序，而非氨基酸序列或电荷。
在本研究中，我们希望确定获得最高aPL亲和力所需残基之间的关联。
基于表位R39 - R43和我们确定的基序，我们生成了包含676种不同肽的打印肽微阵列。然后筛选这些肽与11例特征明确的APS患者血浆相互作用的能力，并通过表面等离子体共振分析进行确认。
我们鉴定出一种肽，它与APS患者来源的免疫球蛋白G（IgG）选择性结合，其亲和力比β2GP1、结构域I或表位R39 - R43高100倍。该肽能够在体外抑制APS患者来源的IgG的活性。我们还制备了针对该肽的单克隆IgG抗体。使用该肽和单克隆抗体，我们能够开发出一种具有高灵敏度的完全标准化的间接比色免疫测定法。优化肽的鉴定为APS的诊断提供了一种新的标准化且准确的工具。此外，由于对aPL具有更高的亲和力，该肽可作为APS预防策略的有用工具以及抗凝剂使用的替代方法。
