Hein Da Rosa Thales, Ramsey Sterling H, Lee Judy J, Kozlova Natalia Y, Zhao Zixuan, Kim Jaeyeon, Schneider Ava C, Li Sheng, Katiki Madhusudhanarao, Firestein Gary S, Murali Ramachandran, Santelli Eugenio, Stanford Stephanie M, Bottini Nunzio
Department of Medicine, Kao Autoimmunity Institute, Cedars-Sinai Medical Center, Los Angeles, California, USA.
Department of Medicine, University of California, San Diego, La Jolla, California, USA.
J Biol Chem. 2025 Jul 10;301(8):110470. doi: 10.1016/j.jbc.2025.110470.
Protein tyrosine phosphatases (PTPs) receptor type II A (R2A) are negatively regulated through oligomerization upon binding of their extracellular domains to glycosaminoglycans (GAGs) on heparan sulfate proteoglycans (HSPGs). Inactivation of receptor PTP sigma (PTPRS) by HSPGs promotes the aggressive behavior of fibroblast-like synoviocytes (FLS) in rheumatoid arthritis (RA). Blocking the binding of its N-terminal, membrane-distal immunoglobulin-like 1 and 2 (Ig1&2) domains to its GAG ligands on the HSPG syndecan-4 (SDC4) promotes PTPRS activity and reverses the pathogenic phenotype of FLS. The potential for therapeutically leveraging other PTPRS ectodomain regions is, however, unknown. We show targeting the membrane-proximal fibronectin type III-like 9 (Fn9) domain offers a novel avenue to activate PTPRS. We mapped PTPRS Fn9 as the binding site of three antibodies (Abs) (13G5, 22H8, 49F2) and characterized their effects on cells. Despite sharing similar epitopes, we found large differences in the ability of these Abs to regulate PTPRS activity. One of these, 13G5, reduced PTPRS-dependent cell migration, PTPRS co-localization with SDC4, and PTPRS oligomerization. Single-chain variable fragment Abs of 13G5 and 22H8 were similarly effective at activating cellular PTPRS as 13G5. Replacing the entire 13G5 constant region enhanced its binding and cellular activity, indicating the Ab's potency can be optimized via isotype engineering. Treatment of cells with recombinant Fn9 protein acted as a decoy, disrupting PTPRS colocalization with SDC4 and oligomerization, and inhibiting FLS migration. Finally, significant disease mitigation in mice using 13G5-derived Abs suggests a viable strategy for the generation of novel drugs for RA therapy.
蛋白酪氨酸磷酸酶（PTP）IIA型受体（R2A）在其细胞外结构域与硫酸乙酰肝素蛋白聚糖（HSPG）上的糖胺聚糖（GAG）结合后，通过寡聚化受到负调控。HSPG使受体型蛋白酪氨酸磷酸酶σ（PTPRS）失活，促进类风湿关节炎（RA）中成纤维样滑膜细胞（FLS）的侵袭性行为。阻断其N端、膜远端免疫球蛋白样结构域1和2（Ig1&2）与HSPG syndecan-4（SDC4）上的GAG配体的结合，可促进PTPRS活性，并逆转FLS的致病表型。然而，利用其他PTPRS胞外结构域区域进行治疗的潜力尚不清楚。我们发现靶向膜近端纤连蛋白III样结构域9（Fn9）为激活PTPRS提供了一条新途径。我们将PTPRS Fn9定位为三种抗体（Abs）（13G5、22H8、49F2）的结合位点，并表征了它们对细胞的影响。尽管这些抗体具有相似的表位，但我们发现它们调节PTPRS活性的能力存在很大差异。其中一种抗体13G5可减少PTPRS依赖性细胞迁移、PTPRS与SDC4的共定位以及PTPRS寡聚化。13G5和22H8的单链可变片段抗体在激活细胞PTPRS方面与13G5同样有效。替换整个13G5恒定区可增强其结合和细胞活性，表明可通过同种型工程优化抗体的效力。用重组Fn9蛋白处理细胞可起到诱饵作用，破坏PTPRS与SDC4的共定位和寡聚化，并抑制FLS迁移。最后，使用源自13G5的抗体在小鼠中显著减轻疾病，这为开发用于RA治疗的新型药物提供了一种可行策略。
