Paula David, Tom Macleod, Ala Altaie, Yu Shi, Kerem Abacar, Sami Giryes, Gabrielle de Mello Santos, Payal Ganguly, Mark Harland, Chi Wong, Andrew Scarsbrook, Paul Emery, Kulveer Mankia, Shouvik Dass, Andrea Di Matteo, Benazir Saleem, Cédric Duval, Robert Ariëns, Dennis McGonagle
{"title":"Augmentation of immunothrombosis as a key mechanism underlying JAK inhibition associated hypercoagulability in rheumatoid arthritis.","authors":"Paula David, Tom Macleod, Ala Altaie, Yu Shi, Kerem Abacar, Sami Giryes, Gabrielle de Mello Santos, Payal Ganguly, Mark Harland, Chi Wong, Andrew Scarsbrook, Paul Emery, Kulveer Mankia, Shouvik Dass, Andrea Di Matteo, Benazir Saleem, Cédric Duval, Robert Ariëns, Dennis McGonagle","doi":"10.1016/j.ard.2025.09.002","DOIUrl":null,"url":null,"abstract":"<p><strong>Objectives: </strong>Venous thromboembolism (VTE), following Janus kinase inhibitors treatment (JAKi), is poorly understood in rheumatoid arthritis (RA). We investigated whether JAKi augmented immune cell-driven clotting or immunothrombosis in RA.</p><p><strong>Methods: </strong>Peripheral blood leukocytes (PBLs) isolated from patients with RA and healthy controls were treated with various JAKi classes, before stimulation with Toll-like receptor (TLR)-4 (lipopolysaccharide [LPS]) or TLR3 (polyinosinic-polycytidylic acid-poly (I:C)) agonists. Conditioned supernatants were used in plasma turbidity assays to evaluate clot formation and lysis dynamics, while bulk RNA sequencing, enzyme-linked immunosorbent assay, and bead-based immunoassays were used to explore immunothrombosis mechanisms.</p><p><strong>Results: </strong>Turbidity analyses showed that conditioned media from PBLs treated with LPS and tofacitinib significantly accelerated clot formation when compared to LPS alone, and this effect was tissue factor pathway dependent and accompanied by elevations in immunothrombotic cytokines, including tumour necrosis factor α, interleukin (IL)-1β, and IL-6. PBLs from patients with active RA exhibited significantly greater immunothrombotic potential compared to those with low disease activity, despite comparable baseline cytokine levels. RNA sequencing analysis revealed significant pathway enrichment in tofacitinib/LPS-treated PBLs, including activation of Nuclear Factor (NF)-κB pathways, increased tissue factor expression, and reduced levels of anticoagulant factors such as protein S. Pharmacological inhibition assays with 5 JAK therapies suggested that JAK1/tyrosine kinase 2-dependent effect underscored increased thrombosis but selective JAK3 inhibition did not reproduce the prothrombotic effects. Finally, patients with RA with JAK-associated pulmonary embolism showed interstitial changes compatible with immunothrombosis in 4/6 (67%).</p><p><strong>Conclusions: </strong>Immunothrombosis offers a novel explanation for JAKi-associated VTE in RA.</p>","PeriodicalId":8087,"journal":{"name":"Annals of the Rheumatic Diseases","volume":" ","pages":""},"PeriodicalIF":20.6000,"publicationDate":"2025-09-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Annals of the Rheumatic Diseases","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.1016/j.ard.2025.09.002","RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"RHEUMATOLOGY","Score":null,"Total":0}
引用次数: 0
Abstract
Objectives: Venous thromboembolism (VTE), following Janus kinase inhibitors treatment (JAKi), is poorly understood in rheumatoid arthritis (RA). We investigated whether JAKi augmented immune cell-driven clotting or immunothrombosis in RA.
Methods: Peripheral blood leukocytes (PBLs) isolated from patients with RA and healthy controls were treated with various JAKi classes, before stimulation with Toll-like receptor (TLR)-4 (lipopolysaccharide [LPS]) or TLR3 (polyinosinic-polycytidylic acid-poly (I:C)) agonists. Conditioned supernatants were used in plasma turbidity assays to evaluate clot formation and lysis dynamics, while bulk RNA sequencing, enzyme-linked immunosorbent assay, and bead-based immunoassays were used to explore immunothrombosis mechanisms.
Results: Turbidity analyses showed that conditioned media from PBLs treated with LPS and tofacitinib significantly accelerated clot formation when compared to LPS alone, and this effect was tissue factor pathway dependent and accompanied by elevations in immunothrombotic cytokines, including tumour necrosis factor α, interleukin (IL)-1β, and IL-6. PBLs from patients with active RA exhibited significantly greater immunothrombotic potential compared to those with low disease activity, despite comparable baseline cytokine levels. RNA sequencing analysis revealed significant pathway enrichment in tofacitinib/LPS-treated PBLs, including activation of Nuclear Factor (NF)-κB pathways, increased tissue factor expression, and reduced levels of anticoagulant factors such as protein S. Pharmacological inhibition assays with 5 JAK therapies suggested that JAK1/tyrosine kinase 2-dependent effect underscored increased thrombosis but selective JAK3 inhibition did not reproduce the prothrombotic effects. Finally, patients with RA with JAK-associated pulmonary embolism showed interstitial changes compatible with immunothrombosis in 4/6 (67%).
Conclusions: Immunothrombosis offers a novel explanation for JAKi-associated VTE in RA.
期刊介绍：
Annals of the Rheumatic Diseases (ARD) is an international peer-reviewed journal covering all aspects of rheumatology, which includes the full spectrum of musculoskeletal conditions, arthritic disease, and connective tissue disorders. ARD publishes basic, clinical, and translational scientific research, including the most important recommendations for the management of various conditions.
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