0593: A New Cytokine Interleukin 40 Is Elevated in the Serum of Patients with Early Rheumatoid Arthritis and Associates with Autoantibodies and Neutrophil Activation

Adéla Navrátilová¹, Viktor Bečvář¹, Hana Hulejová¹, Michal Tomcik², Heřman Mann², Olga Růžičková¹, Olga Šléglová³, Jakub Závada¹, Karel Pavelka⁴, Jiří Vencovský⁵, Ladislav Šenolt⁵ and Lucie Andrés Cerezo¹, ¹Institute of Rheumatology, Prague, Czech Republic, ²Institute of Rheumatology, Department of Rheumatology, 1st Faculty of Medicine, Charles University, Prague, Czech Republic, Prague, Czech Republic, ³Institute of Rheumatology, Prague, ⁴Institute of Rheumatology, Department of Rheumatology, 1st Faculty of Medicine, Charles University, Prague, Czech Republic, Praha, Czech Republic, ⁵Institute of Rheumatology and Department of Rheumatology, First Faculty of Medicine, Charles University, Prague, Czech Republic

Background/Purpose: IL-40 is a newly described cytokine related to malignant transformation and immune system function. In our previous study, we showed that IL-40 is elevated in rheumatoid arthritis (RA) and is associated with disease activity, autoantibodies, and neutrophil extracellular traps (NETs) (1). Since neutrophils and antibodies are associated with RA development, we aimed to investigate the role of IL-40 in early stages of RA (ERA).

Methods: The IL-40 expression was analysed in the serum of treatment naïve patients with ERA (n=60) at baseline and three months after initiation of pharmacological conventional treatment. The patients met the ACR/EULAR criteria for the disease (2). The levels of IL-40 were determined in sex- and age- matched healthy controls (n=60). Concentration of IL-40, cytokines and NETosis markers (proteinase 3, PR3 and neutrophil elastase, NE) was determined by commercially available ELISA kits.Concentration of autoantibodies anti-cyclic citrullinated peptides (anti-CCP) and rheumatoid factor (RF(IgM)) was determined by routine laboratory techniques. Peripheral blood from patients with ERA (n=15) was used to obtain the neutrophils and evaluate the relationship between IL-40 and neutrophils in vitro.

Results: IL-40 was upregulated in ERA patients at baseline compared to healthy controls (p< 0.0001) and normalised after three months of the conventional pharmacological treatment (p< 0.0001). IL-40 was associated with RF (IgM), anti-CCP (both p< 0.01) and NETosis (PR3 and NE) at baseline (both p< 0.0001). The decreases in the serum IL-40 following the therapy correlated with the decrease of NETosis markers PR3 (p< 0.01) and NE (p< 0.05). Our in vitro experiments show that neutrophils from patients with ERA significantly enhanced the release of IL-40 following NETosis (p< 0.05)
or pro-inflammatory stimulation (IL-1β, IL-8 (both p< 0.05), TNF (p< 0.01) or to LPS (p< 0.05)). Neutrophils from ERA patients influenced by recombinant IL-40 upregulated the secretion of IL-1β (p< 0.05) and TNF (p< 0.05).

Conclusion: Here we show for the first time that IL-40 is elevated in ERA patients and the levels decrease following three months of conventional pharmacological therapy. Importantly, we show that neutrophils are the source of IL-40 in RA and that IL-40 is released in response to inflammation and NETosis. Our results indicate that IL-40 could be implicated in early stages of RA.

Acknowledgement: Supported by AZV-NU21-05-00276, MHCR 023728, SVV 260 523 and BBMRI-CZ LM2018125

References:
1. Navrátilová A, Andrés Cerezo L, Hulejová H, Bečvář V, Tomčík M, Komarc M, et al. IL-40: A New B Cell-Associated Cytokine Up-Regulated in Rheumatoid Arthritis Decreases Following the Rituximab Therapy and Correlates With Disease Activity, Autoantibodies, and NETosis. Front Immunol. 2021 Oct 21;12:745523.
2. Aletaha D, Neogi T, Silman AJ, et al., 2010 Rheumatoid arthritis classification criteria: an American College of Rheumatology/European League Against Rheumatism collaborative initiative. Arthritis Rheum. 2010 Sep;62(9):2569-81.

Disclosures: A. Navrátilová, None; V. Bečvář, None; H. Hulejová, None; M. Tomcik, None; H. Mann, None; O. Růžičková, None; O. Šléglová, None; J. Závada, AbbVie/Abbott, Eli Lilly, Sandoz, Novartis, Egis, UCB; K. Pavelka, MSD, Pfizer, Roche, Eli Lilly, Medac, UCB, SOBI, Biogen, Sandoz, Viatris; J. Vencovský, Abbvie, Biogen, Boehringer, Eli Lilly, Gilead, Kezar, Merck, Novartis, Octapharma, Pfizer, Takeda, UCB, Werfen, Argenx; L. Šenolt, None; L. Andrés Cerezo, None.