4-OCTYL ITACONATE AS A METABOLITE DERIVATIVE INHIBITS INFLAMMATION IN ARTHRITIS

Background: Rheumatoid arthritis (RA) is the most prevalent autoimmune-mediated joint disease. Fibroblast-like synoviocytes (FLS) support disease pathology through the formation of a hypertrophic and invasive synovial membrane which cannot be sufficiently controlled by neither synthetic nor biologic disease-modifying antirheumatic drugs. In addition, FLS participate in complex crosstalk with immune cells, exacerbating progression of disease. Objectives: This project examined and validated the anti-inflammatory effects of the natural glucose metabolite itaconate-derivative 4-octyl itaconate (4-OI) both in vitro and in vivo models of RA. Methods: RA FLS were harvested from the synovial fluid mononuclear cells (SFMC) of patients with established RA (esRA). Monocultures of synovial fluid derived FLS (SF-FLS) (n=7) and autologous co-cultures of SF-FLS and autologous peripheral blood mononuclear cells (PBMC) were cultured with and without 4-OI, corticosteroids, or anti-TNFα (n=7). Subsequently, analyses by flow cytometry, Western Blotting, MTT assay, and ELISA were performed. The 4-OI downstream target-protein Nrf2 was genetically deleted in immortalized FLS (FLS-KO) generated by CRISPR. Furthermore, 4-OI or vehicle-treated control was administrated systemically via intraperitoneal injections (IP) in a collagen induced arthritis (CIA) mouse model (n=7) or intra-articular in the monoarticular mBSA antigen-induced arthritis model (n=5). Arthritis severity was determined by clinical scores and caliper measurements of swollen joints. At day 71, CIA-mice were evaluated by clinical assessment, micro-CT, and mesoscale analyses of plasma. P-values <0.05 were considered statistically significant. Results: In vitro , 4-OI treatment mediated a 30 % decrease in MCP-1 secretion (p<0.05) in cultured SFMC and SF-FLS, and a 50% concomitant increase in the gene expression of the downstream protein HO-1. Furthermore, KO of Nrf2, the target of 4-OI, eliminated the effect of 4-OI addition in SF-FLS cultures. 4-OI reduced the MCP-1 secretion by 84%, anti-TNF by 41%, and corticosteroids by 54% (all p<0.01) in autologous RA SF-FLS and FLS+PBMC co-cultures. The changes in cytokine production were not attributable to a reduction in the proportion of inflammatory FLS (CD34-PDPN+THY1+), decreased viability or proliferation. We further investigated the potential effect of 4-OI treatment in two in vivo models of inflammatory arthritis. In vivo , systemic (IP) administration of 4-OI mediated a significant decrease in plasma IL-6 (p<0.05) without significantly ameliorating the number of swollen joints or erosive status over 71 days. However, when 4-OI was administrated directly into the joint (intraarticular), it reduced joint swelling as early as 2 days after administration (p<0001). Conclusion: Taken together, these data suggest that 4-OI treatment ameliorates inflammation in models of inflammatory arthritis mainly in FLS driven disease pathology. In FLS, 4-OI treatment led to a Nrf2-dependent decreased chemokine production. Overall, human in vitro data and murine in vivo data supports that Nrf2-targeting could be a useful and promising therapeutic strategy in RA, directly affecting pathological FLS. REFERENCES: NIL. Acknowledgements: We thank Karin Skovgård Sørensen, Sulaiman Hussein Kassem, and Vivien Shack (Department of Biomedicine, Aarhus University). Medical doctors and nurses at the Department of Rheumatology, Aarhus University Hospital for helping to collect the patient samples. The authors kindly acknowledge a generous grant from the Danish Rheumatoid Association (R177-A6156) and the Kennedy Trust for Rheumatology Research Senior Research Fellowship. Disclosure of Interests: None declared. DOI: 10.1136/annrheumdis-2024-eular.3341 Keywords: Synovium, Anti-Inflammatory Agents, Non-Steroidal, Fibroblasts, Animal Models Citation: , volume 83, supplement 1, year 2024, page 591Session: Inflammatory arthritis (Poster View)

Synovium, Anti-Inflammatory Agents, Non-Steroidal, Fibroblasts, Animal Models