Ab0067 mode of action and target investigation of the new anti-rheumatic drug rabeximod.

Background Rheumatoid arthritis (RA) is a chronic systemic inflammatory autoimmune disease which primarily affects the joints [1] . Numerous agents are currently used to modify the natural course of the disease, however, several RA patients still do not reach clinical remission [2] . A recently developed quinoxaline analogue named Rabeximod (RBM) showed clinical benefits comparable to TNF-α blockers in a mouse model of collagen antibody-induced arthritis [3] . Moreover, in a phase IIa clinical trial a significantly higher clinical efficacy of RBM + Methotrexate versus Methotrexate alone has also been confirmed. Objectives Since RBM’s exact biologic target has not been entirely established yet, our objective is to investigate its mode of action at the cellular and molecular level. Methods Two in vitro models were developed: i) Toll-like receptor (TLR)-4 and TLR-2–stimulated peripheral blood mononuclear cells (PBMCs) cultured for 24-hours and ii) Monocytes derived-macrophages (MDMs), cultured for 96-hours in persisting inflammation conditions mimicking the different phases of ACPA-positive rheumatoid synovial inflammation: recruitment, initiation, development, and persistence of the inflammatory process. PBMCs-transcriptomic analysis was performed using a NanoString nCounter SPRING Profiler (customised panel). Cytokine production was measured by ELISA. Results RBM induced dose-dependent cell mortality. Among PBMCs, monocyte viability was preferentially affected compared to lymphocytes. RBM treatment reduced IL-6 and TNF-α production both in TLR4/TLR2-stimulated PBMCs and MDMs supernatants at 24h (PBMCs) and 10-, 24-, 48- and 72-hour time-points (MDMs). RBM-induced down-regulation of IL-6 and TNF-α in PBMCs was also obtained when PBMCs were pre-incubated for 7 hours prior to TLR activation (without additional RBM), suggesting an internalisation process. After 24h, RBM induced the downregulation of several pro-inflammatory genes in TLR-4 and-2 stimulated PBMCs, including IL-6, TNF-α, CSF1, FCN1, and SPP1. Finally, after 24 hours, MDMs treated with RBM developed an altered morphology independently of the frequency of treatment exposure and the stage of inflammation. Conclusion Overall, our data suggest that RBM acts predominantly on monocytes/macrophages and can inhibit the production of pro-inflammatory molecules playing a crucial role in RA. References [1]Scherer HU, Häupl T, Burmester GR. The etiology of rheumatoid arthritis. J Autoimmun. Published online 2020. doi:10.1016/j.jaut.2019.102400 [2]Guo Q, Wang Y, Xu D, Nossent J, Pavlos NJ, Xu J. Rheumatoid arthritis: Pathological mechanisms and modern pharmacologic therapies. Bone Res. 2018;6(1). doi:10.1038/s41413-018-0016-9 [3]Hultqvist M, Nandakumar KS, Björklund U, Holmdahl R. Rabeximod reduces arthritis severity in mice by decreasing activation of inflammatory cells. Ann. Rheum Dis. 452010;69(8):1527-1532. doi:10.1136/ard.2009.121178 Acknowledgements: NIL. Disclosure of Interests Giulia Maria Ghirardi: None declared, Alessandra Nerviani: None declared, Liliane Fossati-Jimack: None declared, Elena Pontarini: None declared, Paola Italiani: None declared, Federico Pratesi: None declared, Sally Abdelmoaty Employee of: Cyxone AB, Carl Högerkorp Employee of: Cyxone AB, Costantino Pitzalis: None declared.