A8.08 The role of epigenetics in determining the clinical response to methotrexate for the treatment of rheumatoid arthritis. (24th February 2016) - Cregan, S
Abstract : Background and objectives: Rheumatoid Arthritis (RA) is a chronic inflammatory autoimmune disease which leads to progressive joint destruction. RA is characterised by synovial hyperplasia and invasion of the synovium into the adjacent bone and cartilage. The precise aetiology of RA is not known, but both genetic and environmental influences play a role. Methotrexate (MTX) is the most commonly used disease modifying anti-rheumatic drug in the management of RA, however its mechanism of action remains poorly understood. In this study, we aim to identify the functional effects of MTX on primary RA synovial fibroblasts (RASFC) and to determine if the epigenetic signature of RASFC/CD14 + monocytes can predict clinical response to MTX. Materials and methods: Primary RASFC were isolated from synovial biopsies obtained from patients undergoing arthroscopic examination. RASFC were cultured in the presence or absence of MTX (10–100 μM). Cell migration, invasion, viability/proliferation, adhesion and pro-inflammatory cytokine expression were assessed by: wound repairs assays, transwell matrigel TM invasion chambers, MTT/ Crystal violet cell growth assay, xCELLigence Real Time Cell Analyser and ELISA respectively. In parallel, CD14 + monocytes were isolated from peripheral blood mononuclear cells from RA patients both pre- and 12 weeks post-MTX therapy and global methylation was assessed. Results: MTX (10 μM) inhibited RASFC repopulation of the wound margins in comparison to vehicle control where migration across the wound was clearly evident (n = 7, p = 0.03). In parallel, MTX (100 μM)significantly decreased RASFC invasion (n = 7, p = 0.01) and altered cytoskeletal dynamics through inhibition of lamellopodia and filopodia formation, which are indicative of cell movement. MTX had no inhibitory effect on the expression of IL-6, IL-8 and RANTES. Importantly, we determined that the effect of MTX on RASFC migration and invasion were independent of both cell viability (p = 0.37) and proliferation (p = 0.18). Finally preliminary results demonstrated altered DNA methylation in CD14 + monocytes from RA patients (n = 11) at 3 months post MTX treatment. Conclusions: MTX inhibits RASFC migration, invasion and cytoskeletal re-arrangement, mechanisms which are critically involved in the pathogenesis of RA. Further preliminary data suggests that the effect of MTX on pro-inflammatory mechanisms in RA, maybe be mediated through alterations in epigenome.