OBJECTIVES: Macrophages possess widespread pro-inflammatory, destructive, and remodelling capabilities that can critically contribute to acute and chronic diseases, such as rheumatoid arthritis (RA). Continuous monitoring and measurement of selective counteraction of macrophage activity in patients require a sensitivity and non-invasive marker. We characterised the VICM (citrullinated and MMP degraded vimentin fragment) biomarker by investigating the release from in vitro activated macrophages and by monitoring the change in serum levels after treatment with the anti-GM-CSFRα-mAb (mavrilimumab).
METHODS: Peripheral blood mononuclear cells were isolated, and lipopolysaccharide (LPS) was used to activate the macrophages and calcium chloride (CaCl2) was used to facilitate the citrullination process of vimentin. Supernatants, cell lysates, was collected and analysed by ELISA, and western blotting. RA patients were treated with mavrilimumab+methotrexate or methotrexate alone in a phase 2b study (NCT01706926) once every two weeks for 24 weeks. Serum levels of VICM were measured at baseline and multiple time points post-treatment. In addition, whole blood expression of peptidylarginine deiminase-2 (PAD-2) and matrix metalloproteinase-9 (MMP-9) transcripts were tested by quantitative reverse transcriptase PCR assays at day 0 and day 169 post-treatment.
RESULTS: VICM levels were significantly higher at day 5 and 8 in supernatants of activated macrophages compared to controls (p\u003c0.01), which was confirmed by Western blot. In RA patients, VICM correlated with disease activity (DAS28), modified total sharp score (mTSS), joint space narrowing (JSN), joint erosions and CRP at baseline. VICM was dose-dependently and significantly (p\u003c0.01) inhibited by mavrilimumab. This suppression of VICM serum levels was supported by a decreased expression of PAD2 and MMP9 transcripts in patients treated with mavrilimumab.
CONCLUSIONS: These data verified that VICM is released by activated macrophages. Treatment of RA patients with mavrilimumab significantly reduced release of VICM and peptidylarginine deiminases-2 (PAD-2) gene expression indicating that mavrilimumab indeed is targeting activated macrophages and that VICM may be a novel blood-based marker of anti-GM-CSF response.
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