Sustained xanthine oxidase inhibitor treat to target urate lowering therapy rewires a tight inflammation serum protein interactome

Abstract

Background: Effective xanthine oxidoreductase inhibition (XOI) urate-lowering treatment (ULT) to target significantly reduces gout flare burden and synovitis between 1-2 years therapy, without clearing all monosodium urate crystal deposits. Paradoxically, treat to target ULT is associated with increased flare activity for at least 1 year in duration on average, before gout flare burden decreases. Since XOI has anti-inflammatory effects, we tested for biomarkers of sustained, effective ULT that alters gouty inflammation. Methods: We characterized the proteome of febuxostat-treated murine bone marrow macrophages. Blood samples (baseline and 48 weeks ULT) were analyzed by unbiased proteomics in febuxostat and allopurinol ULT responders from two, independent, racially and ethnically distinct comparative effectiveness trial cohorts (n=19, n=30). STRING-db and multivariate analyses supplemented determinations of significantly altered proteins via Wilcoxon matched pairs signed rank testing. Results: The proteome of cultured IL-1b-stimulated macrophages revealed febuxostat-induced anti-inflammatory changes, including for classical and alternative pathway complement activation pathways. At 48 weeks ULT, with altered purine metabolism confirmed by serum metabolomics, serum urate dropped >30%, to normal (<6.8 mg/dL) in all the studied patients. Overall, flares declined from baseline. Treated gout patient sera and peripheral blood mononuclear cells (PBMCs) showed significantly altered proteins (p<0.05) in clustering and proteome networks. CRP was not a useful therapy response biomarker. By comparison, significant serum proteome changes included decreased complement C8 heterotrimer C8A and C8G chains essential for C5b-9 membrane attack complex assembly and function; increase in the NLRP3 inflammasome activation promoter vimentin; increased urate crystal phagocytosis inhibitor sCD44; increased gouty inflammation pro-resolving mediator TGFB1; decreased phagocyte-recruiting chemokine PPBP/CXCL7, and increased monocyte/macrophage-expressed keratin-related proteins (KRT9,14,16) further validated by PBMC proteomics. STRING-db analyses of significantly altered serum proteins from both cohorts revealed a tight interactome network including central mediators of gouty inflammation (eg, IL-1B, CXCL8, IL6, C5). Conclusions: Rewiring of inflammation mediators in a tight serum protein interactome was a biomarker of sustained XOI-based ULT that effectively reduced serum urate and gout flares. Monitoring of the serum and PBMC proteome, including for changes in the complement pathway could help determine onset and targets of anti-inflammatory changes in response to effective, sustained XOI-based ULT. Trial Registration: ClinicalTrials.gov Identifier: NCT02579096