Proteomic and methylation profiles of liquid biopsies in early and advanced rheumatoid arthritis

Secreted proteins and cell-free DNA (cfDNA) methylation profiles show promise as liquid biomarkers for rheumatoid arthritis (RA). We integrated proteomic and cfDNA promoter-methylation data from serum and synovial fluid (SF) to identify candidate features associated with early RA (eRA; disease duration < 1 year) and advanced RA (aRA; > 2 years). The cohort comprised 21 patients with RA (13 eRA, 8 aRA) and 21 healthy controls (HC), with assay-specific sample subsets. Proteins were profiled by label-free LC–MS/MS, and promoter-level cfDNA methylation was assessed using enzymatic whole-genome methylation sequencing. Promoter-methylation features corresponding to proteomic candidates were evaluated using MCFS-ID. Exploratory serum screening identified 158 protein–contrast associations representing 133 proteins; effects were generally small, and none of 457 proteins remained significant after Benjamini–Hochberg correction. In aRA SF versus an external osteoarthritis SF dataset, 114 nominal candidates were identified, including 97 meeting q < 0.05 and |log2FC|≥ 0.5. Integrated analyses prioritized lecithin–cholesterol acyltransferase (LCAT) and angiotensinogen (AGT), with highest promoter methylation in aRA SF. These cross-sectional findings identify LCAT and AGT as candidate multi-omic liquid-biopsy markers but do not establish disease progression or causal relationship between cfDNA methylation and circulating protein abundance. Independent longitudinal cohorts, balanced SF comparators, and orthogonal assays are required for validation.