Background/Purpose: Methotrexate (MTX) is the first-line disease modifying anti-rheumatic drug for the treatment of rheumatoid arthritis (RA). However, many patients do not respond adequately or experience adverse effects, therefore identifying blood-based biomarkers that predict treatment response is a clinical priority. DNA methylation is an epigenetic marker that modifies, but does not alter, DNA sequence, and it is thought that MTX may act, at least in part, by inhibiting intracellular methyl donor status leading to DNA hypomethylation. We aimed to identify differential DNA methylation signatures in whole blood, which may be predictive of response to MTX in patients with RA.

Methods: DNA methylation was measured using the HumanMethylation450 BeadChip in DNA samples from individuals recruited to the Rheumatoid Arthritis Medication Study (RAMS), a one year observational study in the UK including patients with RA starting MTX for the first time. In RAMS, demographic and clinical data are collected prior MTX start (baseline) and at 6 months after commencing MTX. DNA was extracted from whole blood samples collected baseline and at 4 weeks from patients who, at 6 months, had a EULAR good response (n=36) or EULAR poor response (n=36) to MTX (test cohort). Differentially methylated positions (DMPs) between the baseline and 4 weeks, and between good and poor response were identified using linear regression, adjusting for gender, age, cell composition, baseline disease activity score (DAS28), and smoking status. Analyses also compared methylation with changes in DAS28 and the individual DAS28 components over 6 months. DMPs that showed significant differences in the test cohort were selected for replication by pyrosequencing in an independent group of 100 patients with both baseline and 4 week samples (replication cohort).

Results: In the test cohort, differential methylation at 2 CpG sites in samples taken at 4 weeks was associated with response status determined at 6 months (p-value <10-5). Three additional DMPs were associated with change in tender joint count, whilst three other DMPs were associated with change in swollen joint count, and a further four DMPs associated with change in C-reactive protein. One of the 4 DMPs associated with change in CRP, cg04334751, showed a trend to association in the independent replication cohort (Spearmans Rho p-value =0.058). This CpG site is located close to microRNA, mir182.

Conclusion: These preliminary results suggest DNA methylation may provide a biomarker of MTX response but requires additional replication in other cohorts and testing in a prospective study of patients starting MTX for the first time.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/investigation-of-differential-methylation-as-a-potential-biomarker-of-methotrexate-response-in-patients-with-rheumatoid-arthritis/