Methotrexate Treatment of Newly Diagnosed RA Patients Is Associated With DNA Methylation Differences at Genes Relevant for Disease Pathogenesis and Pharmacological Action

Front Immunol. 2021 Nov 18;12:713611. doi: 10.3389/fimmu.2021.713611. eCollection 2021.


Background: Methotrexate (MTX) is the first line treatment of rheumatoid arthritis (RA), and methylation changes in bulk T cells have been reported after treatment with MTX. We have investigated cell-type specific DNA methylation changes across the genome in naïve and memory CD4+ T cells before and after MTX treatment of RA patients. DNA methylation profiles of newly diagnosed RA patients (N=9) were assessed by reduced representation bisulfite sequencing.

Results: We found that MTX treatment significantly influenced DNA methylation levels at multiple CpG sites in both cell populations. Interestingly, we identified differentially methylated sites annotated to two genes; TRIM15 and SORC2, previously reported to predict treatment outcome in RA patients when measured in bulk T cells. Furthermore, several of the genes, including STAT3, annotated to the significant CpG sites are relevant for RA susceptibility or the action of MTX.

Conclusion: We detected CpG sites that were associated with MTX treatment in CD4+ naïve and memory T cells isolated from RA patients. Several of these sites overlap genetic regions previously associated with RA risk and MTX treatment outcome.

Keywords: CD4 memory; CD4 naïve; DNA methylation; RRBS; Rheumatoid arthritis; T cells; epigenetics; methotrexate.

Publication types

  • Observational Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adult
  • Aged
  • Antirheumatic Agents / pharmacology
  • Antirheumatic Agents / therapeutic use*
  • Arthritis, Rheumatoid / drug therapy*
  • Arthritis, Rheumatoid / genetics
  • Arthritis, Rheumatoid / immunology
  • CD4-Positive T-Lymphocytes / drug effects
  • CD4-Positive T-Lymphocytes / immunology
  • CpG Islands
  • DNA Methylation / drug effects*
  • DNA-Binding Proteins / genetics
  • Female
  • Humans
  • Intracellular Signaling Peptides and Proteins / genetics
  • Male
  • Memory T Cells / drug effects
  • Memory T Cells / immunology
  • Methotrexate / pharmacology
  • Methotrexate / therapeutic use*
  • Middle Aged
  • Receptors, CCR6 / genetics
  • Receptors, Cell Surface / genetics
  • STAT3 Transcription Factor / genetics
  • Synaptogyrins / genetics


  • Antirheumatic Agents
  • CCR6 protein, human
  • DNA-Binding Proteins
  • Intracellular Signaling Peptides and Proteins
  • Receptors, CCR6
  • Receptors, Cell Surface
  • SORCS2 protein, human
  • STAT3 Transcription Factor
  • STAT3 protein, human
  • SYNGR1 protein, human
  • Synaptogyrins
  • WDFY4 protein, human
  • tripartite motif-containing protein 15, human
  • Methotrexate