Methotrexate accumulation in target intestinal mucosa and white blood cells differs from non‐target red blood cells of patients with Crohn's disease

Abstract Background Intracellular methotrexate polyglutamates (MTX‐PGs) concentrations are measurable in red blood cells (RBCs) during MTX treatment. MTX‐PG 3 concentrations correlate with efficacy in patients with Crohn's disease (CD). Since RBCs are not involved in pathogenesis of CD and lack extended MTX metabolism, we determined MTX‐PGs accumulation in peripheral blood mononuclear cells (PBMCs: effector cells) and intestinal mucosa (target cells) and compared those with RBCs as a potential more precise biomarker. Methods In a multicentre prospective cohort study, blood samples of patients with CD were collected during the first year of MTX therapy. Mucosal biopsies were obtained from non‐inflamed rectum and/or inflamed intestine. MTX‐PGs concentrations in mucosa, PBMCs and RBCs were measured by liquid chromatography–tandem mass spectrometry. Results From 80 patients with CD, a total of 27 mucosal biopsies, 9 PBMC and 212 RBC samples were collected. From 12 weeks of MTX therapy onwards, MTX‐PG 3 was the most predominant species (33%) in RBCs. In PBMCs, the distribution was skewed towards MTX‐PG 1 (48%), which accounted for an 18 times higher concentration than in RBCs. Long‐chain MTX‐PGs were highly present in mucosa: 21% of MTX‐PG total was MTX‐PG 5 . MTX‐PG 6 was measurable in all biopsies. Conclusions MTX‐PG patterns differ between mucosa, PBMCs and RBCs of patients with CD.