Determinants of erythrocyte methotrexate polyglutamate levels in rheumatoid arthritis

Objective. Low-dose methotrexate (MTX) is the anchor drug in the treatment for rheumatoid arthritis (RA). Response to MTX is related to the intracellular MTX-polyglutamate (MTX-PG) levels and little is known about its determinants. We aimed to define the determinants of erythrocyte MTX-PG concentrations in 2 prospective cohorts of patients with RA. Methods. Patients with RA treated with MTX from 2 longitudinal cohorts were included: 93 from the MTX-R study (Rotterdam, the Netherlands derivation cohort), and 247 from the treatment in Rotterdam Early Arthritis Cohort study (validation cohort). MTX-PG concentrations were measured at 3 months of treatment using liquid chromatography/mass spectrometry. The MTX-PG were used as outcome measure. Various sociodemographic, clinical, biochemical, and genetic factors were assessed at baseline. Associations with MTX-PG levels were analyzed using multivariate regression analysis. Results. Age was positively associated with MTX-PG1 (stβ 0.23, p = 0.033) and total MTX-PG (stβ 0.23, p = 0.018) in the derivation cohort, and with all MTX-PG in the validation cohort (MTX-PG1: stβ 0.13, p = 0.04; MTX-PG2: stβ 0.21, p = 0.001; MTX-PG3: stβ 0.22, p < 0.001; MTX-PG4+5: stβ 0.25, p < 0.001; and total MTX-PG: stβ 0.32, p < 0.001). Erythrocyte folate levels were positively associated with MTX-PG3 (stβ 0.3, p = 0.021) and total MTX-PG levels (stβ 0.32, p = 0.022) in the derivation cohort, which was replicated for MTX-PG3 (stβ 0.15, p = 0.04) in the validation cohort. Patients with the folylpolyglutamate synthase (FPGS) rs4451422 wild-type genotype had higher concentrations of MTX-PG3 (p < 0.05), MTX-PG4+5 (p < 0.05), and total MTX-PG (p < 0.05) in both cohorts. In the combined cohort, MTX dose was positively associated with levels of MTX-PG3 (stβ 0.23, p < 0.001), MTX-PG4+5 (stβ 0.30, p < 0.001), and total MTX-PG (stβ 0.20, p = 0.002), but negatively associated with MTX-PG2 levels (stβ -0.22, p < 0.001). Conclusion. Our prospective study shows that higher age, higher MTX dose, higher erythrocyte folate status, and the FPGS rs4451422 wild-type genotype are associated with higher MTX-PG concentrations. While only up to 21% of interpatient variability can be explained by these determinants, this knowledge may aid in the development of personalized treatment in RA.