Arthritis & Rheumatism, Volume 60,
October 2009 Abstract Supplement
The 2009 ACR/ARHP Annual Scientific Meeting
Philadelphia October 16-21, 2009.
Analysis of Intracellular Methotrexate Polyglutamates in Juvenile Idiopathic Arthritis
Becker1, M. L., van Haandel2, L., Lasky1, A. S., Hoeltzel1, M. F., Neal1, J. E., Palmer1, C. N., Stobaugh2, J. F.
Recently, intracellular MTX polyglutamates (MTXglun) have been shown to be potentially useful biomarkers of response in adult RA patients. We aimed to measure total intracellular MTX gluTOT concentrations in a cohort of JIA patients and determine predictors of MTXgluTOT variability and association with outcome.
After obtaining informed consent, 5 ml of blood was obtained from JIA patients maintained on a stable dose of MTX for 3 months or longer. Demographic characteristics, efficacy and toxicity outcomes were collected by chart review. MTXPG17 in RBC lysates were quantitated using an innovative ion-pairing chromatographic procedure with mass spectrometric detection with improved sensitivity and specificity over existing HPLC separation followed by photochemical/fluorescence detection methods.
Patients (n=85; 61 female) from a single center aged 10.1 ± 4.5 years (mean ± SD; range 1.5 to 19.5 yr) were included in the analysis. Mean MTX dose was 0.51 ± 0.25 mg/kg/dose (range 0.11 to 1.05 mg/kg/dose) for a median of 19 months (range 3 to 156 months). MTX was given SC in 54 (64%) subjects; 50 (59%) subjects had active arthritis.
Total intracellular MTXglun (MTXgluTOT) concentrations varied 40-fold, from 4.80nmol/L to 210.3nmol/L (median 75.5nmol/L), in our cohort. Concentrations of MTXglu17 were measured individually and as a percentage of each patient's MTXgluTOT. MTXglu6 and MTXglu7 concentrations were detected in 68 and 15 patients, respectively. MTXglu3 was the most prominent subtype identified, comprising 41±10.3% of MTXgluTOT, and was most highly correlated with MTXgluTOT (r =0.96). Principal components and cluster analyses revealed that MTXglu1+2 and MTXglu36 formed two internally correlated subgroups, and MTXglu36 correlated highly with MTXgluTOT (r=0.94). MTXglu1+2 constituted 38.1 ± 19.5% of MTXgluTOT (range 5.2 to 92.9%).
Bi-variable analyses revealed the following were associated with MTXgluTOT: age, weight, mg/kg dose, months on MTX, NSAID use and MTX route. Multivariable analyses with inclusion of these variables in the model resulted in only mg/kg dose (p=0.03), route (p=0.01), and months of MTX (p=0.03) remaining significant predictors of MTXgluTOT variability. Months on MTX had a negative association. Response to MTX (no active arthritis) was not associated with MTXgluTOT or any individual absolute MTXglun concentration, however, responders tended to have higher proportions of MTXglu1 (p=0.04) and lower proportions of MTXglu4 (p=0.04).
In our cohort of JIA patients, MTXgluTOT varied 40-fold, but only 32% of this variability was attributed to dose, route and duration of treatment. Longer use of MTX was associated with lower intracellular MTXglun levels. Up to MTXglu7 was detected by our analysis, not previously reported. Unlike adults, long chain MTXglun absolute concentrations were not associated with response to MTX, however, the proportion of glutamate substrates differ between responders and non responders and may be worthy of further investigation.
To cite this abstract, please use the following information:
Becker, M. L., van Haandel, L., Lasky, A. S., Hoeltzel, M. F., Neal, J. E., Palmer, C. N., et al; Analysis of Intracellular Methotrexate Polyglutamates in Juvenile Idiopathic Arthritis [abstract]. Arthritis Rheum 2009;60 Suppl 10 :228