Comparative Quantification of MTX and Its Metabolites Post Glucarpidase

Acidification (i.e., addition of hydrochloric acid) of plasma samples from patients who have received Glucarpidase post high-dose MTX treatment is regarded as a necessary preanalytic step to avoid further in vitro enzymatic cleavage of MTX. However, it is unclear whether this acidification step is essential. A comparative study, which evaluates concentrations of MTX and its metabolites in paired (acidified versus non-acidified) plasma samples, has not yet been performed. Processing plasma samples without acidification would facilitate quantification of MTX, including plasma samples from patients treated at centers without adequate laboratory facilities.

Stored plasma samples (paired - acidified and non-acidified) from the specimens collection ('Tumorbiobank') of the Charité Comprehensive Cancer Center will be analysed by HPLC/MS. A total of 136 samples from patients post Voraxaze across 3 levels of MTX concentrations: 30 without additive, 10 with 0.9% NaCl, 30 with HCl (stored at -80C) 9 without additive, 9 with HCl (stored at +4C and analysed after 3-5 days) 9 without additive, 9 with HCl (stored at -20C and analysed after 3-5 days) 30 with HCl (stored at +4C versus ambient temperature)