Methotrexate resistance in relapsed childhood acute lymphoblastic leukaemia

Treatment failure in childhood acute lymphoblastic leukaemia (ALL) might be associated with methotrexate (MTX) resistance, Little is known about MTX r esistance in relapsed ALL. In this study, we determined Ex vivo MTX resista nce in precursor-B ALL at relapse (rALL) and determined possible defects in MTX membrane transport and polyglutamylation. Using the in situ thymidylat e synthase inhibition assay, 21 rALL samples were threefold more MTX resist ant than 63 initial precursor-B ALL samples, both after short-term and afte r continuous MTX exposure (P less than or equal to 0.01). [H-3]-MTX membran e transport did not differ between eight rALL and 25 precursor-B ALL sample s. Incubation for 24 h with 1 mu M [H-3]-MTX resulted in a trend towards a lower accumulation of MTX in 20 relapsed than in 83 initial samples of prec ursor-B ALL samples (906 vs. 1364 pmol/10(9) cells: P = 0.07). Accumulation ;of long-chain MTX polyglutamates (MTX-Glu(4-6)) did not differ between re lapsed and newly diagnosed samples (746 and 889 pmol/10(9) cells; P = 0.1). Activities of the enzymes involved in polyglutamylation (folylpolyglutamat e synthetase and folylpolyglutamate hydrolase) did not differ between rALL and untreated c/pre-B-ALL. This study demonstrates that leukaemic cells of children with relapsed precursor-B ALL are relatively MTX resistant, but th at this MTX resistance is not associated with major impairments in MTX upta ke or polyglutamylation.