Characterization of resistance to cytosine arabinoside (Ara-C) in NALM-6 human B leukemia cells

BackgroundCytosine arabinoside (1-β-d-arabinofuranosylcytosine;Ara-C) is the most important antimetabolite used for acute leukemia. We established Ara-C (0.003–1 μmol/l)-resistant NALM-6 leukemia cells, and attempted the characterization of their resistance.MethodsThe Ara-C-resistant cell lines were developed by stepwise increases in the drug. The mRNA expressions were analyzed by reverse transcription-polymerase chain reaction (RT-PCR). The uptake of Ara-C, deoxycytidine kinase (dCK) activity and cytidine deaminase (CDA) activity were measured using radioisotope methods. Cytotoxicity was evaluated using the MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] assay.ResultsThe mRNA expression of human equilibrative nucleoside transporter-1 (hENT-1), which is an uptake transporter of Ara-C, was initially decreased during the acquisition of resistance to Ara-C. The expression of dCK, an activation enzyme, and of CDA, an inactivation enzyme, was decreased and increased in the late phase, respectively. The cytotoxic effect of Ara-C on parental NALM-6 cells was ameliorated by hENT-1 inhibitors. There were no differences in the cytotoxic effect of other anticancer drugs, but there was similar resistance to nucleoside analogues via hENT-1 between the parental and resistant cells.ConclusionsDecreased hENT-1 expression and function is causatively responsible for the acquisition of Ara-C resistance and alterations in dCK and CDA contribute to the higher concentration range.