THE EFFECT OF 9-BETA-D-ARABINOFURANOSYL 2-FLUOROADENINE AND 1-BETA-D-ARABINOFURANOSYLCYTOSINE ON THE CELL-CYCLE PHASE DISTRIBUTION, TOPOISOMERASE-II LEVEL, MITOXANTRONE CYTOTOXICITY, AND DNA STRAND BREAK PRODUCTION IN K562 HUMAN LEUKEMIA-CELLS

Antimetabolites and topoisomerase (topo) II-reactive drugs are frequen tly combined in the therapy of acute leukemia. The two types of agents are thought to be synergistic in their actions against malignant blas ts but the mechanism for this synergism is incompletely described. Thi s study sought to determine whether the combination of two rather than one antimetabolite with the topo II-reactive intercalator mitoxantron e would be greater than the effect of the single antimetabolite ara-C on mitoxantrone's cytotoxic actions. We also aimed to determine a mech anism for synergism should it occur. The model system used was K562 hu man leukemia cells. The second antimetabolite selected was F-ara-A, th e active form of fludarabine. The resultant combination (F-ara-A, ara- C, and a topo II-reactive drug) is one currently being tested against acute myelogenous leukemia in clinical trials. F-ara-A itself had litt le effect on the cytotoxicity or the topo II-mediated DNA cleaving act ions of mitoxantrone, while ara-C potentiated these actions as it does those of other topo II-reactive drugs. Surprisingly F-ara-A enhanced the actions of ara-C on mitoxantrone-associated cytotoxicity by at lea st an order of magnitude. The effect of the addition of F-ara-A to ara -C on mitoxantrone-induced DNA cleavage was considerably smaller, but present. Antimetabolite treatment did not increase the amount of topo II within cells measured directly by immunoblotting or indirectly by q uantifying the maximum number of topo II-DNA complexes stabilized by m itoxantrone. Rather, the antimetabolites altered the distribution of t he cells in the cell cycle. Antimetabolite treatment caused a large in crease in S-phase cells, a phase in which cells are more sensitive to topo II-reactive drugs than the associated topo II-mediated DNA cleava ge would predict. Therefore, it is likely that this shift in the distr ibution of the cells within the cell cycle accounts for both the enhan ced cytotoxicity of mitoxantrone in antimetabolite pretreated cells an d the discrepancy between the magnitude of antimetabolite action on to po II-mediated DNA cleavage.