Bromodeoxyuridine enhancement of 1-beta-D-arabinofuranosylcytosine metabolic activation and toxicity in HL-60 leukemic cells.
We tested whether bromodeoxyuridine (BrdUrd), an analogue of thymidine (dThd), enhances 1-beta-D-arabinofuranosylcytosine (ara-C) metabolic activation, as does dThd. HL-60 cells were exposed to 10, 100, or 1000 nM ara-C for 3 h. Simultaneous exposure of log phase HL-60 cells to BrdUrd (1-1000 microM) and ara-C for 3 h resulted in enhancement of ara-C incorporation into DNA, with a doubling of incorporation in response to 10 nM ara-C occurring at concentrations of BrdUrd greater than 100 microM. Preexposure of cells to BrdUrd for 16 h followed by addition of ara-C for 3 h resulted in even greater ara-C incorporation into DNA. This increase was most marked at the lower concentrations of ara-C (10 and 100 nM), where approximately 3-fold enhancement of ara-C incorporation was observed in response to BrdUrd concentrations greater than 100 microM. Intracellular pools of 1-beta-D-arabinofuranosyl-CTP increased significantly (up to 3-fold) following 16-h exposure to BrdUrd (30, 100, or 300 microM) at all concentrations of ara-C tested. The ara-C phosphorylating activity of cell-free extracts obtained following 16-h exposure of cells to BrdUrd increased 1.5- to 2.3-fold over control. Intracellular dCTP pools fell to approximately 50% of control after exposure to 750 microM BrdUrd or dThd. Exposure to BrdUrd for 16 h caused a concentration-dependent increase in cells with S-phase DNA content, as assessed by flow cytometry, with a doubling of cells in S phase (to 60%) observed in response to 500 microM BrdUrd. HL-60 cells exposed to identical conditions of BrdUrd for 3 h showed no significant alteration in cell cycle phase distribution. Thus, although BrdUrd does increase cells in S phase, the increased ara-C incorporation caused by BrdUrd cannot be explained solely on a cytokinetic basis since enhancement of incorporation was observed after a 3-h exposure of cells to BrdUrd and ara-C. The combination of ara-C (100 nM) and BrdUrd (100-1000 microM) exhibited cytotoxic synergism, as measured by the fluorescein diacetate/propidium iodide method. These data demonstrate a clear potential for BrdUrd modulation of ara-C metabolism in human leukemia. Additionally, the interaction of BrdUrd and ara-C should be considered in the interpretation of studies of the effects of ara-C on DNA synthesis as measured by flow cytometric quantification of incorporated BrdUrd.
Ross
D D
DD
Division of Developmental Therapeutics, University of Maryland Cancer Center, Baltimore 21201.
Joneckis
C C
CC
Song
T H
TH
Wu
R K
RK
D001085:Arabinofuranosylcytosine Triphosphate / Q000378:metabolism; D001711:Biotransformation; D001973:Bromodeoxyuridine / Q000494:pharmacology*; D002453:Cell Cycle; D002470:Cell Survival / Q000187:drug effects; D003561:Cytarabine / Q000378:metabolism* / Q000633:toxicity; D004273:DNA, Neoplasm / Q000378:metabolism; D003845:Deoxycytosine Nucleotides / Q000378:metabolism; D003848:Deoxyguanine Nucleotides / Q000378:metabolism; D006801:Humans; D010766:Phosphorylation; D014407:Tumor Cells, Cultured / Q000187:drug effects* / Q000378:metabolism
D004273:DNA, Neoplasm; D003845:Deoxycytosine Nucleotides; D003848:Deoxyguanine Nucleotides; D003561:Cytarabine; D001085:Arabinofuranosylcytosine Triphosphate; C024107:2'-deoxycytidine 5'-triphosphate; C029603:deoxyguanosine triphosphate; D001973:Bromodeoxyuridine