Metabolism and actions of 2-chloro-9-(2-deoxy-2-fluoro-beta-D- arabinofuranosyl)-adenine in human lymphoblastoid cells.
2-Chloro-9-(2-deoxy-2-fluoro-beta-D-arabinofuranosyl)adenine (Cl-F-ara-A) is a new deoxyadenosine analogue that is resistant to phosphorolytic cleavage and deamination. Studies with a variety of cell lines demonstrated that Cl-F-ara-A is a potent cytotoxic agent; in cell-free systems, its triphosphate (Cl-F-ara-ATP) inhibited DNA polymerase alpha and ribonucleotide reductase. To further characterize its mechanism of cytotoxicity, the present study investigated the cellular metabolism of Cl-F-ara-A and the actions of its nucleotide metabolites in human T-lymphoblast leukemia CCRF-CEM cells. The mono-, di-, and triphosphates of Cl-F-ara-A accumulated in cells, with the monophosphate as its major metabolite. After washing cells into drug-free medium, the elimination of each Cl-F-ara-A nucleotide was nonlinear with a prolonged terminal phase. Incubation of CCRF-CEM cells with Cl-F-ara-A resulted in the incorporation of Cl-F-ara-AMP into DNA; a much lesser amount was associated with RNA, suggesting that Cl-F-ara-A is a more DNA-directed compound. The site of Cl-F-ara-AMP in DNA was related to the ratio of the cellular concentrations of the analogue triphosphate and the natural substrate dATP. At low Cl-F-ara-ATP:dATP values, incorporation was mainly in phosphodiester linkages at internal sites, whereas at higher Cl-F-ara-ATP:dATP values, Cl-F-ara-AMP was principally detected at terminal sites. Clonogenicity assays showed a strong inverse correlation between cell survival and Cl-F-ara-AMP incorporation into DNA. These results suggest that the incorporation of Cl-F-ara-A monophosphate into DNA is critical for the cytotoxicity of Cl-F-ara-A.
Xie
C
C
Department of Clinical Investigation, University of Texas M. D. Anderson Cancer Center, Houston 77030, USA.
Plunkett
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W
D000227:Adenine Nucleotides; D000964:Antimetabolites, Antineoplastic / Q000378:metabolism; D001087:Arabinonucleosides / Q000378:metabolism* / Q000494:pharmacology; D002453:Cell Cycle; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D000077866:Clofarabine; D004247:DNA / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D012313:RNA / Q000378:metabolism; D013601:T-Lymphocytes / Q000378:metabolism*
D000227:Adenine Nucleotides; D000964:Antimetabolites, Antineoplastic; D001087:Arabinonucleosides; D012313:RNA; D000077866:Clofarabine; D004247:DNA