title: A new high-performance liquid chromatography method determines low production of 9-beta-D-arabinofuranosylguanine triphosphate, an active metabolite of nelarabine, in adult T-cell leukemia cells.
abstract: The 9-beta-D-arabinofuranosylguanine (ara-G), an active compound of nelarabine, demonstrates potent cytotoxicity specifically on T-cell malignancies. In cells, ara-G is phosphorylated to ara-G triphosphate (ara-GTP), which is subsequently incorporated into DNA, thereby inhibiting DNA synthesis. Because ara-GTP is crucial to ara-G's cytotoxicity, the determination of ara-GTP production in cancer cells is informative for optimizing nelarabine administration. Here, we developed a new, sensitive isocratic-elution HPLC method for quantifying ara-GTP. Samples were eluted isocratically by using phosphate buffer at a constant flow rate. Ara-GTP was clearly separated from other nucleotides by using an anion-exchange column and it was quantitated by its peak area at 254 nm. The standard curve was linear with low variability and a sensitive detection limit (10 pmol). Furthermore, due to ara-G's specificity to T-cells we hypothesized that nelarabine might be effective against adult T-cell leukemia (ATL). The ara-GTP production was compared between T-lymphoblastic leukemia CCRF-CEM and ATL cell lines in vitro. When CEM cells were incubated with ara-G, the ara-GTP production increased in a concentration- and time-dependent manner. In contrast, 5 ATL cell lines accumulated lower ara-GTP in the same condition. While ara-G inhibited the growth of CEM cells with a 50% growth inhibition concentration of 2 microM, the inhibitory-concentration values were >1 mM in 8 of the 12 ATL cell lines. This ineffectiveness appeared to correspond with the low ara-GTP production. The present study is the first to evaluate the potential of ara-G against ATL cells; our results suggest that nelarabine would not be effective against ATL.
authors.affiliation: Department of Hematology and Oncology, Faculty of Medical Sciences, University of Fukui, Matsuoka, Fukui, Japan. tyamauch@u-fukui.ac.jp
authors.forename: Takahiro
authors.initials: T
authors.lastname: Yamauchi
authors.forename: Rie
authors.initials: R
authors.lastname: Nishi
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Kitazumi
authors.forename: Tsuyoshi
authors.initials: T
authors.lastname: Nakano
authors.forename: Takanori
authors.initials: T
authors.lastname: Ueda
mesh_terms: D000970:Antineoplastic Agents / Q000378:metabolism / Q000627:therapeutic use; D001087:Arabinonucleosides / Q000378:metabolism / Q000493:pharmacokinetics* / Q000494:pharmacology / Q000627:therapeutic use; D001088:Arabinonucleotides / Q000032:analysis* / Q000378:metabolism; D015415:Biomarkers / Q000032:analysis / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D005591:Chemical Fractionation / Q000379:methods; D002851:Chromatography, High Pressure Liquid / Q000191:economics / Q000379:methods / Q000592:standards; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives* / Q000032:analysis / Q000378:metabolism; D006801:Humans; D015459:Leukemia-Lymphoma, Adult T-Cell / Q000175:diagnosis / Q000188:drug therapy / Q000378:metabolism*; D011355:Prodrugs / Q000378:metabolism / Q000493:pharmacokinetics; D012015:Reference Standards; D012680:Sensitivity and Specificity; D016896:Treatment Outcome; D014407:Tumor Cells, Cultured
chemical_list: D000970:Antineoplastic Agents; D001087:Arabinonucleosides; D001088:Arabinonucleotides; D015415:Biomarkers; D011355:Prodrugs; C028771:9-arabinofuranosylguanine; C104457:nelarabine; C022206:9-beta-D-arabinofuranosylguanosine 5'-triphosphate; D006160:Guanosine Triphosphate