title: High-performance liquid chromatography method for the determination and quantitation of arabinosylguanine triphosphate and fludarabine triphosphate in human cells.
abstract: A gradient anion-exchange high-performance liquid chromatographic assay was developed for the simultaneous determination and quantitation of the cytotoxic triphosphates of arabinosylguanine (ara-GTP) and fludarabine (F-ara-ATP). The method was validated with respect to selectivity, recovery, linearity, precision, and accuracy using authentic standards. To test this assay in a more complex biological matrix, perchloric acid extracts of circulating human leukemia cells spiked with known concentrations of ara-GTP and F-ara-ATP were examined. Finally, to assess the clinical utility of our method, perchloric acid extracts of circulating human leukemia cells isolated from patients treated with fludarabine and nelarabine were analyzed. The range of quantitation was 0.0125-10 nmol for the ara- and native NTPs in cellular extracts. This assay should be helpful in establishing the mechanistic rationales for drug scheduling and combinations of nelarabine and fludarabine, and for correlating the therapeutic efficacy and levels of the cytotoxic triphosphates in target cells.
authors.affiliation: Department of Experimental Therapeutics, The University of Texas M.D. Anderson Cancer Center, Houston 77030, USA.
authors.forename: C O
authors.initials: CO
authors.lastname: Rodriguez
authors.forename: W
authors.initials: W
authors.lastname: Plunkett
authors.forename: M T
authors.initials: MT
authors.lastname: Paff
authors.forename: M
authors.initials: M
authors.lastname: Du
authors.forename: B
authors.initials: B
authors.lastname: Nowak
authors.forename: P
authors.initials: P
authors.lastname: Ramakrishna
authors.forename: M J
authors.initials: MJ
authors.lastname: Keating
authors.forename: V
authors.initials: V
authors.lastname: Gandhi
mesh_terms: D001088:Arabinonucleotides / Q000032:analysis*; D002138:Calibration; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives* / Q000032:analysis*; D006801:Humans; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013056:Spectrophotometry, Ultraviolet; D014407:Tumor Cells, Cultured; D014740:Vidarabine / Q000031:analogs & derivatives* / Q000032:analysis*
chemical_list: D001088:Arabinonucleotides; C022206:9-beta-D-arabinofuranosylguanosine 5'-triphosphate; D006160:Guanosine Triphosphate; D014740:Vidarabine; C024352:fludarabine