title: Kinetics of carboplatin-DNA binding in genomic DNA and bladder cancer cells as determined by accelerator mass spectrometry.
abstract: Cisplatin and carboplatin are platinum-based drugs that are widely used in cancer chemotherapy. The cytotoxicity of these drugs is mediated by platinum-DNA monoadducts and intra- and interstrand diadducts, which are formed following uptake of the drug into the nucleus of cells. The pharmacodynamics of carboplatin display fewer side effects than for cisplatin, albeit with less potency, which may be due to differences in rates of DNA adduct formation. We report the use of accelerator mass spectrometry (AMS), a sensitive detection method often used for radiocarbon quantitation, to measure both the kinetics of [14C]carboplatin-DNA adduct formation with genomic DNA and drug uptake and DNA binding in T24 human bladder cancer cells. Only carboplatin-DNA monoadducts contain radiocarbon in the platinated DNA, which allowed for calculation of kinetic rates and concentrations within the system. The percent of radiocarbon bound to salmon sperm DNA in the form of monoadducts was measured by AMS over 24 h. Knowledge of both the starting concentration of the parent carboplatin and the concentration of radiocarbon in the DNA at a variety of time points allowed calculation of the rates of Pt-DNA monoadduct formation and conversion to toxic cross-links. Importantly, the rate of carboplatin-DNA monoadduct formation was approximately 100-fold slower than that reported for the more potent cisplatin analogue, which may explain the lower toxicity of carboplatin. T24 human bladder cancer cells were incubated with a subpharmacological dose of [14C]carboplatin, and the rate of accumulation of radiocarbon in the cells and nuclear DNA was measured by AMS. The lowest concentration of radiocarbon measured was approximately 1 amol/10 microg of DNA. This sensitivity may allow the method to be used for clinical applications.
authors.affiliation: Biosciences Directorate and Center for Accelerator Mass Spectrometry, Lawrence Livermore National Laboratory, 7000 East Avenue, L-441, Livermore, California 94551, USA. hah2@llnl.gov
authors.forename: Sang Soo
authors.initials: SS
authors.lastname: Hah
authors.forename: Kristen M
authors.initials: KM
authors.lastname: Stivers
authors.forename: Ralph W
authors.initials: RW
authors.lastname: de Vere White
authors.forename: Paul T
authors.initials: PT
authors.lastname: Henderson
mesh_terms: D000970:Antineoplastic Agents / Q000493:pharmacokinetics* / Q000633:toxicity; D016190:Carboplatin / Q000032:analysis / Q000493:pharmacokinetics* / Q000633:toxicity; D045744:Cell Line, Tumor; D004247:DNA / Q000187:drug effects / Q000378:metabolism; D018736:DNA Adducts / Q000032:analysis*; D004273:DNA, Neoplasm / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D013058:Mass Spectrometry; D001749:Urinary Bladder Neoplasms
chemical_list: D000970:Antineoplastic Agents; D018736:DNA Adducts; D004273:DNA, Neoplasm; C096276:carboplatin-DNA adduct; D004247:DNA; D016190:Carboplatin