title: Measurement of DNA adducts in cells exposed to cisplatin.
abstract: The two main cisplatin-induced DNA lesions, G--G and A--G, have been measured in cells exposed to the drug. (G--G and A--G denote the intrastrand bifunctional adducts formed between adjacent purine bases.) It has proven feasible, using liquid chromatography-tandem mass spectrometry (LC-MS/MS), observe the G--G and A--G lesions in mouse fibroblast cells exposed for 1 h to a 120 microM concentration of cisplatin. After extraction of the DNA from the cells, the lesions were enzymatically isolated from the DNA in the form of modified dinucleoside monophosphates with the phosphodiester bond intact. MS/MS detection of the modified dinucleoside monophosphates in the negative ion mode manifests two transitions; from the negative ion to the loss of one NH(3) group and from the ion less one NH(3) group to the loss of both NH(3) groups. The multiple reaction monitoring capability of LC-MS/MS was used to measure the three most abundant isotopes of the two main lesions for both transitions of each lesion (i.e., 12 MS/MS values in toto). Ion currents could be detected for all 12 pairs of MS/MS values in the DNA from exposed cells. Although this protocol results in some overlap of MS/MS values between the two lesions, a slight difference in elution times clearly distinguishes between them.
authors.affiliation: Roswell Park Cancer Institute, Buffalo, NY 14263, USA.
authors.forename: Herbert
authors.initials: H
authors.lastname: Iijima
authors.forename: Helen B
authors.initials: HB
authors.lastname: Patrzyc
authors.forename: Jean B
authors.initials: JB
authors.lastname: Dawidzik
authors.forename: Edwin E
authors.initials: EE
authors.lastname: Budzinski
authors.forename: Han-Chun
authors.initials: HC
authors.lastname: Cheng
authors.forename: Harold G
authors.initials: HG
authors.lastname: Freund
authors.forename: Harold C
authors.initials: HC
authors.lastname: Box
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D002945:Cisplatin / Q000494:pharmacology*; D018736:DNA Adducts / Q000032:analysis* / Q000378:metabolism; D004249:DNA Damage / Q000187:drug effects*; D003853:Deoxyribonucleosides / Q000737:chemistry / Q000378:metabolism; D013058:Mass Spectrometry; D051379:Mice
chemical_list: D000970:Antineoplastic Agents; D018736:DNA Adducts; D003853:Deoxyribonucleosides; D002945:Cisplatin