Nuclease digestion and mass spectrometric characterization of oligodeoxyribonucleotides containing 1,2-GpG, 1,2-ApG, and 1,3-GpXpG cisplatin intrastrand cross-links

作者：Williams Renee T; Nalbandian Jenifer N; Tu Audrey; Wang Yinsheng^*

来源：Clinica Chimica Acta, 2013, 420: 160-170.

DOI：10.1016/j.cca.2012.12.010

• Mass spectrometry
• HPLC
• Stable isotope labeling
• DNA digestion
• Cisplatin

Background: The primary mode of action for cis-diamminedichloroplatinum (II), referred to as cisplatin, toward the treatment of solid malignancies is through formation of cross-links with DNA at purine sites, especially guanines. %26lt;br%26gt;Methods: We prepared oligodeoxyribonucleotides (ODNs) containing a 1,2-GpG, 1,2-ApG,or 1,3-GpXpG cisplatin intrastrand cross-link and the corresponding ODNs modified with N-15(2)-labeled cisplatin, and characterized these ODNs with electrospray ionization mass spectrometry (ESI-MS) and tandem MS (MS/MS). We also employed LC-MS/MS to characterize the digestion products of these ODNs after treatment with a cocktail of 4 enzymes (nuclease P1, phosphodiesterases I and II, and alkaline phosphatase). %26lt;br%26gt;Results: 1,2-GpG was released from the ODNs as a dinucleoside monophosphate or a dinucleotide. Analyses of the digestion products of ODNs containing a 1,2-GpG cross-link on the 5%26apos; or 3%26apos; terminus revealed that the dinucleotide carries a terminal 5%26apos; phosphate. On the other hand, digestion of the 1,3-GpXpG intrastrand cross-link yielded 3 dinucleoside products with 0, 1, or 2 phosphate groups. %26lt;br%26gt;Conclusion: The availability of the ODNs carrying the stable isotope-labeled lesions, MS/MS analyses of the cisplatin-modified ODNs, and the characterization of the enzymatic digestion products of these ODNs set the stage for the future LC-MS/MS quantification of the 1,2-GpG, 1,2-ApG, and 1,3-GpXpG lesions in cellular DNA.

• 出版日期2013-5