Mechlorethamine-induced DNA-protein cross-linking in human fibrosarcoma (HT1080) cells

Erin D. Michaelson-Richie, Xun Ming, Simona G. Codreanu, Rachel L. Loeber, Daniel C. Liebler, Colin Campbell, Natalia Y. Tretyakova

Research output: Contribution to journalArticlepeer-review

48 Scopus citations

Abstract

Antitumor nitrogen mustards, such as bis(2-chloroethyl)methylamine (mechlorethamine), are useful chemotherapeutic agents with a long history of clinical application. The antitumor effects of nitrogen mustards are attributed to their ability to induce DNA-DNA and DNA-protein cross-links (DPCs) that block DNA replication. In the present work, a mass spectrometry-based methodology was employed to characterize in vivo DNA-protein cross-linking following treatment of human fibrosarcoma (HT1080) cells with cytotoxic concentrations of mechlorethamine. A combination of mass spectrometry-based proteomics and immunological detection was used to identify 38 nuclear proteins that were covalently cross-linked to chromosomal DNA following treatment with mechlorethamine. Isotope dilution HPLC-ESI+-MS/MS analysis of total proteolytic digests revealed a concentration-dependent formation of N-[2-(S-cysteinyl)ethyl]-N-[2-(guan-7-yl)ethyl]methylamine (Cys-N7G-EMA) conjugates, indicating that mechlorethamine cross-links cysteine thiols within proteins to N-7 positions of guanine in DNA.

Original languageEnglish (US)
Pages (from-to)2785-2796
Number of pages12
JournalJournal of Proteome Research
Volume10
Issue number6
DOIs
StatePublished - Jun 3 2011

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