Inactivation of human O6-alkylguanine-DNA alkyltransferase by modified oligodeoxyribonucleotides containing O6-benzylguanine

Anthony E. Pegg, Karina Goodtzova, Natalia A. Loktionova, Sreenivas Kanugula, Gary T. Pauly, Robert C. Moschel

Research output: Contribution to journalArticlepeer-review

12 Scopus citations

Abstract

Inactivation of the DNA repair protein O6-alkylguanine-DNA alkyltransferase (AGT) enhances tumor cell killing by therapeutic alkylating agents. O6-Benzylguanine (b6G) can inactivate AGT and is currently in clinical trials to enhance therapy. Short oligodeoxyribonucleotides containing b6G are much more effective inactivators, but their use for therapeutic purposes is likely to be compromised by metabolic instability. We have therefore examined the ability to inactivate AGT of an 11-mer oligodeoxyribonucleotide containing b6G (11-mpBG) when modified with terminal methylphosphonate linkages to protect it from nucleases. This modification did not reduce the ability to serve as a substrate/inactivator for AGT, and 11-mpBG had an ED50 value of 1.3 nM, more than 300-fold lower than that for b6G. A similar oligodeoxyribonucleotide containing O6-methylguanine (m6G) was also found to be a good substrate (ED50 value of 10 nM), but the benzylated form was repaired more rapidly and preferentially. When added to HT29 cell cultures, 5 μM 11-mpBG was able to cause a prolonged inactivation of cellular AGT for at least 72 h and to greatly sensitize the cells to killing by 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU). The 11-mpMG was ineffective at up to 20 μM, suggesting that the benzyl group allows better uptake into the cell. However, even with 11-mpBG, the 1000-fold decrease in potency toward AGT in HT29 cells compared to that toward the protein in vitro suggests that uptake may be a limiting factor. These results suggest that oligodeoxyribonucleotides such as 11-mpBG may prove to be useful drugs for potentiation of alkylating agent chemotherapy if uptake can be improved.

Original languageEnglish (US)
Pages (from-to)958-965
Number of pages8
JournalJournal of Pharmacology and Experimental Therapeutics
Volume296
Issue number3
StatePublished - 2001

All Science Journal Classification (ASJC) codes

  • Molecular Medicine
  • Pharmacology

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