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Role of glutathione and nucleotide excision repair in modulation of cisplatin activity with O6-benzylguanine

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Abstract

Purpose

Modulation of platinating agent cytotoxicity has important clinical implications as a result of their widespread use in the treatment of many different cancers. O6-Benzylguanine (BG) enhances the cytotoxicity of cisplatin against several human tumor lines. The purpose of our work was to elucidate whether BG affects pathways prior to DNA damage (i.e., glutathione, GSH) or following DNA damage (i.e., nucleotide excision repair, NER).

Methods

In efforts to determine the mechanism of enhancement we: (1) evaluated whether different sequences of BG plus cisplatin treatment differed in their ability to enhance cisplatin-induced cytotoxicity and DNA platination; (2) determined the effect of BG on GSH and glutathione S-transferase (GST) activity and; (3) determined whether BG enhanced cisplatin-induced cytotoxicity in cells lacking specific enzymes in the NER pathway. Colony-forming assay, atomic absorption spectroscopy and HPLC were employed to measure tumor cell growth inhibition, quantitate the amount of platinum on DNA, and determine intracellular GSH concentrations, respectively.

Results

Increased cytotoxicity and platination of DNA was observed when cells were exposed to BG prior to and/or during cisplatin treatment and not when BG followed cisplatin treatment. BG did not significantly alter GST activity with minimal depletion of GSH. In contrast, buthionine sulfoximine (BSO) caused a much more dramatic decrease in GSH than BG that was not accompanied by a dramatic increase in sensitivity to cisplatin. Furthermore, BG enhanced the cytotoxicity of cisplatin in a series of cell lines deficient in NER.

Conclusions

Overall, our results suggest that the mechanism of enhancement involves neither the GSH nor the NER pathways, but triggers an event prior to DNA platination damage that ultimately results in increased cytotoxicity, apoptosis and increased platination levels.

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Abbreviations

AGT:

O6-alkylguanine-DNA alkyltransferase

BG:

O6-Benzylguanine

BSO:

Buthionine sulfoximine

CDK:

Cyclin-dependent kinase

CSA:

Cockayne syndrome A

CSB:

Cockayne syndrome B

ERCC-1/XPF:

Excision repair cross-complementing 1/xeroderma pigmentosum F

GSH:

Glutathione

GST:

Glutathione S-transferase

HPLC:

High-pressure liquid chromatography

NER:

Nucleotide excision repair

TC-NER:

Transcription-coupled NER

XP:

Xeroderma pigmentosum

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Acknowledgements

We are grateful to Terry McManus for his assistance with the XPA cell survival assays and Kristen Kasza for her assistance on the statistical analyses. This work was supported in part by NIH grants CA81485 (M.E.D.) 5T32 CA09594 (M.L.F.) and 2 P30 CA47904 (M.J.E.).

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Authors and Affiliations

  1. Department of Medicine, Committee on Cancer Biology and Cancer Research Center, University of Chicago, Chicago, IL, 60637, USA

    Melissa L. Fishel, Shannon M. Delaney & M. Eileen Dolan

  2. Duke University Medical Center, Durham, NC, 27701, USA

    Michael P. Gamcsik

  3. University of Pittsburgh Cancer Institute, Pittsburgh, PA, 15213, USA

    Eleanor G. Zuhowski & Merrill J. Egorin

  4. The Carcinogenesis Laboratory, Michigan State University, East Lansing, MI, 48824, USA

    Veronica M. Maher

  5. Department of Health Studies, University of Chicago, Chicago, IL, 60637, USA

    Theodore Karrison

  6. Laboratory of Comparative Carcinogenesis, National Cancer Institute at Frederick, Frederick, MD, 21702-1201, USA

    Robert C. Moschel

  7. University of Chicago, 5841 S. Maryland Ave., P.O. Box MC2115, Chicago, IL, 60637, USA

    M. Eileen Dolan

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  1. Melissa L. Fishel
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  2. Michael P. Gamcsik
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  9. M. Eileen Dolan
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Correspondence to M. Eileen Dolan.

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Fishel, M.L., Gamcsik, M.P., Delaney, S.M. et al. Role of glutathione and nucleotide excision repair in modulation of cisplatin activity with O6-benzylguanine. Cancer Chemother Pharmacol 55, 333–342 (2005). https://doi.org/10.1007/s00280-004-0901-3

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  • DOI: https://doi.org/10.1007/s00280-004-0901-3

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