Role of glutathione and nucleotide excision repair in modulation of cisplatin activity with O6-benzylguanine.
OBJECTIVE
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.
Fishel
Melissa L
ML
Department of Medicine, Committee on Cancer Biology and Cancer Research Center, University of Chicago, Chicago, IL, 60637, USA.
Gamcsik
Michael P
MP
Duke University Medical Center, Durham, NC, 27701, USA.
Delaney
Shannon M
SM
Department of Medicine, Committee on Cancer Biology and Cancer Research Center, University of Chicago, Chicago, IL, 60637, USA.
Zuhowski
Eleanor G
EG
University of Pittsburgh Cancer Institute, Pittsburgh, PA, 15213, USA.
Maher
Veronica M
VM
The Carcinogenesis Laboratory, Michigan State University, East Lansing, MI, 48824, USA.
Karrison
Theodore
T
Department of Health Studies, University of Chicago, Chicago, IL, 60637, USA.
Moschel
Robert C
RC
Laboratory of Comparative Carcinogenesis, National Cancer Institute at Frederick, Frederick, MD, 21702-1201, USA.
Egorin
Merrill J
MJ
University of Pittsburgh Cancer Institute, Pittsburgh, PA, 15213, USA.
Dolan
M Eileen
ME
Department of Medicine, Committee on Cancer Biology and Cancer Research Center, University of Chicago, Chicago, IL, 60637, USA. edolan@medicine.bsd.uchicago.edu.|University of Chicago, 5841 S. Maryland Ave., P.O. Box MC2115, Chicago, IL, 60637, USA. edolan@medicine.bsd.uchicago.edu.
D000970:Antineoplastic Agents / Q000494:pharmacology*; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D002945:Cisplatin / Q000494:pharmacology*; D004249:DNA Damage; D004260:DNA Repair / Q000187:drug effects*; D004357:Drug Synergism; D005978:Glutathione / Q000378:metabolism*; D006147:Guanine / Q000031:analogs & derivatives* / Q000494:pharmacology*; D006801:Humans; D010984:Platinum / Q000378:metabolism; D014407:Tumor Cells, Cultured
D000970:Antineoplastic Agents; C064976:O(6)-benzylguanine; D010984:Platinum; D006147:Guanine; D005978:Glutathione; D002945:Cisplatin