Increasing the DNA damage threshold in breast cancer cells.

The biochemical role of estrogens in the development of estrogen-dependent breast cancer remains to be elucidated, and the involvement of estrogens in tumor initiation remains controversial. Reports regarding estrogen-mediated DNA damage include the induction of 8-oxo-2'-deoxyguanosine (8-oxo-dG) in vitro and in vivo, indicating a role for oxidative stress in tumor initiation and/or progression. However, DNA isolation, cellular DNA repair, and high antioxidant status have made the measurement of 8-oxo-dG in vivo and in cell culture somewhat challenging. In this regard, a potentiation in DNA damage can be achieved by depleting cellular stores of glutathione. We chose to deplete glutathione in the estrogen receptor (ER)-positive MCF-7 breast cancer cell line with a gamma-glutamylcysteine transpeptidase enzyme inhibitor buthionine sulphoximine (BSO) for the purpose of studying estrogen-induced DNA damage. Treatment of GSH-depleted MCF-7 cells with 10 microM 2-OH-E2 or 4-OH-E2 for 30 min resulted in a statistically significant increase in 8-oxo-dG/10(5) dG of 127 and 160%, respectively. A potentiation in catechol estrogen-induced DNA damage was observed with the addition of copper(II) chloride for both 2-OH-E2 and 4-OH-E2 by 165 and 200%, respectively. In addition, 100 nM and 1.0 microM estradiol increased DNA damage in a dose-response-like fashion by 145 and 189%, respectively. The depletion of GSH by BSO may prove to be an advantageous technique for the study of DNA damage in cells otherwise resistant to oxidative stress and/or alkylating agents and has proven useful in the study of estrogen-induced oxidative DNA damage in a highly reproducible and sensitive manner.
AuthorsJames A Mobley, Robert W Brueggemeier
JournalToxicology and applied pharmacology (Toxicol Appl Pharmacol) Vol. 180 Issue 3 Pg. 219-26 (May 1 2002) ISSN: 0041-008X [Print] United States
PMID12009861 (Publication Type: Journal Article, Research Support, U.S. Gov't, Non-P.H.S., Research Support, U.S. Gov't, P.H.S.)
CopyrightCopyright 2002 Elsevier Science (USA).
Chemical References
  • Enzyme Inhibitors
  • Receptors, Estrogen
  • Estradiol
  • Buthionine Sulfoximine
  • 8-oxo-7-hydrodeoxyguanosine
  • L-Lactate Dehydrogenase
  • Deoxyguanosine
  • Glutathione
  • Buthionine Sulfoximine (pharmacology)
  • DNA Damage (drug effects)
  • Deoxyguanosine (analogs & derivatives, metabolism)
  • Dose-Response Relationship, Drug
  • Enzyme Inhibitors (pharmacology)
  • Estradiol (metabolism, pharmacology)
  • Glutathione (deficiency, physiology)
  • L-Lactate Dehydrogenase (metabolism)
  • Oxidative Stress (drug effects)
  • Receptors, Estrogen (drug effects, metabolism)
  • Tumor Cells, Cultured

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