Characterization of an etoposide-glutathione conjugate derived from metabolic activation by human cytochrome p450

Curr Drug Metab. 2006 Dec;7(8):897-911. doi: 10.2174/138920006779010638.


Etoposide (VP-16), a DNA topoisomerase II poison widely used as an antineoplastic agent is also known to cause leukemia. One of its major metabolic pathways involves O-demethylation to etoposide catechol (etoposide-OH) by cytochrome P450 3A4 (CYP3A4). The catechol metabolite can undergo sequential one- and two-electron oxidations to form etoposide semi-quinone (etoposide-SQ) and etoposide quinone (etoposide-Q), respectively, which have both been implicated as cytotoxic metabolites. However, etoposide-Q is known to react with glutathione (GSH), which can protect DNA from oxidative damage by this reactive metabolite. In this study, etoposide-Q was reacted with GSH and the two etoposide-GSH conjugates were characterized. The major conjugate was etoposide-OH-6'-SG and the minor product was etoposide-OH-2'-SG. Etoposide-OH-6'-SG, which arose from Michael addition of GSH to etoposide-Q, was characterized by mass spectrometry and 2-D NMR. It was identified as the sole product from in vitro metabolism experiments using recombinant human CYP3A4 or liver microsomes incubated with etoposide in the presence of GSH. Etoposide-OH-6'-SG was also detected from incubations of etoposide-OH and GSH alone. Therefore, the presence of etoposide-OH, which can be formed from etoposide metabolism by CYP3A4, is essential for formation of the GSH conjugate. The oxidation of etoposide-OH to a quinone intermediate is likely the precursor in the formation of etoposide-OH-6'-SG.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Antineoplastic Agents, Phytogenic / metabolism
  • Chromatography, Liquid
  • Cytochrome P-450 CYP3A
  • Cytochrome P-450 Enzyme System / metabolism*
  • Etoposide / chemistry
  • Etoposide / metabolism
  • Glutathione / chemistry
  • Glutathione / metabolism*
  • Humans
  • In Vitro Techniques
  • Magnetic Resonance Spectroscopy
  • Mass Spectrometry
  • Microsomes, Liver / metabolism


  • Antineoplastic Agents, Phytogenic
  • Etoposide
  • Cytochrome P-450 Enzyme System
  • Cytochrome P-450 CYP3A
  • CYP3A4 protein, human
  • Glutathione