Benzene metabolites antagonize etoposide-stabilized cleavable complexes of DNA topoisomerase IIα by Ronda K. Baker, Ebba U. Kurz, David W. Pyatt, Richard D. Irons, and David J. Kroll Blood Volume 98(3):830-833 August 1, 2001 ©2001 by American Society of Hematology

Catalytic inhibition of topo II and abrogation of etoposide-stabilized, enzyme–DNA complexes by hydroquinone, p-benzoquinone, and catechol. Catalytic inhibition of topo II and abrogation of etoposide-stabilized, enzyme–DNA complexes by hydroquinone, p-benzoquinone, and catechol. The pBR322 plasmid DNA (300 ng) was combined with topo II (6 U) following a 10-minute incubation with compound at indicated concentrations in the absence or presence of 100 μM etoposide. The dose-dependent loss of the relaxed band is indicative of inhibition of overall topo II catalytic activity. Although etoposide alone stabilizes enzyme-linked DNA complexes (indicated by the linear band), coincubation with increasing hydroquinone (A) and p-benzoquinone (B) concentrations antagonizes this effect in a dose-dependent fashion, whereas catechol (C) does not. Ronda K. Baker et al. Blood 2001;98:830-833 ©2001 by American Society of Hematology

Enhancement of the catalytic inhibition of topo II by hydroquinone; 4,4′-biphenol; and catechol following peroxidase activation.The pBR322 plasmid DNA (300 ng) was combined with topo II (6 U) following a 10-minute incubation with activated compound (1, 10, ... Enhancement of the catalytic inhibition of topo II by hydroquinone; 4,4′-biphenol; and catechol following peroxidase activation.The pBR322 plasmid DNA (300 ng) was combined with topo II (6 U) following a 10-minute incubation with activated compound (1, 10, 30, 100, 300 μM) in the absence or presence of 100 μM etoposide. (A) DNA cleavage assay performed with peroxidase-activated hydroquinone. (B) DNA cleavage assay performed with peroxidase-activated 4,4′-biphenol. (C) DNA cleavage assay performed with peroxidase-activated catechol. All samples, including untreated controls, contain identical concentrations of activating agents. Reactions were interpreted as described in Figure 1. Ronda K. Baker et al. Blood 2001;98:830-833 ©2001 by American Society of Hematology

Benzene metabolites inhibit topo II–DNA binding Benzene metabolites inhibit topo II–DNA binding.Oligonucleotides, containing a strong topo II binding site from positions 87 to 126 of pBR322, were annealed and end-labeled with [α-32P]dCTP. Benzene metabolites inhibit topo II–DNA binding.Oligonucleotides, containing a strong topo II binding site from positions 87 to 126 of pBR322, were annealed and end-labeled with [α-32P]dCTP. Nuclear extract (1 μg protein) from HeLa cells was assayed for binding activity to the 32P-labeled binding site in the presence of 0.1 μg poly(dI:dC) • (dI:dC). The DNA-protein complex (bound) was separated from free probe (free) by electrophoresis through a nondenaturing, 4% polyacrylamide gel in 0.25 × Tris borate–EDTA buffer. Nuclear extract was incubated with increasing concentrations (1, 10, 30, 100, and 300 μM) ofp-benzoquinone (A), hydroquinone (B), and peroxidase-activated hydroquinone (C) prior to initiating the reaction with labeled oligonucleotide. Ronda K. Baker et al. Blood 2001;98:830-833 ©2001 by American Society of Hematology