INHIBITION OF HUMAN DNA TOPOISOMERASE-II BY HYDROQUINONE AND P-BENZOQUINONE, REACTIVE METABOLITES OF BENZENE

Chronic exposure of humans to benzene (BZ) causes acute myeloid leukem ia (AML). Both BZ and therapy-related secondary AML are characterized by chromosomal translocations that may occur by inappropriate recombin ational events. DNA topoisomerase II (topo II) is an essential sulfhyd ryl (SH)-dependent endonuclease required for replication, recombinatio n, chromosome segregation, and chromosome structure. Topo II cleaves D NA at purine(R)/pyrimidine(Y) repeat sequences that have been shown to be highly recombinogenic in vivo. Certain antineoplastic drugs stabil ize topo II-DNA cleavage complexes at RY repeat sequences, which leads to translocations of the type observed in leukemia. Hydroquinone (HQ) is metabolized to p-benzoquinone (BQ) in a peroxidase-mediated reacti on in myeloid progenitor cells. BQ interacts with SH groups of SH-depe ndent enzymes. Consequently, the aims of this research were to determi ne whether HQ and BQ are topo II inhibitors. The ability of the compou nds to inhibit the activity of topo II was tested using an assay syste m that depends on the conversion, by homogeneous human topo II, of cat enated kinetoplast DNA into open and/or nicked open circular DNA that can be separated from the catenated DNA by electrophoresis in a 1% aga rose-ethidium bromide gel. We provide preliminary data that indicate t hat both HQ and BQ cause a time and concentration (mu M)-dependent inh ibition of topo II activity. These compounds, which potentially can fo rm adducts with DNA, have no effect on the migration of the supercoile d and open circular forms in the electrophoretic gradient, and BQ-addu cted KDNA can be decatenated by topo II. Using a pRYG plasmid DNA with a single RY repeat as a cleavage site, it was determined that BQ does not stimulate the production of linear DNA indicative of an inhibitio n of topo II religation of strand breaks by stabilization of the coval ent topo II-DNA cleavage complex. Rather, BQ most probably inhibits th e SH-dependent topo II by binding to an essential SH group. The inhibi tion of topo II by BQ has implications for the formation of deleteriou s translocations that may be involved in BZ-induced initiation of leuk emogenesis.