Teniposide-resistant CEM cells, which express mutant DNA topoisomerase II alpha, when treated with non-complex-stabilizing inhibitors of the enzyme, display no cross-resistance and reveal aberrant functions of the mutant enzyme.
We have examined the effects of a group of DNA topoisomerase II (topo II) inhibitors, merbarone, aclarubicin, SN22995, RP60475F, and fostriecin, in CCRF-CEM cells and two sublines, CEM/VM-1 and CEM/VM-1-5, that were selected for increasing resistance to teniposide (VM-26). The teniposide-resistant sublines have been termed "at-MDR" for altered topo II-associated multidrug resistance. These topo II inhibitors differ from the "classic" inhibitors such as teniposide in that they do not stabilize DNA-topo II complexes. In this study, we found that our at-MDR cell lines express little or no cross-resistance to these "non-classic" topo II inhibitors. Merbarone and SN22995 inhibited VM-26-mediated DNA-topo II complexes in CEM cells only when they were added before VM-26. Since they did not deplete topo II protein, it suggested that these drugs may inhibit topo II activity before the enzyme binds to DNA, thereby preventing stabilization of VM-26-mediated topo II-DNA complexes. Continuous exposure of CEM cells to merbarone, SN22995, or VM-26 caused G2 arrest, as determined by flow cytometry. Likewise, at-MDR cells continuously treated with VM-26 also arrested in G2. By contrast, treatment of at-MDR cells with either merbarone or SN22995 produced a qualitatively different pattern; the at-MDR cells first accumulated in G2 but then escaped the G2 block and proceeded into mitosis with elongated and intertwined chromosomes but failed to divide. Their DNA was re-replicated, however, and the cells eventually accumulated at the 8N DNA stage. Given that both wild-type and mutant topo II alpha alleles are expressed in the at-MDR cells (B. Y. Bugg, M. K. Danks, W. T. Beck, and D. P. Suttle. Expression of a mutant DNA topoisomerase II in CCRF-CEM human leukemic cells selected for resistance to teniposide. Proc. Natl. Acad. Sci. USA, 88: 7654-7658, 1991), we hypothesize that drugs such as merbarone may inhibit the activity of wild-type topo II alpha, allowing the aberrant activity of the mutant enzyme to be revealed during chromosome condensation and sister chromatid segregation.
Chen
M
M
Department of Molecular Pharmacology, St. Jude Children's Research Hospital, Memphis, Tennessee 38101.
Beck
W T
WT
D002453:Cell Cycle / Q000187:drug effects; D002875:Chromosomes / Q000187:drug effects / Q000648:ultrastructure; D004247:DNA / Q000378:metabolism; D004250:DNA Topoisomerases, Type II / Q000235:genetics; D004351:Drug Resistance; D006801:Humans; D007938:Leukemia / Q000188:drug therapy / Q000473:pathology; D009154:Mutation; D011506:Proteins / Q000378:metabolism; D013713:Teniposide / Q000494:pharmacology*; D013858:Thiobarbiturates / Q000494:pharmacology; D059005:Topoisomerase II Inhibitors; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000648:ultrastructure
D011506:Proteins; D013858:Thiobarbiturates; D059005:Topoisomerase II Inhibitors; D004247:DNA; D013713:Teniposide; D004250:DNA Topoisomerases, Type II; C045459:merbarone