DIFFERENTIAL POISONING OF TOPOISOMERASES BY MENOGARIL AND NOGALAMYCINDICTATED BY THE MINOR GROOVE-BINDING NOGALOSE SUGAR

The effect of DNA binding on poisoning of human DNA TOP1 has been stud ied using a pair of related anthracyclines which differ only by a noga lose sugar ring. We show that the nogalose sugar ring of nogalamycin, which binds to the minor groove of DNA, plays an important role in aff ecting topoisomerase specific poisoning, Using purified mammalian topo isomerases, menogaril is shown to poison topoisomerase II but not topo isomerase I. By contrast, nogalamycin poisons topoisomerase I but not topoisomerase II. Consistent with the biochemical studies, CEM/VM-1 ce lls which express drug-resistant TOP2 alpha are cross-resistant to men ogaril but not nogalamycin. The mechanism by which nogalamycin poisons topoisomerase I has been studied by analyzing a major topoisomerase I -mediated DNA cleavage site induced by nogalamycin. This site is mappe d to a sequence embedded in an AT-rich region with four scattered GC b ase pairs (bps) (at -10, -6, +2, and +12 positions). GC bps embedded i n AT-rich regions are known to be essential for nogalamycin binding. S urprisingly, DNase I footprinting analysis of nogalamycin-DNA complexe s has revealed a drug-free region from -2 to +9 encompassing the major cleavage site. Our results suggest that nogalamycin, in contrast to c amptothecin, may stimulate TOP1 cleavage by binding to a site(s) dista l to the site of cleavage.