Examination of the Impact of Copper(II) α-(N)-Heterocyclic Thiosemicarbaione Complexes on DNA Topoisomerase IIα

Type II DNA topoisomerases resolve topological knots and tangles in DNA that result from routine cellular processes and are effective targets for anticancer therapeutics. To this end, thiosemicarbazones have been identified as having the ability to kill cancer cells from several cell lines: Literature evidence suggests that at least some thiosemicarbazones have an impact on topoisomerase II activity. However, the mechanism is not as clearly defined. Therefore, we set out to analyze the activity of four alpha-(N)-heterocyclic thiosemicarbazone compounds against topoisomerase II alpha. The ligands, acetylpyridine-ethylthiosemicarbazone (APY-ETSC) and acetylpyrazine-methylthiosemi-carbazone (APZ-MTSC), and their copper(II) [Cu(II)] complexes [Cu(APY-ETSC)Cl] and [Cu(APZ-MTSC)Cl] were examined for the ability to impact the catalytic cycle of human topoisomerase Ha. Both [Cu(APY-ETSC)Cl] and [Cu(APZ-MTSC)Cl] were More effective at inhibiting DNA relaxation compared with the ligands alone. Further, both [Cu(APY-ETSC)Cl] and [Cu(APZ-MTSC)Cl] increased double-stranded DNA cleavage levels without inhibiting topoisomerase II alpha-mediated DNA ligation. The Cu(H) complexes inactivate enzyme activity over time suggesting a I critical interaction with the enzyme. Additionally, we found that the Cu(II)-thiosemicarbazone complexes do not significantly impact DNA cleavage by the catalytic core of the enzyme. This evidence is supported by the fact that both [Cu(APY-ETSC)Cl] and [Cu(APZ-MTSC)Cl], and to a lesser extent the ligands, inhibit topoisomerase II alpha-mediated ATP hydrolysis. Based upon kinetic analysis, the Cu(II) complexes appear to be noncompetitive inhibitors of the ATPase domain of topoisomerase Ha. Taken together, our results provide evidence that Cu(II) complexes of alpha-(N)-heterocyclic thiosemicarbazones catalytically inhibit the enzyme through the ATPase domain but also promote double-stranded DNA cleavage by the enzyme.