Institute for Cell and Molecular Biosciences, Newcastle University, Newcastle upon Tyne, United Kingdom (M.A., R.L.S., C.R., K.C.L., I.G.C., C.A.A.) and Institute of Genetic Medicine, Newcastle University, International Centre for Life, Central Parkway, Newcastle upon Tyne, United Kingdom (D.C.L., L.A.).
Institute for Cell and Molecular Biosciences, Newcastle University, Newcastle upon Tyne, United Kingdom (M.A., R.L.S., C.R., K.C.L., I.G.C., C.A.A.) and Institute of Genetic Medicine, Newcastle University, International Centre for Life, Central Parkway, Newcastle upon Tyne, United Kingdom (D.C.L., L.A.)
Mol Pharmacol. 2019 Oct;96(4):475-484. doi: 10.1124/mol.119.117259. Epub 2019 Aug 9.
Topoisomerase II (TOP2) poisons are effective cytotoxic anticancer agents that stabilize the normally transient TOP2-DNA covalent complexes formed during the enzyme reaction cycle. These drugs include etoposide, mitoxantrone, and the anthracyclines doxorubicin and epirubicin. Anthracyclines also exert cell-killing activity via TOP2-independent mechanisms, including DNA adduct formation, redox activity, and lipid peroxidation. Here, we show that anthracyclines and another intercalating TOP2 poison, mitoxantrone, stabilize TOP2-DNA covalent complexes less efficiently than etoposide, and at higher concentrations they suppress the formation of TOP2-DNA covalent complexes, thus behaving as TOP2 poisons at low concentration and inhibitors at high concentration. We used induced pluripotent stem cell (iPSC)-derived human cardiomyocytes as a model to study anthracycline-induced damage in cardiac cells. Using immunofluorescence, our study is the first to demonstrate the presence of topoisomerase II (TOP2B) as the only TOP2 isoform in iPSC-derived cardiomyocytes. In these cells, etoposide robustly induced TOP2B covalent complexes, but we could not detect doxorubicin-induced TOP2-DNA complexes, and doxorubicin suppressed etoposide-induced TOP2-DNA complexes. In vitro, etoposide-stabilized DNA cleavage was attenuated by doxorubicin, epirubicin, or mitoxantrone. Clinical use of anthracyclines is associated with cardiotoxicity. The observations in this study have potentially important clinical consequences regarding the effectiveness of anticancer treatment regimens when TOP2-targeting drugs are used in combination. These observations suggest that inhibition of TOP2B activity, rather than DNA damage resulting from TOP2 poisoning, may play a role in doxorubicin cardiotoxicity. SIGNIFICANCE STATEMENT: We show that anthracyclines and mitoxantrone act as topoisomerase II (TOP2) poisons at low concentration but attenuate TOP2 activity at higher concentration, both in cells and in in vitro cleavage experiments. Inhibition of type II topoisomerases suppresses the action of other drugs that poison TOP2. Thus, combinations containing anthracyclines or mitoxantrone and etoposide may reduce the activity of etoposide as a TOP2 poison and thus reduce the efficacy of drug combinations.
拓扑异构酶 II（TOP2）抑制剂是有效的细胞毒性抗癌药物，可稳定酶反应循环过程中形成的正常瞬时 TOP2-DNA 共价复合物。这些药物包括依托泊苷、米托蒽醌和蒽环类药物阿霉素和表阿霉素。蒽环类药物还通过 TOP2 非依赖性机制发挥细胞杀伤活性，包括 DNA 加合物形成、氧化还原活性和脂质过氧化。在这里，我们表明蒽环类药物和另一种嵌入 TOP2 抑制剂米托蒽醌不如依托泊苷有效地稳定 TOP2-DNA 共价复合物，并且在较高浓度下，它们抑制 TOP2-DNA 共价复合物的形成，因此在低浓度下表现为 TOP2 抑制剂，在高浓度下表现为 TOP2 抑制剂。我们使用诱导多能干细胞（iPSC）衍生的人心肌细胞作为模型来研究心脏细胞中蒽环类药物引起的损伤。通过免疫荧光，我们的研究首次证明在 iPSC 衍生的心肌细胞中只有拓扑异构酶 II（TOP2B）作为唯一的 TOP2 同工酶的存在。在这些细胞中，依托泊苷强烈诱导 TOP2B 共价复合物，但我们无法检测到阿霉素诱导的 TOP2-DNA 复合物，并且阿霉素抑制依托泊苷诱导的 TOP2-DNA 复合物。在体外，阿霉素稳定的 DNA 断裂被阿霉素、表阿霉素或米托蒽醌减弱。蒽环类药物的临床应用与心脏毒性有关。这项研究的观察结果对于在使用 TOP2 靶向药物联合治疗时，抗癌治疗方案的有效性具有潜在的重要临床意义。这些观察结果表明，TOP2B 活性的抑制，而不是 TOP2 中毒引起的 DNA 损伤，可能在阿霉素心脏毒性中发挥作用。 意义陈述：我们表明，蒽环类药物和米托蒽醌在低浓度下作为拓扑异构酶 II（TOP2）抑制剂起作用，但在较高浓度下会减弱 TOP2 活性，无论是在细胞中还是在体外切割实验中。II 型拓扑异构酶抑制剂抑制其他 TOP2 中毒药物的作用。因此，含有蒽环类药物或米托蒽醌和依托泊苷的组合可能会降低依托泊苷作为 TOP2 抑制剂的活性，从而降低药物组合的疗效。
