Nitiss J L, Liu Y X, Hsiung Y
Division of Hematology/Oncology, Children's Hospital of Los Angeles, California 90027.
Cancer Res. 1993 Jan 1;53(1):89-93.
We have developed a simple system for determining the specific contribution of topoisomerase II targeting to the cytotoxic activity of a drug. We have constructed yeast strains that are permeable to anti-topoisomerase II drugs, carry a DNA repair mutation, rad52, and also have a temperature sensitive topoisomerase II mutation, top2-1. Strains carrying the top2-1 mutation have nearly normal topoisomerase II activity at 25 degrees C but less than 10% of wild type activity at 36 degrees C. We find that at a semi-permissive temperature (30 degrees C), there is sufficient topoisomerase II activity for viability; but since the topoisomerase II activity is greatly reduced, the cells are very resistant to anti-topoisomerase II drugs. Conversely, such cells are hypersensitive to the topoisomerase I inhibitor camptothecin. These results provide strong support for the model that drug stabilized DNA cleavage, rather than a lack of enzyme activity, is responsible for cell killing by eukaryotic anti-topoisomerase II agents. They also show that there is a minimum level of topoisomerase II activity in yeast that is consistent with viability but also allows a high degree of resistance to anti-topoisomerase II agents.
我们开发了一个简单的系统，用于确定靶向拓扑异构酶II对药物细胞毒性活性的具体贡献。我们构建了对抗拓扑异构酶II药物具有通透性、携带DNA修复突变rad52且具有温度敏感型拓扑异构酶II突变top2-1的酵母菌株。携带top2-1突变的菌株在25摄氏度时具有近乎正常的拓扑异构酶II活性，但在36摄氏度时活性不到野生型的10%。我们发现，在半允许温度（30摄氏度）下，有足够的拓扑异构酶II活性维持细胞存活；但由于拓扑异构酶II活性大幅降低，细胞对抗拓扑异构酶II药物具有很强的抗性。相反，这类细胞对拓扑异构酶I抑制剂喜树碱高度敏感。这些结果为以下模型提供了有力支持：药物稳定的DNA切割而非酶活性的缺乏是真核抗拓扑异构酶II药物导致细胞死亡的原因。它们还表明，酵母中存在拓扑异构酶II活性的最低水平，这一水平既与细胞存活一致，又使细胞对抗拓扑异构酶II药物具有高度抗性。
