Olive P L, Banáth J P
British Columbia Cancer Research Centre, Vancouver, British Columbia, Canada.
Cancer Res. 1997 Dec 15;57(24):5528-33.
Multicell spheroids were exposed to DNA-damaging agents with the aim of determining whether prompt DNA damage could be predictive for cell killing and drug resistance. Chinese hamster V79 cells, SiHa human cervical carcinoma cells, and WiDr human colon carcinoma cells were grown as spheroids and exposed to N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), 4-nitroquinoline-1-oxide (4NQO), doxorubicin, etoposide, actinomycin D, 1-(2-nitro-1-imidazolyl)-3-aziridino-2-propanol (RSU 1069), 3-amino-1,2,4-benzotriazine-1,4-dioxide (tirapazamine), and nitrogen mustard. Average DNA damage measured using the alkali comet assay generally correlated with cell killing irrespective of exposure times or drug concentration. However, better predictive power was achieved by using DNA damage levels in individual cells to identify the fraction of cells containing sufficient numbers of DNA strand breaks to cause death. Using this concept of a "threshold" for DNA damage, cell survival could be predicted for exposure to 4NQO, tirapazamine, nitrogen mustard, RSU 1069, and actinomycin D and was largely independent of cell type. The threshold value varied for each drug. For 4NQO, tirapazamine, and RSU 1069, DNA damage equivalent to about 10,000 strand breaks/cell was not toxic to cells of any spheroid type. Conversely, for actinomycin D, any DNA damage above background levels (approximately 100 breaks) was toxic for all three cell types. For some DNA-damaging drugs, the lack of correlation between DNA damage and cell killing was also informative. For etoposide and doxorubicin, no common threshold for cell killing could be determined, consistent with the hypothesis that DNA damage is only one of the actions of these drugs leading to cell death. For MNNG, the tail moment threshold varied significantly for the different spheroid types, probably indicating differences in repair. Overall, for five of the eight drugs, DNA damage measured using the comet assay was an effective and quantitative method of predicting drug cytotoxicity in complex multicelled systems.
将多细胞球体暴露于DNA损伤剂，目的是确定即时DNA损伤是否可预测细胞杀伤和耐药性。将中国仓鼠V79细胞、SiHa人宫颈癌细胞和WiDr人结肠癌细胞培养成球体，并暴露于N-甲基-N'-硝基-N-亚硝基胍（MNNG）、4-硝基喹啉-1-氧化物（4NQO）、阿霉素、依托泊苷、放线菌素D、1-(2-硝基-1-咪唑基)-3-氮丙啶基-2-丙醇（RSU 1069）、3-氨基-1,2,4-苯并三嗪-1,4-二氧化物（替拉扎明）和氮芥。使用碱性彗星试验测量的平均DNA损伤通常与细胞杀伤相关，与暴露时间或药物浓度无关。然而，通过使用单个细胞中的DNA损伤水平来确定含有足够数量DNA链断裂以导致死亡的细胞分数，可以获得更好的预测能力。使用这种DNA损伤“阈值”的概念，可以预测暴露于4NQO、替拉扎明、氮芥、RSU 1069和放线菌素D时的细胞存活情况，并且在很大程度上与细胞类型无关。每种药物的阈值不同。对于4NQO、替拉扎明和RSU 1069，相当于约10000个链断裂/细胞的DNA损伤对任何球体类型的细胞都无毒。相反，对于放线菌素D，任何高于背景水平（约100个断裂）的DNA损伤对所有三种细胞类型都有毒。对于一些DNA损伤药物，DNA损伤与细胞杀伤之间缺乏相关性也提供了信息。对于依托泊苷和阿霉素，无法确定细胞杀伤的共同阈值，这与DNA损伤只是这些药物导致细胞死亡的作用之一的假设一致。对于MNNG，不同球体类型的尾矩阈值差异显著，可能表明修复存在差异。总体而言，对于八种药物中的五种，使用彗星试验测量的DNA损伤是预测复杂多细胞系统中药物细胞毒性的一种有效且定量的方法。
