Long B H, Musial S T, Brattain M G
Cancer Res. 1986 Aug;46(8):3809-16.
Evidence suggests that the anticancer agents etoposide (VP16-213) and teniposide (VM26) produce DNA breaks and cytotoxicity by interaction with type II topoisomerase. Therefore, levels of type II topoisomerase may influence sensitivity to VP16-213 and VM26. We have characterized four lung carcinoma-derived cell lines for natural sensitivity or resistance to VP16-213 and VM26. Included in this study were two small cell lung carcinoma lines (SW900 and SW1271), an adenocarcinoma line (A549), and a large cell carcinoma (H157). SW1271 was the most sensitive line with a median inhibitory concentration for cell proliferation of 0.5 microM for VM26 and 2.7 microM for VP16-213, and SW900 was the most resistant with median inhibitory concentration values of 2.0 and 16 microM, respectively. A549 and H157 cells were intermediate in sensitivity to these drugs. Alkaline elution techniques were used to study in vivo formation and repair of single and double strand DNA breaks. Single strand DNA breaks were observed in SW1271 cells exposed to as little as 10 nM VM26 or 100 nM VP16-213 for 1 h, whereas SW900 cells required exposure to 10-fold higher concentrations of VM26 or VP16-213 to produce similar results. Single strand DNA breaks predominated only in SW1271 and A549 cells and then, only at low drug concentrations, whereas the ratios between single and double strand DNA breaks decreased at higher drug concentrations. Plots of cytotoxicity versus single and double strand DNA breakage revealed that cytotoxicity produced by both drugs was more closely related to double strand DNA break formation in all four cell lines. DNA breaks appeared rapidly upon addition of drug, reaching plateaus in DNA breaks within 30 min, and repair of both single and double strand DNA breaks occurred rapidly with time to repair one-half of the DNA breaks of 20 to 60 min in all four cell lines upon removal of drug, arguing against repair as a mechanism for drug resistance. DNA breakage was also observed in nuclei isolated from SW900 and SW1271 cells in similar magnitude to that observed in the respective cells. Results indicate that DNA breakage plateaus may reflect a steady-state equilibrium established between the drug and its nuclear target, possibly type II topoisomerase, and suggest that natural resistance to VP16-213 and VM26 may be due to different enzyme levels in sensitive and naturally resistant cells.
有证据表明，抗癌药物依托泊苷（VP16 - 213）和替尼泊苷（VM26）通过与II型拓扑异构酶相互作用产生DNA断裂并具有细胞毒性。因此，II型拓扑异构酶的水平可能会影响对VP16 - 213和VM26的敏感性。我们已对四种源自肺癌的细胞系进行了表征，以确定它们对VP16 - 213和VM26的天然敏感性或抗性。本研究包括两个小细胞肺癌细胞系（SW900和SW1271）、一个腺癌细胞系（A549）和一个大细胞癌细胞系（H157）。SW1271是最敏感的细胞系，VM26对细胞增殖的半数抑制浓度为0.5微摩尔，VP16 - 213为2.7微摩尔；SW900是最耐药的，VM26和VP16 - 213的半数抑制浓度值分别为2.0和16微摩尔。A549和H157细胞对这些药物的敏感性处于中等水平。采用碱性洗脱技术研究体内单链和双链DNA断裂的形成及修复情况。在暴露于低至10纳摩尔VM26或100纳摩尔VP16 - 213 1小时的SW1271细胞中观察到单链DNA断裂，而SW900细胞需要暴露于高10倍浓度的VM26或VP16 - 213才能产生类似结果。仅在SW1271和A549细胞中，且仅在低药物浓度下，单链DNA断裂占主导，而在较高药物浓度下单链和双链DNA断裂的比例下降。细胞毒性与单链和双链DNA断裂的关系图显示，在所有四个细胞系中，两种药物产生的细胞毒性与双链DNA断裂形成的相关性更强。加入药物后DNA断裂迅速出现，30分钟内达到DNA断裂的平台期，并且在去除药物后，所有四个细胞系中，单链和双链DNA断裂的修复都随着时间迅速发生，修复一半DNA断裂所需时间为20至60分钟，这表明修复不是耐药的机制。在从SW900和SW1271细胞中分离出的细胞核中也观察到了类似程度的DNA断裂，与在相应细胞中观察到的情况相似。结果表明，DNA断裂平台期可能反映了药物与其核靶点（可能是II型拓扑异构酶）之间建立的稳态平衡，并表明对VP16 - 213和VM26的天然抗性可能是由于敏感细胞和天然耐药细胞中酶水平的差异。
