Dote Hideaki, Cerna David, Burgan William E, Carter Donna J, Cerra Michael A, Hollingshead Melinda G, Camphausen Kevin, Tofilon Philip J
Molecular Radiation Therapeutics Branch, National Cancer Institute, Bethesda, Maryland, USA.
Clin Cancer Res. 2005 Jun 15;11(12):4571-9. doi: 10.1158/1078-0432.CCR-05-0050.
Aberrant DNA hypermethylation is a frequent finding in tumor cells, which has suggested that inhibition of DNA methylation may be an effective cancer treatment strategy. Because DNA methylation affects gene expression and chromatin structure, parameters considered to influence radioresponse, we investigated the effects of the DNA methylation inhibitor zebularine on the radiosensitivity of human tumor cells. Three human tumor cell lines were used in this study (MiaPaCa, DU145, and U251) and the methylation status of three genes frequently hypermethylated in tumor cells (RASSF1A, HIC-1, and 14-3-3sigma) was determined as a function of zebularine exposure. Zebularine resulted in DNA demethylation in a time-dependent manner, with the maximum loss of methylation detected by 48 hours. Treatment of cells with zebularine for 48 hours also resulted in an increase in radiosensitivity with dose enhancement factors of >1.5. As a measure of radiation-induced DNA damage, gammaH2AX expression was determined. Whereas zebularine had no effect on radiation-induced gammaH2AX foci at 1 hour, the number of gammaH2AX foci per cell was significantly greater in the zebularine-treated cells at 24 hours after irradiation, suggesting the presence of unrepaired DNA damage. Zebularine administration to mice reactivated gene expression in U251 xenografts; irradiation of U251 tumors in mice treated with zebularine resulted in an increase in radiation-induced tumor growth delay. These results indicate that zebularine can enhance tumor cell radiosensitivity in vitro and in vivo and suggest that this effect may involve an inhibition of DNA repair.
异常的DNA高甲基化在肿瘤细胞中很常见，这表明抑制DNA甲基化可能是一种有效的癌症治疗策略。由于DNA甲基化会影响基因表达和染色质结构，而这些参数被认为会影响放射反应，因此我们研究了DNA甲基化抑制剂泽布勒林对人肿瘤细胞放射敏感性的影响。本研究使用了三种人肿瘤细胞系（MiaPaCa、DU145和U251），并确定了肿瘤细胞中经常发生高甲基化的三个基因（RASSF1A、HIC-1和14-3-3sigma）的甲基化状态与泽布勒林暴露量的关系。泽布勒林以时间依赖性方式导致DNA去甲基化，48小时时检测到最大甲基化损失。用泽布勒林处理细胞48小时也导致放射敏感性增加，剂量增强因子>1.5。作为辐射诱导DNA损伤的指标，测定了γH2AX的表达。虽然泽布勒林在1小时时对辐射诱导的γH2AX焦点没有影响，但在照射后24小时，经泽布勒林处理的细胞中每个细胞的γH2AX焦点数量明显更多，这表明存在未修复的DNA损伤。给小鼠施用泽布勒林可重新激活U251异种移植物中的基因表达；照射用泽布勒林处理的小鼠中的U251肿瘤会导致辐射诱导的肿瘤生长延迟增加。这些结果表明，泽布勒林可以在体外和体内增强肿瘤细胞的放射敏感性，并表明这种作用可能涉及对DNA修复的抑制。
