Clinical Pharmacy, Institute of Pharmacy, Hamburg University, Bundesstrasse 45, Hamburg, Germany.
Apoptosis. 2011 Apr;16(4):359-69. doi: 10.1007/s10495-010-0564-y.
For DNA targeting anticancer drugs, cellular DNA repair mechanisms may cause resistance and hamper the therapeutic outcome. DNA damage induced by topoisomerase IIα inhibitors like etoposide and anthracyclines, which are a mainstay of cancer therapy, is also repaired in many cell types, but the impact and precise mechanisms of this repair are still obscure. To investigate the DNA damage response of human adenocarcinoma HT29-cells to doxorubicin and to compare the involvement of Ku70 and Rad51 in the repair of doxorubicin- versus etoposide-induced DNA damage, we assessed cell cycle distribution and cell death, DNA damage, proteins relevant for repair by homologous recombination and non-homologous end-joining, and clonogenicity following exposure to doxorubicin at clinically achievable concentrations. Also, we assessed changes in the repair kinetics after siRNA-mediated attenuation of Ku70 or Rad51 expression. We found that exposure to doxorubicin for 24 h induced a substantial amount of DNA damage that was largely repaired when doxorubicin was removed and the cells were maintained in drug-free medium. Nevertheless, a pronounced G(2)/M arrest occurred at times when repair was maximal. This was followed by a distinct increase in cell death and loss of clonogenicity. In this regard, responses to doxorubicin and etoposide were similar. However, distinct differences in the repair process following doxorubicin versus etoposide were seen in concentration dependency, time-course and requirement of Ku70 and Rad51 proteins. In spite of the shared molecular target of doxorubicin and etoposide, DNA lesions induced by these compounds are repaired differently.
对于针对 DNA 的抗癌药物，细胞 DNA 修复机制可能会导致耐药性并影响治疗效果。拓扑异构酶 IIα 抑制剂（如依托泊苷和蒽环类药物）诱导的 DNA 损伤也在许多细胞类型中得到修复，这些药物是癌症治疗的主要药物，但这种修复的影响和精确机制仍不清楚。为了研究人结肠癌 HT29 细胞对阿霉素的 DNA 损伤反应，并比较 Ku70 和 Rad51 在修复阿霉素与依托泊苷诱导的 DNA 损伤中的作用，我们评估了细胞周期分布和细胞死亡、DNA 损伤、同源重组和非同源末端连接修复相关的蛋白质，以及在临床可达到的浓度下暴露于阿霉素后的集落形成能力。此外，我们还评估了在 Ku70 或 Rad51 表达被 siRNA 下调后修复动力学的变化。我们发现，暴露于阿霉素 24 小时会引起大量的 DNA 损伤，当阿霉素被去除且细胞在无药物的培养基中培养时，大部分的损伤会被修复。然而，在修复达到最大值时，会出现明显的 G2/M 期阻滞。随后，细胞死亡和集落形成能力明显下降。在这方面，阿霉素和依托泊苷的反应相似。然而，在阿霉素与依托泊苷修复过程中，在浓度依赖性、时间过程和 Ku70 和 Rad51 蛋白的需求方面存在明显的差异。尽管阿霉素和依托泊苷的分子靶标相同，但这些化合物诱导的 DNA 损伤的修复方式不同。
