Clinical application of the methods to detect enzyme activities of proteins involved in repair of DNA double strand breaks

细胞中基因组不稳定性的存在在多种器官的多阶段癌变中起着重要作用。DNA双链断裂(DSB)修复途径通过抑制染色体重排来维持基因组的完整性。DNA依赖蛋白激酶(DNA-PK)在DNA双链断裂修复中起重要作用。这项研究的目的是确定未经治疗的癌症患者和无癌症的正常健康志愿者之间DNA-PK活性的差异，以及这如何影响癌症风险。正常人外周血淋巴细胞DNA-PK活性为15.84±4.87pmol，癌症患者为12.28±5.10pmol。两组比较差异有统计学意义(P=0.012)。年龄、吸烟与DNA-PK活性无关。DNA-PK活性与外周血淋巴细胞染色体畸变率之间存在一定的关系。DNA双链断裂修复所必需的蛋白质NBS1在电离辐射诱导DNA损伤后重新定位到亚核结构中，形成电离辐射诱导灶(IRIF)。我们比较了46例散发性乳腺癌患者和30例健康志愿者外周血中的NBS1IRIF。浸润性癌患者照射后24小时每个核的持续NBS1IRIF数显著高于正常健康志愿者。染色体自发畸变率随NBS1IRIF数目的增加而增加，提示NBS1IRIF数目的增加可能与染色体不稳定性有关。NBS1IRIF数与DNA-PK活性呈负相关，在DNA DSB修复的另一种机制--非同源末端连接途径(NHEJ)中起重要作用，提示HR与NHEJ在DNA DSB修复机制中密切相关。

The presence of genomic instability in cells is known to play an important role in the multistage carcinogenesis of various organs. The DNA double strand breaks (DSBs) repair pathway has been implicated in maintaining genomic integrity via suppression of chromosomal rearrangements. DNA-dependent protein kinase (DNA-PK) has an important role in DNA DSBs repair. The purpose of this study was to determine how DNA-PK activity varies among untreated cancer patients and cancer-free normal healthy volunteers and how this affects cancer risk. DNA-PK activities of peripheral blood lymphocytes (PBL) in normal volunteers were 15.84 ± 4.87 pmol and those of cancer patients were 12.28 ± 5.10 pmol. There was the significant difference between them (p=0.012). Age and smoking had no association with DNA-PK activity A relationship between DNA-PK acitvity and chromosome aberration in PBL was shown to exist. The frequency of excess fragment increased as the DNA-PK activity decreased.NBS1, a protein essential for DNA double strand break repair, re-localizes into subnuclear structures upon the induction of DNA damage by ionizing radiation, forming ionizing radiation-induced foci (IRIF). We compared NBS1 IRIF in PBL taken from 46 sporadic breast cancer patients and 30 cancer-free healthy volunteers. The number of persistent NBS1 IRIF per nucleus at 24 hours after irradiation in invasive cancer patients was significantly higher than in normal healthy volunteers. The frequency of spontaneous chromosome aberration increased as the number of persistent NBS1 IRIF increased, indicating that number of persistent NBS1 IRIF might be associated with chromosome instability. There was also an inverse correlation between NBS1 IRIF number and the activity of DNA-PK, which plays an important role in the non-homologous end-joining pathway (NHEJ), another mechanism of DNA DSB repair, indicating close interrelationship between HR and NHEJ in DNA DSB repair mechanism.