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Imaging of DNA damage repair using 53BP1 protein and establishment of therapeutic strategy for cancer treatment.

使用 53BP1 蛋白进行 DNA 损伤修复成像并建立癌症治疗策略。

基本信息

批准号：
23659587
负责人：
KURIMASA Akihiro
金额：
$ 2.33万
依托单位：
Tottori University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Challenging Exploratory Research
财政年份：
2011
资助国家：
日本
起止时间：
2011 至 2012
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-23659587/
关键词：
放射線腫瘍学バイオセンサー DNA修復細胞周期抗がん剤放射線

项目摘要

Tumor suppressor p53 Binding Protein 1(53BP1) is recruited rapidly to sites of DNA double-strand breaks (DSBs) and participates in the cellular response to DNA damage. We exploited property of 53BP1 that accumulate in DSB sites and developed biosensor detecting DNA damage using the specical form of 53BP1-GFP fusion protein that could express stably in the cell. Futhermore, we built a system that can identify the difference of cell cycle stage by intranuclear localization patterns of PCNA-DsRed. U2OS cell lines (U2RDP-LE53-21) expressing both these fluorescent fusion proteins make it possible to observe DSBs occurred in different cell cycle stage, these cells remain living. Here, we show how 53BP1 foci are observed in normal culturing conditions, and also how DSBs 53BP1 foci are induced by chemical agents such as Neocarcinostatine (NCS) or DNA Topoisomerase I inhibitor Camptothesin (CPT) that are well-known DSBs inducers. The number of 53BP1 foci was increased by these chemical agents and cell cycle distribution was prolonged compare to cells in normal culture condition. U2OS cell lines may be available for the screening chemotherapy drugs and radiosensitizers that induce DSBs in cancer cells.

肿瘤抑制因子p53结合蛋白1（53 BP 1）被迅速募集到DNA双链断裂（DSB）位点，并参与细胞对DNA损伤的反应。利用53 BP 1在DSB位点的聚集特性，利用53 BP 1-GFP融合蛋白在细胞中稳定表达的特异性形式，研制了检测DNA损伤的生物传感器。此外，我们建立了一个可以通过PCNA-DsRed的核内定位模式来识别细胞周期阶段差异的系统。表达这两种荧光融合蛋白的U2 OS细胞系（U2 RDP-LE 53 -21）使观察不同细胞周期阶段发生的DSB成为可能，这些细胞保持存活。在这里，我们展示了如何在正常培养条件下观察到53 BP 1病灶，以及DSB 53 BP 1病灶如何被化学试剂如新癌抑制素（NCS）或DNA拓扑异构酶I抑制剂Camptothesin（CPT）诱导，这些化学试剂是众所周知的DSB诱导剂。与正常培养条件下的细胞相比，这些化学试剂增加了53 BP 1病灶的数量，并延长了细胞周期分布。U2 OS细胞系可用于筛选诱导癌细胞DSB的化疗药物和放射增敏剂。