Inhibition of DNA-PK activity by phosphorothioate oligonucleotides

硫代磷酸酯寡核苷酸对 DNA-PK 活性的抑制

• 批准号: 12470184
• 负责人: HOSOI Yoshio
• 金额: $ 8.38万
• 依托单位: The University of Tokyo
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-12470184/
• 关键词: Ionizing radiation; DNA repair; Cancer; DNA-PK; 放射線感受性

Phosphorothioate oligonudeotides and suramin bind to heparin binding proteins including DNA polymerases, and inhibit their functions. In the present study, we report inhibition of DNA-dependent protein kinase (DNA-PK) activity by phosphorothioate oiigonucleotides, suramin and heparin. Inhibitory effect of phosphorothioate oligonudeotides on DNA-PK activity was increased with length and reached a plateau at 36-mer. The base composition of phosphorothioate digonucleotides did not affect the inhibitory effect The inhibitory effect by phosphorothioate oligodeoxycytidine 36-mer can be about 200-fold greater than that by the phosphodiester oligodeoxycytidine 36-mer. The inhibitory effect was also observed with purified DNA-PK, which suggests direct interaction between DNA-PK and phosphorothioate oligonucleotides. DNA-PK will have different binding positions for double-stranded DNA and phosphorothioate oligodeoxycytidine 36-mer because they were not competitive in DNA-PK activation. Suramin and heparin inhibited DNA-PK activity with IC_<50> of 1.7 μM and 0.27 μg/ml respectively. DNA-PK activities and DNA double-stranded breaks (DSBs) repair in cultured cells were significantly suppressed by the treatment with suramin in vivo. Our present observations suggest that suramin mav possibly result in sensitization of cells to ionizing radiation bv inactivation of DNA-PK and the impairment of DSBs repair.

硫代寡核苷酸和苏拉明与包括DNA聚合酶在内的肝素结合蛋白结合，并抑制其功能。在本研究中，我们报道了硫代寡核苷酸、苏拉明和肝素对DNA依赖蛋白激酶(DNA-PK)活性的抑制作用。硫代寡核苷酸对DNA-PK活性的抑制作用随时间的延长而增强，在36-mer时达到平台期。硫代双核苷酸的碱基组成不影响抑制作用，硫代寡脱氧胞苷36-聚体的抑制作用约为磷酸二酯脱氧胞苷36-聚体的200倍。纯化的DNA-PK也有抑制作用，提示DNA-PK与硫代寡核苷酸有直接的相互作用。DNA-PK与双链DNA和硫代寡聚脱氧胞苷36-聚体的结合位置不同，因为它们在DNA-PK的激活过程中不具有竞争性。苏拉明和肝素抑制DNA-PK活性的IC_(50)和IC_(50)分别为1.7μM和0.27μg/ml。苏拉明可显著抑制培养细胞的DNA-PK活性和DNA双链断裂(DSB)修复。我们目前的观察表明，苏拉明MAV可能通过使DNA-PK失活和DSBs修复受损而导致细胞对电离辐射的敏化。

项目成果

Yoshida, M.: "Roles of DNA-dependent protein kinase and ATM in cell-cycle dependent radiation sensitivity in human cells"International Journal of Radiation Biology. 78. 503-512 (2002)

Yoshida, M.：“DNA 依赖性蛋白激酶和 ATM 在人类细胞细胞周期依赖性辐射敏感性中的作用”国际放射生物学杂志。

Enomoto, A.: "Decreased c-Myc expression involved in X-ray-induced apoptotic cell death of human T-cell leukemia cell line MOLT-4"International Journal of Radiation Biology. in press.

Enomoto, A.：“c-Myc 表达降低参与 X 射线诱导的人 T 细胞白血病细胞系 MOLT-4 的细胞凋亡”国际放射生物学杂志。

Tomita, M.: "Wortmannin enhanced X-ray-induced apoptotic cell death through JNK/SAPK activation in human T-cell leukemia MOLT-4 cells"Radiation Research. in press.

Tomita, M.：“渥曼青通过人 T 细胞白血病 MOLT-4 细胞中的 JNK/SAPK 激活增强 X 射线诱导的细胞凋亡”辐射研究。

Hosoi, Y.: "Phosphorothioate oligonucleotides, suramin and heparin inhibit DNA-dependent protein kinase activity"British Journal of Cancer. (in press).

Hosoi, Y.：“硫代磷酸酯寡核苷酸、苏拉明和肝素抑制 DNA 依赖性蛋白激酶活性”英国癌症杂志。

Enomoto, A.: "Decreased c-Myc expression involved in X-ray-induced apoptotic cell death of human T-cell leukemia cell line MOLT-4"International Journal of Radiation Biology. (in press).

Enomoto, A.：“c-Myc 表达降低参与 X 射线诱导的人 T 细胞白血病细胞系 MOLT-4 的细胞凋亡”国际放射生物学杂志。