Laboratory of Chemical Carcinogenesis, Chulabhorn Research Institute, Bangkok, Thailand.
Laboratory of Environmental Toxicology, Chulabhorn Research Institute, Bangkok, Thailand.
J Pharm Pharmacol. 2021 Mar 1;73(1):40-51. doi: 10.1093/jpp/rgaa050.
To investigate whether the inhibitions of ataxia-telangiectasia mutated (ATM) and ATM- and Rad3-related (ATR) kinases by their specific inhibitors, KU-55933 and VE-821, respectively, are able to promote the cytotoxic activity of genotoxic agents including gemcitabine, 5-Fluorouracil, cisplatin and doxorubicin, in cholangiocarcinoma (CCA) and immortalized cholangiocyte cell lines.
Cell viability of cells treated with DNA damaging agents, alone and in combination with KU-55933 and VE-821, was determined by MTT assay. The changes of cell cycle distribution were evaluated by flow cytometry analysis. Colony formation was conducted to assess the effects of KU-55933 and VE-821 on cell proliferation. The levels of protein expression and phosphorylation were examined by western blot analysis.
The cytotoxic effects of DNA damaging agents varied among CCA cell lines. Each DNA damaging drug induced different phases of the cell cycle in CCA cells. The combinations of both KU-55933 and VE-821 with DNA damaging agents promoted more cytotoxic activity than single inhibition in some CCA cell lines. ATM and ATR inhibitors decreased the effects of DNA damaging agent-induced ATM-Chk2 and ATR-Chk1 activations in CCA cells.
Inhibitions of ATM and ATR potentiated the cytotoxic effects of DNA damaging agents in CCA cells, especially p53 defective HuCCA1 and RMCC1 cell lines.
研究特异性抑制 ATM 和 ATR 激酶（分别为 KU-55933 和 VE-821）是否能够增强包括吉西他滨、5-氟尿嘧啶、顺铂和阿霉素在内的致瘤药物对胆管癌细胞系和永生化胆管细胞系的细胞毒性作用。
通过 MTT 法测定单独和联合使用 KU-55933 和 VE-821 处理的细胞中 DNA 损伤剂的细胞活力。通过流式细胞术分析评估细胞周期分布的变化。通过集落形成实验评估 KU-55933 和 VE-821 对细胞增殖的影响。通过 Western blot 分析检测蛋白表达和磷酸化水平。
在胆管癌细胞系中，DNA 损伤剂的细胞毒性作用存在差异。每种 DNA 损伤药物在胆管癌细胞中诱导不同的细胞周期阶段。在一些胆管癌细胞系中，KU-55933 和 VE-821 与 DNA 损伤药物的联合应用比单独抑制作用产生了更强的细胞毒性作用。ATM 和 ATR 抑制剂降低了 DNA 损伤剂诱导的 ATM-Chk2 和 ATR-Chk1 在胆管癌细胞中的激活作用。
在 p53 缺陷型 HuCCA1 和 RMCC1 细胞系等胆管癌细胞中，抑制 ATM 和 ATR 增强了 DNA 损伤剂的细胞毒性作用。
