新型苯并噻二唑衍生物对KRAS突变非小细胞肺癌的辐射增敏作用及机制研究  

Radiosensitizing effects of novel benzothiadiazole derivatives on KRAS-mutant non-small cell lung cancers

作　　者：朱知梅 田红旗[1] Zhu Zhimei;Tian Hongqi(Institute of Radiation Medicine, Chinese Academy of Medical Sciences ＆ Peking Union Medical Colleg;Tianjin Key Laboratory of Radiation Medicine and Molecular Nuclear Medicine, Tianjin 300192, China)

机构地区：[1]中国医学科学院北京协和医学院放射医学研究所天津市放射医学与分子核医学重点实验室,天津300192

出　　处：《国际生物医学工程杂志》2018年第2期138-147,共10页International Journal of Biomedical Engineering

基　　金：中国医学科学院北京协和医学院“中央级公益类科研院所基本科研业务费”（2016ZX310199）;中国医学科学院医学与健康科技创新工程（2017-12M-3-019）;中国医学科学院放射医学研究所科研创新团队项目（创新1649）

摘　　要：目的 探究新型苯并噻二唑衍生物HL-095对3种KRAS突变非小细胞肺癌（NSCLC）细胞（H358、A549和H460）的辐射增敏作用及其相关机制.方法 以AZD6244为先导化合物设计合成苯并噻二唑衍生物HL-095,采用丝裂原活化的细胞外信号调节激酶（MEK）1激酶实验检测HL-095对MEK1激酶活性的抑制能力.将H358、A549和H460细胞分别以适宜密度接种培养,分为对照组、HL-095组、照射组、HL-095联合照射组.采用137Csγ射线一次性照射,剂量率为1.02 Gy/min.蛋白质印迹法（Western Blot）检测H358、A549和H460细胞中磷酸化细胞外调节蛋白激酶（p-ERK）的表达水平,克隆形成实验检测细胞增殖,彗星实验分析DNA损伤程度,流式细胞仪检测G2/M期细胞百分比,Western Blot检测A549和H358细胞中检查点激酶1（CHK1）蛋白及其磷酸化水平.结果 HL-095能抑制MEK1的活性.与对照组相比,HL-095组3种细胞（H358、A549和H460）中的p-ERK蛋白表达水平降低,G2/M期细胞百分比降低,其差异均具有统计学意义（均P〈0.01）;A549和H358细胞中DNA损伤修复关键蛋白CHK1的表达及其磷酸化下调.与照射组相比,HL-095联合照射组3种细胞的增殖均受到抑制,DNA损伤亦更为严重,Olive尾距、尾距、尾长和尾部DNA百分比均明显升高,差异均具有统计学意义（均P〈0.05）.结论 作为MEK抑制剂,新型苯并噻二唑衍生物HL-095可通过抑制DNA损伤修复和减少G2/M期阻滞来增强KRAS突变NSCLC细胞的辐射敏感性.Objective To investigate the radiosensitization effects and mechanisms of novel benzothiadiazole derivatives HL-095 to KRAS-mutant non-small cell lung cancer （ NSCLC ） H358 , A549 , and H460 cell lines . Methods The benzothiadiazole derivative HL-095 was designed and synthesized with AZD6244 as the lead compound. The ability of HL-095 to inhibit the activity of MEK kinase was detected by the mitogen-activated extracellular signal-regulated kinase 1 （MEK1）. H358, A549 and H460 cells were inoculated and cultured at appropriate density, and divided into control group, HL-095 group, irradiation group and HL-095 combined irradiation group. One-time irradiation with 137Cs gamma rays was used with a dose rate of 1.02 Gy/min. The expression of phosphorylated extracellular regulatory protein kinase （ p-ERK ） in H358 , A549 and H460 cells was detected by Western Blot. The cell proliferation was detected by cloning assay. The degree of DNA damage was analyzed by Comet assay. The G2/M phase cells were detected by flow cytometry. The level of the checkpoint kinase 1 （CHK1） protein and its phosphorylation in A549 and H358 cells was detected by Western Blot. Results HL-095 can inhibit the activity of MEK1. Compared with the control group, the expression levels of p-ERK protein of H358, A549, and H460） cells in HL-095 group were decreased, the percentages of cells in the G2/M phase were also decreased, and the differences were statistically significant （all P〈0.01）. The expression and phosphorylation of CHK1, a key protein＆nbsp;of DNA damage repair, was down-regulated in A549 and H358 cells. Compared with the irradiated group, the proliferation of the three kinds of cell in HL-095 combined irradiation group was inhibited, the DNA damage was more serious, the Olive tail moment, tail length, tail length and tail DNA percentage were significantly increased, and the differences were statistically significant （all P〈0.05）. Conclusion As a MEK inhibitor, the novel benzothiadiazole derivative HL-095 can

关 键 词：HL-095 MEK抑制剂 非小细胞肺癌 辐射增敏 DNA损伤 

分 类 号：R734.2[医药卫生—肿瘤]