The combination of temsirolimus and chloroquine increases radiosensitivity in colorectal cancer cells
The combination of temsirolimus and chloroquine increases radiosensitivity in colorectal cancer cells
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DOI：
10.3892/or.2019.7134
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发表时间：
2019-07-01
期刊：
影响因子：
4.2
通讯作者：
Nozawa, Hiroaki
中科院分区：
文献类型：
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作者：
Shiratori, Hiroshi;Kawai, Kazushige;Nozawa, Hiroaki
PI 3 K/AKT/mTOR通路和自噬在肿瘤放射抗性中起重要作用。本研究的目的是研究mTOR抑制剂坦罗莫司（TEM）和自噬抑制剂氯喹（CQ）的组合是否可以增加结直肠癌（CRC）细胞的放射敏感性。使用CRC细胞系SW 480和HT-29中的克隆形成测定来检查TEM和/或CQ作为放射增敏剂的功效。通过蛋白质印迹分析评估了mTOR下游蛋白S6和4 E-BP 1的磷酸化亚型的表达水平，以及自噬相关蛋白p62和LC 3的表达水平。吖啶橙染色的细胞中检测到酸性细胞器的形成。流式细胞术检测细胞凋亡和caspase活性。结果表明，电离辐射（IR）可激活mTOR下游蛋白，诱导自噬。在克隆形成试验中，TEM和CQ均不影响IR的功效，而它们的组合显著增加IR的剂量依赖性功效。TEM抑制mTOR下游蛋白的磷酸化并诱导自噬。CQ在晚期抑制自噬，而不影响mTOR的下游蛋白。TEM和CQ抑制mTOR下游蛋白的磷酸化和自噬。细胞死亡分析显示，TEM和CQ的组合强烈诱导细胞凋亡暴露于IR。总之，TEM和CQ的组合通过共同抑制mTOR和自噬增加CRC细胞的放射敏感性。
The PI3K/AKT/mTOR pathway and autophagy are known to play important roles in cancer radioresistance. The aim of the present study was to investigate whether the combination of temsirolimus (TEM), an mTOR inhibitor, and chloroquine (CQ), an autophagy inhibitor, can increase radiosensitivity in colorectal cancer (CRC) cells. The efficacies of TEM and/or CQ as radiosensitizers were examined using clonogenic assays in CRC cell lines SW480 and HT-29. The expression levels of the phosphorylated isoforms of S6 and 4E-BP1, downstream proteins of mTOR, as well as the expression levels of p62 and LC3, autophagy-related proteins, were assessed by western blot analysis. The formation of acidic organelles was detected in acridine orange-stained cells. Apoptosis and caspase activity were assessed using flow cytometry. The results revealed that ionizing radiation (IR) activated the downstream proteins of mTOR and induced autophagy. In the clonogenic assays, neither TEM nor CQ influenced the efficacy of IR, whereas their combination significantly increased the dose-dependent efficacy of IR. TEM inhibited phosphorylation of the downstream proteins of mTOR and induced autophagy. CQ inhibited autophagy in the late phase and did not influence the downstream proteins of mTOR. TEM and CQ inhibited both the phosphorylation of downstream proteins of mTOR and autophagy. Cell death analysis revealed that the combination of TEM and CQ strongly induced apoptosis in cells exposed to IR. In conclusion, the combination of TEM and CQ increased radiosensitivity in CRC cells through co-inhibition of mTOR and autophagy.