Chloroquine Sensitizes Nasopharyngeal Carcinoma Cells but Not Nasoepithelial Cells to Irradiation by Blocking Autophagy

Background
Treatment of nasopharyngeal carcinoma requires the application of high dosages of radiation, leading to severe long-term complications in the majority of patients. Sensitizing tumor cells to radiation could be a means to increase the therapeutic window of radiation. Nasopharyngeal carcinoma cells display alterations in autophagy and blockade of autophagy has been shown to sensitize them against chemotherapy.
Methods
We investigated the effect of chloroquine, a known inhibitor of autophagy, on sensitization against radiation-induced apoptosis in a panel of five nasopharyngeal carcinoma cell lines and a SV40-transformed nasoepithelial cell line. Autophagy was measured by immunoblot of autophagy-related proteins, immunofluorescence of autophagosomic microvesicles and electron microscopy. Autophagy was blocked by siRNA against autophagy-related proteins 3, 5, 6 and 7 (ATG3, ATG5, ATG6 and ATG7).
Results
Chloroquine sensitized four out of five nasopharyngeal cancer cell lines towards radiation-induced apoptosis. The sensitizing effect was based on the blockade of autophagy as inhibition of ATG3, ATG5, ATG6 and ATG7 by specific siRNA could substitute for the effect of chloroquine. No sensitization was seen in nasoepithelial cells.
Conclusion
Chloroquine sensitizes nasopharyngeal carcinoma cells but not nasoepithelial cells towards radiation-induced apoptosis by blocking autophagy. Further studies in a mouse-xenograft model are warranted to substantiate this effect in vivo.

背景
鼻咽癌的临床治疗需采用高剂量放射线照射，这会导致多数患者出现严重的长期并发症。使肿瘤细胞对放射线产生增敏，或是扩大放疗治疗窗口的可行策略。鼻咽癌细胞存在自噬（autophagy）异常，而阻断自噬已被证实可使其对化疗产生增敏作用。

方法
本研究以氯喹（chloroquine，一种经典自噬抑制剂）为研究对象，在5株鼻咽癌细胞系及1株SV40转化的鼻咽上皮细胞系中，探究其对放射线诱导细胞凋亡的增敏效应。自噬水平通过自噬相关蛋白免疫印迹（immunoblot）、自噬体微囊泡免疫荧光（immunofluorescence）及电子显微镜（electron microscopy）进行检测。通过靶向自噬相关蛋白3、5、6、7（ATG3、ATG5、ATG6、ATG7）的小干扰RNA（siRNA）阻断自噬过程。

结果
氯喹可使5株鼻咽癌细胞系中的4株对放射线诱导的细胞凋亡产生增敏作用。该增敏效应依赖于自噬阻断：通过特异性小干扰RNA抑制ATG3、ATG5、ATG6、ATG7的表达，可模拟氯喹的增敏效果。而在鼻咽上皮细胞系中未观察到此类增敏效应。

结论
氯喹可通过阻断自噬，使鼻咽癌细胞而非鼻咽上皮细胞对放射线诱导的细胞凋亡产生增敏作用。后续需通过小鼠异种移植模型开展体内研究，以验证该效应。