Background Autophagy is an evolutionarily conserved process through which cells degrade and recycle cytoplasm. The relation among autophagy, apoptosis and tumor is highly controversial until now and the molecular mechanism is poorly understood. Methods Cell viability and apoptosis were detected by CCK8, crystal violet staining, Hoechst333342 staining and flow cytometry. The expression of AMPK and ULK1 was analyzed by western blotting. Colon cancer growth suppression by NVP-BEZ235 or CQ in vivo was studied in a tumor xenograft mouse model. Results Our previous study revealed that NVP-BEZ235 suppressed colorectal cancer growth via inducing apoptosis, however later, we found it also initiated autophagy simultaneously. In this present study, our results show that NVP-BEZ235 induced autophagy through AMPK/ULK1 pathway in colon cancer cells. Blocking autophagy by knocking down AMPK or ULK1 inhibited cell proliferation and further promoted NVP-BEZ235 induced apoptosis. Meantime, the autophagy inhibitor chloroquine (CQ) shows obvious effect on inhibiting cell proliferation but not on inducing apoptosis, while it significantly increased NVP-BEZ235 induced apoptosis. Furthermore, the combinational therapy of NVP-BEZ235 and CQ shows synergistic antitumor effects in colon cancer in vivo. Conclusion NVP-BEZ235 induced AMPK/ULK1-dependent autophagy. Targeting this autophagy suppressed colon cancer growth through further promoting apoptosis, which is a potential therapeutic option for clinical patients.

中文翻译：

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阻断 AMPK/ULK1 依赖性自噬促进细胞凋亡并抑制结肠癌生长。

背景 自噬是一个进化上保守的过程，细胞通过该过程降解和回收细胞质。自噬、细胞凋亡和肿瘤之间的关系至今仍存在很大争议，其分子机制知之甚少。方法采用CCK8、结晶紫染色、Hoechst333342染色和流式细胞仪检测细胞活力和凋亡。通过蛋白质印迹分析AMPK和ULK1的表达。在肿瘤异种移植小鼠模型中研究了 NVP-BEZ235 或 CQ 在体内对结肠癌生长的抑制作用。结果我们之前的研究表明，NVP-BEZ235通过诱导细胞凋亡来抑制结直肠癌的生长，但后来我们发现它也同时启动了自噬。在本研究中，我们的结果表明 NVP-BEZ235 通过 AMPK/ULK1 通路诱导结肠癌细胞自噬。通过敲低 AMPK 或 ULK1 来阻断自噬可抑制细胞增殖并进一步促进 NVP-BEZ235 诱导的细胞凋亡。同时，自噬抑制剂氯喹（CQ）对细胞增殖有明显的抑制作用，但对诱导细胞凋亡没有作用，而对NVP-BEZ235诱导的细胞凋亡有显着的促进作用。此外，NVP-BEZ235 和 CQ 的联合治疗在体内结肠癌中显示出协同抗肿瘤作用。结论 NVP-BEZ235诱导AMPK/ULK1依赖性自噬。针对这种自噬通过进一步促进细胞凋亡来抑制结肠癌的生长，这是临床患者的潜在治疗选择。自噬抑制剂氯喹（CQ）对细胞增殖有明显抑制作用，但对诱导细胞凋亡没有作用，而对NVP-BEZ235诱导的细胞凋亡有显着增强作用。此外，NVP-BEZ235 和 CQ 的联合治疗在体内结肠癌中显示出协同抗肿瘤作用。结论 NVP-BEZ235诱导AMPK/ULK1依赖性自噬。针对这种自噬通过进一步促进细胞凋亡来抑制结肠癌的生长，这是临床患者的潜在治疗选择。自噬抑制剂氯喹（CQ）对细胞增殖有明显抑制作用，但对诱导细胞凋亡没有作用，而对NVP-BEZ235诱导的细胞凋亡有显着增强作用。此外，NVP-BEZ235 和 CQ 的联合治疗在体内结肠癌中显示出协同抗肿瘤作用。结论 NVP-BEZ235诱导AMPK/ULK1依赖性自噬。针对这种自噬通过进一步促进细胞凋亡来抑制结肠癌的生长，这是临床患者的潜在治疗选择。