Chair of Medical Biochemistry, Jagiellonian University Medical College, ul. Kopernika 7, 31-034 Kraków, Poland.
Int J Mol Sci. 2021 Apr 1;22(7):3682. doi: 10.3390/ijms22073682.
The twofold role of autophagy in cancer is often the therapeutic target. Numerous regulatory pathways are shared between autophagy and other molecular processes needed in tumorigenesis, such as translation or survival signaling. Thus, we have assumed that ILK knockdown should promote autophagy, and used together with chloroquine, an autophagy inhibitor, it could generate a better anticancer effect by dysregulation of common signaling pathways. Expression at the protein level was analyzed using Western Blot; siRNA transfection was done for ILK. Analysis of cell signaling pathways was monitored with phospho-specific antibodies. Melanoma cell proliferation was assessed with the crystal violet test, and migration was evaluated by scratch wound healing assays. Autophagy was monitored by the accumulation of its marker, LC3-II. Our data show that ILK knockdown by siRNA suppresses melanoma cell growth by inducing autophagy through AMPK activation, and simultaneously initiates apoptosis. We demonstrated that combinatorial treatment of melanoma cells with CQ and siILK has a stronger antitumor effect than monotherapy with either of these. It generates the synergistic antitumor effects by the decrease of translation of both global and oncogenic proteins synthesis. In our work, we point to the crosstalk between translation and autophagy regulation.
自噬在癌症中的双重作用通常是治疗的靶点。自噬与肿瘤发生过程中所需的其他分子过程(如翻译或存活信号)之间存在许多共同的调节途径。因此,我们假设 ILK 敲低应该会促进自噬,并且与自噬抑制剂氯喹一起使用,通过共同信号通路的失调,可以产生更好的抗癌效果。使用 Western Blot 分析蛋白质水平的表达;进行了针对 ILK 的 siRNA 转染。用磷酸化特异性抗体监测细胞信号通路的分析。用结晶紫试验评估黑素瘤细胞增殖,用划痕愈合试验评估迁移。通过其标记物 LC3-II 的积累来监测自噬。我们的数据表明,通过 siRNA 敲低 ILK 通过激活 AMPK 诱导自噬从而抑制黑色素瘤细胞生长,并同时引发细胞凋亡。我们证明,用 CQ 和 siILK 联合治疗黑素瘤细胞比单独使用这两种药物中的任何一种都具有更强的抗肿瘤作用。它通过降低两种全球蛋白和致癌蛋白的翻译来产生协同的抗肿瘤作用。在我们的工作中,我们指出了翻译和自噬调节之间的串扰。