Ray Anasuya, Vasudevan Smreti, Sengupta Suparna
Division of Cancer Research, Rajiv Gandhi Centre for Biotechnology, Thiruvananthapuram - 695014, India.
PLoS One. 2015 Sep 10;10(9):e0137614. doi: 10.1371/journal.pone.0137614. eCollection 2015.
Cancer stem cells (CSCs) pose a serious obstacle to cancer therapy as they can be responsible for poor prognosis and tumour relapse. In this study, we have investigated inhibitory activity of the ginger-derived compound 6-shogaol against breast cancer cells both in monolayer and in cancer-stem cell-like spheroid culture. The spheroids were generated from adherent breast cancer cells. 6-shogaol was effective in killing both breast cancer monolayer cells and spheroids at doses that were not toxic to noncancerous cells. The percentages of CD44+CD24-/low cells and the secondary sphere content were reduced drastically upon treatment with 6-shogaol confirming its action on CSCs. Treatment with 6-shogaol caused cytoplasmic vacuole formation and cleavage of microtubule associated protein Light Chain3 (LC3) in both monolayer and spheroid culture indicating that it induced autophagy. Kinetic analysis of the LC3 expression and a combination treatment with chloroquine revealed that the autophagic flux instigated cell death in 6-shogaol treated breast cancer cells in contrast to the autophagy inhibitor chloroquine. Furthermore, 6-shogaol-induced cell death got suppressed in the presence of chloroquine and a very low level of apoptosis was exhibited even after prolonged treatment of the compound, suggesting that autophagy is the major mode of cell death induced by 6-shogaol in breast cancer cells. 6-shogaol reduced the expression levels of Cleaved Notch1 and its target proteins Hes1 and Cyclin D1 in spheroids, and the reduction was further pronounced in the presence of a γ-secretase inhibitor. Secondary sphere formation in the presence of the inhibitor was also further reduced by 6-shogaol. Together, these results indicate that the inhibitory action of 6-shogaol on spheroid growth and sustainability is conferred through γ-secretase mediated down-regulation of Notch signaling. The efficacy of 6-shogaol in monolayer and cancer stem cell-like spheroids raise hope for its therapeutic benefit in breast cancer treatment.
癌症干细胞（CSCs）对癌症治疗构成了严重障碍，因为它们可能导致预后不良和肿瘤复发。在本研究中，我们研究了生姜衍生化合物6-姜辣素对单层培养和癌症干细胞样球体培养中的乳腺癌细胞的抑制活性。球体由贴壁乳腺癌细胞生成。6-姜辣素能有效杀死乳腺癌单层细胞和球体，且这些剂量对非癌细胞无毒。用6-姜辣素处理后，CD44+CD24-/低细胞的百分比和二次球体含量大幅降低，证实了其对癌症干细胞的作用。6-姜辣素处理导致单层和球体培养中细胞质空泡形成以及微管相关蛋白轻链3（LC3）的切割，表明它诱导了自噬。对LC3表达的动力学分析以及与氯喹的联合处理表明，与自噬抑制剂氯喹相比，自噬通量促使6-姜辣素处理的乳腺癌细胞死亡。此外，在氯喹存在的情况下，6-姜辣素诱导的细胞死亡受到抑制，即使长时间处理该化合物后也仅表现出非常低水平的凋亡，这表明自噬是6-姜辣素诱导乳腺癌细胞死亡的主要方式。6-姜辣素降低了球体中切割的Notch1及其靶蛋白Hes1和细胞周期蛋白D1的表达水平，在γ-分泌酶抑制剂存在的情况下，这种降低更为明显。6-姜辣素还进一步减少了抑制剂存在时的二次球体形成。总之，这些结果表明，6-姜辣素对球体生长和可持续性的抑制作用是通过γ-分泌酶介导的Notch信号下调实现的。6-姜辣素在单层培养和癌症干细胞样球体中的疗效为其在乳腺癌治疗中的治疗益处带来了希望。
