背景:严重的内质网(ER)应激引起细胞凋亡抑制肺癌。我们之前的研究发现,西法兰碱(CEP),一种植物药,具有强大的抗癌功效,其潜在的机制仍然被发现。在这里,我们调查了CEP如何诱导ER应激并对抗肺癌.
方法:通过RNA序列检测差异表达基因(DEGs)和富集。通过细胞热转移测定(CETSA)和分子对接分析CEP和NRF2的亲和力。用蛋白质印迹法测定肺癌细胞的功能,流式细胞术,免疫荧光染色,和铁凋亡抑制剂。
结果:CEP治疗在铁中毒和内质网应激中富集了DEGs。进一步的分析表明目标是NRF2。体外和体内实验表明,CEP诱导了明显的铁凋亡,以铁离子升高为特征,ROS,COX-2表达,GPX4的下调和萎缩性线粒体。此外,增强型Grp78,CHOP,p-eIF2A表达,β-淀粉样蛋白肿块,CEP组观察到ER平行堆叠结构消失,表明ER压力被唤醒。CEP表现出优异的抗肺癌疗效,细胞凋亡增加证明了这一点,减少扩散,细胞干性减弱,并在动物模型中突出抑制肿瘤移植物。此外,添加铁凋亡抑制剂可减弱CEP诱导的ER应激和细胞凋亡。
结论:总之,我们的发现证明CEP通过抑制NRF2来诱导强烈的内质网应激,从而导致肺癌细胞的凋亡和干性减弱。当前的工作提出了天然化合物CEP的抗肿瘤功效的新机制。
BACKGROUND: Severe endoplasmic reticulum (ER) stress elicits apoptosis to suppress lung cancer. Our previous research identified that Cepharanthine (CEP), a kind of phytomedicine, possessed powerful anti-cancer efficacy, for which the underlying mechanism was still uncovered. Herein, we investigated how CEP induced ER stress and worked against lung cancer.
METHODS: The differential expression genes (DEGs) and enrichment were detected by RNA-sequence. The affinity of CEP and NRF2 was analyzed by cellular thermal shift assay (CETSA) and molecular docking. The function assay of lung cancer cells was measured by western blots, flow cytometry, immunofluorescence staining, and ferroptosis inhibitors.
RESULTS: CEP treatment enriched DEGs in ferroptosis and ER stress. Further analysis demonstrated the target was NRF2. In vitro and in vivo experiments showed that CEP induced obvious ferroptosis, as characterized by the elevated iron ions, ROS, COX-2 expression, down-regulation of GPX4, and atrophic mitochondria. Moreover, enhanced Grp78, CHOP expression, β-amyloid mass, and disappearing parallel stacked structures of ER were observed in CEP group, suggesting ER stress was aroused. CEP exhibited excellent anti-lung cancer efficacy, as evidenced by the increased apoptosis, reduced proliferation, diminished cell stemness, and prominent inhibition of tumor grafts in animal models. Furthermore, the addition of ferroptosis inhibitors weakened CEP-induced ER stress and apoptosis.
CONCLUSIONS: In summary, our findings proved CEP drives ferroptosis through inhibition of NRF2 for induction of robust ER stress, thereby leading to apoptosis and attenuated stemness of lung cancer cells. The current work presents a novel mechanism for the anti-tumor efficacy of the natural compound CEP.