Novel Insights of Herbal Remedy into NSCLC Suppression through Inducing Diverse Cell Death Pathways via Affecting Multiple Mediators

Artemisia species previously have been reported to have antimicrobial, antioxidant, an-tiulcer, and anticancer properties. In this study, we investigated the prospective antitumor effects of Artemisia santolinifolia ethanol extract (ASE) against two non-small cell lung cancer (NSCLC) cell lines and their molecular mechanisms of action. Morphological observations and flow cytometric analyses showed that ASE induced cell death in A549 and H23 cells but with different action features. Further studies by Western blotting showed that ASE induced caspase-3 cleavage in H23 cells, sug-gesting caspase-dependent apoptosis was predominantly involved in H23 cell death. Contrarily, ASE treatment selectively altered the glutathione peroxidase (GPX4) protein expression, reactive oxygen species (ROS) generation, and lipid peroxidation in A549 cells, all of which are linked to ferropto-sis. Using a ferroptosis inhibitor (desferrioxamine (DFO)), further study showed that DFO could significantly rescue ASE-induced cell death. All these results implied that ASE induced ferroptosis predominately in A549 cells. Several studies have demonstrated that the nuclear factor erythroid 2–related factor 2 (NRF2) can be dual-selectively targeted depending on the cell line. Subsequently, it can exert opposing effects until either being activated or suppressed. This was consistent with our data, which might explain inconsistent observations of the cell death type in this study. In addition, after ASE treatment, signal transduction and activator of transcription 3 (STAT3) were inhibited in both cell lines. Consequently, downstream prosurvival proteins, including heat shock protein 70 (HSP70) and survivin, which play pivotal roles in the STAT3 pathway, decreased after ASE administration. Our findings revealed that ASE inhibited NSCLC cell proliferation by simultaneously downregulating prosurvival protein expressions and activating multiple cell death pathways.