Nrf2 inhibition and NCOA4-mediated ferritinophagy activation synergistically exacerbated S-3'-hydroxy-7', 2', 4'-trimethoxyisoxane induced ferroptosis in lung cancer cells.

S-3'-hydroxy-7', 2', 4'-trimethoxyisoxane (ShtIX) is a novel isoflavane compound that exhibits significant anticancer activity against a variety of cancer cells. Our previous studies have confirmed that ShtIX induced ferroptosis by inhibiting Nr2/HO-1 pathway in non-small cell lung cancer (NSCLC) cells, both in vitro and vivo. Recent research has increasingly recognized ferroptosis as an autophagy-dependent form of cell death. However, it has not been previously explored whether ShtIX can activate autophagy during ferroptosis and its relationship with ferroptosis. In the present study, we discovered that ShtIX was able to trigger autophagy, and the activation of autophagy is essential for ShtIX-induced ferroptosis. These findings demonstrated that ShtIX induced an autophagy-dependent form of ferroptosis in NSCLC cells. Intriguingly, the autophagy triggered by ShtIX is independent of ferroptosis. Furthermore, our results indicated that ShtIX degraded ferritin through autophagy and promoted NCOA4-mediated ferritinophagy, which contributed significantly to ShtIX-induced ferroptosis in NSCLC cells. Additionally, the knockdown Nrf2 reinforced ShtIX-induced NCOA4-mediated ferritinophagy, while the inhibition of autophagy attenuated the suppressive effect of ShtIX on Nrf2 and HO-1. Taken together, our work uncovers a new mechanism by which ShtIX induced ferroptosis through inhibition the Nrf2 pathway and activation of NCOA4-mediated ferritinophagy in NSCLC cells. Targeting ferritinophagy to regulate ferroptosis offers a novel therapeutic strategy for the treatment of lung cancer with ShtIX.