SMAD4 endows TGF-β1-induced highly invasive tumor cells with ferroptosis vulnerability in pancreatic cancer

Pancreatic ductal adenocarcinoma (PDAC) is an extremely aggressive malignancy prone to recurrence and metastasis. Studies show that tumor cells with increased invasive and metastatic potential are more likely to undergo ferroptosis. SMAD4 is a critical molecule in the transforming growth factor β (TGF-β) pathway, which affects the TGF-β-induced epithelial-mesenchymal transition (EMT) status. SMAD4 loss is observed in more than half of patients with PDAC. In this study, we investigated whether SMAD4- positive PDAC cells were prone to ferroptosis because of their high invasiveness. We showed that SMAD4 status almost determined the orientation of transforming growth factor β1 (TGF-β1)-induced EMT via the SMAD4-dependent canonical pathway in PDAC, which altered ferroptosis vulnerability. We identified glutathione peroxidase 4 (GPX4), which inhibited ferroptosis, as a SMAD4 down-regulated gene by RNA sequencing. We found that SMAD4 bound to the promoter of GPX4 and decreased GPX4 transcription in PDAC. Furthermore, TGF-β1-induced high invasiveness enhanced sensitivity of SMAD4-positive organoids and pancreas xenograft models to the ferroptosis inducer RAS-selective lethal 3 (RSL3). Moreover, SMAD4 enhanced the cytotoxic effect of gemcitabine combined with RSL3 in highly invasive PDAC cells. This study provides new ideas for the treatment of PDAC, especially SMAD4-positive PDAC.