Tumor heterogeneity in autophagy-dependent ferroptosis

Macroautophagy (hereafter referred to as “autophagy”) is a lysosome-mediated degradation process that plays a complex role in cellular stress, either promoting survival or triggering death. Early studies suggest that ferroptosis, an iron-dependent form of regulated cell death, is not related to autophagy. Conversely, recent evidence indicates that the molecular machinery of autophagy facilitates ferroptosis through the selective degradation of anti-ferroptosis regulators. However, the mechanism of autophagy-dependent ferroptosis remains incompletely understood. Here, we examine the early dynamic change in protein expression of autophagic (e.g., MAP1LC3B and SQSTM1) or ferroptotic (e.g., SLC7A11 and GPX4) regulators in 60 human cancer cell lines in response to two classical ferroptosis activators (erastin and RSL3) in the absence or presence of the lysosomal inhibitor chloroquine. Compared to erastin, RSL3 exhibits wider and stronger activity in the upregulation of MAP1LC3B-II or downregulation of SQSTM1 in 80% (48/60) or 63% (38/60) of cell lines, respectively. Both RSL3 and erastin failed to affect SLC7A11 expression, but they led to GPX4 downregulation in 12% (7/60) and 3% (2/60) of cell lines, respectively. Additionally, the intracellular iron exporter SLC40A1/ferroportin-1 was identified as a new substrate for autophagic elimination, and its degradation by SQSTM1 promoted ferroptosis in vitro and in xenograft tumor mouse models. Together, these findings show tumor heterogeneity in autophagy-dependent ferroptosis, which might have different biological behaviors with regard to the dynamic characteristics of cell death.Abbreviations: ATG: Autophagy-related; CQ: Chloroquine; GPX4: Glutathione peroxidase 4; MAP1LC3B/LC3: Microtubule-associated protein 1 light chain 3 beta: NCOA4: Nuclear Receptor Coactivator 4; ROS: Reactive Oxygen Species; SLC40A1/ferroportin-1: Solute Carrier family 40 Member 1; SLC7A11: Solute Carrier Family 7 Member 11; SQSTM1/p62: Sequestosome 1