Erastin triggers oxidative, iron-dependent cell death. Treatment of NRAS-mutant HT-1080 fibrosarcoma cells with the RSL molecule erastin (10 µM) results in a time-dependent increase in cytosolic and lipid ROS beginning at 2 hours. Cell death triggered by erastin is significantly inhibited by antioxidants (e.g., α-tocopherol, butylated hydroxytoluene, and β-carotene) and iron chelators, suggesting that ROS- and iron-dependent signaling is required for erastin-induced ferroptosis. Erastin can directly bind to VDAC2/3 in BJeLR cells. Knockdown of VDAC2 and VDAC3, but not VDAC1, leads to erastin resistance. Erastin has the ability to reduce glutathione level by directly inhibiting cystine/glutamate antiporter system Xc− activity, with activation of the ER stress response. Erastin potently inhibits HT-29 cell survival. Erastin shows a dose-dependent effect, and 30 μM of erastin displays the most dramatic effect.
Erastin treatment significantly decreased intracellular GSH levels, indicating the depletion of a hallmark antioxidant of ferroptosis, while concurrently markedly increasing malondialdehyde (MDA) content, a product of lipid peroxidation. Notably, these changes were significantly more pronounced in Lpin3-KO cells than in WT cells, confirming that Erastin effectively triggers the core biochemical reactions of ferroptosis[1].
Western blot analysis further revealed that Erastin treatment substantially upregulated the expression of ACSL4, a positive regulator of ferroptosis, and downregulated the expression of GPX4, a negative regulator. These molecular alterations were also more prominent in Lpin3-KO cells, demonstrating that Erastin precisely targets the ferroptosis-related molecular pathways[1].

