Abstract

Ferroptosis is a regulated non-apoptotic cell death process characterized by iron-dependent lipid peroxidation. Peroxidation of polyunsaturated fatty acid-containing phospholipids (PUFA-PLs) is necessary for the execution of ferroptosis. Glutathione peroxidase 4 (GPX4) suppresses ferroptosis by reducing lipid hydroperoxides to lipid alcohols. GPX4 may be a useful target for drug development, highlighting the need to identify factors that govern GPX4 inhibitor sensitivity. Here, we found that reduced GPX4 expression was sufficient to induce ferroptosis in multiple adherent (2D) cancer cell cultures. However, lower GPX4 protein levels did not consistently affect tumor xenograft growth in mice. Culturing cells as spheroids (3D) was sufficient to reduce sensitivity to pharmacological GPX4 inhibition. Mechanistically, growth in 3D versus 2D conditions upregulated expression of the monounsaturated fatty acid (MUFA) biosynthetic gene stearoyl-CoA desaturase (SCD), altering the ratio of MUFA-PLs to PUFA-PLs in a direction favoring ferroptosis resistance. Similar shifts in MUFA-PL to PUFA-PL ratios were observed in xenograft tumors. Thus, lipidome remodeling in 3D growth conditions and in vivo may limit GPX4 inhibitor efficacy.