ALKBH4 confers ferroptosis resistance and drives tumorigenesis via dysregulation of GPX4 in breast cancer cells.

Abstract

Breast cancer (BC) remains a predominant source of cancer-related fatalities among women globally. While the AlkB homolog family member ALKBH4 has been implicated in cancer, its role and mechanism in BC are unknown. Its role within BC and its correlation with ferroptosis-a type of iron-mediated regulated cell death, were examined in this work. Data from The Cancer Genome Atlas (TCGA), Gene Expression Omnibus (GEO), and immunohistochemistry (IHC) analyses indicate ALKBH4 overexpression in BC, further associating its high expression levels with unfavorable clinical outcomes. Functional experiments in BC cells demonstrated that ALKBH4 knockdown exerted inhibitory effects on proliferation, migration, and invasion. Concurrently, it promoted apoptosis. Transcriptomic and biochemical investigations revealed that knocking down ALKBH4 sets off a ferroptotic cascade, with telltale signs including heightened levels of malondialdehyde (MDA) and ferrous iron (Fe²⁺), a serious drain on reduced glutathione (GSH) reserves, and a buildup of reactive oxygen species (ROS). Fortunately, this ferroptotic cell death was prevented by the ferroptosis inhibitor Ferrostatin-1 (Fer-1) and enhanced by the inducer Erastin. Mechanistically, we demonstrated that ALKBH4 maintains GPX4 protein levels in a demethylase activity-dependent manner, and GPX4 overexpression reversed the ferroptosis and growth defects caused by ALKBH4 loss. Combined application of ALKBH4 knockdown and GPX4 overexpression resulted in enhanced anti-tumor effects both in cellular and animal models. In conclusion, our findings establish that ALKBH4 is a novel driver of BC progression, which confers ferroptosis resistance to cancer cells by governing GPX4 levels in an enzyme activity-dependent way. Therefore, targeting ALKBH4 to disrupt its regulatory effect on GPX4 may offer a new therapeutic strategy for BC.