GSTM5 suppresses ovarian cancer progression in association with RASSF2A promoter hypomethylation and ferroptosis.

OBJECTIVE

Glutathione S-transferase Mu 5 (GSTM5) is a detoxification-related enzyme whose function in ovarian cancer (OC) remains incompletely understood. This study investigated the association of GSTM5 with OC progression and examined whether mitochondrial dysfunction, ferroptosis-related injury, and Ras association domain family member 2 isoform A (RASSF2A) promoter methylation contribute to its effects.

METHODS

Gain- and loss-of-function experiments were performed in OC cells. Cell proliferation, migration, invasion, mitochondrial function, reactive oxygen species (ROS) production, cytotoxicity, and ferroptosis-related markers were evaluated. Ferrostatin-1 (Fer-1) was used for functional rescue, and RAS-selective lethal 3 (RSL3) served as a ferroptosis positive control. RASSF2A expression and promoter methylation were analyzed by reverse transcription-quantitative PCR (RT-qPCR) and methylation-specific PCR (MSP). Promoter-associated chromatin was assessed by DNA pull-down and chromatin immunoprecipitation (ChIP).

RESULTS

GSTM5 expression was reduced in OC tissues and was lower in an independent metastatic cohort than in primary tumors. GSTM5 overexpression suppressed OC cell proliferation, migration, and invasion, attenuated xenograft growth and intraperitoneal dissemination, increased intracellular ROS, and decreased mitochondrial membrane potential (MMP). It also elevated intracellular Fe2+, lipid peroxidation, and malondialdehyde (MDA), while reducing glutathione (GSH) and glutathione peroxidase 4 (GPX4). Fer-1 partially attenuated GSTM5-induced cell death and lipid peroxidation, whereas RSL3 produced the expected ferroptosis-associated changes. GSTM5 expression correlated with reduced RASSF2A promoter methylation, and elevated RASSF2A expression. RASSF2A knockdown partially reversed the phenotypic, mitochondrial, and ferroptosis-related effects of GSTM5 overexpression.

CONCLUSION

GSTM5 acts as a tumor-suppressive regulator in the OC models examined and its effects are accompanied by ferroptosis-related cell injury and mitochondrial dysfunction. RASSF2A partially mediates these effects, and GSTM5 expression is associated with reduced RASSF2A promoter methylation. The molecular intermediates linking GSTM5 to locus-specific methylation remain to be established.