Senescence is a critical cellular process in both aging and cancer, marked by irreversible growth arrest, DNA damage, and the secretion of pro-inflammatory cytokines, collectively known as the senescence-associated secretory phenotype (SASP). Senescent cells play tumor-suppressive roles but can also alter the tumor microenvironment, promoting cellular proliferation. Ferroptosis, a form of iron-dependent cell death driven by lipid peroxidation, has emerged as a promising therapeutic strategy against chemotherapy-resistant cancer cells. This study investigates the effects of senescence and ferroptosis inducers on two distinct epithelial ovarian cancer (EOC) cell lines, SKOV-3 and OVCAR-3, using qPCR to assess the expression of various biomarkers.

SKOV-3 and OVCAR-3 EOC cell lines were treated with ferroptosis inducers (RSL3, Erastin, FSP1i, Brequinar) and senescence inducers (Etoposide, Olaparib, Carboplatin, Cisplatin). Cells were treated with varying drug concentrations (0.25–2 µM) and harvested for RNA extraction at Day 7. Gene expression of biomarkers associated with ferroptosis (NCOA4, CP) and senescence (p16 (CDKN2A), p21 (CDKN1A), IL6, IL8) was assessed by quantitative PCR. Cell viability was measured by crystal violet staining.

In response to RSL3 (1 µM) and FSP1i (2 µM), both SKOV-3 and OVCAR-3 cells showed ferroptosis induction. After treatment with RSL3, SKOV-3 showed an increase in CDKN1A and a decrease in CDKN2A, while no significant modulation was observed in other genes. In contrast, in OVCAR-3, there was an increase in CDKN1A and IL8, along with a decrease in NCOA4 and CP. Notably, these ferroptosis-related biomarkers were modulated independently of the cell line, indicating a common response to RSL3 treatment across both SKOV-3 and OVCAR-3. For SKOV-3, treatment with Etoposide (1 µM) led to a reduction in CDKN1A, CDKN2A, and IL8, with a further decrease in IL8 at 2 µM, suggesting that IL8 downregulation could serve as a genetic marker of senescence in SKOV-3. In OVCAR-3, Etoposide (1 µM) increased CDKN1A, CDKN2A, IL8, and NCOA4, and at 2 µM, CDKN1A and CDKN2A were further elevated. This suggests that p16 and p21 could be markers of senescence induction in OVCAR-3 cells.

This study highlights the potential of targeting both senescence and ferroptosis in epithelial ovarian cancer. Ferroptosis-related biomarkers, such as NCOA4 and CP, were modulated independently of cell line, indicating a common response to ferroptosis induction. Senescence was more cell line-dependent, with CDKN1A and CDKN2A serving as markers of senescence induction in OVCAR-3. These findings provide valuable insights into the molecular pathways governing ferroptosis and senescence in ovarian cancer, suggesting that combining therapies that target both processes could be a promising strategy to overcome chemotherapy resistance and improve treatment outcomes in EOC.