The Role of the MAPK Pathway in High-Grade Serous Ovarian Cancer

Ovarian cancer (OC) is the deadliest gynecological malignancy and the 7th most common female cancer worldwide. Due to its rapid progression and nonspecific symptoms, OC is often diagnosed at advanced stages, with a five-year survival rate of just 31%. High-grade serous ovarian cancer (HGSOC), the most aggressive subtype, accounts for ~80% of OC-related deaths. Standard treatment includes cytoreductive surgery combined with chemotherapy using carboplatin, paclitaxel, or PARP inhibitors. However, over 70% of patients experience relapse, primarily due to the transcoelomic spread of cancer cells forming spheroids. These spheroids exhibit replicative dormancy, evade mainline therapies, and contribute to chemotherapy resistance and disease recurrence. Understanding the molecular mechanisms that support spheroid survival is critical for identifying new therapeutic targets. Our lab previously conducted a genome-wide CRISPR knockout screen, identifying essential pathways for spheroid viability, including axon guidance and the MAPK pathway. Notably, netrins—key components of the axon guidance pathway—are overexpressed in HGSOC tumors and correlate with poor patient survival. This study investigates the role of netrin signaling and the downstream MAPK pathway in the survival of HGSOC spheroids under suspension conditions. Using in vitro spheroid formation and reattachment assays, we demonstrated that MEK inhibition, a key node in the MAPK pathway, significantly reduces spheroid viability across multiple HGSOC cell lines. Additionally, trypan blue viability assays revealed that while MEK inhibition induces cytostasis in adherent OVCAR8 cells, it triggers cytotoxicity in OVCAR8 HGSOC spheroids. However, analysis of cell death mechanisms using Western blots and DNA fragmentation assays showed no evidence of apoptotic markers, suggesting that apoptosis is not the primary cytotoxic mechanism in HGSOC spheroids. In conclusion, this study highlights the critical role of netrin signaling and the MAPK pathway in HGSOC spheroid survival, providing insights into potential molecular targets to overcome therapy resistance in HGSOC.