Genetic Alterations in KEAP1-NRF2 system Induced by Anti-Cancer Treatments Contribute to Cisplatin Resistance in Head and Neck Squamous Cell Carcinoma

Cisplatin (CDDP) is widely used in the chemotherapy of head-and-neck squamous cell carcinoma (HNSCC), but its efficacy is limited in recurrent cases. This highlights the need for a deeper understanding of the mechanisms underlying the CDDP resistance. We hypothesized that chemoradiotherapy with CDDP may induce hyperactivation of NRF2, contributing to CDDP resistance. To investigate this, we first examined NRF2 expression in clinical HNSCC samples by immunohistochemistry and assessed the occurrence of somatic mutations in KEAP1 and NRF2 genes using public databases. We found that high NRF2 expression was associated with poorer prognosis compared to low expression. Furthermore, recurrent HNSCC cases following post-operative chemoradiotherapy exhibited higher NRF2 accumulation than their corresponding primary tumors. Database analysis also revealed occurrence of frequent mutations in KEAP1 associated with CDDP treatment. We then assessed the role of NRF2 in HNSCC malignancy utilizing seven parent (P) HNSCC cell lines and their CDDP-resistant (CR) derivatives. Of these P-CR pairs, three CR cell lines exhibited NRF2 upregulation. Two of them carried KEAP1 mutations and one carried an NRF2 mutation, both present in P and CR lines. These NRF2-high CR lines showed elevated expression of NRF2 target gene and activation of xenobiotic metabolism and reactive oxygen species pathways. Treatment with mitomycin C (MMC), known for its synthetic lethal activity against NRF2-activated cancer cells, showed strong cytotoxicity in NRF2-high CR lines. These findings indicate that the KEAP1-NRF2 system plays a critical role in the development of CDDP resistance and that MMC represents a promising therapeutic option for NRF2-high/CDDP-resistant HNSCC.