Hypoxia and Oxidation Pathway Crosstalk in Head and Neck Squamous Cell Carcinoma

OBJECTIVE: Treatment resistance remains a significant challenge in head and neck squamous cell carcinoma (HNSCC). Alterations in oxidative stress and hypoxia pathways predict poor therapeutic outcomes. The nuclear factor erythroid 2-related factor 2 pathway is the key regulator of oxidative stress response and an established mediator of treatment resistance. There is emerging evidence suggesting that its activation may influence hypoxia-inducible factor 1α (HIF-1α) signaling. However, this relationship remains poorly defined in HNSCC. STUDY DESIGN: Translational research. SETTING: University-based research laboratory. METHODS: This study utilized bioinformatic and experimental approaches to investigate the relationship between NRF2 and HIF-1α expression. Correlations among KEAP1, NFE2L2 (NRF2), and HIF1A (HIF-1α) mRNA were examined using The Cancer Genome Atlas (TCGA) and RNA-sequencing data from patient-derived HNSCC cell lines. Functional validation was performed using siRNA-mediated KEAP1 knockdown in vitro, followed by quantitative polymerase chain reaction (PCR) and Western blot analysis. RESULTS: TCGA analysis revealed that HIF1A mRNA positively correlated with NFE2L2 (NRF2), though the expression did not differ between NRF2 pathway-mutant and wild-type tumors. No significant correlation between HIF1A and NFE2L2 or KEAP1 mRNA was observed in HNSCC cell lines. KEAP1 knockdown achieved >90% transcript reduction and significantly increased the expression of the NRF2 target gene NQO1. Although NFE2L2 and HIF1A transcripts were unchanged, Western blot analysis demonstrated increased NRF2 and HIF-1α protein levels, suggesting a post-translational relationship. CONCLUSION: KEAP1 knockdown promotes NRF2 upregulation and HIF-1α protein accumulation in HNSCC independent of mRNA changes, suggesting protein-level or redox-mediated crosstalk between oxidative stress and hypoxia pathways.