KRT6A mediates immune exclusion and anti-PD-1 resistance in head and neck squamous cell carcinoma

Head and neck squamous cell carcinoma (HNSCC) is one of the most common malignancies worldwide and is currently the third leading cause of cancer-related death among Taiwanese men. Immune checkpoint blockade (ICB) therapies, including the PD-1-blocking monoclonal antibodies pembrolizumab and nivolumab, have been approved as first-line and salvage treatments for PD-L1-positive recurrent or metastatic (R/M) HNSCC. However, only a minority of patients derive durable benefit from anti-PD-1 therapy. Both innate and acquired resistance to ICB remain major obstacles to effective treatment. In this study, we investigated potential mechanisms underlying ICB resistance using a syngeneic mouse HNSCC model, MOCL2-1. Serial in vivo and in vitro passaging under anti-PD-1 treatment pressure generated an anti-PD-1-resistant subline, L2-1-R. L2-1-R tumors were immunologically “cold,” characterized by minimal CD8⁺ T-cell infiltration and resistance to anti-PD-1 therapy. RNA sequencing of sequential sublines, along with the anti-PD-1-sensitive subline L2-1-S, revealed distinct transcriptional reprogramming in resistant tumors. KRT6A , a stress-induced keratin, was markedly upregulated in L2-1-R. Knockdown of KRT6A in resistant cells increased CD8⁺ T-cell infiltration and restored responsiveness to anti-PD-1 therapy in vivo. Furthermore, analysis of a clinical cohort of HNSCC patients treated with pembrolizumab demonstrated that high KRT6A expression was associated with poor therapeutic response. Together, these findings identify KRT6A as a key mediator of immune exclusion and resistance to PD-1 blockade in HNSCC, suggesting that targeting KRT6A-related pathways may enhance immunotherapy efficacy.