Abstract: Objective To identify potential key genes associated with radiotherapy response in head and neck squamous cell carcinoma (HNSCC) and validate their functional. Methods Differentially expressed genes (DEGs) between the radiotherapy response group and the non⁃response group were screened using the GSE39366 dataset. Prognostically relevant genes were selected through Cox proportional hazards regression analysis within the cancer genome atlas (TCGA). The key genes were identified through a Venn diagram analysis, followed by functional enrichment analysis. The immune microenvironment, including the ESTIMATE score and immune cell infiltration analysis, as well as interactions with immune⁃related genes, were performed on these key genes. A prognostic prediction model was developed by integrating these key genes with the clinical characteristics of patients. The prognostic impact of the key genes was validated using the external dataset GSE41613. Differences in radiosensitivity among cells with varying levels of gene expression were assessed via CCK⁃8 assays and flow cytometry. Changes in the expression of key genes under different radiotherapy doses were detected by RT⁃qPCR. Results A total of 519 DEGs and 81 prognostic⁃related genes were identified, with CCR7, MS4A1, TBC1D10C and CTTN being determined as key genes influencing radiosensitivity in HNSCC. These genes exhibited significant correlations with tumor immune infiltration, and the nomogram model constructed based on them demonstrated good predictive performance. External dataset validation indicated that CCR7 and CTTN were significantly associated with patient prognosis (all P<0.05), whereas MS4A1 and TBC1D10C did not show significant prognostic value (all P>0.05). In vitro experiments further confirmed that HNSCC cells with elevated CCR7 expression and reduced CTTN expression exhibited increased sensitive to radiotherapy. The expression levels of CCR7 and CTTN were observed to change dynamically with varying radiotherapy doses.  Conclusions The expression levels of CCR7 and CTTN are closely related to radiosensitivity and prognosis in HNSCC, potentially throungh mechanisms involving the regulation of immune microenvironment. CRRT and CTTN may serve as predictive biomarkers for HNSCC radiosensitivity and potential therapeutic targets.

Key words: Head and neck squamous cell carcinoma, Radiosensitivity, Prognosis, Immune cell infiltration, Biomarker