Objective To investigate the prognostic significance of p16 protein expression in patients with locally advanced laryngeal and hypopharyngeal carcinoma who are undergoing comprehensive treatment. Methods A retrospective analysis was carried out on the clinical data of 118 patients diagnosed with locally advanced （Stage Ⅲ - ⅣA） laryngeal or hypopharyngeal carcinoma， who underwent comprehensive treatment at our institution from January 2021 to January 2024. The p16 protein expression was detected via immunohistochemistry. Patients were stratified according to their p16 protein expression into positive and negative groups to compare their clinicopathological characteristics. The Kaplan-Meier method was employed to plot survival curves and compute overall survival （OS） and progression-free survival （PFS）， and inter-group comparisons were conducted using the Log-rank test. Independent prognostic factors were identified by means of a Cox proportional hazards regression model. Results The p16-positive rate was 26.27% （31 out of 118）. When compared with the p16-negative group， the p16-positive group exhibited a significantly higher proportion of patients with Stage IV disease， poor differentiation， or lymph node metastasis （all P < 0.05）. Patients in the p16-negative group had a shorter median overall survival （OS） of 36.90 months （95%CI： 33.29 - 40.51）， whereas the median OS was not reached in the p16-positive group （Log-rank P = 0.002）. The p16-positive group also demonstrated a superior median progression-free survival （PFS） ［39.12 months （95%CI： 35.88 - 42.39） compared to 34.68 months （95%CI： 31.56 - 37.81）， Log-rank χ2 = 9.347， P = 0.002］. In the concurrent chemoradiotherapy （CCRT） subgroup （n = 66）， p16-positive patients had a better median OS than p16-negative patients （median OS not reached versus 37.5 months， log-rank χ2 = 8.854， P = 0.003）. Multivariate Cox regression analysis revealed that smoking for ≥ 10 years， alcohol consumption ≥ 40 g/day， lymph node metastasis， tumor differentiation grade， and p16 protein expression were independent prognostic factors influencing both OS and PFS. Conclusion The p16 protein expression serves as an independent prognostic factor in locally advanced laryngeal and hypopharyngeal carcinoma. Conducting tests for the p16 protein expression facilitates prognostic stratification and offers a reference for individualized treatment.