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Despite advances in laryngeal squamous cell carcinoma (LSCC) treatment, patient survival has not improved in the last two decades. Novel, more effective strategies should be based on receptor-mediated LSCC-targeted therapy. The epidermal growth factor receptor (EGFR) is the most widely studied molecular target. MASPIN multifaceted anti-tumor effects have been rarely evaluated in LSCC. The aims of this study were to investigate EGFR and MASPIN expression and the role of sub-cellular MASPIN localization in LSSC. MASPIN and EGFR expression and the sub-cellular localization of MASPIN were assessed using a computerized image analysis system in 108 consecutive cases of operable LSCC. The rates of occurrence of lymph node metastases and recurrent disease were lower in patients with a nuclear pattern of MASPIN expression (p = 0.004, p = 0.0028). As expected, the loco-regional recurrence rate was lower in N0 patients (p = 0.009), but the disease recurrence rate was even lower in N0 patients with a nuclear localization of MASPIN (p = 0.020). Disease-free survival was longer in cases of LSCC with a nuclear MASPIN pattern (p = 0.003). The intensity of EGFR reactivity and the EGFR area fraction were unrelated to the clinico-pathological and prognostic parameters in LSCC. A nuclear MASPIN pattern is a promising prognostic indicator in LSCC, but further evidence is needed before elective neck dissection can be considered for cN0 LSCCs with a non-nuclear MASPIN pattern. Although a better understanding of the mechanisms behind sub-cellular MASPIN localization is mandatory, re-activating nuclear MASPIN in association with specific chemotherapeutic agents may be an important goal in the treatment of LSCC.