AIM: To evaluate the importance of epidermal growth factor receptor (EGFR) protein overexpression and gene amplification in carcinogenesis of glottic cancer. METHOD: Immunohistochemical (IHC) analysis and fluorescent in situ hybridization (FISH) were used on tissue microarrays of laryngeal tissue (145 samples), ranging from normal mucosa (38 samples), hyperplastic (46 samples) and cancerous lesions (61 samples), in order to evaluate EGFR at protein and gene level. RESULTS: Membranous (mEGFR) and cytoplasmic (cEGFR) EGFR expression was statistically different among the analyzed groups. The differences were most striking in the suprabasal-transforming zone. IHC evaluation indicated that high and low mEGFR staining contributed to the differentiation of dysplastic lesions versus simple hyperplasia and cancerous tissue as well as between different degrees of atypia among hyperplastic lesions (P<0.050). Gene amplification was not found in simple and abnormal hyperplastic lesions. It was confirmed in 2/21 atypical hyperplasias, indicating that gene amplification can facilitate better identification of malignant potential in hyperplastic lesions. In cancerous tissue, EGFR gene amplification was found in 8/50 samples. EGFR gene amplification was found in preinvasive cancer in one patient. In invasive carcinomas, gene amplification was not associated with stage or grade. Carcinomas with gene amplification showed significantly higher scores of cEGFR expression (basal layer P=0.003; suprabasal layer P=0.002). CONCLUSIONS: This study confirmed an increase of EGFR protein expression and gene amplification with the increase in biological aggressiveness of glottic lesions. A correlation between EGFR gene amplification and protein expression was established. Gene amplification proved to be an early event in glottic carcinogenesis, indicating its importance for glottic cancer prevention, early detection and protocol selection.