Deleted in liver cancer (DLC1), a Rho GTPase-activating protein, was observed to be differentially expressed in oral squamous cell carcinoma in comparison with normal tissues using tissue proteomics. In the current study, we investigated the clinical significance of loss of DLC1 expression in different stages of development of oral squamous cell carcinoma to determine its potential as a biomarker for oral dysplasia and prognosis of oral squamous cell carcinoma. Immunohistochemical analysis of DLC1 expression was carried out in oral squamous cell carcinoma patients (n=214), dysplasia (n=51), hyperplastic squamous mucosa (n=45), and histologically normal oral tissues (n=80), and correlated with clinicopathological parameters and disease prognosis over 91 months for oral squamous cell carcinomas. Loss of DLC1 expression was observed in oral squamous cell carcinoma (64%), oral dysplasia (31%), hyperplastic squamous mucosa (22%), and normal mucosa (16%). Significant loss of DLC1 expression was observed in oral squamous cell carcinomas as compared with dysplasia (P<0.001, odds ratio=3.8, 95% CI=2.0-7.3), suggesting it may be an important event involved in cancer progression. Among oral squamous cell carcinomas, the loss of DLC1 expression was significantly associated with poor prognosis (P=0.021, hazards ratio (HR)=1.8, 95% CI=1.1-2.9). Multivariate analysis revealed loss of DLC1 (P=0.023, HR=2.1, 95% CI=1.2-3.9) and histopathological grade (P=0.015, HR=1.7, 95% CI=1.1-2.7) to be independent predictors for disease-free survival in oral squamous cell carcinoma patients in comparison with known prognostic factors, viz. tumor stage, nodal status, and overall stage. Loss of DLC1 expression emerged as an important biomarker for predicting patients diagnosed with oral dysplasia at high risk of transformation upon future validation in longitudinal studies. Loss of DLC1 expression is a poor prognostic marker for oral squamous cell carcinoma patients.