Molecular prognostic indicators for oropharyngeal squamous cell carcinoma (OSCC), including HPV-DNA detection, epidermal growth factor receptor (EGFR) and p16 expression, have been suggested in the literature, but none of these are currently used in clinical practice. To compare these predictors, 106 newly diagnosed OSCC for the presence of HPV-DNA and expression of p16 and EGFR were analyzed. The 5-year disease-free survival (DFS) and overall survival (OS) were calculated in relation to these markers and a multivariate Cox analysis was performed. Twenty-eight percent of the cases contained oncogenic HPV-DNA and 30% were positive for p16. The p16 expression was highly correlated with the presence of HPV-DNA (p < 0.001). Univariate analysis of the 5-year DFS revealed a significantly better outcome for patients with p16-positive tumors (84% vs. 49%, p = 0.009). EGFR-negative tumors showed a tendency toward a better prognosis in DFS (74% vs. 47%, p = 0.084) and OS (70% vs. 45%, p = 0.100). Remarkable and highly significant was the combination of p16 and EGFR expression status, leading to 5-year DFS of 93% for p16+/EGFR− tumors vs. 39% for p16−/EGFR+ tumors (p = 0.003) and to a 5-year OS of 79% vs. 38%, respectively (p = 0.010). In multivariate analysis p16 remained a highly significant prognostic marker for DFS (p = 0.030) showing a 7.5-fold increased risk for relapse in patients with p16-negative tumors. Our data indicate that p16 expression is the most reliable prognostic marker for OSCC and further might be a surrogate marker for HPV-positive OSCC. HPV+/p16+ tumors tended to have decreased EGFR expression, but using both immunohistological markers has significant prognostic implications. © 2007 Wiley-Liss, Inc.