Using this system for classification, we define the molecular profile of HPV+ OSCC with favorable prognosis, namely HPV+/p16 high (class III). This study defines a novel classification scheme that may have value for patient stratification for clinical trials testing HPV-targeted therapies.

Background: Several lines of evidence support the epidermal growth factor receptor (EGFR) as a molecular target for therapy in head and neck squamous cell carcinomas (HNSCC). Determination of tumor EGFR levels by conventional immunohistochemistry has not always predicted antitumor efficacy. Quantitative assays may provide more accurate assessment of the level of EGFR receptor in the tumor, which may thus provide more reliable prognostic and predictive information. We studied the prognostic value of quantitative assessment of EGFR in oropharyngeal squamous cell cancers treated with radiotherapy. Experimental Design:We studied EGFR protein expression on a tissue microarray composed of 95 oropharyngeal cancer cases using an in situ molecular-based method of quantitative assessment of protein expression (AQUA) and correlated those with clinical and pathologic data. Automated, quantitative analysis uses cytokeratin to define pixels as cancer (tumor mask) within the array spot and measures intensity of EGFR expression using a Cy5-conjugated antibody within the mask. A continuous index score is generated, which is directly proportional to the number of molecules per unit area, and cases were defined as high expressing if they were above the median expression level. Results: The mean follow-up time for survivors was 44.9 months, and for the entire cohort was 34.8 months. Patients with high tumor EGFR expression levels had a local recurrence rate of 58% compared with 17% for patients with low EGFR tumor expression (P < 0.01). Similarly, patients with high nuclear EGFR expression had a local recurrence rate of 54% compared with 21% for patients with low EGFR nuclear expression (P < 0.05). Additionally, patients with high tumor and nuclear EGFR levels had inferior disease-free survival compared with low expressors (19% versus 43% and 19% versus 45%, respectively. P < 0.05 for each). In multivariate analysis adjusting for well-characterized prognostic variables, high tumor and nuclear EGFR expression levels retained their prognostic significance. Conclusion:The AQUA system provides a continuous measurement of EGFR on paraffinembedded tissue and was able to reveal the association between EGFR expression and outcome expected from the biological role of EGFR. In the future, EGFR AQUA score may be useful in predicting response to EGFR-targeted therapies.

Purpose: Functional inactivation of p16 is an early and frequent event in head and neck squamous cell cancers. In this study, we sought to determine whether p16 expression is of prognostic importance in oropharyngeal squamous cell carcinoma.Experimental Design: p16 protein expression was evaluated by immunohistochemistry in a tissue microarray composed of 123 oropharyngeal squamous cell cancers with a mean patient follow-up time of 33 months.Results: p16 overexpression was associated with more advanced Tumor-Node-Metastasis stage and higher histologic grade. Despite this association with unfavorable features, p16 overexpression was associated with decreased 5-year local recurrence rates (11 versus 53%) and increased 5-year disease-free survival (62 versus 19%) and overall survival (60 versus 21%). In multivariate analysis, p16 expression status remained an independent prognostic factor for local recurrence, disease-free survival, and overall survival.Conclusions: In patients with oropharyngeal squamous cell carcinoma, overexpression of p16 as determined by immunohistochemistry is associated with significantly improved prognosis and lower local recurrence rates.

Background: Several lines of laboratory evidence support the epidermal growth factor receptor (EGFR) as an adverse prognostic indicator in ovarian cancers. However, different methods of immunohistochemical assessment have yielded conflicting results. Here, we sought to determine the prognostic value of EGFR in ovarian cancer using a novel method of compartmentalized in situ protein analysis. Methods: A tissue array composed of 150 advanced-stage ovarian cancers uniformly treated, with surgical debulking followed by platinum-paclitaxel combination chemotherapy, was constructed. For evaluation of EGFR protein expression, we used an immunofluorescence-based method of automated in situ quantitative measurement of protein analysis (AQUA). Results: Mean follow-up time for the entire cohort was 34.4 months. Eighty-one of 150 cases had sufficient tissue for AQUA analysis. High tumor EGFR expression was associated with poor outcome for overall survival (P = 0.0001) and disease-free survival (P = 0.0005) at 3 years. In multivariable analysis, adjusting for well-characterized prognostic variables, EGFR expression status was the most significant prognostic factor for disease-free and overall survival. Conclusion: The conflicting results in the literature regarding the prognostic value of EGFR may be due to the technical difficulties inherent in assessing EGFR with immunocytochemistry. In the present study, we show that measurement of EGFR protein levels in ovarian cancer using AQUA is feasible and can give important prognostic information.

To examine the expression of c-Met, c-Erb-b2 (HER2/neu), and epidermal growth factor (EGFR) in a cohort of 12 chordomas, based on the current and future availability of targeted molecular inhibitors. Design: Protein expression levels were analyzed by immunohistochemical analysis from archival tissue using multitumored tissue microarray and by Spearman rank correlation test.