Proteômica do carcinoma epidermóide de cabeça e pescoço: identificação e validação de biomarcadores potenciais

Abstract

Epidermoid tumors of the oral cavity, larynx and pharynx, collectively known as head and neck squamous cell carcinomas, are strongly related to tobacco and alcohol consumption and still result in high mortality rates. The development of a primary tumor antecipates a greater risk of second cancers occurring in the contiguous epithelium, which can be a consequence of the abnormal response of the microenvironment to the carcinogen exposure in the upper aerodigestive tract. The prediction of tumor behavior for patients with these carcinomas still represents a challenge for clinicians and researchers. The presence of regional lymph node metastasis is their most important prognostic factor but is limited in predicting local recidive or survival. In addition, early metastatic disease is often missed by clinical, histological and radiological analysis. This emphasizes the need for identifying biomarkers which may effectively contribute to early diagnosis and prediction of tumor progression. Objectives: In this study, we analyzed oral squamous cell carcinomas (OSCCs) from different anatomic subsites and their matched adjacent normal mucosa aiming to compare the protein expression profiles in the context of a known prognosticator, namely, the presence of neoplastic cells in regional lymph nodes and of a refined classification in regard to prognosis, grouping into two groups small but already metastatic and large non-metastatic tumors. We also aimed to investigate the proteome of the tumor microenvironment to identify markers of aggressiveness that may be relevant for prognosis and therapy. Materials and Methods: To reach these objectives, we performed one- and two-dimensional electrophoresis (1-DE and 2-DE), fluorescent two-dimensional differential in-gel electrophoresis (2-D DIGE), mass spectrometry, Western blot and immunohistochemistry to analyze the protein expression in OSCCs. Results and Discussion: A variety of factors influence the protein sample preparation and the optimization of protein solubilization and 2-DE protocols was fundamental for the acquisition of consistent results. Many protein differences between metastatic and non-metastatic tumors and between tumor and normal cells including abnormal expression of annexin A1, calgranulin-B, cofilin-1, galectin-7, glutathione Stransferase P, cytokeratin-4 and creatine kinase were observed. These proteins are involved in cell signaling, inflammatory response, apoptosis, cell motility and adhesion, development, cell differentiation and proliferation and metabolic process, and may be related to the aggressive phenotype. Tumor microenvironment may also contribute to the neoplastic process. Our results on paracrine factors synthetyzed in vitro by stromal cells showed altered gene and protein levels in neoplastic cells after treatment with conditioned medium from tumor-associated fibroblasts, and vice-versa. Conclusions: The data may help to understand the mechanisms governing aggressiveness in OSCCs at the molecular level and the contributions of the tumor microenvironment to carcinogenic process, providing new insights into signaling and metabolic pathway abnormalities that could be useful to prognosis.