Análise comparativa das proteínas expressas na saliva de pacientes com leucoplasia oral e carcinoma de boca: uma abordagem proteômica

Abstract

Oral cancer is a worldwide problem. More than 90% of oral cancer cases are oral squamous cell carcinomas (OSCC), which is aggressive and particularly affects chronic smokers and drinkers. Tumor pathogenesis in non-smokers/drinkers is not well understood. Currently, OSCC is diagnosed through physical examination and biopsies. Visual inspection or dye staining of the mouth is useful for early detection of oral cancer and precancerous lesions, such as leukoplakia, but still the majority of OSCC are diagnosed at late stages. 2D-gel protein profiling has been used to identify proteins in OSCC tissue and saliva specimens. Analysis of saliva has the advantages of being safe, easy to access and non-invasive. The objective of this study was to evaluate proteins present in saliva of patients with oral leukoplakia (OL)and patients with tongue and floor of mouth (TF)SCC compared to a control group using proteomic techniques to better understand the molecular pathogenesis of OSCC, enabling the tumor to be identified at an early stage in individuals with or without risk factors. A total of 39 primary TFSCC in clinical stage I and II were identified during 2007 and 2008 at Instituto Nacional de Câncer (INCA), and 15 patients with OL were gathered from Odontoclínica Central do Exército and INCA. Patients were submitted to anamnesis and oral inspection. Paraffin embedded tissues were analyzed for diagnosis confirmation, WHO dysplasia evaluation, tumor grading, and histologic risk assessment. Whole saliva samples from ten healthy individuals were employed as a control group and compared to patients with OL as well as TFSCC patients with (n=10) and without (n=10) cancer associated oral habits(smoking and drinking). Cancer groups were compared with each other. Saliva was submitted to centrifugation and the supernatant stored at 80 °C. Saliva from each group was pooled (n=10) and precipitated with cold acetone. After 2D electrophoresis the differential protein components were analyzed with Image Master Software and tandem matrix assisted laser desorption/ionization time-of-flight mass spectrometry for protein identification. Descriptive statistics were calculated and a p-value of less than.05 was considered significant. TFSCC patients who smoked and drinked were mainly man, over 45 years old, while non-smokers/drinkers were female with high education level and tongue tumors. Among clinicopathologic features analyzed, only tumoral lymphocytic infiltrate showed association with neck metastasis absence. A total of 48 protein spots representing 20 proteins were identified in LO and TFSCC. Cytokeratin 10 and lysozyme were leukoplakia specific while apolipoprotein-A1 was significantly raised in LO. Beta actin was increased in smokers/drinkers TFSCC. Proapolipoprotein was specific for non-drinkers/smokers TFSCC compared to control, as well as tropomyosin in the same group when compared to smokers/drinkers TFSCC. In conclusion, both usually high- and low-abundant proteins were identified in saliva of TFSCC and OL patients and can be used in the future as a panel for early detection of OSCC. Differences in TFSCC in patients with and without oral habits must be further investigated