Predictive value of DNA ploidy combined with malignant transformation biomarkers in oral potentially malignant diseases

Abstract

Oral squamous cell carcinoma (OSCC) is among the commonest types of cancer worldwide, with approximately 600,000 new cases per year, 120,000 deaths and a 5-year survival of approximately 50%. Early diagnosis and treatment are the most important factors with an impact on patient survival and quality of life. Most OSCCs are preceded by asymptomatic changes of the oral mucosa known as oral potentially malignant diseases (OPMD). There are no pathognomonic clinical or microscopic features to date able to predict malignant transformation. It is suggested that a panel of potential surrogate markers of biological processes relating to tumorigenesis, such as aneuploidy, cell cycle alterations, autophagy, as well as metabolic alterations may have a higher prognostic value than isolated markers alone. The objectives of this study will therefore be: 1) Review the OPMD according to their degree of epithelial dysplasia and to identify cases that have undergone malignant transformation; 2) Evaluate the DNA ploidy of the identified lesions and their correlation with the expression of proteins involved in the process of tumorigenesis, such as cyclin D1, Glut-1, galectin 3, LC3B and Beclin-1; 3) Calculate the predictive values of malignant transformation, sensitivity and specificity of each of the markers as well as combinations of markers.Formalin fixed and paraffin embedded biopsy tissue from the archive of the São Leopoldo Mandic Researh Center will be used. The H&E stained sections form each case will be reviewed for the diagnosis of epithelial dysplasia and divided into 6 groups (N = 240, n = 40): leukoplakia without dysplasia, leukoplakia with mild dysplasia, leukoplakia with moderate dysplasia, leukoplakia with severe dysplasia, carcinoma in situ (positive control) and inflammatory fibrous hyperplasias (negative control). The determination of DNA ploidy will be performed using suspensions of epithelial nuclei obtained from enzymatic digestion, propidium iodide staining and flow cytometry. The biomarkers cyclin D1, Glut-1, galectin 3, LC3B and Beclin-1 will be investigated by immunohistochemistry. Data from ploidy and other biomarkers will be analyzed descriptively and statistically using the Kaplan-Meier survival test for both individual marker and combination of markers. The results are expected to yield a diagnostic and prognostic matrix based on such lesions that could be able to predict malignant transformation of OPMD with high sensitivity and specificity. (AU)