Biopsia liquida per la diagnosi precoce del carcinoma orale: ruolo dei microRNA come nuovi biomarkers

Oral cancer is the most frequent and aggressive tumor of the oral cavity. Due to its highly aggressive behavior, oral cancer represents a serious problem for human health. In addition, despite the availability of screening programs and the accessible anatomical sites of oral lesions, the incidence, prevalence and mortality rates are high as the diagnosis is often made when the neoplasm is in an advanced state. Therefore, advanced oral cancer patients have often a poor prognosis. Therefore, it is evident how the discovery of novel biomarkers for the management of this pathology is necessary to early detect oral cancer and improve patients’ clinical outcomes. At present there are no effective diagnostic or prognostic biomarkers for oral cancer, on this basis, the aim of the present study was to identify and validate a set of microRNAs (miRNAs) involved in the development and progression of oral cancer thus useful as predictive biomarkers for this tumor. For this purpose, both computational and experimental analyses were performed in order to establish the diagnostic and prognostic potential of the miRNAs here identified. Through the analysis of miRNA expression data and clinical data obtained from The Cancer Genome Atlas Head and Neck Cancer (HNSC) and GEO DataSets databases a set of eleven miRNAs significantly involved in oral cancer was identified. Through further computational analyses, the functional roles of these 11 miRNAs were evaluated and four miRNAs, the up-regulated hsa-miR-196a- 5p and hsa-miR-503-5p, and the two down-regulated miRNAs hsa-miR-133a- 3p and hsa-miR-375-3p, were selected for the validation analyses performed on liquid biopsy samples collected from oral cancer patients, individual at risk for this tumor and healthy controls. The validation analyses on liquid biopsy samples collected from patients and controls were performed by using the high sensitivity droplet digital PCR (ddPCR) amplification system in order to evaluate the expression levels of selected miRNAs in serum and saliva samples obtained from oral cancer patients, oral lichen planus (OLP) patients and healthy controls. The ddPCR results obtained in serum samples revealed the high diagnostic and predictive value of hsa-miR-503-5p and hsa-miR-133a-3p. Of note, the diagnostic potential of miRNAs was higher in saliva samples which showed a higher significant difference in the expression levels of both hsa-miR-503-5p and hsa-miR-133a-3p. More in detail, by analyzing the miRNA expression levels of hsa-miR-503-5p, hsa-miR-133a-3p and hsa-miR-375-3p in normal individuals, OLP and oral cancer patients the expression levels of hsa-miR-503- 5p observed in both serum and saliva samples were strongly predictive for OLP and oral cancer development. Also in this case, saliva samples were more reliable compared to serum samples in correctly diagnosing oral lesions. Further statistical analyses confirmed the high diagnostic value of the miRNAs identified. Also the prognostic value of the selected miRNAs was confirmed with further computational analyses. Overall, the results obtained revealed that the evaluation of hsa-miR-196a-5p, hsa-miR-503-5p, hsa-miR-133a-3p and hsa-miR-375-3p in serum and saliva samples of patients with suspicious lesions may be helpful for the early identification of oral cancer. Therefore, the use of ddPCR and liquid biopsy samples may represent an innovative strategy to improve the management of oral cancer.