Inflammatory response elements, glycan and proteome profiles as salivary biomarkers for the early diagnosis of oscc

Resumen

Oral cancer, predominantly oral squamous cell carcinoma (OSCC), is the most common malignant neoplasia in the oral cavity characterized by poor prognosis and a low survival rate, when diagnosed at advanced disease. It can be treated effectively if detected at initial stages with better prognosis. Therefore, early diagnosis has become an ultimate goal in the management of oral cancer after prevention, considered to have a positive impact on survival and quality of life of these patients. The shortage of symptoms at the disease onset often remain unnoticed, ultimately leading to discovery at advanced phases, so improvement of the diagnostic manner is required. Reliable biomarkers for OSCC are yet unavailable in the routine clinical setting, emphasizing the emerging need of a practical and simple tool to be used for definitive diagnosis, as well as for screening programs. OSCC tumours appear through a series of molecular mutations leading to uncontrolled cellular growth from hyperplasia areas to dysplastic lesions, to carcinoma in situ and is finally followed by invasive carcinoma. Besides, the development of many OSCC cases have been correlated to cancerous transformation of oral potentially malignant disorders (OPMDs) such as oral leukoplakia. The latter exhibits heterogeneous subtypes with varying modification potentials, among which proliferative verrucous leukoplakia (PVL) stands out with high risk of malignant conversion. Diagnosis of OSCC requires sensitive and specific indicative tools that can support untraceable and difficult to access sites of the oral mucosa for which the most recommended discovery medium are biological fluids, such as saliva. Sputum derived biomarkers offer easy sampling with reduced risk for the patients, cost and diagnosis time, encompassing a variety of detectable and measurable parameters that can discriminate health from disease. In an attempt to identify novel salivary biomarkers of OSCC, we herein combined multi- omics analytical strategies to profile different types of molecules with potential diagnostic utility. As inflammation has previously been linked to oral pathologies, research so far indicates the possibility of using salivary pro and anti-inflammatory proteins for screening of oral disorders. Altered cytokine responsiveness has been tightly associated with the development of OSCC, as well as detected in patients with OPMDs. To reveal changes related to oral carcinogenesis, a multiplex immune bead-based assay was used to survey the levels of 8 different cytokines in saliva of patients with PVL, at early and advanced OSCC stages and their healthy counterparts. Results indicated altered cytokine expressions associated with the pathology groups, the predictive models for sensitivity and specificity of which showed high ROC AUC values. It is now well established that defective glycosylation accompanies many chronic and infectious conditions and is a common feature of tumour cells that may affect N-glycans on glycoproteins. To compile a list of candidate biomarkers, another type of molecules have been studied. Saliva derived N-glycans of healthy volunteers, PVL and OSCC patients were analysed by the means of liquid chromatography (LC) coupled to mass spectrometry (MS). Comparative N-glycome profiling revealed several differentially expressed fucosylated bi- and tri-antennary glycans among the studied groups, providing a reasonable platform to further investigate the utility of salivary glycosylation for diagnosis of OSCC. Lastly, to get an insight into the complex molecular alterations associated with cancer, LC-MS generated proteomic profiles revealed more than 600 quantified proteins in saliva of healthy and pre/cancerous lesions. Comparative analysis resulted in a list of differentially expressed proteins, characterized in OSCC, indicating significantly altered mechanisms implicating the immune system, inhibition of enzymatic activities, and cell adhesion. Among the identified candidate markers, some had previously been described in saliva. In addition, several newly OSCC-associated proteins have been annotated. In summary, in this discovery phase of biomarker identification, we provide a molecular panel of potential indicators of malignant transformation and/or early OSCC diagnosis. Further validation is needed to verify its clinical utility.