Villano, Valentina (2016) Searching for informative biomarkers for Gastric Cancer. [Tesi di dottorato]

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Gastric cancer (GC) is still one of the prevalent leading causes of cancer-related deaths worldwide and high mortality rate is mainly due to late-stage diagnosis. GC is characterized by two distinct histological type of adenocarcinomas each having different epidemiological and pathophysiological features. The intestinal-type generally evolves through a relatively well-defined sequence of histological lesions. The diffuse-type has instead a poorer prognosis and develops through unknown genetic and morphological events from normal gastric epithelium. The pathogenesis of GC remains poorly understood however several environmental factors, such as Helicobacter pylori (H. pylori) infection can be the cause leading to this disease. This risk is probably the result of a combination of genetic and environmental factors in which the infection by H. pylori is of particular relevance. To search for gastric cancer biomarkers, we have analyzed the protein profile of malignant and normal gastric tissues and identified a novel stomach specific protein gastrokine 1 (GKN1) whose expression is reduced in H. pylori infected gastric mucosa and down-regulated or completely absent in GC tissues and precancerous lesions so this protein might play an important role as biomarker in carcinogenic process. On the basis of these evidences, we are investigating on the possible utility of GKN1 as an informative biomarker. To do this, we have analyzed by Western blot the expression profile of GKN1 in several biological fluids of normal individual in order to highlight the presence of GKN1 at the protein level. However, no GKN1 expression was possible to detect even if the search was performed by ELISA assay on serum of healthy individuals. We then have searched for the presence of GKN1 mRNA in the serum of healthy subjects by RT-PCR using several primers spanning over the entire GKN1 cDNA. This result suggests that qRT-PCR might be a possible media by which to follow the GKN1 expression profile in normal patients and in patients with gastric lesions in order to assess its possible use as preventive gastric cancer biomarker. The second part of my work is focused on identification of potential GKN1 interactors with the aim to add knowledge on its functional role within gastric cell lines and tissues. To this purpose we used a proteomic strategy and the GKN1 interactors were identified by peptide mass fingerprinting using MALDI-TOF mass spectrometry. The results show that GKN1 interacts whit SLC26A3, a protein expressed in apical membrane of intestinal epithelial cells. Therefore, we propose in future, the evalutation of SLC26A3 in human sera as a potential innovative biomarker in CG.

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