Integrated Proteomic And Phosphoproteomic Analysis Of Gastric Adenocarcinoma Discovers Molecular Signatures Capable Of Stratifying Patient Outcome

Gastric cancer is one of a leading cause of cancer-related deaths worldwide,among which gastric adenocarcinoma(GAC)is the most common form.Diagnosis of GAC is generally made at advanced stages when metastatic spreading occurs,which reduces the efficacy of surgical treatment and contributes to poor patient survival.In this study,focusing on lymph node metastasis(LNM)of gastric adenocarcinoma,we collected 11 tumor samples without LNM and 12 samples with LNM as well as para-tumor tissues.We applied proteomic analysis to compare primary tumor tissues from GAC patients with or without LNM,and captured unique molecular features of GAC patients with LNM.We applied fuzzy c-means clustering(Mfuzz)to capture the dynamic protein and phosphorylation sites expression along different stages of GAC,considering both tumor and para-tumor tissues simultaneously.Combined with KEGG and overall survival analysis,we provided a series of biological pathways and biomarkers that play a significant role in gastric cancer progression.We also applied quantitative mass spectrometry to compare the proteome and phosphoproteome of primary tumor tissues between GAC patients with and without lymph node metastasis(LNM).We then performed an integrated analysis of the proteomic and transcriptomic data to reveal the molecular features.We quantified a total of 5536 proteins,with 218 up-regulated and 49 down-regulated proteins in tumor samples from patients with LNM.Focal adhesion and extracellular matrix play an essential role in GAC with LNM.Integrating TCGA data,we found that TNXB and SPON1 are over expressed in primary tumors of GAC patients with LNM comparing to GAC patients without LNM,and over expression of both proteins correlates with poor GAC patient survival.Furthermore,treating gastric cancer cells with anti-TNXB antibody significantly reduced the migration of these cells.Finally,quantitative phosphoproteomic analysis revealed a number of activated kinases including GSK-3,in primary tumor tissues from patients with LNM.Our study provides a snapshot of the proteome and phosphoproteome of GAC tumor tissues that have metastatic potential,and identifies potential biomarkers for GAC spreading.