Integrating endogenous peptides analysis and protease mapping for identification of potential serum biomarkers in gastric adenocarcinoma.

Abstract

e15564 Background: Gastric cancer (GC) is the third cause of cancer death in the world. In Brazil 21.000 new cases had been diagnosed in 2018 and the mortality rate is associated with late diagnosis that contributed to poor prognosis. The objective was to study endogenous peptides in serum of patients with GC. Methods: Fifteen serum samples of patients with diagnosis of gastric adenocarcinoma (TNM stage I-IV) and 15 controls were included. Endogenous peptides were extracted and pooled of random mode in 5 biological replicates (n = 3 samples) for each group. The mixture peptides was submitted to nLC-MS/MS analysis and peptide sequences identified were used to protease mapping. Results: A total of 191 peptides (≥ 7 amino acids) were identified corresponding to 36 proteins involved mainly on metabolic and immune system processes, signal transduction, platelet and neutrophils degranulation. Peptidome-based protease mapping identified 59 proteases (29 serine-, 19 metallo-, 8 cisteine- and 3 aspartic-), in which Prothrombin, Plasminogen, MMP14, MMP7 and MMP3 proteases yielded most of peptides. Twenty sequences derived from Fibrinogen A, Fibrinogen B, Complement C3 (C3f), Apolipoprotein A-I (N-terminal), Prothrombin (C-terminal of Activation peptide fragment 2; N-terminal of Thrombin light chain) and Coagulation factor XIII A (N-terminal) were significantly different (p ≤ 0.5). Between these peptides, 14 were up regulated and six downregulated in GC patients. The phosphopeptide 20ADSpGEGDFLAEGGGVR35 (Fibrinopeptide A) was significantly increased (p< 0.03) in GC serum samples, while the non-phosphorylated Fibrinopeptide A was decreased (p<0.04), suggesting a higher phosphorylation rate in this protein in GC patients. Further, the peptide 31QGVNDNEEGFFSAR44 (Fibrinopeptide B) was downregulated (p<0.04) in GC serum samples; in counterpart, the Fibrinopeptide B derived peptides NDNEEGFF (p<0.0007) and QGVNDNEEGFFS (p<0.0013) was up regulated, suggesting a higher proteolysis in GC serum samples. Conclusions: The investigation of protease/substrate activity relationship may be described such as a novel panorama for discovery of serum biomarkers.