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Background: Gastrokine 1 (GKN1) is a stomach-specific tumor suppressor that is secreted into extracellular space as an exosomal cargo protein. The objective of this study was to investigate the uptake and tumor-suppressive pathways of exosome-associated GKN1 protein in gastric epithelial cells. Methods: Immunofluorescent and Western blot analysis were used to investigate gastric-specific uptake of HFE-145-derived exosomes. Binding affinity of HFE-145 derived exosomes with integrin proteins was examined using protein microarray chip. Tumor suppressor activities of exosome-carrying GKN1 protein were analyzed using transwell co-culture, MTT assay, BrdU incorporation, immunoprecipitation, and Western blot analysis. Results: HFE-145-derived exosomes were internalized only into HFE-145 gastric epithelial cells and gastric cancer cells. Gastric-specific uptake of stomach-derived exosomes required integrin α6 and αX proteins. Clathrin and macropinocytosis increased the uptake of exosomes into gastric epithelial cells, whereas caveolin inhibited the uptake of exosomes. Transwell co-culture of AGS cells with HFE-145 cells markedly inhibited viability and proliferation of AGS cells. Following uptake of HFE-145-derived exosomes in recipient cells, GKN1 protein bound to HRas and inhibited the binding of HRas to b-Raf and c-Raf which subsequently downregulated HRas/Raf/MEK/ERK signaling pathways in AGS, MKN1 cells, and MKN1-derived xenograft tumor tissues. In addition, exosomal GKN1 protein suppressed both migration and invasion of gastric cancer cells by inhibiting epithelial–mesenchymal transition. Conclusions: Gastric-specific uptake of exosomes derived from gastric epithelial cells requires integrin α6 and αX proteins in both gastric epithelial cells and exosomes. Exosomal GKN1 protein inhibits gastric carcinogenesis by downregulating HRas/Raf/MEK/ERK signaling pathways. [ABSTRACT FROM AUTHOR]

Early detection of cancer provides effective treatment and saves lives. The objective of this study was to determine whether serum gastrokine 1 (GKN1) protein is a gastric cancer‐specific diagnostic biomarker. The serum concentration of GKN1 in healthy individuals (median: 6.34 ng/μL, interquartile range (IQR): 5.66‐7.54 ng/μL) was significantly higher compared with the levels in gastric cancer patients (median: 3.48 ng/μL, IQR: 2.90‐4.11 ng/μL; P < .0001). At the optimum cutoff (4.94 ng/μL) of serum GKN1 protein, the sensitivity and specificity were 91.2% and 96.0%, respectively, for gastric cancer. Using serum GKN1 protein as the diagnostic reference, the ROC curve showed a satisfactory diagnostic efficacy with an AUC value of 0.9954 (95% CI 0.9919‐0.9988) and Youden index of 0.8740. In addition, the diagnostic accuracy of the serum GKN1 protein at the optimum cutoff was 0.9675. Interestingly, serum GKN1 concentrations in patients with advanced gastric cancer (AGC; median: 3.11 ng/μL, IQR: 2.72‐3.72 ng/μL) were lower than in patients with early gastric cancer (EGC; median: 4.31 ng/μL, IQR: 3.88‐4.88 ng/μL). The diagnostic accuracies at the optimum serum GKN1 cutoff were 0.8912 and 0.9589 for EGC and AGC, respectively. Furthermore, the serum GKN1 concentrations robustly discriminated the patients with gastric cancer from the patients with colorectal, liver, lung, breast, pancreatic, ovary, and prostatic cancers with AUC values greater than 0.94. These data suggest that serum GKN1 is a promising and highly specific diagnostic biomarker for the prompt detection of early and advanced gastric cancers. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) engage in complex interactions with the machinery that controls the transcriptome and concurrently target multiple mRNAs. Here, we demonstrate that microRNA-495-3p (miR-495-3p) functions as a potent tumor suppressor by governing ten oncogenic epigenetic modifiers (EMs) in gastric carcinogenesis. From the large cohort transcriptome datasets of gastric cancer (GC) patients available from The Cancer Genome Atlas (TCGA) and the National Center for Biotechnology Information (NCBI) Gene Expression Omnibus (GEO), we were able to recapitulate 15 EMs as significantly overexpressed in GC among the 51 EMs that were previously reported to be involved in cancer progression. Computational target prediction yielded miR-495-3p, which targets as many as ten of the 15 candidate oncogenic EMs. Ectopic expression of miRNA mimics in GC cells caused miR-495-3p to suppress ten EMs, and inhibited tumor cell growth and proliferation via caspase-dependent and caspase-independent cell death processing. In addition, in vitro metastasis assays showed that miR-495-3p plays a role in the metastatic behavior of GC cells by regulating SLUG, vimentin, and N-cadherin. Furthermore, treatment of GC cells with 5-aza-2'-deoxcytidine restored miR-495-3p expression; sequence analysis revealed hypermethylation of the miR-495-3p promoter region in GC cells. A negative regulatory loop is proposed, whereby DNMT1, among ten oncogenic EMs, regulates miR-495-3p expression via hypermethylation of the miR-495-3p promoter. Our findings suggest that the functional loss or suppression of miR-495-3p triggers overexpression of multiple oncogenic EMs, and thereby contributes to malignant transformation and growth of gastric epithelial cells. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Oxidative stress is accepted as one of the main factors involved in the development and progression of cancer. Superoxide dismutases ( SODs) are important in avoiding oxidative stress by eliminating reactive oxygen species ( ROS). To determine whether single nucleotide polymorphisms at G7958A within SOD1 and at T5482C within SOD2 are associated with an increased susceptibility to gastric cancer, we investigated the genotype and allele frequencies of the genes from 294 gastric cancer patients and 300 healthy individuals. A polymerase chain reaction-single strand conformation polymorphism assay was used to identify the SOD1 G7958A and the SOD2 T5482C genotypes. Statistically significant differences in the genotype and allele frequencies of SOD2 T5482C were found between the healthy controls and gastric cancer patients (p = 0.0001 and p < 0.0001, respectively). When the data were stratified according to gastric cancer histological subtypes, the risk of both diffuse- and intestinal-type gastric cancer was statistically higher for carriers of the C allele compared with carriers of the T allele. However, there were no statistically significant differences in genotype distribution (p = 0.5069) and allele frequencies (p = 0.3714) of SOD1 G7958A between gastric cancer patients and controls. Our findings suggest that polymorphism of the SOD2 T5482C may be closely associated with an increased susceptibility to the development and differentiation of gastric cancer in the Korean population. [ABSTRACT FROM AUTHOR]

Helicobacter pylori increases the risk of gastric cancer development and triggers aberrant expression of activation-induced cytidine deaminase (AID). The goal of the present study was to investigate whether AID expression is involved in the development or progression of gastric cancer and the nuclear expression of p53 protein in cancer cells. We examined the expression pattern of the AID and p53 proteins in 186 gastric adenocarcinomas by immunohistochemistry. In order to investigate the molecular mechanism of AID expression, we also searched for mutations in the AID gene by single-strand conformational polymorphism and sequencing methods. In 186 sporadic gastric cancers, AID expression was detected in the 73 corresponding normal gastric mucosa and in 50 gastric cancers. Statistically, the expression of AID protein was not associated with clinicopathological parameters, including tumor size, location, differentiation and lymph node metastasis (p > 0.05). Interestingly, a significant association was observed between AID and the nuclear expression of p53 (p = 0.0094). Mutational analysis revealed no mutation in the AID gene in the gastric cancers. These results suggest that aberrant expression of the AID protein may contribute to the development of gastric cancers and induce p53 nuclear expression. Copyright © 2008 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Elevated levels of the calcium-binding protein S100A4 cause metastasis of benign rat mammary tumor cells. To investigate whether S100A4 plays an important role in the invasion and metastasis of gastric cancers, we examined the gene mutations in the coding regions and expression patterns of the S100A4 in gastric adenocarcinoma in Korea. Moderate to strong expression of S100A4 was found in 53 (68.8%) of the 77 gastric adenocarcinomas, whilst normal gastric epithelium either failed to stain or showed weak staining. Interestingly, S100A4 expression was more frequently observed in gastric cancer patients with advanced gastric cancer (p=0.039), positive lymph node metastasis (p=0.001), and peritoneal dissemination (p=0.022). No gene mutations were found in the analyzed genomic area in 77 gastric adenocarcinomas and 15 gastric cancer cell lines. We found one single nucleotide polymorphism without an amino acid change, A99G, in two cases. These data suggest that the overexpression of S100A4 may be closely related to the aggressiveness of gastric cancer in Korea. [ABSTRACT FROM AUTHOR]

We investigated whether Helicobacter pylori ( H. pylori ) CagA contributes to the DNA copy change and mRNA transcript expression of the HER-2 gene and, consequently, affects HER-2 protein expression to evaluate the significance of CagA and HER-2 amplification in gastric cancer. We used the AGS and MKN1 gastric cancer and HFE-145 immortalized non-neoplastic gastric mucosa cell lines. We also confirmed the effects of CagA on HER-2 expression in human gastric cancer tissues and gastric mucosal tissues of H. pylori infected C57BL/6 mice. Ectopic CagA expression in AGS, MKN1 and HFE-145 cells showed a significant increase in HER-2 gene copy number and expression. The gastric mucosae of H. pylori infected C57BL/6 mice also showed increased HER-2 DNA copy number and protein expression. In addition, CagA expression was detected in 17 (56.7%) of 30 gastric cancer tissues, and eight (47%) of them showed HER- 2 DNA amplification of more than two-fold. In immunohistochemistry, HER-2 overexpression was detected in 12 (40%) of 30 gastric cancers and a positive correlation was observed among DNA copy number, the mRNA transcript, and protein expression of the HER- 2 gene in gastric cancer ( P < 0.05). These results suggest that H. pylori CagA may induce overexpression of the HER-2 protein by increasing HER-2 DNA and mRNA copy number. [ABSTRACT FROM AUTHOR]

Abstract: Fas-associated death domain (FADD) plays a crucial role during death receptor-mediated apoptosis. In addition, FADD possesses apoptosis-independent activities, including cell-cycle regulation and cell proliferation regulated by the phosphorylation of FADD at Ser194. The aim of this study was to explore the possibility whether alteration of phosphorylated FADD (p-FADD) expression might be a characteristic of gastric cancer. We analyzed the expression of p-FADD protein in 60 gastric adenocarcinomas by immunohistochemistry using a tissue microarray approach. In the normal gastric mucosal cells, surface and glandular epithelial cells evenly expressed p-FADD in the nuclei but not in the cytoplasm. In the cancers, p-FADD expression was detected in 38 cases (63%) of the gastric carcinomas, but there was no p-FADD immunostaining in the remaining 22 cancers (37%). Of note, p-FADD immunostaining was observed in cytoplasm/nuclei (20 cancers; 33%) and cytoplasm (18 cancers; 30%). There was no significant association of p-FADD expression with clinocopathological characteristics, including invasion, metastasis, and stage. Our data showed that the expression of p-FADD in gastric cancers was heterogenous in its location compared to the uniform nuclear expression of p-FADD in normal gastric cells. Many of the cancers (67%) were devoid of nuclear p-FADD, suggesting that p-FADD functions in the nucleus may be perturbed in the cancers. Also, p-FADD expression in the cytoplasm in a large fraction of the cancers (63%), not seen in the normal cells, suggested that the cell death functions of p-FADD could be altered in the cancer cells. [Copyright &y& Elsevier]