Well-differentiated Gastric Cancer Research Articles

Clinicopathological characteristics of well-differentiated gastric cancer missed by endoscopy.

The objective was to explore the clinicopathological features of moderately well-differentiated gastric cancer missed by endoscopy. The clinicopathological data of 88 patients who were diagnosed as moderately well-differentiated gastric cancer by biopsy and pathology from January 2019 to December 2020 in Anyang Tumor Hospital were analyzed retrospectively. The clinicopathological features and immunophenotypic characteristics of these gastric cancers were analyzed together with the literature review. Seventy males and 18 females were included in this study. There were 59 cases of gastritis, 14 cases of ulcer and 15 cases of polyp under endoscope. Based on mucous phenotype, there were 27 gastric cases, 18 intestinal cases, 39 gastrointestinal cases and 4 untyping cases. 54 cases expressed mutant p53 and 34 cases had wild type p53. None of the indicators could yield reliablediagnosis, which requires comprehensive morphological and immunohistochemical analysis by the pathologist. Meanwhile, the clinician should give the patient a systematic treatment plan incuding endoscopy, magnifying endoscopy, NBI, linear endoscopic ultrasound and other examinations. Regardless of the diagnostic criteria, once the pathology suggests a tumor lesion, local resection should be performed first. Additional surgery can be performed if there is submucosal invasion.

The Intestinal Stem Cell Marker SMOC2 Is an Independent Prognostic Marker Associated With Better Survival in Gastric Cancer.

SPARC-related modular calcium-binding protein 2 (SMOC2), a secreted matricellular protein, is reported to be involved in cancer progression such as cell cycle, angiogenesis, and invasion. In this study, we aimed to investigate the expression of SMOC2 in various gastric lesions and assessed its prognostic value in a large cohort of gastric cancer (GC) patients. SMOC2 mRNA levels were measured by quantitative real-time PCR using 26 matched fresh-frozen GC samples. SMOC2 protein expression was determined by immunohistochemistry on tissue microarrays including 734 GC specimens and its correlations with clinicopathological features and survival were evaluated. The transcription level of SMOC2 was higher in GC samples compared to normal mucosa (p=0.006). Its expression levels were associated with the intestinal stem cell (ISC) marker, LGR5, but there were no correlations with EPHB2 and OLFM4 or the candidate cancer stem cell markers CD133 and CD44. SMOC2 expression was significantly increased in the intestinal metaplasia and was further increased in gastric adenomas and early gastric cancers (EGC). In total, 34% of GCs were positive for SMOC2, and SMOC2 positivity was higher in old (p=0.001) and male (p<0.001) patients, and in well-differentiated GC (p<0.001). SMOC2 expression had a negative association with perineural invasion (p<0.001) and tumor stage (p<0.001). In survival analysis, SMOC2-positive GC patients had much better clinical outcomes in overall survival rates (p<0.001) compared to SMOC2-negative GC patients. The prognostic impact of SMOC2 remained significant both in intestinal (p<0.001) and diffuse-type GC (p<0.001). Remarkably, a multivariate analysis demonstrated SMOC2 as an independent prognostic marker [hazard ratio (HR)=0.732, p=0.045] along with venous invasion (p=0.012), tumor stage (p<0.001) and CDX2 (p=0.028). Our results suggest that SMOC2 can be a prognostic marker for better clinical outcomes in GC.

Transcriptome Analysis and the Prognostic Role of NUDC in Diffuse and Intestinal Gastric Cancer.

Introduction:There have been few studies about gene differences between patients with diffuse-type gastric cancer and those with intestinal-type gastric cancer. The aim of this study was to compare the transcriptomes of signet ring cell gastric cancer (worst prognosis in diffuse-type) and well-differentiated gastric cancer (best prognosis in intestinal-type); NUDC was identified, and its prognostic role was studied.Materials and Methods:We performed next-generation sequencing with 5 well-differentiated gastric cancers and 3 of signet ring cell gastric cancer surgical samples. We performed gene enrichment and functional annotation analysis using the Database for Annotation, Visualization and Integrated Discovery bioinformatics resources. Immunohistochemistry was used to validate NUDC expression.Results:Overall, 900 genes showed significantly higher expression, 644 genes showed lower expression in signet ring cell gastric cancer than in well-differentiated gastric cancers, and there was a large difference in adhesion, vascular development, and cell-to-cell junction components between the 2 subtypes. We performed variant analysis and found 52 variants and 30 cancer driver genes, including NUDC. We analyzed NUDC expression in gastric cancer tissue and its relationship with prognosis. Cox proportional hazard analysis identified T stage, N stage, and NUDC expression as independent risk factors for survival (P < 0.05). The overall survival of the NUDC-positive group was significantly higher (53.2 ± 0.92 months) than that of the NUDC-negative group (44.6 ± 3.7 months) (P = 0.001) in Kaplan-Meier survival analysis.Conclusion:We found 30 cancer driver gene candidates and found that the NUDC-positive group showed significantly better survival than the NUDC-negative group via variant analysis.

Gastric adenocarcinoma of the fundic gland: A review of clinicopathological characteristics, treatment and prognosis.

Gastric adenocarcinoma of the fundic gland is a rare, well-differentiated gastric cancer entity, and very few patients transition to poorly differentiated tubular adenocarcinoma during progression. Gastric adenocarcinoma of the fundic gland originates from the mucosa of the gastric fundic gland, usually without chronic gastritis or intestinal metaplasia. Histologically, the tumor cells are closely arranged to form anastomosing tubular glands, and more than 95% of tumor cells differentiate towards chief cells. Most gastric adenocarcinoma of the fundic gland cases are characterized by submucosal involvement, but the tumor volume is usually small, with lymphatic and vascular invasion rarely observed. Therefore, endoscopic submucosal dissection can be an ideal treatment, leading to a favorable prognosis, and recurrence and metastasis of the disease are uncommon.

Accumulation of stabilin-1 positive macrophages in the early stage of gastric cancer is associated with short cumulative survival.

The scavenger receptor stabilin-1 has been reported to be expressed by tumor-associated macrophages (TAMs) and to facilitate tumor growth and metastasis in mouse models of breast carcinoma and melanoma. However, to the best of our knowledge, its expression and association with prognosis in human gastric cancer has not been evaluated. The present study investigated the expression of stabilin-1 and its association with clinicopathological parameters in patients with gastric cancer. The expression of stabilin-1 was evaluated by immunohistochemical staining of gastric cancer tissue samples of 371 Chinese patients with primary gastric adenocarcinoma. Confocal laser scanning microscopy was used to determine the cellular source of stabilin-1 in the gastric cancer tissues using anti-CD68, anti-CD163, anti-stabilin-1 and anti-secreted protein acidic and rich in cysteine antibodies. A higher number of stabilin-1-positive cells were observed in the cancer tissues of primary gastric adenocarcinoma compared with adjacent non-cancerous tissues of primary gastric adenocarcinoma (P<0.001); the majority of stabilin-1-positve cells were CD68+/CD163+ macrophages. Poorly-differentiated gastric cancer tissue had fewer stabilin-1-positive cells compared with medium- and well-differentiated gastric cancer (P=0.030). A higher number of stabilin-1-positive cells were found in the early Tumor-Node-Metastasis (TNM) stage (TNM I stage) of primary gastric adenocarcinoma (P=0.038) compared with TNM stage IV. For patients with TNM stage I disease, a higher number of stabilin-1-positive TAMs was associated with shorter cumulative survival (P<0.05). Overall, stabilin-1 was found to be expressed by CD68+ TAMs in human gastric cancer. The high expression of stabilin-1 in TNM stage I disease was associated with poor patient survival, indicating the clinical significance of stabilin-1 in gastric cancer.

Increased Sulfiredoxin Expression in Gastric Cancer Cells May Be a Molecular Target of the Anticancer Component Diallyl Trisulfide.

Sulfiredoxin (Srx) is a newly discovered antioxidant enzyme playing a role in the catalytic reduction of oxidative modifications. Srx is overexpressed in a variety of cancers. It may promote carcinogenesis as well as tumor progression. In this study, we report for the first time that Srx expression might be positively associated with the development of gastric cancer and tumor malignancy. Immunohistochemistry showed that, compared to normal tissues (42%, 20/47), Srx expression in gastric tumors (85%, 40/47) was much more common (chi-square test, p<0.01). In addition, the staining of Srx was stronger in poorly differentiated gastric cancer than in well-differentiated gastric cancer. Western blotting showed that, in the gastric tumor cell line BGC823, the Srx protein was upregulated in response to H2O2 treatment, although it was inadequate to counteract the increased oxidative stress, as indicated by the gradually increasing level of malondialdehyde (MDA). In addition, Srx expression, MDA levels, and ROS levels in BGC823 cells were markedly inhibited upon treatment with diallyl trisulfide (DATS), a major constituent of garlic oil with proven anticancer effects. These results suggest that Srx may be an oxidative stress marker. Antioxidation may account for the anticancer potential of garlic.

Simultaneous Analysis of Sulfated and Phosphorylated Glycans by Serotonin-Immobilized Column Enrichment and Hydrophilic Interaction Chromatography.

Changes in the structures and quantities of sulfated glycans play important roles in inflammatory and neurological diseases and cancer. Therefore, sulfated glycans are expected to become diagnostic markers for a variety of diseases such as Alzheimer's disease and cancer. On the other hand, structural abnormalities in the phosphorylated glycans on lysosomal enzymes cause a number of lysosomal diseases, while novel phosphorylated glycans have been found in other proteins. As with sulfated glycans, structural and quantitative changes in these phosphorylated glycans and their associations with disease are also of interest. In this article, we introduce a new method for the simultaneous analysis of sulfated and phosphorylated glycans. We first employ a serotonin-immobilized column to enrich these glycans. Glycans obtained in this manner were sequentially subjected to other analytical techniques without desalting. We employed hydrophilic interaction chromatography to distinguish the sulfate and phosphate groups of the glycans and were able to analyze sulfated and phosphorylated N-glycans expressed on thyroglobulin, ovalbumin, and myozyme. We showed that our method not only analyzes sulfated and phosphorylated glycans, but also glycans containing the GlcNAc-HPO3 residue. We comparatively analyzed sulfated glycans, phosphorylated glycans, and GlcNAc-HPO3-residue-containing glycans expressed on MKN7 cells (well-differentiated gastric cancer cells) and MKN45 cells (poorly differentiated gastric cancer cells). To the best of our knowledge, this is the first report of a method for the simultaneous and quantitative analysis of sulfated and phosphorylated glycans.

Survivin and XIAP expression in distinct tumor compartments of surgically resected gastric cancer: XIAP as a prognostic marker in diffuse and mixed type adenocarcinomas.

There is considerable evidence that the inhibitor of apoptosis protein (IAP) family serves a role in tumorigenesis. The most studied IAP family members, survivin and X-linked inhibitor of apoptosis (XIAP), have been demonstrated to serve as biomarkers in distinct tumor entities. Thus, the present study aimed to investigate the expression levels of both IAPs in the tumor center, invasion front and lymph node metastases of surgically resected gastric cancer (GC) specimens. Tissue microarrays containing samples from 201 primary GCs were analyzed. IAP expression was detected using immunohistochemistry in different tumor compartments, normal mucosa and lymph node metastases. In addition, the association between the expression levels of these proteins, and clinicopathological parameters and overall survival was investigated. High levels of survivin and XIAP were evident in GC, when compared with normal mucosa, and were correlated with intestinal-type and well-differentiated GC, as well as low International Union Against Cancer stages. Increased XIAP expression was detected in lymph node metastases as compared with corresponding primary tumors. XIAP overexpression was identified to be an independent negative prognostic marker in diffuse and mixed type GC. These results suggest a potential role of survivin and XIAP in the early phase of gastric carcinogenesis. In addition, increased XIAP expression in lymph node metastases supports the observation that IAPs serve an essential role in metastatic tumor disease. Since XIAP expression was identified to be associated with poor survival in diffuse and mixed type GC, XIAP may serve as a novel therapeutic target in these types of GC.

Dual role of carcinoembryonic antigen-related cell adhesion molecule 6 expression in predicting the overall survival of gastric cancer patients

Carcinoembryonic antigen-related cell adhesion molecule 6 (CEACAM6) is a member of the glycosylphosphatidylinositol-linked immunoglobulin superfamily that is implicated in many human cancers. Here, we aimed to investigate the role of CEACAM6 expression in predicting the overall survival (OS) in gastric cancer (GC). The impact of CEACAM6 on the survival of patients with GC (n = 876) was assessed using an online Kaplan-Meier plotter. Findings were validated using the OS data of patients (n = 160) recruited from Ruijin Hospital. We found that high CEACAM6 expression was associated with a better OS in early-stage or well-differentiated GC, or who were treated without 5-fluorouracil (5-FU). Conversely, high CEACAM6 expression was associated with a poor OS in advanced-stage GC, poorly differentiated tumors, or who were treated with 5-FU. Furthermore, CEACAM6 may serve as a better marker for predicting OS in GC than CEA. In addition, CEACAM6 overexpression in GC cells increased apoptotic resistance to 5-FU. Moreover, CEACAM6 induced cluster of differentiation 4- and 8-positive lymphocytes were detected in early-stage GC. In conclusion, CEACAM6 plays a contradictory role in predicting the OS in GC. In early-stage GC, high CEACAM6 expression is associated with improved OS. However, in advanced-stage GC, high CEACAM6 expression is associated with a poor OS.

Incidence of human epidermal growth factor receptor-2 expression and its correlation with clinicopathological characteristics in gastric cancer at tertiary care centre at IGMC Shimla, Himachal Pradesh, India

Background: Trastuzumab has been approved for the treatment of patients with HER-2 positive metastatic/locally advanced adenocarcinoma of the stomach by FDA.1 Most of gastric cancer patients present at an advanced stage in our country, all patients of gastric adenocarcinoma should be subjected to HER-2 testing at initial diagnosis. Immunohistochemistry should be the initial testing methodology and Fluorescence in-situ hybridization should be used to retest immunohistochemistry 2+ samples.Methods: We conducted this study with the aim to establish the expression profile of HER-2 and its correlation to clinicopathological characteristics in resectable cases of gastric adenocarcinoma, in patients presenting with gastric adenocarcinoma at IGMC Shimla. The study was conducted over a year from 1st July 2014 to 30th June 2015. Sample size included 40 diagnosed cases of resectable gastric adenocarcinoma. All patients with operable and resectable disease were subjected to surgery and the specimen were sent for detailed histopathological examination and immunohistochemistry . χ 2 statistical analysis was performed to assess the HER-2 positivity rate . All statistics were performed using 2-sided analysis, with a significance level of p&lt; 0.05.Results: In our study , 7 cases (17.5%) were scored as strongly positive for HER-2 membrane staining (3+), 4 cases (10%) were moderately positive (2+) and 29 cases (72.5%) were HER-2 negative (0/1+) on IHC. The positivity rates in diffuse type (22.2%) and poorly-differentiated (25%) gastric cancer were higher than those in intestinal type (9.9%) and well-differentiated (14.3%) gastric cancer respectively.Conclusions: There is a strong association of human epidermal growth factor receptor-2 positivity with Gastric Malignancy in our country especially in poorly differentiated cancers. Targeted biological therapy with Trastuzumab hence becomes an attractive addition to the multimodality therapy usually required in the treatment of locally advanced /metastatic gastric cancers.

Assessment of histological differentiation in gastric cancers using whole-volume histogram analysis of apparent diffusion coefficient maps.

To investigate the efficacy of histogram analysis of the entire tumor volume in apparent diffusion coefficient (ADC) maps for differentiating between histological grades in gastric cancer. Seventy-eight patients with gastric cancer were enrolled in a retrospective 3.0T magnetic resonance imaging (MRI) study. ADC maps were obtained at two different b values (0 and 1000 sec/mm2 ) for each patient. Tumors were delineated on each slice of the ADC maps, and a histogram for the entire tumor volume was subsequently generated. A series of histogram parameters (eg, skew and kurtosis) were calculated and correlated with the histological grade of the surgical specimen. The diagnostic performance of each parameter for distinguishing poorly from moderately well-differentiated gastric cancers was assessed by using the area under the receiver operating characteristic curve (AUC). There were significant differences in the 5th , 10th , 25th , and 50th percentiles, skew, and kurtosis between poorly and well-differentiated gastric cancers (P < 0.05). There were correlations between the degrees of differentiation and histogram parameters, including the 10th percentile, skew, kurtosis, and max frequency; the correlation coefficients were 0.273, -0.361, -0.339, and -0.370, respectively. Among all the histogram parameters, the max frequency had the largest AUC value, which was 0.675. Histogram analysis of the ADC maps on the basis of the entire tumor volume can be useful in differentiating between histological grades for gastric cancer. 4 J. Magn. Reson. Imaging 2017;45:440-449.

Gastric cancer cell proliferation is suppressed by frizzled-2 short hairpin RNA.

In order to identify novel targets for the molecular therapy of gastric cancer (GC), we investigated the mRNA and protein expression of frizzled-2 (Fz2), a Wnt signaling pathway receptor. Reverse-transcriptase polymerase chain reaction (PCR) amplification was utilized to determine the expression patterns of Fz genes in normal stomach and in the GC cell lines MKN45 and MKN74. Immunostaining was performed on surgical specimens of GC using an antibody against Fz2. The 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2- (4-sulfophenyl)-2H-tetrazolium inner salt (MTS) assay was performed on MKN45 cells and MKN74 cells transfected with Fz2 short-hairpin (sh) RNA. Cell motility was analyzed by scratch assay following Fz2 shRNA. Real-time quantitative PCR was performed to analyze the expression levels of cyclin D1 and matrix metallopeptidase 9 (MMP-9). Fz1, 3, 6 and 8 were expressed in normal stomach, and in MKN45 and MKN74 cells. Fz2 was expressed in normal stomach and in MKN45, but not in MKN74 cells. Well-differentiated GC tissue was weakly positive for Fz2 in cell membranes. Fz2 was positive in both the cell membrane and cytoplasm of GC tissues of moderately differentiated and poorly differentiated adenocarcinoma. Signet ring cells were positive for cytoplasmic Fz2. Proliferation of MKN45 and MKN74 cells was suppressed by Fz2 shRNA, and a scratch assay demonstrated that Fz2 shRNA suppressed also MKN45 and MKN74 cell motility. Furthermore, Fz2 shRNA application led to downregulated mRNA expression of both cyclin D1 and MMP-9. Fz2, 3, 6 and 8 were expressed in normal stomach, and in MKN45 and MKN74 GC cells. Fz2 shRNA suppressed cell proliferation and motility of MKN45 and MKN74 cells, and downregulated cyclin D1 and MMP-9 expression in these GC cell lines.

Abstract 546: CD44 variant expression is a poor prognostic marker in early gastric cancer and well-differentiated gastric cancer

Abstract The present study is to investigate the significance of the expression of CD44 variant (CD44v), a functional cancer stem cell marker, in the primary gastric cancer. We first tested whether CD44v is selectively expressed in gastric cancer tissue. With the tissue microarray samples containing the 6 normal tissue, 6 H. pylori associated gastritis, 12 intestinal metaplasia, 12 low-grade adenoma, 12 high-grade adenoma, 25 early gastric cancer and 26 advanced gastric cancer. The positive expression rates in tumor were 65.3% (49/75) whereas the positive rates in normal and gastritis tissue were 30.4% (p = 0.003). Next, we investigated the clinical significance of gastric cancer with the tissue microarray containing 333 gastric cancer tissue derived from the patients who underwent radical gastrectomy for gastric cancer from 1999 to 2007. The positive expression rates of CD44v (55%, 134/242) were higher in intestinal type gastric cancer than in diffuse type (28%, 21/76). Intestinal type gastric cancer among the CD44v-positive gastric cancer accounted for 82% (134/164). The 5-year survival rate of CD44v-positive gastric cancer was 67.7% whereas that of CD44v-negative gastric cancer was 62.9%. The survival difference was higher as time passed, but it was statistically insignificant (p = 0.219). However, in the subgroup analysis we found that the survival differences were significantly higher in early gastric cancer and well-differentiated gastric cancer (p = 0.013, 5-YSR ; p = 0.046, 5-YSR , respectively). We also investigated the correlation between the expressions of CD44v and EGFR along with the well-known proliferative molecular biomarkers, cylin D1 and Ki-67. Only the expression of Ki-67 were some correlation with CD44v expression in gastric cancer; CD44v-positive rates of the cancers with negative or 1 positive Ki-67 expression were 37% (47/127) whereas those with 2 or 3 positive Ki-67 expression were 57.4% (116/202). This study demonstrates that CD44v can be a good biomarker for early gastric cancer and well-differentiated gastric cancer. [This study was supported by a grant from the National R&amp;D Program for Cancer Control, Ministry for Health, Welfare &amp; Family Affairs, Republic of Korea. (0820050).] Citation Format: Won Sup Lee, Se-Il Go, Jeong-Hee Lee, Sang-Ho Jeong, Young-Joon Lee, Soon Chan Hong, Woo Song Ha, Gyung Hyuck Ko. CD44 variant expression is a poor prognostic marker in early gastric cancer and well-differentiated gastric cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 546. doi:10.1158/1538-7445.AM2014-546

OCT-1 overexpression is associated with poor prognosis in patients with well-differentiated gastric cancer

Octamer transcription factor-1 (OCT-1) is a well-known transcription factor that is reportedly overexpressed in intestinal metaplasia and gastric carcinoma in the intestine. In this study, we investigated OCT-1 overexpression as a prognostic factor for gastric cancer. The association between OCT-1 overexpression (detected using immunohistochemistry) and clinicopathological features including survival was evaluated. In vitro gain-of-function approaches were utilized to assess the function of OCT-1 in malignancy. Analysis of OCT-1 expression in patients with gastric cancer with well-differentiated carcinoma as per the World Health Organization classification showed that OCT-1 overexpression was correlated with advanced tumor invasion (58.8 % of patients with advanced tumor invasion vs. 21.2 % of patients with early tumor invasion; p<0.01), lymph node metastasis (63.9 % of patients with metastasis vs. 24.1 % of those without; p=0.015), and cancer recurrence (83.3 % of patients with recurrence vs. 25.4 % of those without; p<0.01), as well as a lower survival rate (62.8 vs. 87.9 Mo; p<0.01). However, there were no significant differences in the levels of OCT-1 expression in gastric cancer patients with other carcinoma types (p>0.05). Furthermore, we found that the proliferation rate of OCT-1-overexpressing MKN-45 cells was higher than that of the control cells. OCT-1 overexpression may be a marker for poor prognosis in patients with well-differentiated gastric adenocarcinoma.

MiR-7 inhibits the invasion and metastasis of gastric cancer cells by suppressing epidermal growth factor receptor expression

The present study profiled differentially expressed microRNAs (miRs) in gastric cancer cell lines and then investigated miR-7 expression in gastric cancer tissue specimens and the effects of miR-7 on the growth, invasion and metastasis of gastric cancer cells and the underlying molecular events. A microRNA microarray was used to profile differentially expressed miRNAs in human gastric cancer cell lines relative to a normal stomach mucosal epithelial cell line. The miRNA miR-7 was selected for further investigation, which included real-time reverse-transcription PCR (qRT-PCR) analysis of miR-7 levels in different gastric cancer cell lines and tissues and distant non-tumor tissues from patient resections. Cell counting kit-8 (CCK-8), Transwell migration and invasion, and western blot assays were performed to assess tumor cell viability, invasion and gene expression, respectively, after miR-7 transfection. The miRNA microarray profiling revealed 14 upregulated miRNAs (including miR-21, miR-26b and miR-30b) and 19 downregulated miRNAs (including let-7i, miR-7 and miR-622) between gastric cancer and normal cell lines. The qRT-PCR analysis confirmed that reduced miR-7 expression occurred more frequently in poorly and moderately differentiated gastric cancer MGC-803, MKN-45 and SGC-7901 cell lines than in the well-differentiated gastric cancer NCI-N87 cell line, which was consistent with the results for gastric cancer tissues. Expression of miR-7 was downregulated in 86.9% (20/23) of the gastric cancer tissues compared with that in the distant non-tumor tissues. Restoration of miR-7 expression significantly inhibited tumor cell viability, invasiveness and migration when compared with the control cells. Luciferase assay confirmed the epidermal growth factor receptor (EGFR) as a target gene of mR-7, and expression of miR-7 significantly suppressed EGFR expression at both the mRNA and protein levels. The data from the present study demonstrated that reduced miR-7 expression contributes to gastric cancer development and progression. Further study will investigate miR-7 in the regulation of EGFR expression in vitro and in vivo.

RASAL1 influences the proliferation and invasion of gastric cancer cells by regulating the RAS/ERK signaling pathway

The aim of this study was to investigate the biological characteristics of the RASAL1 gene in a well-differentiated gastric cancer cell line MKN-28 and a poorly differentiated gastric cancer cell line BGC-823 cells, using RNA interference and gene transfection technology, respectively. MKN-28 cells were transfected with the shRNA of RASAL1 and BGC-823 cells were transfected with the pcDNA 3.1 plasmid vector containing RASAL1. RT-PCR and western blotting were then used to detect the expression of RASAL1 mRNA and protein. The activities of RAS and extracellular signal-regulated kinase 1/2 were analyzed by the pull-down method and western blotting. The proliferate capacity, apoptosis rate, invasive and migratory potentials of MKN-28 or BGC-823 cells were also measured by Cell Counting Kit-8 cell proliferation assay, propidium iodide/Annexin V staining coupled with flow cytometry, and transwell chamber assays, respectively. Measurement of RASAL1 mRNA and protein expression in two cells revealed successful transfection of the shRNA of RASAL1 and RASAL1-pcDNA3.1 plasmid into these two cells. Moreover, decreased expression of RASAL1 in MKN-28 cells resulted in increased expression of RAS-GTP and p-ERK1/2. Interestingly, decreased expression of RASAL1 inhibited apoptosis and facilitated cell proliferation, invasion and migration. The increased expression of RASAL1 in BGC-823 cells caused declined expression of RAS-GTP and p-ERK1/2, as well as promotedapoptosis and restrained cell proliferation, invasion and migration. The down-regulation of RASAL1 promoted the proliferation, invasion and migration of gastric cancer MKN-28 cells, and up-regulation of RASAL1 inhibited the proliferation, invasion and migration of BGC-823 gastric cancer cells by regulating the RAS/ERK signaling pathway. Thus, our results suggest that RASAL1 may play an important role as a tumor suppressor gene in gastric cancer.

Regulatory mechanism of ZNF139 in multi-drug resistance of gastric cancer cells

Our previous study found increased zinc finger protein 139 (ZNF139) expression in gastric cancer (GC) cells. Purpose of the study is to further clarify the role and mechanism of ZNF139 in multi-drug resistance (MDR) of GC cells. MTT assay, RT-PCR, Western blotting were employed to detect susceptibility of GC cells to chemotherapeutic agents (5-FU, L-OHP) in vitro, and expressions of ZNF139 and MDR associated genes MDR1/P-gp, MRP1, Bcl-2, Bax were also detected. siRNA specific to ZNF139 was transfected into MKN28 cells, then chemosensitivity of GC cells as well as changes of ZNF139 and MDR associated genes were detected. It's found the inhibition rate of 5-FU, L-OHP to well-differentiated GC tissues and cell line was lower than that in the poorly differentiated tissues and cell line; expressions of ZNF139 and MDR1/P-gp, MRP1 and Bcl-2 in well-differentiated GC tissues and cell line MKN28 were higher, while Bax expression was lower. After ZNF139-siRNA was transfected into MKN28, ZNF139 expression in GC cells was inhibited by 90%; inhibition rate of 5-FU, L-OHP to tumor cells increased, and expressions of MDR1/P-gp, MRP1 and Bcl-2 were down-regulated, while Bax was up-regulated. ZNF139 was involved in GC MDR by promoting expressions of MDR1/P-gp, MRP1 and Bcl-2 and inhibiting Bax simultaneously.

Effects of Bcl-2, Bax and P53 Gene Protein Expressions in Gastric Cancer Tissue on the Apoptosis of Cancer Cells

Objective: To investigate the expression of p53 gene in gastric cancer tissue and its correlation with bcl-2 and bax protein expression in relationship and discuss the significance of their correlation in clinic. Methods: Pathological specimens from 100 gastric cancer patients with complete medical data and 24 normal lung tissue specimens were selected. Immunohistochemical streptavidin-biotin-peroxidase assay was used to detect the expression of bcl-2, bax and p53 protein. Results: p53 expression was positively correlated with bcl-2 expression (Pearson's R =0.491, P &lt;0.05). The positive expression of p53 was no significantly correlated with bax expression (P&gt; 0.05). The positive rate of p53 expression in the well-differentiated gastric cancer tissues was 17.6%, the positive rate in the differentiated tissues was 90.9%, the positive rate in the poorly differentiated tissues was 72.7%, and There was a significant difference in the expression of p53 in the three tissues (P &lt;0.05). The expression of p53 in the TNM staging showed that the positive rate in stage Iand IIwas 27.3%, the positive rate in stage III and stage IVwas 89.3%, the difference between them was significant (P &lt;0.05). Conclusions: The expression of p53 gene has an obvious clinical significance for the assessment of degree of malignancy and the prognosis of gastric cancer. Bcl-2 and p53 gene expression could jointly promote the development of gastric cancer in a synergistic way by acting on different stages of apoptosis, or bcl-2s participating in p53-induced mitochondria-mediated apoptosis signaling pathways.

Interaction effects of DNA methylation and histone modification key enzymes in gastric cancer

Objective To study the expression and interactions of DNA methyltransferase 1 （DNMT1）, enhancer of zeste homolog 2 （EZH2） and histone deacetylase 1 （HDAC1） in gastric cancer cell lines and tissue specimens. Methods The expression of DNMT1, EZH2 and HDAC1 was detected at mRNA and protein level in gastric cancer lines MKN28, $GC7901, BGC823, AGS, normal gastric epithelium cell line GES-1 and 10 pairs of fresh gastric cancer tissues and corresponding normal gastric tissues by real-time polymerase chain reaction and Western blot. Whether DNMT1, EZH2 and HDAC1 forming complex or not was detected by co-immunoprecipitation （Co-IP） in well-differentiated gastric cancer cell line MKN28, medium-differentiated gastric cancer cell line SGC7901, low- differentiated gastric cancer cell line BCG823, normal gastric epithelium cell line GESq, medium- differentiated, medium to low-differentiated, low-differentiated gastric cancer tissues and corresponding normal gastric tissues. Results Compared with that of normal gastric epithelium cell and gastric tissue, the expression of DNMT1, EZH2 and HDAC1 in gastric cancer cell lines and gastric tissue was higher. The results of Co-IP indicated that DNMT1, EZH2 and HDAC1 formed complex in the high, medium, and poor differentiated gastric cancer cells and the medium, medium- low, poor differentiated gastric cancer tissues, but not in normal gastric epithelium cell and tissue.Conclusion DNMT1, EZH2 and HDAC1 highly expressed in gastric cancer and there was interaction effects among them, which might be an important mechanism in the correlation between DNA methylation and histone modifications in gastric cancer. Key words: DNA （cytosine-5-）-methyltransferase; Methyltransferases; Histones; Histonedeacetylases; Stomach neoplasms

Correlation of human epidermal growth factor receptor 2 expression with clinicopathological characteristics and prognosis in gastric cancer

To investigate human epidermal growth factor receptor 2 (HER2) gene amplification and protein expression in Chinese patients with resectable gastric cancer and the association with clinicopathological characteristics and survival. One hundred and ninety-seven gastric cancer patients who underwent curative surgery procedures were enrolled into this study. HER2 gene amplification and protein expression were examined using fluorescence in-situ hybridization (FISH) and immunohistochemistry (IHC) analysis on formalin-fixed paraffin-embedded gastric cancer samples from all patients. For scoring, Hofmann's HER2 gastric cancer scoring system was adopted. All cases showing IHC3+ or FISH positivity were defined as HER2 positive. Patient clinicopathological data and survival information were collected. Finally, χ² statistical analysis was performed to analyze the HER2 positivity rate amongst the subgroups with different clinicopathological characteristics including; gender, age, tumor location, Lauren classification, differentiation, TNM staging, depth of invasion, lymph node metastases and distant metastasis. The probability of survival for different subgroups with different clinicopathological characteristics was calculated using the Kaplan-Meier method and survival curves plotted using log rank inspection. According to Hofmann's HER2 gastric cancer scoring criteria, 31 cases (15.74%) were identified as HER2 gene amplified and 19 cases (9.64%) were scored as strongly positive for HER2 membrane staining (3+), 25 cases (12.69%) were moderately positive (2+) and 153 cases (77.66%) were HER2 negative (0/1+). The concordance rate between IHC and FISH analyses was 88.83% (175/197). Thirty-six cases were defined as positive for HER2 gene amplification and/or protein expression, with 24 of these cases being eligible for Herceptin treatment according to United States recommendations, and 29 of these cases eligible according to EU recommendations. Highly consistent results were detected between IHC3+, IHC0/1 and FISH (73.68% and 95.42%), but low consistency was observed between IHC2+ and FISH (40.00%). The positivity rates in intestinal type and well-differentiated gastric cancer were higher than those in diffuse/mixed type and poorly-differentiated gastric cancer respectively (28.57% vs 13.43%, P = 0.0103; 37.25% vs 11.64%, P < 0.0001), but were not correlated with gender, age, tumor location or TNM stage, depth of invasion, lymph node metastases and distant metastasis. In poorly-differentiated gastric cancer patients, those without lymph node metastasis showed a higher HER2 positivity rate than those with lymph node metastasis (26.47% vs 7.14%, P = 0.0021). This association was not present in those patients with well-differentiated gastric cancer (28.57% vs 43.33%, P = 0.2832). Within our patient cohort, 26 cases were lost to follow-up. The median survival time for the remaining 171 patients was 18 mo. The median survival times of the HER2 positive and negative groups were 17 and 18.5 mo respectively. Overall survival was not significantly different between HER2-positive and negative groups (χ(2) = 0.9157, P = 0.3386), but in patients presenting well-differentiated tumors, the overall survival of the HER2-positive group was significantly worse than that of the HER2-negative group (P = 0.0123). In contrast, patients with poorly differentiated and diffuse/mixed subtype gastric cancers showed no significant differences in overall survival associated with HER2. Furthermore, the median survival time of the HER2 positive group did not show any statistically significant differences when compared to the subgroups of gender, age, tumor location, TNM classification, lymph node metastases and distant metastasis. Patients with intestinal type gastric cancer (GC), well-differentiated GC and poorly-differentiated GC without lymph node metastasis, may all represent suitable candidates for targeted therapy using Herceptin.