EBV-associated Gastric Carcinoma Research Articles

Epstein-Barr virus detection in endoscopic submucosal dissection-proven early gastric cancer with mixed-type histology.

Early gastric cancer (EGC) with mixed-type histology is a significant risk factor for additional surgery after endoscopic submucosal dissection (ESD). On the other hand, Epstein-Barr virus-associated gastric cancer (EBVaGC) with mixed-type histology is a favorable risk factor with regard to lymph node metastasis. We performed EBV detection in 13 ESD-proven lesions in 13 cases of early gastric cancer with mixed-type histology using EBV-encoded small RNA1 in situ hybridization (EBER1 ISH). EBVaGC was diagnosed in only one (7.7%) of the tested lesions. That EBVaGC patient underwent surgery and there was no residual lesion and no lymph metastasis. EBVaGC is not frequent in EGC with mixed-type histology. EBV testing of gastric biopsy specimens seems not to be useful to predict the mixed-type histology results of ESD. However, EBV testing for ESD specimens of EGC with mixed-type histology is expected to be useful for avoiding excessive additional surgery.

Analysis of the relationship between the expression of EBV-related antibodies and ET-1 axis in gastric cancer.

EBV-associated gastric cancer (EBVaGC) is a distinct subtype of GC, and EBV plays an important role in tumor progress. The standard method to identify EBV-positive tumor is determined by in situ hybridization for EBV-encoded EBERs in tumor tissues. The present study aims to detect the serological expression of EBV-related antibodies and ET-1 axis to provide a noninvasive method for diagnosis of EBVaGC. The content of EBV-related antibodies and ET-1 axis in preoperative peripheral blood of GC was performed by Chemiluminescence and ELISA assay. The EBV DNA copy number was measured by qRT-PCR. The results showed that the levels of anti-EBV early antigen (EA) IgG, viral capsid antigen (VCA) IgA, nuclear antigen (NA) IgG, and EBV DNA copy number were significantly higher in EBVaGC. The ET-1 axis level was much lower in EBVaGC than EBVnGC. The combined detection of specific anti-EBV antibodies and ET-1 axis might provide new molecular markers for the identification of EBVaGC.

Tumor Molecular and Microenvironment Characteristics in EBV-Associated Malignancies as Potential Therapeutic Targets: Focus on Gastric Cancer.

Although most people are infected with Epstein-Barr Virus (EBV) during their lifetime, only a minority of them develop an EBV-associated malignancy. EBV acts in both direct and indirect ways to transform infected cells into tumor cells. There are multiple ways in which the EBV, host, and tumor environment interact to promote malignant transformation. This paper focuses on some of the mechanisms that EBV uses to transform the tumor microenvironment (TME) of EBV-associated gastric cancer (EBVaGC) for its benefit, including overexpression of Indoleamine 2,3-Dioxygenase 1 (IDO1), synergism between H. pylori and EBV co-infection, and M1 to M2 switch. In this review, we expand on different modalities and combinatorial approaches to therapeutically target this mechanism.

The value of a nomogram based on clinical data and contrast enhanced CT radiomics in the preoperative prediction of Epstein-Barr virus-associated gastric carcinoma

Objective: To explore the value of a nomogram based on clinical data and enhanced CT radiomics in the prediction of Epstein-Barr virus-associated gastric carcinoma(EBVaGC). Methods: The data of 136 patients, including 100 males and 36 females, aged [M (Q1, Q3)] 65 (53, 71) years, with gastric cancer confirmed by surgery and pathology were retrospectively analyzed. According to Epstein-Barr virus-encoded small RNA (EBER) in situ hybridization, those patients were divided into Epstein-Barr virus (EBV) positive group (n=32) and EBV negative group (n=104). All patients underwent multi-phase enhanced CT scanning before surgery and randomly assigned to the training group (n=95) and validation group (n=41) in a ratio of 7︰3. MaZda software was used to extract radiomics features of enhanced CT images. The intra-group correlation coefficient (ICC), variance analysis and minimum absolute shrinkage and selection algorithm (LASSO) regression were used to reduce the dimensionality of the radiomics features, and then the radiomics score (Radscore) was calculated. The nomogram model was based on combined clinical data, morphological features and Radscore. The predictive power of the nomogram was evaluated according to the area under the receiver operating characteristic curve (AUC), and the net clinical benefit of the nomogram was evaluated by the decision curve and calibration curves were drawn according to the data of the training group and the validation group to analyze the consistency of the nomogram model. Results: After selection, six optimal radiomics features were obtained, including Mean, Skewness, S(1, 0) Sum entropy, S(1, 1) Contrast, 99% percentile and S(2, 2)Angular second moment. Radscore of EBV positive group were higher than that of the EBV negative group (training group: 3.78±0.83 vs 2.80±0.98; validation group: 3.81±0.47 vs 2.94±0.95) (both P<0.05) both in the training group and validation group. The AUC of the radiomics model in training group and validation group were 0.773(95%CI:0.612-0.962)and 0.792(95%CI:0.597-0.927)respectively,and the sensitivity and specificity were 63.6% and 93.1%, 70.0% and 87.1%, respectively. The AUC of the nomogram model based on clinical data and radiomics in the training group and the validation group were 0.883(95%CI:0.644-0.984) and 0.851(95%CI:0.715-0.996), respectively. The nomogram model showed superior predictive performance (both P<0.05). Conclusion: The nomogram model based on clinical data and radiomics has better efficacy in the prediction of Epstein-Barr virus associated gastric cancer.

Prediction of Epstein-Barr Virus Status in Gastric Cancer Biopsy Specimens Using a Deep Learning Algorithm

Epstein-Barr virus (EBV)-associated gastric cancer (EBV-GC) is 1 of 4 molecular subtypes of GC and is confirmed by an expensive molecular test, EBV-encoded small RNA in situ hybridization. EBV-GC has 2 histologic characteristics, lymphoid stroma and lace-like tumor pattern, but projecting EBV-GC at biopsy is difficult even for experienced pathologists. To develop and validate a deep learning algorithm to predict EBV status from pathology images of GC biopsy. This diagnostic study developed a deep learning classifier to predict EBV-GC using image patches of tissue microarray (TMA) and whole slide images (WSIs) of GC and applied it to GC biopsy specimens from GCs diagnosed at Kangbuk Samsung Hospital between 2011 and 2020. For a quantitative evaluation and EBV-GC prediction on biopsy specimens, the area of each class and the fraction in total tissue or tumor area were calculated. Data were analyzed from March 5, 2021, to February 10, 2022. Evaluation metrics of predictive model performance were assessed on accuracy, recall, precision, F1 score, area under the receiver operating characteristic curve (AUC), and κ coefficient. This study included 137 184 image patches from 16 TMAs (708 tissue cores), 24 WSIs, and 286 biopsy images of GC. The classifier was able to classify EBV-GC image patches from TMAs and WSIs with 94.70% accuracy, 0.936 recall, 0.938 precision, 0.937 F1 score, and 0.909 κ coefficient. The classifier was used for predicting and measuring the area and fraction of EBV-GC on biopsy tissue specimens. A 10% cutoff value for the predicted fraction of EBV-GC to tissue (EBV-GC/tissue area) produced the best prediction results in EBV-GC biopsy specimens and showed the highest AUC value (0.8723; 95% CI, 0.7560-0.9501). That cutoff also obtained high sensitivity (0.895) and moderate specificity (0.745) compared with experienced pathologist sensitivity (0.842) and specificity (0.854) when using the presence of lymphoid stroma and a lace-like pattern as diagnostic criteria. On prediction maps, EBV-GCs with lace-like pattern and lymphoid stroma showed the same prediction results as EBV-GC, but cases lacking these histologic features revealed heterogeneous prediction results of EBV-GC and non-EBV-GC areas. This study showed the feasibility of EBV-GC prediction using a deep learning algorithm, even in biopsy samples. Use of such an image-based classifier before a confirmatory molecular test will reduce costs and tissue waste.

Clinicopathological Characteristics of Advanced Epstein-Barr Virus-associated Gastric Cancer Highlighting Aberrant p53 Expression.

Epstein-Barr virus (EBV)-associated gastric cancer is a distinct subtype of stomach adenocarcinoma. Although previous studies have investigated its clinicopathological characteristics, there is a lack of research focusing on advanced EBV-associated gastric cancer. In this study, we performed a comprehensive review of advanced EBV-associated gastric cancer cases. We retrospectively collected 18 consecutive cases of surgically resected advanced EBV-associated gastric cancer. Clinicopathological parameters were investigated using histological review, immunohistochemistry, and a review of the electronic medical records of the hospital. The predominant histological pattern of advanced EBV-associated gastric cancer, according to the Laurén classification, was intestinal-type adenocarcinoma with varying degrees of differentiation. However, focal areas showing conventional gastric carcinoma with a lymphoid stromal pattern were found in all cases except one. In addition to the previously described histological patterns of EBV-associated gastric cancer, one case displayed chronic granulomatous inflammation-like histology with barely identifiable malignant epithelial cells. Another case had a pure signet-ring cell carcinoma component showing nuclear positivity for the EBV-encoded RNA in situ hybridization assay. Remarkably, five out of 18 cases (27.8%) showed aberrant p53 expression on immunostaining, which is known to occur rarely in EBV-associated gastric cancer. All cases with aberrant p53 expression had intestinal-type adenocarcinoma-like components. Advanced EBV-associated gastric cancer had distinct histology and a higher rate of aberrant p53 immunostaining pattern than conventional EBV-associated gastric cancer. Therefore, their biological behavior should be investigated separately.

Molecular mechanism of aquaporin 3 (AQP3) regulating by LMP2A and its crosstalk with 4E-BP1 via ERK signaling pathway in EBV-associated gastric cancer

Aquaporin 3(AQP3) is involved in epithelial-mesenchymal transformation of tumor cells and is closely related to the occurrence and development of tumors. However, the regulatory mechanism and function of AQP3 in EBV-associated gastric cancer (EBVaGC) are still poorly understood. This study aims to explore the regulatory effect of EBV on AQP3 and the cross talk of AQP3 with EIF4E-binding proteins 1(4E-BP1) in EBVaGC. The effect of LMP2A on the expression of AQP3 and 4E-BP1 was analyzed using real-time PCR and western blotting. The biological functions of AQP3 and 4E-BP1 in gastric cancer cells were detected by cell biological experiments. In addition, we examined the role of mTOR and ERK signaling pathways in the LMP2A/AQP3/4E-BP1 regulatory axis. We found that LMP2A could down-regulate AQP3 expression by inhibiting the activation of mTOR signaling pathway, and further promote autophagy and migration of gastric cancer cells. AQP3 up-regulated the expression of 4E-BP1 and its phosphorylated protein by activating ERK signaling pathway, thus promoting the autophagy and proliferation of gastric cancer cells. In conclusion, EBV-encoded LMP2A inhibits AQP3 expression, and further participates in cell proliferation, migration and autophagy through the mTOR/AQP3/ERK/4E-BP1 axis.

MC180295 Inhibited Epstein-Barr Virus-Associated Gastric Carcinoma Cell Growth by Suppressing DNA Repair and the Cell Cycle.

DNA methylation of both viral and host DNA is one of the major mechanisms involved in the development of Epstein–Barr virus-associated gastric carcinoma (EBVaGC); thus, epigenetic treatment using demethylating agents would seem to be promising. We have verified the effect of MC180295, which was discovered by screening for demethylating agents. MC180295 inhibited cell growth of the EBVaGC cell lines YCCEL1 and SNU719 in a dose-dependent manner. In a cell cycle analysis, growth arrest and apoptosis were observed in both YCCEL1 and SNU719 cells treated with MC180295. MKN28 cells infected with EBV were sensitive to MC180295 and showed more significant inhibition of cell growth compared to controls without EBV infection. Serial analysis of gene expression analysis showed the expression of genes belonging to the role of BRCA1 in DNA damage response and cell cycle control chromosomal replication to be significantly reduced after MC180295 treatment. We confirmed with quantitative PCR that the expression levels of BRCA2, FANCM, RAD51, TOP2A, and CDC45 were significantly decreased by MC180295. LMP1 and BZLF1 are EBV genes with expression that is epigenetically regulated, and MC180295 could up-regulate their expression. In conclusion, MC180295 inhibited the growth of EBVaGC cells by suppressing DNA repair and the cell cycle.

Short-term outcomes of three rare cases of intramucosal Epstein-Barr virus-associated gastric cancer treated with endoscopic submucosal dissection.

Epstein–Barr virus-associated gastric cancer (EBVaGC) is a subtype of gastric cancer morphologically characterized by massive lymphocyte infiltration. We herein report the short-term outcomes of three rare cases of intramucosal EBVaGC treated with endoscopic submucosal dissection (ESD). Histologically, the lesions exhibited poorly to moderately differentiated tubular adenocarcinoma without lymphovascular invasion, and in situ hybridization revealed EBV-encoded small RNA. Helicobacter pylori infection was not found in any of the lesions. During the 3 to 12 months of follow-up following ESD, none of the ESD-treated patients showed evidence of local recurrence or distant metastases. Essential characteristics of intramucosal EBVaGC may include lymphocyte infiltration into the mucosal stroma or cancer nests as well as the presence of a lace pattern. We believe that ESD might be a safe and suitable treatment method for intramucosal EBVaGC that avoids needless surgery, particularly in patients with severe comorbidities or a high operational risk.

Transcriptome sequencing of LMP2A-transfected gastric cancer cells identifies potential biomarkers in EBV-associated gastric cancer

Epstein-barr virus (EBV) is a well-known human oncogenic virus. However, its molecular mechanisms in the initiation and development of EBV-associated gastric cancer (EBVaGC) remain poorly understood. Latent membrane protein 2A (LMP2A) is an EBV latency-associated protein expressed in part of EBVaGC cases. This study analyzed the effect of LMP2A on the gene expression of gastric cancer cells by transcriptome sequencing on the gastric cancer cell line SGC7901 that expresses LMP2A. The study monitored a total of 238 genes with significant differences in expression, including 101 upregulated genes and 137 downregulated genes. Using the KEGG pathway analysis, it was found that more genes were enriched in the Steroid biosynthesis, Axon guidance, and Terpenoid backbone biosynthesis pathway, and there were 5 genes each enriched in PI3K-Akt and AMPK signaling pathway, all of which were significant. This indicates that LMP2A may be involved in cell biosynthesis, and affects downstream genes and cell biological behavior through AKT and AMPK signaling pathway. Further evaluation confirmed that LMP2A induces ETV5 transcription, but repress GATA6 and NOTCH3 expression. ETV5, GATA6 and NOTCH3 are the candidate targets of LMP2A in gastric cancer.

Clinical and pathologic characteristics of gastric adenocarcinoma associated with Epstein-Barr virus in a region with a high incidence of gastric cancer in Colombia

Introduction and aimsEpstein-Barr virus (EBV) infection is an etiologic factor in EBV-associated gastric carcinoma (EBVaGC). The aim of our study was to describe the clinical and histopathologic characteristics of EBV infection in intestinal-type gastric adenocarcinoma samples. Material and methodsOf 180 paraffin-embedded gastrectomy samples, 28 were studied. Chromogenic in situ hybridization was performed to detect EBV. Sociodemographic and histopathologic data were obtained from the patients’ clinical histories. ResultsA total of 21.4% of the samples were positive for EBV. The predominant morphologic characteristic was the lace pattern, with dense inflammatory infiltration. Fifty percent of the EBVaGC+ patients were men, and the median age of the positive patients was 59 years (range: 50–75); 77.2% of the EBVaGC− patients were men, and the median age of the negative patients was 66 years (range: 34–89). Helicobacter pylori infection was associated with 10.7% of the EBVaGC+ patients and 53.6% of the EBVaGC− patients. In the EBVaGC+ patients, the cardia was the most frequent tumor location (17.9%), 7.1% had histologic grades 2 and 3, and 17.9% presented with Borrmann classification type III. In the EBVaGC− patients, the cardia and fundus were the most frequent tumor locations (71.4%), 35.7% had histologic grade 2, and 39.3% and 21.4% presented with Borrmann classification type III and IV, respectively. ConclusionsThe present study describes the clinical and histopathologic characteristics associated with EBVaGC positivity. Those data may aid in the selection of cases that are candidates for analysis through molecular methods aimed at identifying EBV infection in intestinal-type gastric adenocarcinoma.

Activation of DNA methyltransferase 3a by Epstein-Barr nuclear antigen 1 in gastric carcinoma.

Epstein-Barr nuclear antigen 1 (EBNA1) is expressed in all Epstein-Barr virus (EBV)-infected cells. It interacts with a variety of cellular proteins and activates the transcription of other EBV latency genes, which plays an important role in the persistence of the EBV genome during latent infection. Several studies have shown that EBV infection induces the expression of DNA methyltransferases (DNMTs) and causes extensive methylation of the whole genome in EBV-associated gastric carcinoma (EBVaGC). However, the specific mechanism by which EBV regulates DNMTs expression is still unclear. EBNA1 plasmid and siRNA were transfected to evaluate the effect of EBNA1 on DNMT3a expression. Molecular biology experiments were used to detect the biological function of DNMT3a and its effect on EBV latency in gastric carcinoma cells. We showed that EBNA1 upregulated DNMT3a expression through the E2F1 transcription factor (E2F1) in EBVaGC. DNMT3a knockdown restrained cell proliferation, induced cell cycle arrest, promoted cell apoptosis and suppressed cell migration in vitro. Our results showed a new mechanism for EBV to regulate the expression of DNMT3a. Targeting the EBNA1/E2F1/DNMT3a axis may provide an alternative therapeutic strategy in the treatment of EBVaGC with high DNMT3a expression.

Clinicopathological features of Epstein-Barr virus-associated superficial early stage gastric cancer treated with endoscopic submucosal dissection.

Epstein-Barr virus (EBV) is known to be involved in gastric carcinogenesis. EBV-associated early gastric carcinoma (EBVEGC) has a lower incidence of lymph node involvement and could be an expanded indication for endoscopic submucosal dissection (ESD) treatment. To clarify the prevalence and clinicopathological features of EBVEGC. This study reviewed 618 lesions in 519 patients treated with ESD between 2014 and 2016. Tissue microarray sections were subjected to in situ hybridization staining for EBV-encoded small RNA transcripts (EBER). Lesions positive for EBER were compared with control lesions and were retrospectively analyzed. 12 (1.9%) of the 618 lesions were EBVEGC. EBVEGCs were more frequently located near the atrophic border than control lesions in the middle or upper stomach and were reddish. EBVEGC invasion was deeper and more often histologically undifferentiated. On narrow-band imaging magnifying endoscopy, the EBVEGC group significantly more often showed an endoscopic lace pattern, defined as an absent or obscure microsurface pattern and a microvascular pattern of a tiny, dense, and irregular subepithelial capillary network. The rate of curative resection was significantly lower in the EBVEGC group. Only 1.9% of the ESD specimens were EBV-positive. Endoscopic features could raise clinical suspicion of EBV infection.

Maintenance of Epstein-Barr virus latency through interaction of LMP2A with CXCR4.

Epstein-Barr virus (EBV) belongs to the subfamily Gammaherpesvirinae and was the first human tumor virus to be discovered. The global rate of EBV infection in adults exceeds 90%. EBV can participate in the regulation of multiple genes and signal pathways through its latency genes. Many studies have shown that CXCR4 is involved in the development of gastric cancer, but there have been few studies on the specific mechanisms involved in EBV-associated gastric cancer (EBVaGC). In this study, we explored the mechanism by which EBV-encoded products maintain latent EBV infection through interaction with CXCR4 and investigated the role of CXCR4 in EBV-positive cells. The results show that there is a positive feedback between the EBV-encoded products and CXCR4, and LMP2A can activate CXCR4 through the NF-κB pathway. In addition, CXCR4 can be fed back to LMP2A and EBNA1 through the ERK signaling pathway. At the same time, CXCR4 can promote the proliferation and migration of EBV-positive cells, reduce the expression of the immediate early protein BZLF1, the late protein EBV gp350, and the viral capsid antigen, and play an important role in maintaining the incubation period of EBV infection. These findings are applicable to the further targeted therapy of EBVaGC.

Abstract 2517: Characterization of the immune microenvironment of Epstein-Barr virus associated gastric cancer

Abstract Consistent with better clinical outcome, Epstein-Barr Virus associated gastric cancer (EBVaGC) displays a particular expression pattern of immunological genes and is highly infiltrated by effector immune cells. To decipher potential mechanisms linking tumor infiltrating lymphocytes (TILs) with an expression of immune-related genes in EBVaGC we utilized RNAseq gene expression data from The Cancer Genome Atlas (TCGA) Stomach Adenocarcinoma (STAD) dataset and performed cellular deconvolution analyses by a mixed approach including two complementary approaches (MIXTURE and xCell). In accordance with previous studies, we observed an increase in CD8+ cells and Interferon activity. Further analysis by gene set variation analysis (GSVA) showed an increased exhausted CD8+ signature accompanied by a higher intratumoral Treg signature. Accordingly, an enrichment in both stimulatory and inhibitory immune checkpoint levels on EBVaGC tumors was observed. We next performed differential expression analysis comparing EBVaGC to other STAD subtypes (CIN, GS, HM-SNV, HM-indel) individually. As a result, from this approach, we generated a list of unique EBVaGC-specific genes that was used to construct EBVaGC-specific cell-cell interaction networks. To infer cell-cell interactions underlying the specific tumor immune microenvironment, we implemented the NicheNetR algorithm by including only differentially expressed ligands and targets from the list of EBVaGC genes obtaining a ligand-interaction matrix from which we constructed a EBVaGC-like gene signature. Functional annotation of this ligand-target network showed ligands and targets principally involved in chemotaxis, T-cell apoptosis, and cellular responses to bacterial infection. This signature correlated with better survival not only in EBVaGC but also in other TCGA cohorts such as colon adenocarcinoma and kidney renal clear cell carcinoma. Our findings identified an increase in exhausted CD8+ signature, high intratumoral Treg signature and enrichment in inhibitory immune checkpoint transcripts. We herein propose a new EBVaGC-like gene expression signature that correlates with better survival in multiple cancer types. In-vitro and in-vivo assays will be required for functional validation of these findings as well as clinical trials to confirm the role of our novel EBVaGC-like gene signature in a precision oncology setting. Grant Support: ANID/FONDECYT/POSTDOCTORADO/3201028, CONICYT-FONDAP 15130011, FONDECYT 1191928 &amp; 1180241, IMII P09/016-F, CONICET and Universidad Católica de Córdoba 80020180100029CC, grant 33620180100993CB from the Universidad Nacional de Córdoba. Citation Format: Keila E. Torres, Charlotte N. Hill, Dario Rocha, Elmer Fernández, Ignacio A. Wichmann, Gareth I. Owen, Alejandro H. Corvalan. Characterization of the immune microenvironment of Epstein-Barr virus associated gastric cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2517.

Abstract 3573: DNA immunotherapy targeting BARF1 demonstrates therapeutic impact in novel murine carcinoma models expressing EBV latent antigens

Abstract Epstein Barr Virus (EBV) latent infection is associated with multiple types of cancer. Several clinical studies have targeted EBV antigens as immune therapeutic targets with limited efficacy of EBV+ malignancies, suggesting additional targets might be important. BamHI-A rightward frame 1 (BARF1) is an EBV antigen that is highly expressed in EBV-associated gastric carcinoma (EBVaGC) and EBV+ nasopharyngeal carcinoma (NPC) and has not been evaluated as an antigen in the context of therapeutic immunization. Here, we developed a synthetic DNA-based expression cassette as immunotherapy targeting BARF1 (pBARF1). Immunization with pBARF1 induced potent antigen-specific humoral and polyfunctional effector T cell responses in vivo. We observed significant antigen-specific increases of IFN-γ+, IFN-γ+ TNF-α+, and IFN-γ+ TNF-α+ IL-2+ populations in CD8+ T cells from the pBARF1 immunized animals in both C57BL/6 and BALB/c mice. Novel BARF1 expressing carcinoma lines that grow in immune-competent animals were developed as surrogates for human EBV+ tumors. We confirmed that BARF1 expression in these new cancer cell lines, MC38-BARF1 and CT26-BARF1, was within the range of human EBV+ cancer cell lines, SNU719 and C666-1. Immunization with pBARF1 plasmid demonstrated impact of tumor progression through induction of CD8+ T cell in these BARF1+ carcinoma models. We also observed long-term antitumor immunity, at least up to 446 days, against BARF1 in a CT26-BARF1 rechallenge study. Using IVIS in vivo imaging, we observed that pBARF1 immunized animals rapidly controlled and cleared cancer cells. These studies demonstrate the potential for pBARF1 immunity to induce antigen-specific immune responses impacting tumor progression. Further study of this immune target and these EBV+ tumor models is likely important as part of therapeutic approaches for EBV+ malignancies. Citation Format: Xizhou Zhu, Alfredo Perales-Puchalt, Krzysztof Wojtak, Ziyang Xu, Kun Yun, Pratik S. Bhojnagarwala, Devivasha Bordoloi, Daniel H. Park, Kevin Liaw, Mamadou A. Bah, Paul M. Lieberman, Ebony N. Gary, Ami Patel, David B. Weiner. DNA immunotherapy targeting BARF1 demonstrates therapeutic impact in novel murine carcinoma models expressing EBV latent antigens [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3573.

Rate of Epstein-Barr Virus in Gastric Adenocarcinoma in Egyptian Patients in View of the WHO Classification and Correlation with p16 Immunoreactivity

BACKGROUND AND AIM: Gastric cancer (GC) is one of the top causes of cancer-related deaths worldwide. According to the Cancer Genome Atlas, there are four subtypes of GC, with the Epstein-Barr virus (EBV) subtype accounting for about 10% of cases. EBV infection causes EBV-associated GC (EBVaGC). The previous research suggested that the presence of the EBV viral genome in gastric carcinomas could be used as a surrogate marker for targeted therapy and optimal GC treatment. AIM: We aimed to explore the rate of EBV involvement in gastric carcinogenesis from molecular perspective view and to evaluate the role of the tumor-suppressor protein p16 as a marker for diagnosis in GC Egyptian patients in relation to EBV infection. METHODS: One hundred-four surgically resected GC cases were analyzed. Two methods including quantitative real-time polymerase chain reaction (qPCR) for detecting EBV-derived latent membrane protein-1 (LMP-1) and Epstein-Barr nuclear antigen-1 (EBNA-1) genes as well as immunohistochemistry (IHC) detection of LMP-1 protein and p16 protein on paraffinized tissue blocks were applied. RESULTS: Using IHC, p16 protein was presented in 90/104 (86.5%) of the GC cases, and EBV LMP-1 was detected in 4 cases (3.84%). qPCR detected 14 cases positive for EBV (13.46%). In EBV positive cases detected using qPCR, no expression of p16 was detected. CONCLUSION: EBVaGC has a low incidence in Egypt; loss of p16 expression was recognized in EBVaGC and could be considered as a promising biomarker of EBVaGC. The combination of the two methods IHC and qPCR in addition to p16 is recommended for improving the accuracy of identification of infected cancer.

Characterization of the Prognostic Values of CXCL Family in Epstein-Barr Virus Associated Gastric Cancer.

Background CXCL family is a class of secreted growth factors signaling through G-protein-coupled receptors, and abnormal expression is associated with the growth and progression of many tumors. However, their prognostic value has been poorly studied in Epstein–Barr virus- (EBV-) associated gastric cancer (EBVaGC). Therefore, it is of great significance to explore the prognostic value of the CXCL family in EBVaGC. Methods CXCL family mRNA expression was analyzed in STAD data from The Cancer Genome Atlas (TCGA). Kaplan-Meier Plotter was used to assess the prognostic value of the CXCL family. Transcription factors (TFs) and miRNAs associated with the CXCL family were identified by TFCheckpoint, miRWalk, and ViRBase databases. The prognostic model was evaluated using the EBVaGC patient cohort GSE51575. Results The mRNA expression of CXCL1/3/5/6/8/9/10/11/16 was significantly upregulated, while the expression of CXCL12/14 was downregulated in EBVaGC compared with normal tissues from TCGA-STAD. The mRNA expressions of CXCL9, CXCL10, CXCL11, and CXCL17 in EBVaGCs were higher than those in EBVnGCs, but the mRNA expressions of CXCL6, CXCL12, and CXCL17 were lower than those in EBVnGCs. The mRNA expression levels of CXCL9, CXCL10, and CXCL11 in EBVaGCs were higher than those in EBVnGCs regardless of the tumor stage. High mRNA expression of CXCL8 was associated with better OS in patients with EBVaGC, while high expression of CXCL9 was associated with better OS in patients with EBVnGC. We obtained 10 candidate potential transcription factors (TFs) associated with CXCLs: OTOP3, NKX6-2, NKX2-2, FEV, SMYD1, TRIMSO, TBX10, CDX1, SLC26A3, and ARC. 576 miRNA-mRNA interactions were obtained. Among them, 65 miRNAs were predicted to be correlated with CXCL6, CXCL9, CXCL10, and CXCL11. Similar to the results of TCGA-STAD, the GSE51575 dataset also showed that the mRNA expression levels of CXCL1/3/9/10/11/16 were markedly enhanced in EBVaGC tissues compared with corresponding normal gastric mucosa tissues, while the mRNA expression levels of CXCL12/14 were significantly reduced. The mRNA expression levels of CXCL3/9/10/11/13/17 were increased in EBVaGC compared with EBVnGC tissues. Conclusions The expression differences of CXCL family members are closely associated with the progression of EBVaGC. Expression of CXCL9/10/11/17 mRNA may be a promising prognostic indicator for EBVaGC patients.

The promoter aberrant methylation status of TMEM130 is associated with gastric cancer.

Gastric cancer (GC) is a malignant tumor that seriously affects human health and Epstein-Barr virus (EBV)-associated gastric cancer (EBVaGC) is a molecular subtype of GC. This study aims to determine the relationship between the methylation status of the TMEM130 gene and GC, and to explore the influence of EBV infection. qRT-PCR was conducted to investigate the transcriptional expression of TMEM130 in GC. BSP and MSP assays were used to detect the methylation level of the TMEM130 promoter. The cell migration ability was detected by Transwell and western blot after transfection of TMEM130 plasmids in GC cells. The transcriptional expression of TMEM130 decreased in GC with hypermethylation of the promoter region. The DNA methyltransferase inhibitor could increase the mRNA expression of TMEM130. Moreover, hypermethylation of the TMEM130 promoter in GC tissues was associated with EBV infection. Overexpression of TMEM130 in GC cell lines suppresses cell migration ability. This study was the first to research the expression and function of TMEM130 and found that TMEM130 gene hypermethylation might contribute to GC migration and EBV infection as a cause of hypermethylation of the TMEM130 gene. TMEM130 is a promising biomarker for the diagnosis of GC.

Características clínico-patológicas del adenocarcinoma gástrico asociado al virus de Epstein-Barr en una región de alta incidencia de cáncer gástrico en Colombia

Introduction and aimsEpstein-Barr virus (EBV) infection is an etiologic factor in EBV-associated gastric carcinoma (EBVaGC). The aim of our study was to describe the clinical and histopathologic characteristics of EBV infection in intestinal-type gastric adenocarcinoma samples. Material and methodsOf 180 paraffin-embedded gastrectomy samples, 28 were studied. Chromogenic in situ hybridization was performed to detect EBV. Sociodemographic and histopathologic data were obtained from the patients’ clinical histories. ResultsA total of 21.4% of the samples were positive for EBV. The predominant morphologic characteristic was the lace pattern, with dense inflammatory infiltration. Fifty percent of the EBVaGC + patients were men, and the median age of the positive patients was 59 years (range: 50-75); 77.2% of the EBVaGC– patients were men, and the median age of the negative patients was 66 years (range: 34-89). Helicobacter pylori infection was associated with 10.7% of the EBVaGC + patients and 53.6% of the EBVaGC– patients. In the EBVaGC + patients, the cardia was the most frequent tumor location (17.9%), 7.1% had histologic grades 2 and 3, and 17.9% presented with Borrmann classification type III. In the EBVaGC– patients, the cardia and fundus were the most frequent tumor locations (71.4%), 35.7% had histologic grade 2, and 39.3% and 21.4% presented with Borrmann classification type III and IV, respectively. ConclusionsThe present study describes the clinical and histopathologic characteristics associated with EBVaGC positivity. Those data may aid in the selection of cases that are candidates for analysis through molecular methods aimed at identifying EBV infection in intestinal-type gastric adenocarcinoma.