The chromogenic in situ hybridization (CISH) test is the gold standard for detecting Epstein-Barr virus (EBV)-associated gastric carcinoma (GC). Real-time (RT) PCR method is also a sensitive test that can detect the viral load in samples. As such, three EBV oncogenes were investigated in this study. RNA extraction and cDNA synthesis were performed on GC tissues of nine patients, who were previously confirmed to have EBVGC subtype. In addition, 44 patients that had positive RT-PCR but negative CISH results were also included as the control group. TaqMan RT-PCR analysis was performed to determine the expression of EBV-ENCODED microRNAs, and the expression of EBV-ENCODED dUTPase, as well as LMP2A, was analyzed by SYBR Green RT-PCR. EBV-ENCODED microRNAs and LMP2A were identified in 2 out of 9 (22%) EBVGC subtypes. In addition, EBV-ENCODED dUTPase was detected in 4 out of 9 (44. 5%) EBVGC subtypes. EBV-ENCODED dUTPase was also expressed in a sample of the control group. The expression of LMP2A, EBV-ENCODED microRNAs, and EBV-ENCODED dUTPase viral oncogenes in patients with high EBV viral loads indicates that these expressions correlate with viral loads. Our findings indicate that the EBV-ENCODED dUTPase gene may have a role in EBVGC patients’,non-response to treatment and might be considered a Biomarker-targeted therapy.

Background: Diffuse large B-cell lymphoma (DLBCL) is the most common type of lymphoma. There are various types of DLBCL including immunoblastic and centroblastic. Epstein-Barr virus (EBV) is a member of Herpes virus family found in all human populations inducing different lymphoproliferative disorders. The role of EBV in the development of DLBCL is known. Multiple laboratory methods are available for detecting EBV. This study was conducted to determine the correlation of EBV with DLBCL in samples referred to pathology ward in Shariati and Sina Hospitals by chromogenic in situ hybridization (CISH) method.Methods: In this case/control study, pathological specimens of 50 patients with DLBCL as well as 50 reactive lymph nodes and tonsils (control group) were collected from archives of Shariati and Sina Hospitals and were evaluated for EBV ENCODED RNA ((EBER)) expression based on CISH method. A peptide nucleic acid (PNA) EBV probe (Dakocytomatin) was used while all the processes were done in RNAase-free conditions using RNAase-free water, sterile gloves and samplers.Results: Out of fifty specimens in the case group, eight were positive for (EBER) in comparison with two in the control group (P=0.046). No statistically significant difference was observed between intranodal or extranodal samples (P=0.736) or between males and females (P=0.0746).Conclusion: Our study showed that EBV positivity for (EBER) in patient with DLBCL could be determined more effectively by CISH method than immunohistochemistry (IHC). Comparative analysis between CISH, PCR and IHC methods is recommended.

Background: Gastric cancer is the fourth leading cause of cancer-related deaths in the world. The identification of gastric cancer subtypes related to recognizable microbial agents may play a pivotal role in the targeted prevention and treatment of this cancer. The current study is conducted to define the frequency of Epstein-Barr virus (EBV) infection in gastric cancers of four major provinces, with different incidence rates of gastric cancers, in Iran. Methods: Paraffin blocks of 682 cases of various types of gastric cancer from Tehran, South and North areas of Iran were collected. Twelve tissue microarray (TMA) blocks were constructed from these blocks. Localization of EBV in tumors was assessed by in situ hybridization (ISH) for EBV-ENCODED RNA ((EBER)). Chi-squared test was used to evaluate the statistical significance between EBV-associated gastric cancer (EBVaGC) and clinicopathologic tumor characteristics. Results: Fourteen out of 682 cases (2.1%) of gastric adenocarcinoma were (EBER)-positive. (EBER) was positive in 8 out of 22 (36.4%) of medullary carcinomas and 6 out of 660 (0.9%) of non-medullary type, which was a statistically significant difference (P<0.001). The EBVaGCs were more frequent in younger age (P=0.009) and also showed a trend toward the lower stage of the tumor (P=0.075). Conclusion: EBV-associated gastric adenocarcinoma has a low prevalence in Iran. This finding can be due to epidemiologic differences in risk factors and exposures, and the low number of gastric medullary carcinomas in the population. It may also be related to gastric tumor heterogeneity not detected with the TMA technique.

Background and purpose: Gastric carcinoma is the second leading cause of cancer death worldwide and its incidence varies considerably according to geographical location. There are multiple etiologic factors such as infection with EBV that play a role in carcinogenesis of gastric cancer, yet there is not enough evidence about pathogenesis of EBV and its relationship with gastric cancer. Thus, recognizing pathogenic genes can help improve the prognosis of the patients. The purpose of this study is to explore the relationship between EBV and EBV-ENCODED BARF-1 oncogene in gastric adenocarcinoma pathogenesis.Materials and methods: In this analytical case-control study, 200 paraffin-embedded blocks of biopsies of patients with gastric cancer and those of healthy people were selected. After DNA extraction, polymerase chain reaction (PCR) was used to determine EBV genome and BARF-1 oncogene by specific primers of BARF-1. Grade and stage of the gastric cancer biopsies were also examined.Results: The results of PCR indicated that EBV genome was detected in three cases in the case group, whereas all samples in the control group were negative for EBV. Thus, no significant relationship was observed between the two groups (P>0.05). The EBV positive gastric cancers were negative for BARF-1 oncogene. It was also found that 41% of the cancers were grade I, 52% grade II, and 7% grade III.Conclusion: The findings suggest that gastric cancer has a high prevalence in the geographical region under study; however, EBV plays a minor role in the carcinogenesis of gastric adenocarcinoma.

Diffuse Large B-cell Lymphoma (DLBCL), the most common type of primary central nervous system lymphoma (PCNSL), is a rare aggressive subtype of DLBCL with a poorly understood biology. This study aimed to investigate the prevalence of O-6-Methylguanine-DNA Methyltransferase (MGMT), C-MYC and Epstein-Barr virus ENCODED RNA ((EBER)) positivity in CNS-DLBCLs. Using tissue microarray method, formalin-fixed paraffin-embedded blocks of 76 cases of confirmed PCNS-DLBCL and 2 cases of immunodeficiency-related CNS DLBCL were examined for (EBER) and C-MYC by chromogenic in situ hybridization (CISH), and for MGMT, CD10, BCL2, BCL6, MUM1 and Ki67 by Immunohistochemistry (IHC). The results were analyzed in association with histopathologic and demographic characteristics. The majority of the tumors were of non-germinal center B-cell (non-GCB) type. Loss of MGMT expression on IHC, as a surrogate marker of MGMT methylation, was detected in about 68.9% of PCNSLs. Preserved MGMT expression was found to occur more frequently in males and in MUM1-negative and GCB-type tumors. (EBER) positivity was exclusively seen in immunodeficient cases. Low C-MYC amplification was detected in 18% of cases and showed association with BCL2 and Ki67 expression. We concluded that loss of MGMT expression is a common phenomenon in PCNSLs. Epstein-Barr virus (EBV) may not be commonly detected in PCNS-DLBCL as frequently as in systemic DLBCL, but its expression is inevitable in CNS-DLBCLs of immunocompromised ones. Maintained MGMT expression is associated with less aggressive histopathologic features. Further studies are warranted to confirm the prognostic significance of loss of MGMT expression in PCNSLs and its potential use for predicting therapeutic response to alkylating agents in PCNSLs.

Context: Wiskott-Aldrich syndrome (WAS) is an X-linked primary immunodeficiency characterized by microthrombocytopenia, eczema, recurrent infections, and a significant predisposition to autoimmune diseases and malignancies, particularly B-cell lymphomas. Case presentation: We report the case of a 14-year-old male with a confirmed diagnosis of WAS since infancy, who later developed Crohn’s disease. He presented with vegetative lesions in the rectum and transverse colon, which were diagnosed as diffuse large B-cell lymphoma (DLBCL) based on immunohistochemistry and positive EBV-ENCODED small RNAs ((EBER)). The patient received 12 cycles of chemotherapy, including Rituximab and Etoposide, along with intravenous immunoglobulin (IVIG) support. Conclusion: This is a rare case of EBV-associated DLBCL in a WAS patient from Iran. The case highlights the importance of early and routine malignancy screening in WAS patients, even in the absence of significant symptoms. Molecular and immunophenotypic assessments, including PCR and immunohistochemistry, are essential for accurate diagnosis and management.

Background and Objectives: Gastric cancer (GC) is the third most common cause of cancer-related mortality. Epstein-Barr virus (EBV) is associated with several human tumors. The present research was performed to investigate the prevalence of EBV-associated gastric cancer (EBVaGC) among Iranian patients. Materials and Methods: Seventy cases of gastric cancer and 30 cases of gastric ulcer, all preserved in formalin-fixed paraffin-embedded (FFPE), were examined in a case-control study conducted between 2011 and 2018. The specimens underwent analysis to detect the presence of the EBV genome using a Nested-PCR method targeting EBNA1. Subsequently, samples testing positive for the EBNA1 underwent further testing for the presence of the (EBER) gene using PCR. Finally, Positive samples were subjected to sequencing. Results: Five out of 70 cases (7%) were found to be positive for EBV based on EBNA1 testing, while all EBNA1 positive samples were negative for (EBER). Notably, EBV was not detected in patients with gastric ulcer. The mean age of EBV-positive gastric carcinomas pateints was 64.5 years. Within this group, 60% were male and 40% were female. A higher prevalence of EBV association was observed in diffuse-type cases, with 60% (3 out of 24) testing positive, compared to intestinal-type cases where 40% (2 out of 46) were EBV-positive. Most cases of EBVaGC belonged to grade Ⅰ. Conclusion: This research demonstrates a low prevalence of EBVaGC in Iran. Discrepancies in EBVaGC occurrence among countries could be attributed to epidemiological variables and dietary practices. A comprehensive studies will provide significant contributions to understanding of its etiology.

Background & Objectives: Epstein-Barr virus (EBV) is a tumorigenic herpes virus and belong to DNA viruses that infects and persists in B lymphocytes with in the majority of humans and is also associated with some of cases of Hodgkin lymphoma (HL). The aim of the present study was to investigate the presence of Epstein-Barr virus in Hodgkin's lymphoma in children.Materials & Methods: In this study, 16 formalin-fixed paraffin embedded blocks of Hodgkin's lymphoma tissue samples collected from the archive of pathology departments. The presence of RNA transcripts ENCODED by the virus were examined by (EBER)s in situ hybridization ((EBER)s-ISH).Results: From investigated sixteen formalin-fixed, paraffin-embedded samples of Hodgkin's disease (HD) by (EBER)s in situ hybridization, 12 samples were from male (75%) and 4 samples were from female (25%). Age range of patients was 4 -12 years (with a median of 8 years). In 12 specimens (75%) Epstein-Barr virus was present. The positive cases included 9 male samples (75%) and 3 female samples (75%). Epstein-Barr virus was present in different subtypes of Hodgkin's lymphoma.80% of Mixed Cellularity (MC), 67% of Nodular Sclerosis (NS) and 100% of Lymphocyte Predominance (LP) subtypes were positive. The presence of Epstein-Barr virus in the age groups of 4-7 and 8-12 years were 71.5% and 77.8%, respectively.Conclusion: Results show strong association between Epstein-Barr virus and Hodgkin lymphoma in children. Epstein-Barr virus and Hodgkin lymphoma association in Iranian children show similar pattern with other developing countries.

Nasopharyngeal carcinoma, particulary tumors endemic to the Far East, commonly harbors Epstein-Barr virus. The detection of nuclear antigen associated with EBV and viral DNA in NPC cells have revealed that EBV can infect epithelial cells and is associated with transformation. Human papilloma virus is an epitheliotrophic oncogenic virus that has been detected in a variety of head and neck tumors. This retrospective study was undertaken to investigate the prevalence of EBV and HPV infection subtypes 6/11 and 16/18. in 20 patients with nasopharyngeal carcinoma by in situ hybridization. 16 case (80%) were classified as undifferentiated carcinoma (WHO type III) and 4 (20%) as non keratinizing SCC (WHO type II). According to AJCC (American Joint Committee of Cancer) system the clinical stage at presentation was available for 20 patients: 10% presented with stage I, 5% presented with stage II, 25% presented with stage III and 60% presented with stage IV. 55% of the cases presented with palpable cervical lymph node metastasis. In situ hybridization for (EBER) was performed with a fluorescien-conjugated PNA probe. (EBER) expression was detected in 19 of the 20 evaluated nasopharyngeal carcinoma (95%). Thyramid signal amplification of ISH was performed for HPV DNA subtype 6/11 and 16/18 with a biotinylated DNA probe. Two of 20 NPC (10%) contained HPV 6/11 sequences and two of 20 NPC (10%) contained HPV 16/18 sequences and combined EBV and HPV infection was detected in 3 of the 20 (15%) patients. In this study a high correlation of almost 95% was observed between EBV and NPC type II & III, in WHO classification. Combined EBV and HPV infection was detected only in 15% of patients. There seems to be a geographic difference in positive rates between our results and those obtained from high-risk area.