Abstract

This study reports an initial analysis of an EBV-infected B cell line (TCC36B), established from an urothelial carcinoma (UC) lesion of the renal pelvis.

MATERIALS AND METHODS: Cytofluorometric and G-banding analyses were performed for phenotyping and cytogenetics. PCR was used to detect EBV DNA, and sequence analysis to investigate mutations and deletions of the latent membrane protein (LMP)-1 gene of EBV.

RESULTS: TCC36B cells proliferated in vitro and showed positivity for surface CD19, CD20, HLA-DR and IgG(λ), indicating that they belong to B-cells. Cytogenetic analysis showed 46,XX with a unique clonal abnormality of dup(2)(p13p25). EBV DNA was detected in TCC36B cells. Sequence analysis revealed a 30-bp deletion and 7 point mutations on the LMP-1 gene in TCC36B cells.

CONCLUSION: These results suggest the involvement of an EBV variant in the pathogenesis of UC. This cell line should thus facilitate further investigations on the aetiological role of EBV in urothelial cancer.

Publication Types, MeSH Terms, Substances

Abstract

AIMS: To correlate the pre-treatment plasma Epstein-Barr virus (EBV) DNA with tumour burden and to explore the prognostic implications of pre- and post-treatment plasma EBV DNA load in nasopharyngeal carcinoma patients treated with radiotherapy.

MATERIALS AND METHODS: Plasma EBV DNA load was measured using a real-time quantitative polymerase chain reaction assay in 69 patients with nasopharyngeal carcinoma before and after radiation treatment and correlated with tumour volume and treatment outcome. Tumour volume was calculated by multiplying the sum of the areas of gross extent of the primary tumour and regional lymph nodes shown by computed tomography images and/or magnetic resonance imaging. Prognostic models for distant metastasis and overall survival were constructed using a multivariable fractional polynomial algorithm.

RESULTS: The pre-treatment plasma EBV DNA concentration was significantly associated with tumour volume (Spearman correlation coefficient, 0.61; P<0.001). The multivariable fractional polynomial algorithm selected post-treatment EBV DNA and administration of chemotherapy as prognostic factors for distant metastasis (P<0.001, P=0.021, respectively), as well as for overall survival (P<0.001, P=0.018, respectively).

CONCLUSIONS: Pre- and post-treatment plasma EBV DNA load have important clinical significance. Pre-treatment plasma EBV DNA concentration reflects tumour burden, whereas clearance of circulating plasma EBV DNA after treatment predicts the risk of distant metastasis and overall survival.

Copyright © 2010 The Royal College of Radiologists. Published by Elsevier Ltd. All rights reserved.

Abstract

ABSTRACT:

BACKGROUND: To clarify the immunological alterations leading to classical Kaposi sarcoma (cKS) among people infected with KS-associated herpesvirus (KSHV).

METHODS: In a population-based study of 119 cKS cases, 105 KSHV-seropositive controls, and 155 KSHV-seronegative controls, we quantified plasma soluble cluster of differentiation (sCD) levels and antibodies against Epstein-Barr virus nuclear antigen-1 (anti-EBNA-1) and viral capsid antigen (anti-VCA). Differences between groups in prevalence of low-tertile anti-EBNA-1 and high-tertile anti-VCA were compared by logistic regression. Continuous levels between groups and by presence of cKS co-factors among controls were compared by linear regression and Mann-Whitney-Wilcoxon methods.

RESULTS: Comparisons of cKS cases to seropositive controls and of seropositive to seronegative controls revealed no significant differences. However, controls with known cKS cofactors (male sex, nonsmoking, diabetes and cortisone use) had significantly lower levels of anti-EBNA (P = 0.0001 – 0.07) and anti-VCA (P = 0.0001 – 0.03). Levels of sCD26 were significantly lower for male and non-smoking controls (Padj ≤ 0.03), and they were marginally lower with older age and cortisone use (Padj ≤ 0.09).

CONCLUSIONS: Anti-EBV and sCD26 levels were associated with cofactors for cKS, but they did not differ between cKS cases and matched controls. Novel approaches and broader panels of assays are needed to investigate immunological contributions to cKS.

Abstract

This study reports an initial analysis of an EBV-infected B cell line (TCC36B), established from an urothelial carcinoma (UC) lesion of the renal pelvis.

MATERIALS AND METHODS: Cytofluorometric and G-banding analyses were performed for phenotyping and cytogenetics. PCR was used to detect EBV DNA, and sequence analysis to investigate mutations and deletions of the latent membrane protein (LMP)-1 gene of EBV.

RESULTS: TCC36B cells proliferated in vitro and showed positivity for surface CD19, CD20, HLA-DR and IgG(λ), indicating that they belong to B-cells. Cytogenetic analysis showed 46,XX with a unique clonal abnormality of dup(2)(p13p25). EBV DNA was detected in TCC36B cells. Sequence analysis revealed a 30-bp deletion and 7 point mutations on the LMP-1 gene in TCC36B cells.

CONCLUSION: These results suggest the involvement of an EBV variant in the pathogenesis of UC. This cell line should thus facilitate further investigations on the aetiological role of EBV in urothelial cancer.

Publication Types, MeSH Terms, Substances

Abstract

AIMS: To correlate the pre-treatment plasma Epstein-Barr virus (EBV) DNA with tumour burden and to explore the prognostic implications of pre- and post-treatment plasma EBV DNA load in nasopharyngeal carcinoma patients treated with radiotherapy.

MATERIALS AND METHODS: Plasma EBV DNA load was measured using a real-time quantitative polymerase chain reaction assay in 69 patients with nasopharyngeal carcinoma before and after radiation treatment and correlated with tumour volume and treatment outcome. Tumour volume was calculated by multiplying the sum of the areas of gross extent of the primary tumour and regional lymph nodes shown by computed tomography images and/or magnetic resonance imaging. Prognostic models for distant metastasis and overall survival were constructed using a multivariable fractional polynomial algorithm.

RESULTS: The pre-treatment plasma EBV DNA concentration was significantly associated with tumour volume (Spearman correlation coefficient, 0.61; P<0.001). The multivariable fractional polynomial algorithm selected post-treatment EBV DNA and administration of chemotherapy as prognostic factors for distant metastasis (P<0.001, P=0.021, respectively), as well as for overall survival (P<0.001, P=0.018, respectively).

CONCLUSIONS: Pre- and post-treatment plasma EBV DNA load have important clinical significance. Pre-treatment plasma EBV DNA concentration reflects tumour burden, whereas clearance of circulating plasma EBV DNA after treatment predicts the risk of distant metastasis and overall survival.

Copyright © 2010 The Royal College of Radiologists. Published by Elsevier Ltd. All rights reserved.

Abstract

ABSTRACT:

BACKGROUND: To clarify the immunological alterations leading to classical Kaposi sarcoma (cKS) among people infected with KS-associated herpesvirus (KSHV).

METHODS: In a population-based study of 119 cKS cases, 105 KSHV-seropositive controls, and 155 KSHV-seronegative controls, we quantified plasma soluble cluster of differentiation (sCD) levels and antibodies against Epstein-Barr virus nuclear antigen-1 (anti-EBNA-1) and viral capsid antigen (anti-VCA). Differences between groups in prevalence of low-tertile anti-EBNA-1 and high-tertile anti-VCA were compared by logistic regression. Continuous levels between groups and by presence of cKS co-factors among controls were compared by linear regression and Mann-Whitney-Wilcoxon methods.

RESULTS: Comparisons of cKS cases to seropositive controls and of seropositive to seronegative controls revealed no significant differences. However, controls with known cKS cofactors (male sex, nonsmoking, diabetes and cortisone use) had significantly lower levels of anti-EBNA (P = 0.0001 – 0.07) and anti-VCA (P = 0.0001 – 0.03). Levels of sCD26 were significantly lower for male and non-smoking controls (Padj ≤ 0.03), and they were marginally lower with older age and cortisone use (Padj ≤ 0.09).

CONCLUSIONS: Anti-EBV and sCD26 levels were associated with cofactors for cKS, but they did not differ between cKS cases and matched controls. Novel approaches and broader panels of assays are needed to investigate immunological contributions to cKS.

Abstract

Background: Epstein-Barr virus (EBV)-associated gastric carcinomas (GC) constitute a distinct clinicopathological entity of gastric cancer. In order to determine underlying distinct aberrant promoter methylation we tested cardiac and non-cardiac GC with regard to the presence of EBV.Methods: One hundred GC were tested by RNA-in situ hybridization for the presence of EBV by EBV-encoded small RNA (EBER). Aberrant promoter methylation was investigated by methylation-specific real-time PCR for p16, p14, APC and hMLH1. P16 protein expression was assessed by immunohistochemistry.Results: In our selected study cohort, EBER-transcripts were detected in 19.6% (18/92) of GC. EBV-positive GC revealed significantly more often gene hypermethylation of p16, p14 and APC (p<0.0001, p<0.0001 and p=0.02, respectively) than EBV-negative GC. The majority of GC with p16 hypermethylation showed a p16 protein loss (22/28). In contrast, no correlation between the presence of EBV and hMLH1 hypermethylation was found (p=0.7). EBV-positive GC showed a trend towards non-cardiac location (p=0.06) and lower stages (I/II) according to the WHO (p=0.05).Conclusions: Hypermethylation of tumor suppressor genes is significantly more frequent in EBV-associated GC compared to EBV-negative GC. Our data add new insights to the role of EBV in gastric carcinogenesis and underline that EBV associated GC comprise a distinct molecular-pathologic as well as a distinct clinicopathological entity of GC.

Abstract

Anorectal Hodgkin lymphoma (HL) is rare, mainly described in human immunodeficiency virus (HIV) patients with exceptional cases reported in immunocompetents. We report the case of a middle age HIV male, presenting with intestinal occlusion. Rectosigmoidoscopy showed multiple anorectal nodular and ulceronecrotic masses. The biopsy specimens revealed a diffuse polymorphous inflammatory infiltrate in the lamina propria, associated with CD30, CD20, CD3, CD15, and ALK1 scattered large Hodgkin and/or Reed Sternberg -like cells stained by LMP1 antibody and EBER. A diagnosis of EBV-associated atypical lymphoproliferative disease mimicking HL was made. These lesions remained stable for 2 years without treatment then disappeared leaving a mucosal scar. A later control biopsy showed a condylomatous lesion, without lymphoid lesion, suggesting a sexually acquired infection. Eight years later, the complete resolution of the lesion without any treatment is a strong argument against a malignant lymphoid process and raises doubts as to the reality of isolated anorectal HL in immunocompetent participants.

Publication Types, MeSH Terms, Substances

Abstract

AIMS AND BACKGROUND: The mechanisms of Epstein-Barr virus (EBV)-associated tumor development are incompletely understood. The aim of this study was to investigate the gene expression of EBV-associated lymphomas in hu-PBL/SCID mice.

METHODS: Human peripheral blood lymphocytes (hu-PBL) from EBV-seropositive donors were transplanted into severe combined immunodeficiency (SCID) mice. In situ hybridization was used to detect EBV-encoded small RNA- 1 (EBER1) in tumor tissues. Mutation of TP53 exons 5-8 in EBV-induced lymphomas was analyzed by PCR-SSCP. Immunohistochemical staining was used to examine EBV gene products and cellular oncoproteins.

RESULTS: Twenty-one of 29 mice developed tumors. EBER1 was positive in the nuclei of almost all tumor cells. Immunohistochemistry showed positive staining of LMP1, EBNA2 and ZEBRA in a small number of tumor cells. Immunohistochemically detectable p53 protein expression was common (85.7%), but TP53 gene mutations were identified in only four cases (19.1%) of EBV-associated lymphomas. Positivity rates of C-myc, Bcl-2 and Bax expression were 100%, 95.2%, and 90.5%, respectively, in the 21 cases of EBV-associated lymphomas.

CONCLUSIONS: Our preliminary findings suggest that EBV-associated lymphomas in hu-PBL/SCID chimeras show EBV infection, expression of oncogenic viral genes, and overexpression of cellular oncogenes. TP53 gene mutations are rare but p53 protein is commonly expressed in EBV-associated lymphomas.

Publication Types, MeSH Terms, Substances

Abstract

BACKGROUND: Epstein-Barr Virus (EBV)-encoded RNAs (EBERs) are non-polyadenylated RNA molecules transcribed from the EBV genome by RNA polymerase III (pol III). EBERs are the most abundant viral latent gene products, although the precise mechanisms by which EBV is able to achieve such high levels of EBER expression are not fully understood. Previously EBV has been demonstrated to induce transcription factors associated with EBER expression, including pol III transcription factors and ATF-2. We have recently demonstrated that EBV-encoded nuclear antigen-1 (EBNA1) induces cellular transcription factors, and given these findings, we investigated the role of EBNA1 in induction of EBER-associated transcription factors.

RESULTS: Our data confirm that in epithelial cells EBNA1 can enhance cellular pol III transcription. Transient expression of EBNA1 in Ad/AH cells stably expressing the EBERs led to induction of both EBER1 and EBER2 and conversely, expression of a dominant negative EBNA1 led to reduced EBER expression in EBV-infected Ad/AH cells. EBNA1 can induce transcription factors used by EBER genes, including TFIIIC, ATF-2 and c-Myc. A variant chromatin precipitation procedure showed that EBNA1 is associated with the promoters of these genes but not with the promoters of pol III-transcribed genes, including the EBERs themselves. Using shRNA knock-down, we confirm the significance of both ATF-2 and c-Myc in EBER expression. Further, functional induction of a c-Myc fusion protein led to increased EBER expression, providing c-Myc binding sites upstream of EBER1 were intact. In vivo studies confirm elevated levels of the 102 kD subunit of TFIIIC in the tumour cells of EBV-positive nasopharyngeal carcinoma biopsies.

CONCLUSIONS: Our findings reveal that EBNA1 is able to enhance EBER expression through induction of cellular transcription factors and add to the repertoire of EBNA1’s transcription-regulatory properties.

Publication Types, Grant Support

Abstract

Cell immortalization is regarded as an early and pre-requisite step in tumor development. Defining the specific genetic events involved in cell immortalization may provide insights into the early events of carcinogenesis. Nasopharyngeal carcinoma is common among the Southern Chinese population. Epstein-Barr virus (EBV) infection is associated closely with nasopharyngeal carcinoma. The involvement of LMP1 (an EBV-encoded oncogene) has been implicated in the pathogenesis of nasopharyngeal carcinoma. In this study, LMP1 expression, in combination with ectopic expression of hTERT (catalytic unit of human telomerase), was shown to extend the life span of primary cultures of nasopharyngeal epithelial cells and facilitate the immortalization of one of the cell lines (NP446). This is the first report on the successful immortalization of nasopharyngeal epithelial cells involving LMP1. The events associated with the immortalization of nasopharyngeal epithelial cells by LMP1/hTERT were characterized. Expression of c-Myc, Bmi-1, and Id-1 were upregulated at an early stage of immortalization. At a later stage of immortalization, downregulation of p21 and p16 expression were observed. Upregulation of EGFR expression and activation of MAPK signaling pathway were observed in LMP1/hTERT-immortalized nasopharyngeal epithelial cells. The LMP1/hTERT-immortalized NP446 cells were non-tumorigenic in immunosuppressed nude mice and retained anchorage-dependent growth, suggesting that additional events are required for tumorigenic transformation. The ability of the EBV-encoded LMP1, in the presence of hTERT expression, to extend the life span and immortalize primary cultures of nasopharyngeal epithelial cells supports the involvement of EBV infection and its viral products in the early stage of pathogenesis of nasopharyngeal carcinoma.

(c) 2010 Wiley-Liss, Inc.

Publication Types

Abstract

The Epstein-Barr virus (EBV) has an important and multifaceted role in liver pathology. As a member of the herpes virus family, EBV establishes a persistent infection in more than 90% of adults. Besides acute hepatitis during primary infection, many clinical syndromes of interest for the hepatologist are associated with EBV infection. The role of EBV in the evolution of chronic hepatitis from hepatotropic viruses is considered. Chronic EBV-associated hepatitis is suspected in immunocompetent adults with compatible serology, suggestive histology and detection of the viral genome in the liver and/or increase of specific circulating cytotoxic T-lymphocytes. EBV is the main cause of post-transplant lymphoproliferative disorders which occur in up to 30% of cases. EBV-driven lymphoproliferative diseases are also recognized in non-immunocompromised patients and liver is involved in up to a third of the cases. Directly implicated in the pathogenesis of different tumors, EBV has a disputable role in hepatocellular carcinoma carcinogenesis. Further research is required in order to establish or reject the role of EBV in human liver cancer. This paper attempts to discuss the range of EBV-associated chronic liver diseases in immunocompetent patients, from mild, self-limiting mononuclear hepatitis to liver cancer.