EBV-related Tumors Research Articles

Epstein-Barr virus and cancer.

EBV was the first human virus to be directly implicated in carcinogenesis. It infects >90% of the world's population. Although most humans coexist with the virus without serious sequelae, a small proportion will develop tumors. Normal host populations can have vastly different susceptibility to EBV-related tumors as demonstrated by geographical and immunological variations in the prevalence of these cancers. EBV has been implicated in the pathogenesis of Burkitt's lymphoma, Hodgkin's disease, non-Hodgkin's lymphoma, nasopharyngeal carcinoma, and lymphomas, as well as leiomyosarcomas arising in immunocompromised individuals. The presence of this virus has also been associated with epithelial malignancies arising in the gastric region and the breast, although some of this work remains in dispute. EBV uses its viral proteins, the actions of which mimic several growth factors, transcription factors, and antiapoptotic factors, to usurp control of the cellular pathways that regulate diverse homeostatic cellular functions. Recent advances in antiviral therapeutics, application of monoclonal antibodies, and generation of EBV-specific CTLs are beginning to show promise in the treatment of EBV-related disorders.

Complex Genomic Rearrangement of ALK Loci Associated with Integrated Human Epstein-Barr Virus in a Post-Transplant Myogenic Liver Tumor

Epstein-Barr virus (EBV) is a ubiquitous viral agent, well known to be associated with lymphoid, epithelial, and smooth-muscle malignancies in immunocompromised individuals. This report describes a 10-year-old patient with an EBV-related liver tumor occurring after kidney transplantation. The neoplasm presented a phenotypic spectrum, ranging from a smooth-muscle tumor to an inflammatory pseudotumor (IPT). The neoplastic cells failed to disclose CD21, CD35, or ALK expression, the latter confirmed by reverse-transcription polymerase chain reaction. Cytogenetic analysis revealed a single clonal cell population showing 46,XY,del (2)(p23),der(3)t (2;3)(p23;q29),der(21) t(Y;21)(q12;p13) karyotype. By metaphase FISH analysis, the neoplastic cells demonstrated the presence of two molecularly different but related aberrant clones, one with the loss of one ALK allele and the second with translocation of the 3'end of ALK kinase domain on the der(3) chromosome. Using FISH with an EBV-specific and 3'end ALK DNA probes, a co-localization of the viral DNA and the ALK sequences was found on the der(3) chromosome. Metaphases with loss of rearranged ALK did not show integrated virus; instead, viral particles together with an associated 3'end ALK domain formed an ex-chromosomal, episomal-like type configuration. The interphase study, using dual-color 5'/3' end ALK FISH assay, revealed 30% of nuclei with only one fused signal, confirming the total loss of one ALK allele in the subset of tumor cells. A combined immunofluorescence and FISH study indicated this separate clonal variant to correspond to desmin-positive smooth-muscle cells. In contrast, desmin-negative myofibroblasts showed the presence of both normal and rearranged ALK alleles. Our results indicate that ALK locus may be a target of EBV integration, a hitherto unreported finding. Although the sustained clonal expansion in EBV-related smooth-muscle tumors/IPTs may depend on functions provided by the EBV oncogenic proteins, the tumor phenotype may be further modified by the secondary genomic rearrangements imposed by the virus during and/or after the integration event. In this respect, the observed phenotypic heterogeneity most likely reflects divergence during neoplastic progression, with the subsequent expansion of morphologically and molecularly distinct but cytogenetically related clones.

Plasma Epstein-Barr virus DNA and residual disease after radiotherapy for undifferentiated nasopharyngeal carcinoma.

Epstein-Barr virus (EBV) DNA can be detected and quantified in the plasma of patients with EBV-related tumors, such as nasopharyngeal carcinoma (NPC). Although NPC at early stages can be cured by radical radiotherapy, there is a high recurrence rate in patients with advanced NPC. The pretreatment level of circulating EBV DNA is a prognostic factor for NPC, but the prognostic value of post-treatment EBV DNA has not been studied. We designed a prospective study in Hong Kong, China, to investigate the value of plasma EBV DNA as a prognostic factor for NPC. One hundred seventy NPC patients, without metastatic disease at presentation, were treated with a uniform radiotherapy protocol. Circulating EBV DNA was measured by real-time quantitative polymerase chain reaction before treatment and 6-8 weeks after radiotherapy was completed. Risk ratios (RRs) were determined with a Cox regression model, and associations of various factors with progression-free and overall survival and recurrence rates were determined with a stepwise Cox proportional hazards model. All statistical tests were two-sided. Ninety-nine percent of patients achieved complete clinical remission. Levels of post-treatment EBV DNA dominated the effect of levels of pretreatment EBV DNA for progression-free survival. The RR for NPC recurrence was 11.9 (95% confidence interval [CI] = 5.53 to 25.43) for patients with higher post-treatment EBV DNA and 2.5 (95% CI = 1.14 to 5.70) for patients with higher pretreatment EBV DNA. Higher levels of post-treatment EBV DNA were statistically significantly associated with overall survival (P<.001; RR for NPC recurrence = 8.6, 95% CI = 3.69 to 19.97). The positive and negative predictive values for NPC recurrence for a higher level of post-treatment EBV DNA were 87% (95% CI = 58% to 98%) and 83% (95% CI = 76% to 89%), respectively. Levels of post-treatment plasma EBV DNA in patients with NPC appear to strongly predict progression-free and overall survival and to accurately reflect the post-treatment residual tumor load.

Use of adenovirus vectors expressing Epstein-Barr virus (EBV) immediate-early protein BZLF1 or BRLF1 to treat EBV-positive tumors.

The Epstein-Barr virus (EBV) genome is present in a variety of tumor types, including virtually all undifferentiated nasopharyngeal carcinomas (NPC) and a portion of gastric carcinomas. The uniform presence of the EBV genome in certain tumors (versus only a very small number of normal B cells) suggests that novel therapies which specifically target EBV-positive cells for destruction might be effective for treating such tumors. Although the great majority of EBV-positive tumor cells are infected with one of the latent forms of EBV infection, expression of either viral immediate-early protein (BZLF1 or BRLF1) is sufficient to convert the virus to the lytic form of infection. Induction of the lytic form of EBV infection could potentially result in death of the tumor cell. Here we have examined the efficacy of adenovirus vectors expressing the BZLF1 or BRLF1 proteins for treatment of EBV-positive epithelial tumors. The BZLF1 and BRLF1 vectors induced preferential killing of EBV-positive, versus EBV-negative, gastric carcinoma cells in vitro. Infection of C18 NPC tumors (grown in nude mice) with either the BZLF1 or BRLF1 vector, but not a control adenovirus vector, induced expression of early lytic EBV genes in tumor cells. Injection of C18 tumors with the BZLF1 or BRLF1 adenovirus vector, but not the control vector, also significantly inhibited growth of the tumors in nude mice. The addition of ganciclovir did not significantly affect the antitumor effect of the BZLF1 and BRLF1 adenovirus vectors. These results suggest a potential cancer therapy against EBV-related tumors.

Association of p53 and BCL-2 expression with Epstein-Barr virus infection in the cancers of head and neck.

Reports that have both evaluated the site-specific Epstein-Barr virus (EBV) infection and compared it with the expression of the EBV-related proto-oncogenes and tumor suppressor genes in the various cancers of head and neck are scarce. Thirty-eight nasopharyngeal carcinoma (NPC) cases, 32 oropharyngeal or hypopharyngeal carcinoma (OPC/HPC) cases, and 93 laryngeal carcinoma (LC) cases were evaluated with in situ hybridization on EBV-encoded small RNA (EBER) and immunohistochemical assessments of the p53, bcl-2, and epidermal growth factor receptor (EGFR) by use of formalin-fixed paraffin-embedded tissue array slides. The expression of viral EBERs was observed in more than two thirds (71.1%) of the NPC cases. In contrast, only 1 case of OPC and none of the HPC or LC cases exhibited EBV positivity. In the nonkeratinizing NPC, the EBV positivity was significantly associated with both frequent p53 overexpression (p =.033) and bcl-2 expression (p =.001). In the EBV-positive nonkeratinizing NPC, a correlation between p53 overexpression and the tumor infiltration lymphocyte (TIL) density was noted (p =.012). A site-specific expression of viral EBER was demonstrated in the head and neck cancers, which suggests an important role for both p53 and bcl-2 in the carcinogenesis of an EBV-infected NPC. The correlation between p53 overexpression and the TIL density in the EBV-infected NPC suggests that the product of a lymphoepithelial interaction, such as A20, can induce a dysfunctional p53 protein.

Association of Epstein-Barr virus with leiomyosarcomas in young people with AIDS.

Children with the acquired immunodeficiency syndrome (AIDS) have an unusually high incidence of smooth-muscle tumors (leiomyomas and leiomyosarcomas) in addition to malignant lymphomas. We tested the hypothesis that the smooth-muscle tumors in these children are associated with the Epstein-Barr virus (EBV). Tissue specimens of five leiomyosarcomas and two leiomyomas from six children with AIDS were studied for evidence of the human immunodeficiency virus (HIV) and EBV by in situ hybridization and quantitative polymerase chain reaction (PCR). Comparison specimens included samples of leiomyosarcoma and leiomyoma from HIV-negative children. EBV clonality of leiomyosarcomas was determined by Southern blot analysis with oligonucleotide probes for EBV terminal-repeat fragments. Tumor specimens were tested by immunoperoxidase staining for infiltration by B lymphocytes and expression of the EBV receptor. Serologic testing for EBV was performed. In situ hybridization showed EBV genomes in all muscle cells of the five leiomyosarcomas and the two leiomyomas from the six HIV-infected children. Quantitative PCR demonstrated strikingly high levels of EBV in tumor tissue, with as many as 4.3 genome copies per cell. Two colonic leiomyosarcomas obtained from different sites at different times from one patient contained different episomal EBV clones, signifying the presence of distinct monoclonal EBV-related tumors. We found biclonal EBV infection in the leiomyosarcoma of another patient. No EBV was detected in normal muscle or tumor specimens from HIV-negative patients. Immunostaining for the EBV receptor was strongly positive in six of the seven leiomyomas and leiomyosarcomas from the patients with AIDS. EBV can infect smooth-muscle cells, at least in patients with AIDS, and it may contribute to the pathogenesis of leiomyomas and leiomyosarcomas in children with AIDS. EBV seems to play no part in smooth-muscle tumors in HIV-negative children.

Efficient Foreign Gene Expression in Epstein-Barr Virus-Transformed Human B-Cells

Epstein-Barr virus (EBV) is a herpesvirus that transforms B-cells (B-LCL) and has undergone intense scrutiny owing to its association with Burkitt's lymphoma, nasopharyngeal carcinoma, and immunoblastic lymphomas. B-LCL have also proven useful in the study of human immunology. We describe a novel system for inducing efficient foreign gene expression in B-LCL using biotinylated adenovirus as an endosome-disrupting agent. Plasmid DNA is coupled to the exterior of viral particles by streptavidin-polylysine chimeric proteins. Up to 67% of B-LCL may be induced to express foreign genes in vitro in transient expression systems, and gene expression lasts for at least 17 days. We have expressed firefly luciferase, β-galactosidase (β-gal), chloramphenicol acetyltransferase, HIV gag, and env genes, as well as infectious HIV, and the EBV-specific BZLF gene in B-LCL with this system. In vivo delivery of a β-gal reporter gene to B-LCL was documented in a SCID mouse model. Potential applications include study of genetic regulation of EBV infection and transformation events, study of potential gene therapies for EBV-related B-cell tumors, and production of antigen-presenting cells for use in immunologic assays. Because of the high percentage of cells transformed and the length of foreign gene expression, the possibility of examining foreign gene expression in transient assays, without selection for clonal populations, exists.

Epstein-Barr virus related gastric cancer in Japan: a molecular patho-epidemiological study.

Epstein-Barr virus (EBV) involvement in gastric carcinoma has been demonstrated by the presence of EBV genomes and EBV-encoded small RNA (EBER) in the carcinoma cells, monoclonal proliferation of EBV-infected carcinoma cells and elevated antibody titers. The present study was conducted to investigate the prevalence of EBV involvement among gastric carcinomas observed in nine Japanese cities with varying gastric cancer rates. In situ hybridization of EBER-1 was applied to paraffin sections from 1848 carcinomas observed in 1795 cases and EBV involvement was detected based on uniform hybridization in carcinoma cells. Epstein-Barr virus was detected in 6.6% of lesions and 6.7% of cases. The rate of EBV involvement did not vary significantly for each city and there was no correlation with underlying gastric cancer mortality rates. Thus, geographic variation of gastric cancer rates within Japan cannot be explained in terms of EBV involvement. Epstein-Barr virus-related gastric carcinoma is one of the most common EBV-related tumors in Japan. The involvement of EBV was significantly more frequent among males than among females, mainly for cancers occurring in the upper and middle part of the stomach, and exhibited more variation by cell type among males. These observations suggest that other factors yet to be discovered may modulate the causal role of EBV in gastric carcinogenesis.