Epstein–Barr Virus—Associated Malignancies and Immune Escape: The Role of the Tumor Microenvironment and Tumor Cell Evasion Strategies

The detailed mechanisms of Epstein–Barr virus (EBV) infection in the initiation and progression of EBV-associated malignancies are not yet completely understood. During the last years, new insights into the mechanisms of malignant transformation of EBV-infected cells including somatic mutations and epigenetic modifications, their impact on the microenvironment and resulting unique immune signatures related to immune system functional status and immune escape strategies have been reported. In this context, there exists increasing evidence that EBV-infected tumor cells can influence the tumor microenvironment to their own benefit by establishing an immune-suppressive surrounding. The identified mechanisms include EBV gene integration and latent expression of EBV-infection-triggered cytokines by tumor and/or bystander cells, e.g., cancer-associated fibroblasts with effects on the composition and spatial distribution of the immune cell subpopulations next to the infected cells, stroma constituents and extracellular vesicles. This review summarizes (i) the typical stages of the viral life cycle and EBV-associated transformation, (ii) strategies to detect EBV genome and activity and to differentiate various latency types, (iii) the role of the tumor microenvironment in EBV-associated malignancies, (iv) the different immune escape mechanisms and (v) their clinical relevance. This gained information will enhance the development of therapies against EBV-mediated diseases to improve patient outcome.

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Role of BamHI-A Rightward Frame 1 in Epstein–Barr Virus-Associated Epithelial Malignancies

Biology ◽

10.3390/biology9120461 ◽

2020 ◽

Vol 9 (12) ◽

pp. 461

Author(s):

Rancés Blanco ◽

Francisco Aguayo

Keyword(s):

Epstein Barr Virus ◽

Immune Escape ◽

Tumor Formation ◽

Host Proteins ◽

Barr Virus ◽

Functional Aspects ◽

Infected Cells ◽

Epstein Barr ◽

Apoptotic Effects

Epstein–Barr virus (EBV) infection is associated with a subset of both lymphoid and epithelial malignancies. During the EBV latency program, some viral products involved in the malignant transformation of infected cells are expressed. Among them, the BamHI-A rightward frame 1 (BARF1) is consistently detected in nasopharyngeal carcinomas (NPC) and EBV-associated gastric carcinomas (EBVaGCs) but is practically undetectable in B-cells and lymphomas. Although BARF1 is an early lytic gene, it is expressed during epithelial EBV latency, mainly as a secreted protein (sBARF1). The capacity of sBARF1 to disrupt both innate and adaptive host antiviral immune responses contributes to the immune escape of infected cells. Additionally, BARF1 increases cell proliferation, shows anti-apoptotic effects, and promotes an increased hTERT activity and tumor formation in nude mice cooperating with other host proteins such as c-Myc and H-ras. These facts allow for the consideration of BARF1 as a key protein for promoting EBV-associated epithelial tumors. In this review, we focus on structural and functional aspects of BARF1, such as mechanisms involved in epithelial carcinogenesis and its capacity to modulate the host immune response.

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Infection of Epstein–Barr Virus in Type III Latency Modulates Biogenesis of Exosomes and the Expression Profile of Exosomal miRNAs in the Burkitt Lymphoma Mutu Cell Lines

Cancers ◽

10.3390/cancers10070237 ◽

2018 ◽

Vol 10 (7) ◽

pp. 237 ◽

Cited By ~ 13

Author(s):

Asuka Nanbo ◽

Harutaka Katano ◽

Michiyo Kataoka ◽

Shiho Hoshina ◽

Tsuyoshi Sekizuka ◽

...

Keyword(s):

Epithelial Cells ◽

Burkitt Lymphoma ◽

Epstein Barr Virus ◽

Type I ◽

Type Iii ◽

Barr Virus ◽

Ebv Infection ◽

Latently Infected ◽

Infected Cells ◽

Epstein Barr

Infection of Epstein–Barr virus (EBV), a ubiquitous human gamma herpesvirus, is associated with various malignancies in B lymphocytes and epithelial cells. EBV encodes 49 microRNAs in two separated regions, termed the BART and BHRF1 loci. Although accumulating evidence demonstrates that EBV infection regulates the profile of microRNAs in the cells, little is known about the microRNAs in exosomes released from infected cells. Here, we characterized the expression profile of intracellular and exosomal microRNAs in EBV-negative, and two related EBV-infected Burkitt lymphoma cell lines having type I and type III latency by next-generation sequencing. We found that the biogenesis of exosomes is upregulated in type III latently infected cells compared with EBV-negative and type I latently infected cells. We also observed that viral and several specific host microRNAs were predominantly incorporated in the exosomes released from the cells in type III latency. We confirmed that multiple viral microRNAs were transferred to the epithelial cells cocultured with EBV-infected B cells. Our findings indicate that EBV infection, in particular in type III latency, modulates the biogenesis of exosomes and the profile of exosomal microRNAs, potentially contributing to phenotypic changes in cells receiving these exosomes.

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Direct evidence of abortive lytic infection-mediated establishment of Epstein-Barr virus latency during B-cell infection

10.1101/2020.03.23.004440 ◽

2020 ◽

Author(s):

Tomoki Inagaki ◽

Yoshitaka Sato ◽

Jumpei Ito ◽

Mitsuaki Takaki ◽

Yusuke Okuno ◽

...

Keyword(s):

Viral Infection ◽

Epstein Barr Virus ◽

Lytic Infection ◽

Lytic Gene ◽

Barr Virus ◽

Ebv Infection ◽

Infected Cells ◽

Epstein Barr ◽

Viral Responses ◽

Latent Phase

AbstractViral infection induces dynamic changes in transcriptional profiles. Virus-induced and anti-viral responses are intertwined during the infection. Epstein-Barr virus (EBV) is a human gammaherpesvirus that provides a model of herpesvirus latency. To measure the transcriptome changes during the establishment of EBV latency, EBV-negative Akata cells were infected with EBV-EGFP and observed by transcriptome sequencing (RNA-seq) at 0, 2, 4, 7, 10, and 14 days after infection. We found transient downregulation of mitotic division-related genes, reflecting reprograming of cell growth by EBV. Moreover, a burst of viral lytic gene expression was detected in the early phase of infection. Experimental and mathematical investigations demonstrated that infectious virions were not produced in the pre-latent phase, suggesting the presence of an abortive lytic infection. Finally, we conducted fate mapping using recombinant EBV, enabling the noninvasive, continuous observation of infected cells during EBV infection. Our tracking analysis provided direct evidence that the abortive lytic infection in the pre-latent phase converges to latent infection during EBV infection of B-cells, shedding light on novel roles of viral lytic gene(s) in establishing latency.Author summaryViral infection is a complex process that activates both virus-triggered and host anti-viral responses. This process has classically been studied by snapshot analysis such as microarray and RNA-seq at discrete time points as population averages. Snapshot data lead to invaluable findings in host-pathogen interactions. However, these “snapshot” omics, even from a single cell, lack temporal resolution. Because the behavior of infected cells is highly dynamic and heterogenous, continuous analysis is required for deciphering the fate of infected cells during viral infection. Here, we exploited fate mapping techniques with recombinant Epstein-Barr virus (EBV) to track the infected cells and recorded a log of lytic gene expression during EBV infection. Our continuous observation of infected cells revealed that EBV established latency in B-cells via an abortive lytic infection in the pre-latent phase.

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Oncogenic Properties of the EBV ZEBRA Protein

Cancers ◽

10.3390/cancers12061479 ◽

2020 ◽

Vol 12 (6) ◽

pp. 1479

Author(s):

Diego Germini ◽

Fatimata Bintou Sall ◽

Anna Shmakova ◽

Joëlle Wiels ◽

Svetlana Dokudovskaya ◽

...

Keyword(s):

Epstein Barr Virus ◽

Immune Escape ◽

Immune Surveillance ◽

Host Proteins ◽

Barr Virus ◽

Infected Cells ◽

Functional Features ◽

Epstein Barr ◽

Human Herpesviruses ◽

Transcriptional Transactivator

Epstein Barr Virus (EBV) is one of the most common human herpesviruses. After primary infection, it can persist in the host throughout their lifetime in a latent form, from which it can reactivate following specific stimuli. EBV reactivation is triggered by transcriptional transactivator proteins ZEBRA (also known as Z, EB-1, Zta or BZLF1) and RTA (also known as BRLF1). Here we discuss the structural and functional features of ZEBRA, its role in oncogenesis and its possible implication as a prognostic or diagnostic marker. Modulation of host gene expression by ZEBRA can deregulate the immune surveillance, allow the immune escape, and favor tumor progression. It also interacts with host proteins, thereby modifying their functions. ZEBRA is released into the bloodstream by infected cells and can potentially penetrate any cell through its cell-penetrating domain; therefore, it can also change the fate of non-infected cells. The features of ZEBRA described in this review outline its importance in EBV-related malignancies.

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Epstein–Barr virus and risk of breast cancer: a systematic review and meta-analysis

Future Oncology ◽

10.2217/fon-2019-0232 ◽

2019 ◽

Vol 15 (24) ◽

pp. 2873-2885 ◽

Cited By ~ 5

Author(s):

Mohammad Farahmand ◽

Seyed Hamidreza Monavari ◽

Zabihollah Shoja ◽

Hadi Ghaffari ◽

Mehdi Tavakoli ◽

...

Keyword(s):

Breast Cancer ◽

Epstein Barr Virus ◽

Potential Role ◽

Meta Analysis ◽

Case Control ◽

Case Control Studies ◽

Barr Virus ◽

Ebv Infection ◽

Epstein Barr

Despite the numerous publications regarding the role of Epstein–Barr virus (EBV) in breast cancer development, the topic has still remained controversial. The aim of the meta-analysis was to estimate the overall prevalence of EBV in the breast cancer population, and to investigate the association between EBV and breast cancer risk. The overall prevalence of EBV was calculated 26.37% (95% CI: 22–31%) from the 44 included studies. Meta-analysis of 30 case–control studies showed that the pooled association between EBV and risk of breast cancer is odds ratio 4.74 (95% CI: 2.92–7.69; Z = 6.30; p < 0.0001). Our analyses indicate a strong statistical relationship between EBV infection and risk of breast cancer, suggesting a potential role of EBV infection in the development of breast cancer.

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Role of Epstein-Barr virus in the development of nasopharyngeal carcinoma

Open Medicine ◽

10.1515/med-2017-0025 ◽

2017 ◽

Vol 12 (1) ◽

pp. 171-176 ◽

Cited By ~ 6

Author(s):

Hui Zhang ◽

Jing Wang ◽

Dan Yu ◽

Yan Liu ◽

Kai Xue ◽

...

Keyword(s):

Nasopharyngeal Carcinoma ◽

Epstein Barr Virus ◽

Southern China ◽

Latent Membrane Protein 1 ◽

Barr Virus ◽

Ebv Infection ◽

Micro Rnas ◽

Total Cancer ◽

Epstein Barr

AbstractSouthern China experiences larger extent of total cancer pathologies, of which nasopharyngeal carcinoma has the highest incidence under otorhinolaryngeal malignant carcinomas. Risk factor of nasopharyngeal carcinoma varies from hereditary causes to virus infection, among which Epstein-Barr virus (EBV) infection is the mostly investigated. The study into mechanism of EBV in occurrence, development and prognosis of nasopharyngeal carcinoma has been studied for several decades. The pathophysiology in making of EBV into a cancerogen includes proteins as latent membrane protein 1 (LMPs) and nucleic acids as micro-RNAs. In this paper, we reviewed till date studies focusing on relationship between EBV and nasopharyngeal carcinoma.

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Epstein-Barr Virus Infection Is Not the Sole Cause of High Prevalence for Hodgkin’s Lymphoma in Saudi Arabia.

Blood ◽

10.1182/blood.v104.11.3120.3120 ◽

2004 ◽

Vol 104 (11) ◽

pp. 3120-3120

Author(s):

Khawla S. Al-Kuraya ◽

Rajeswari Narayanappa ◽

Fouad Al-Dayel ◽

Hassan El-Solh ◽

Adnan Ezzat ◽

...

Keyword(s):

Saudi Arabia ◽

Virus Infection ◽

Epstein Barr Virus ◽

Susceptibility Genes ◽

Hodgkin's Lymphoma ◽

Barr Virus ◽

Ebv Infection ◽

High Prevalence ◽

Epstein Barr

Abstract Age adjusted incidence of Hodgkin’s Lymphoma (HL) is markedly higher in Saudi Arabia than in the United States. For example, HL accounts for 10.5% of all neoplasia in children 15 years and younger in Saudi Arabia. EBV virus infection, which can induce HL transformation has been suspected to cause high HL incidence in developing countries. To investigate the role of EBV for the high frequency of HL in Saudi Arabia, we analyzed 265 HL from Saudi Arabia and 58 HL from Europe for EBV infection by in situ hybridization with fluoresce in-conjugated Epstein-Barr virus (EBER) on tissue microarray (TMA) sections. All Saudi and European HL were analyzed in one staining run under identical conditions. Unexpectedly, our data show only minor, statistically insignificant differences in EBV infection rates between Saudi (64 out of 150 informative cases; 42.6%) and European HL (11 out of 30 informative cases; 33%; p=0.5). Within the Arabian population, EBV positivity was more frequent in 79 children (53%) than among 133 adults (36%; p=0.015). EBV positivity was also linked to high stage with EBV positivity in 36% of 69 stage I/II and 64% of 73 stage III/IV tumors (p=0.009). EBV infection was most frequently seen in mixed cellularity HL (63% of 27 cases). This was significantly more frequent than in nodular sclerosing HL (39% of 136; p=0.02). Interestingly, EBV positivity was associated with good prognosis in Saudi childhood HL (p=0.016) but with poor prognosis in Saudi adulthood HL (p=0.0048). In conclusion, our data strongly suggest that EBV is not the main cause for the high prevalence of HL in Middle East countries. Among others, this would be consistent with a major role of genetic susceptibility genes for HL in these populations. Saudi Arabia with high consanguinity and large families would be ideal to search for HL susceptibility genes.

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Role of ATM in the Formation of the Replication Compartment during Lytic Replication of Epstein-Barr Virus in Nasopharyngeal Epithelial Cells

Journal of Virology ◽

10.1128/jvi.01437-14 ◽

2014 ◽

Vol 89 (1) ◽

pp. 652-668 ◽

Cited By ~ 31

Author(s):

Pok Man Hau ◽

Wen Deng ◽

Lin Jia ◽

Jie Yang ◽

Tatsuya Tsurumi ◽

...

Keyword(s):

Epithelial Cells ◽

Viral Replication ◽

Epstein Barr Virus ◽

Viral Dna ◽

Barr Virus ◽

Lytic Reactivation ◽

Infected Cells ◽

Epstein Barr ◽

Replication Compartment

ABSTRACTEpstein-Barr virus (EBV), a type of oncogenic herpesvirus, is associated with human malignancies. Previous studies have shown that lytic reactivation of EBV in latently infected cells induces an ATM-dependent DNA damage response (DDR). The involvement of ATM activation has been implicated in inducing viral lytic gene transcription to promote lytic reactivation. Its contribution to the formation of a replication compartment during lytic reactivation of EBV remains poorly defined. In this study, the role of ATM in viral DNA replication was investigated in EBV-infected nasopharyngeal epithelial cells. We observed that induction of lytic infection of EBV triggers ATM activation and localization of DDR proteins at the viral replication compartments. Suppression of ATM activity using a small interfering RNA (siRNA) approach or a specific chemical inhibitor profoundly suppressed replication of EBV DNA and production of infectious virions in EBV-infected cells induced to undergo lytic reactivation. We further showed that phosphorylation of Sp1 at the serine-101 residue is essential in promoting the accretion of EBV replication proteins at the replication compartment, which is crucial for replication of viral DNA. Knockdown of Sp1 expression by siRNA effectively suppressed the replication of viral DNA and localization of EBV replication proteins to the replication compartments. Our study supports an important role of ATM activation in lytic reactivation of EBV in epithelial cells, and phosphorylation of Sp1 is an essential process downstream of ATM activation involved in the formation of viral replication compartments. Our study revealed an essential role of the ATM-dependent DDR pathway in lytic reactivation of EBV, suggesting a potential antiviral replication strategy using specific DDR inhibitors.IMPORTANCEEpstein-Barr virus (EBV) is closely associated with human malignancies, including undifferentiated nasopharyngeal carcinoma (NPC), which has a high prevalence in southern China. EBV can establish either latent or lytic infection depending on the cellular context of infected host cells. Recent studies have highlighted the importance of the DNA damage response (DDR), a surveillance mechanism that evolves to maintain genome integrity, in regulating lytic EBV replication. However, the underlying molecular events are largely undefined. ATM is consistently activated in EBV-infected epithelial cells when they are induced to undergo lytic reactivation. Suppression of ATM inhibits replication of viral DNA. Furthermore, we observed that phosphorylation of Sp1 at the serine-101 residue, a downstream event of ATM activation, plays an essential role in the formation of viral replication compartments for replication of virus DNA. Our study provides new insights into the mechanism through which EBV utilizes the host cell machinery to promote replication of viral DNA upon lytic reactivation.

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Heterogeneous Epstein-Barr virus infection patterns in peripheral T- cell lymphoma of angioimmunoblastic lymphadenopathy type

Blood ◽

10.1182/blood.v80.7.1804.1804 ◽

1992 ◽

Vol 80 (7) ◽

pp. 1804-1812 ◽

Cited By ~ 142

Author(s):

I Anagnostopoulos ◽

M Hummel ◽

T Finn ◽

M Tiemann ◽

P Korbjuhn ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

B Cells ◽

Epstein Barr Virus ◽

Cell Lymphoma ◽

Angioimmunoblastic Lymphadenopathy ◽

Barr Virus ◽

Ebv Infection ◽

Infected Cells ◽

Epstein Barr

Abstract In this study, 32 cases of T-cell lymphoma of angioimmunoblastic lymphadenopathy type (AILD-TCL) were investigated for their association with Epstein-Barr virus (EBV). For this purpose, three different approaches were applied: polymerase chain reaction (PCR) for the presence of EBV-DNA, in situ hybridization (ISH) for EBV-encoded small nuclear RNAs (EBER), and immunohistology for EBV-encoded latent membrane protein (LMP). PCR and EBER-ISH produced almost identical results, showing that all but one case of AILD-TCL contained EBV genomes. Three distinctive patterns of EBV infection were observed after immunophenotypical characterization of EBER-positive cells: (1) in 26% of the cases, B and T cells were infected, the majority of which were B cells of immunoblastic morphology located in the remnants of lymphoid follicles; (2) in 42% of the cases, the vast majority of infected cells were neoplastic T cells diffusely distributed in the lymph nodes, but infected B cells were also present; and (3) in 32% of the cases, there were only a few infected small lymphoid cells. Detectable LMP was frequent in cases exhibiting patterns 1 and 2. These findings suggest that in AILD-TCL patients, B cells and especially T cells are highly susceptible to a persistent EBV infection, which often leads to a growth advantage of the infected cells. Thus EBV, in conjunction with genetic abnormalities and selective defects of the immune system, might be involved in the pathogenesis of AILD-TCL.

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Morphology, immunophenotype, and distribution of latently and/or productively Epstein-Barr virus-infected cells in acute infectious mononucleosis: implications for the interindividual infection route of Epstein-Barr virus

Blood ◽

10.1182/blood.v85.3.744.bloodjournal853744 ◽

1995 ◽

Vol 85 (3) ◽

pp. 744-750 ◽

Cited By ~ 155

Author(s):

I Anagnostopoulos ◽

M Hummel ◽

C Kreschel ◽

H Stein

Keyword(s):

Epithelial Cells ◽

Infectious Mononucleosis ◽

Epstein Barr Virus ◽

Lymphoid Cells ◽

Productive Infection ◽

Barr Virus ◽

Ebv Infection ◽

Infected Cells ◽

Epstein Barr ◽

Acute Infectious Mononucleosis

The present study was undertaken to unequivocally demonstrate the morphology, immunophenotype, and localization of Epstein Barr virus (EBV)-infected cells as well as the type of infection (latent versus productive) in tonsils of acute infectious mononucleosis. Paraffin sections from nine cases with clinical, serologic, and morphologic evidence of EBV infection were analyzed for the detection of small transcripts, designated EBER1 & 2, and BHLF1 by in situ hybridization (ISH) using nonisotopically labeled probes. ISH was combined with immunohistology, employing a broad panel of antibodies against B-, T-, epithelial-, macrophage-, and follicular dendritic cell (FDC)-antigens. All EBER-positive cells could be identified as lymphocytes, as they did not exhibit any morphologic or immunologic characteristics of epithelial cells, macrophages, or FDCs. A preferential accumulation of EBER-positive cells was noted around crypts, within surface squamous epithelium, and in the surroundings of necrosis. The majority of these lymphocytes could be shown to be B cells, which morphologically included Reed-Sternberg (RS)-like cells, immunoblasts, medium-sized lymphoid cells, as well as cells with plasmacytoid differentiation. In all cases, a varying number of EBER-positive T cells could be identified. ISH for BHLF1-RNA detection showed that almost all cases contained single positive small lymphoid cells, indicating a transition from latent to productive infection cycle. Such cells could also be detected within the crypt epithelium reaching up to its surface. Additional screening of 123 oropharyngeal mucosa samples from patients without evidence of acute EBV-infection, using the polymerase chain reaction for EBV-DNA detection combined with EBER- and BHLF1-ISH showed single latently infected lymphocytes in only one case. Our data imply that infected lymphocytes and not epithelial cells are, in fact, the reservoir for EBV infection, and that these are the cells that participate in the interindividual virus transfer.

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