Transcriptional profiling of Epstein-Barr virus (EBV) genes and host cellular genes in nasal NK/T-cell lymphoma and chronic active EBV infection.
ABSTRACT: Nasal NK/T-cell lymphoma is an aggressive subtype of non-Hodgkin lymphoma (NHL) that is closely associated with Epstein-Barr virus (EBV). The clonal expansion of EBV-infected NK or T cells is also seen in patients with chronic active EBV (CAEBV) infection, suggesting that two diseases might share a partially similar mechanism by which EBV affects host cellular gene expression. To understand the pathogenesis of EBV-associated NK/T-cell lymphoproliferative disorders (LPD) and design new therapies, we employed a novel EBV DNA microarray to compare patterns of EBV expression in six cell lines established from EBV-associated NK/T-cell LPD. We found that expression of BZLF1, which encodes the immediate-early gene product Zta, was expressed in SNK/T cells and the expression levels were preferentially high in cell lines from CAEBV infection. We also analyzed the gene expression patterns of host cellular genes using a human oligonucleotide DNA microarray. We identified a subset of pathogenically and clinically relevant host cellular genes, including TNFRSF10D, CDK2, HSPCA, IL12A as a common molecular biological properties of EBV-associated NK/T-cell LPD and a subset of genes, such as PDCD4 as a putative contributor for disease progression. This study describes a novel approach from the aspects of viral and host gene expression, which could identify novel therapeutic targets in EBV-associated NK/T-cell LPD.
Project description:The contribution of Epstein-Barr virus (EBV) to the development of specific types of benign lymphoproliferations and malignant lymphomas has been extensively studied since the discovery of the virus over the last 50 years. The importance and better understanding of the EBV-associated lymphoproliferative disorders (LPD) of B, T or natural killer (NK) cell type has resulted in the recognition of new entities like EBV+ mucocutaneous ulcer or the addition of chronic active EBV (CAEBV) infection in the revised 2016 World Health Organization (WHO) lymphoma classification. In this article, we review the definitions, morphology, pathogenesis, and evolving concepts of the various EBV-associated disorders including EBV+ diffuse large B-cell lymphoma, not otherwise specified (DLBCL, NOS), EBV+ mucocutaneous ulcer, DLBCL associated with chronic inflammation, fibrin-associated DLBCL, lymphomatoid granulomatosis, the EBV+ T and NK-cell LPD of childhood, aggressive NK leukaemia, extranodal NK/T-cell lymphoma, nasal type, and the new provisional entity of primary EBV+ nodal T- or NK-cell lymphoma. The current knowledge regarding the pathogenesis of B-cell lymphomas that can be EBV-associated including Burkitt lymphoma, plasmablastic lymphoma and classic Hodgkin lymphoma will be also explored.
Project description:Nasal T/NK lymphoma is a unique subtype of non-Hodgkin lymphoma (NHL) that is aggressive and incurable closely associated with Epstein-Barr virus (EBV)3. The clonal expansion of EB infected T- or NK cell is also seen in patients with chronic active EBV (CAEBV) infection, suggesting that two diseases might partly share a similar mechanism by which EBV affect host cellular genes. In order to understand the pathogenesis of EBV-associated T/NK lymphoproliferative disorders (LPD) and design new therapies, we employed a novel EBV DNA microarray to compare patterns of EBV expression in SNK/T cells established from EBV-associated T/NK LPD. Keywords: parallel sample
Project description:Epstein-Barr virus-positive T/NK-cell lymphoproliferative disorders (EBV+ T/NK LPD) encompass a heterogeneous group of disorders, including chronic active Epstein-Barr virus infection (CAEBV), Epstein-Barr virus-associated hemophagocytic lymphohistiocytosis (EBV-HLH), systemic EBV+ T-cell lymphoma of childhood and hydroa vacciniforme-like lymphoproliferative disorder (HVLPD) and so on, predominantly affecting children and young adults with high mortality. Patients with EBV+ T/NK LPD have overlapping clinical symptoms as well as histologic and immunophenotypic features. In this review, we summarized the clinical features of EBV+ T/NK LPD in Chinese patients from the published articles.
Project description:Epstein-Barr virus (EBV) has been detected in several T- and NK-cell neoplasms such as extranodal NK/T-cell lymphoma nasal type, aggressive NK-cell leukemia, EBV-positive peripheral T-cell lymphoma, systemic EBV-positive T-cell lymphoma of childhood, and chronic active EBV infection (CAEBV). However, how this virus contributes to lymphomagenesis in T or NK cells remains largely unknown. Here, we examined NF-?B activation in EBV-positive T or NK cell lines, SNT8, SNT15, SNT16, SNK6, and primary EBV-positive and clonally proliferating T/NK cells obtained from the peripheral blood of patients with CAEBV. Western blotting, electrophoretic mobility shift assays, and immunofluorescent staining revealed persistent NF-?B activation in EBV-infected cell lines and primary cells from patients. Furthermore, we investigated the role of EBV in infected T cells. We performed an in vitro infection assay using MOLT4 cells infected with EBV. The infection directly induced NF-?B activation, promoted survival, and inhibited etoposide-induced apoptosis in MOLT4 cells. The luciferase assay suggested that LMP1 mediated NF-?B activation in MOLT4 cells. IMD-0354, a specific inhibitor of NF-?B that suppresses NF-?B activation in cell lines, inhibited cell survival and induced apoptosis. These results indicate that EBV induces NF-?B-mediated survival signals in T and NK cells, and therefore, may contribute to the lymphomagenesis of these cells.
Project description:Extranodal NK/T-cell, nasal type (hereinafter, nasal T/NK lymphoma), is a distinct clinicopahtologic entity highly associated with EBV. Among nasal T/NK lymphoma, some cases are developed from the long-lasting EBV infection termed chronic active EBV (CAEBV) infection.The clonal expansion of EB infected T- or NK cell are seen in patients with both nasal T/NK lymphoma and CAEBV infection, suggesting that two diseases might partly share the similar mechanism by which EBV affect host cellular genes. Question has thus arisen why a subset of patients with CAEB infection develop into nasal T/NK lymphoma. This study aimed to investigate the virus-host interaction in EBV-associated lymphoma. Keywords = nasak NK/T lymphoma Keywords = chronic active EBV infection Keywords: other
Project description:Epstein-Barr virus (EBV) preferentially infects epithelial cells and B lymphocytes and sometimes T and NK lymphocytes. Persistence of EBV-infected cells results in severe lymphoproliferative disorders (LPDs). Diagnosis of EBV-driven T or NK cell LPD and chronic active EBV diseases (CAEBV) is difficult, often requiring biopsies. Herein, we report a flow-FISH cytometry assay that detects cells expressing EBV-encoded small RNAs (EBERs), allowing rapid identification of EBV-infected cells among PBMCs. EBV-infected B, T, and/or NK cells were detectable in various LPD conditions. Diagnosis of CAEBV in 22 patients of Caucasian and African origins was established. All exhibited circulating EBV-infected T and/or NK cells, highlighting that CAEBV is not restricted to native American and Asian populations. Proportions of EBV-infected cells correlated with blood EBV loads. We showed that EBV-infected T cells had an effector memory activated phenotype, whereas EBV-infected B cells expressed plasma cell differentiation markers. Thus, this method achieves accurate and unambiguous diagnoses of different forms of EBV-driven LPD and represents a powerful tool to study their pathophysiological mechanisms.
Project description:<h4>Background</h4>Chronic active Epstein-Barr virus infection (CAEBV) is defined as Epstein-Barr virus (EBV)-positive T/NK cell-related neoplasia, and its major clinical symptom is systemic inflammation presenting as infectious mononucleocytosis, whereas enteritis and diarrhea are minor clinical symptoms. The complex mixture of tumorigenic processes of EBV-positive cells and physical symptoms of systemic inflammatory disease constitutes the varied phenotypes of CAEBV. Herein, we describe a case of CAEBV that was initially diagnosed as Crohn's disease (CD) based on ileal ulcers and clinical symptoms of enteritis.<h4>Case presentation</h4>A 19-year-old woman complained of abdominal pain and fever. Blood examination showed normal blood cell counts without atypical lymphocyte but detected modest inflammation, hypoalbuminemia, slight liver dysfunction, and evidence of past EBV infection. The esophagogastroduodenoscopic findings were normal. However, colonoscopy revealed a few small ulcers in the terminal ileum. The jejunum and ileum also exhibited various forms of ulcers, exhibiting a cobblestone appearance, on capsule endoscopy. Based on these clinical findings, she was strongly suspected with CD. In the course of treatment by steroid and biologics for refractory enteritis, skin ulcers appeared about 50 months after her initial hospital visit. Immunohistology of her skin biopsy revealed proliferation of EBV-encoded small RNA (EBER)-positive atypical lymphocytes. We retrospectively assessed her previous ileal ulcer biopsy before treatment and found many EBER-positive lymphocytes. Blood EBV DNA was also positive. Therefore, she was diagnosed with extranodal NK/T-cell lymphoma with CAEBV-related enteritis rather than CD. She was treated with cyclosporine and prednisolone combination therapy for CAEBV-related systemic inflammation and chemotherapy for malignant lymphoma. Unfortunately, her disease continued to progress, leading to multiple organ failure and death at the age of 23 years.<h4>Conclusion</h4>Clinicians need to remember the possibility of CAEBV as a differential diagnosis of refractory enteritis. Enteritis with intestinal ulcer is a rare symptom of CAEBV, and it is impossible to acquire a definitive diagnosis by ulcer morphology only. In cases where the possibility of CAEBV remains, tissue EBVR expression should be checked by in situ hybridization and blood EBV DNA.
Project description:Limited studies are available on the molecular pathogenesis of Epstein-Barr virus (EBV)-associated T or natural killer (NK) cell lymphoproliferative disorders (EBV+T/NK-LPD). In this retrospective study, we aim to elucidate the mutation profile of EBV+T/NK-LPD using capture-based targeted sequencing with a panel consisting of 64 lymphoma-related genes to identify driver genes associated with the development of EBV+T/NK-LPD. Targeted sequencing of 169 EBV+T/NK-LPD cases was performed using a panel of 64 lymphoma-related genes. Of the 169 EBV+T/NK-LPD cases, 123 had extra-nodal NK/T-cell lymphoma (ENKTL), 12 had aggressive NK-cell leukemia (ANKL) and 34 had EBV+ T-cell lymphoma of childhood (EBV+TL). The mutation profile revealed that all three subtypes of EBV+T/NK-LPDs had high mutation rates in STAT3, KMT2D, DDX3X, NOTCH1 and TET2. Target sequencing revealed that ENKTL, ANKL and EBV+TL were molecularly distinct, the mutation in nasal-ENKTL and extra-nasal-ENKTL are also different. Survival analysis revealed that ENKTL patients with gene mutations or loss of protein expression in either KMT2D or TET2 were significantly correlated with shorter overall survival. And although the EBV+TL and ANKL groups were too small to confirm survival disadvantage, the adverse prognosis trends of KMT2D or TET2 were showed in these two groups. We conclude that EBV+T/NK lymphoproliferative disorders have very distinct molecular profiles. Our findings also suggest the likely involvement of KMT2D and TET2 in the development of ENKTL, and possibly EBV+T/NK-LPDs in general.
Project description:Epstein-Barr virus (EBV), a ubiquitous B-lymphotropic herpesvirus, ectopically infects T or NK cells to cause severe diseases of unknown pathogenesis, including chronic active EBV infection (CAEBV) and EBV-associated hemophagocytic lymphohistiocytosis (EBV-HLH). We developed xenograft models of CAEBV and EBV-HLH by transplanting patients' PBMC to immunodeficient mice of the NOD/Shi-scid/IL-2R?(null) strain. In these models, EBV-infected T, NK, or B cells proliferated systemically and reproduced histological characteristics of the two diseases. Analysis of the TCR repertoire expression revealed that identical predominant EBV-infected T-cell clones proliferated in patients and corresponding mice transplanted with their PBMC. Expression of the EBV nuclear antigen 1 (EBNA1), the latent membrane protein 1 (LMP1), and LMP2, but not EBNA2, in the engrafted cells is consistent with the latency II program of EBV gene expression known in CAEBV. High levels of human cytokines, including IL-8, IFN-?, and RANTES, were detected in the peripheral blood of the model mice, mirroring hypercytokinemia characteristic to both CAEBV and EBV-HLH. Transplantation of individual immunophenotypic subsets isolated from patients' PBMC as well as that of various combinations of these subsets revealed a critical role of CD4+ T cells in the engraftment of EBV-infected T and NK cells. In accordance with this finding, in vivo depletion of CD4+ T cells by the administration of the OKT4 antibody following transplantation of PBMC prevented the engraftment of EBV-infected T and NK cells. This is the first report of animal models of CAEBV and EBV-HLH that are expected to be useful tools in the development of novel therapeutic strategies for the treatment of the diseases.
Project description:PURPOSE:We report an increased incidence of EBV-induced B-cell lymphoproliferative disease (LPD) in patients treated with siplizumab, an anti-CD2 antibody. The development of EBV-LPD has been associated with the use of immunosuppressive agents used in solid organ, bone marrow, and stem cell transplantation and in certain congenital immunodeficiencies. EXPERIMENTAL DESIGN:We conducted a single-institution phase I dose-escalation trial of siplizumab, a humanized monoclonal antibody to CD2, in 29 patients with T-cell malignancies. RESULTS:Although initial responses were encouraging, 4 (13.7%) patients developed EBV-LPD and the trial was stopped. Reductions in CD4(+) and CD8(+) cell count numbers in response to therapy were seen in all patients, but in those patients developing EBV-LPD a significantly greater reduction in natural killer (NK) cell number and CD2 expression on T cells was seen. These findings highlight the importance of NK-cell depletion and CD2 expression in addition to T-cell depletion in the etiology of EBV-LPD. CONCLUSIONS:The emergence of EBV-LPD may be associated with the ability of siplizumab to deplete both T and NK cells without affecting B cells. Agents that deplete T- and NK-cell populations without affecting B cell number should be screened for this potentially serious adverse event.