EBV-associated Lymphoproliferative Diseases Research Articles

Prevention of Oncogenic Gammaherpesvirinae (EBV and HHV8) Associated Disease in Solid Organ Transplant Recipients.

Long-term risk for malignancy is higher among solid organ transplant (SOT) recipients compared to the general population. Four non-hepatitis viruses have been recognized as oncogenic in SOT recipients-EBV, cause of EBV-associated lymphoproliferative diseases; human herpes virus 8 (HHV8), cause of Kaposi sarcoma, primary effusion lymphoma and multicentric Castleman disease; human papilloma virus, cause of squamous cell skin cancers, and Merkel cell polyomavirus, cause of Merkel cell carcinoma. Two of these viruses (EBV and HHV8) belong to the human herpes virus family. In this review, we will discuss key aspects regarding the clinical presentation, diagnosis, treatment, and prevention of diseases in SOT recipients associated with the two herpesviruses.

Clinical and genetic characterization of Epstein-Barr virus–associated T/NK-cell lymphoproliferative diseases

Epstein-Barr virus (EBV)-associated T-/natural killer (T/NK)-cell lymphoproliferative diseases clinically take on various forms, ranging from an indolent course to an aggressive condition. Clinically, failure to establish precise diagnosis and provide proper treatment makes it difficult to help patients. We sought to better understand the underlying pathogenesis and to identify genetic prognostic factors to achieve better treatment efficacy. In this study, 119 cases of EBV-associated lymphoproliferative diseases, including EBV-associated hemophagocytic lymphohistiocytosis (n= 46) and chronic active EBV disease of T/NK cell type (n= 73), were retrospectively examined. Adults aged >20 years at onset accounted for 71.4% of our cohort. About 54.6% patients with unfavorable overall survival developed hemophagocytic lymphohistiocytosis and had higher plasma EBV load. Allogenic hematopoietic stem-cell transplantation was the sole independent favorable factor. We systematically screened germline and somatic aberrations by whole-exome and targeted sequencing. Among 372 antiviral immunity genes, germline variants of 8 genes were significantly enriched. From a panel of 24 driver genes, somatic mutations were frequently identified in dominant EBV-infected T/NK cells. Patients carrying any germline/somatic aberrations in epigenetic modifiers and RIG-I-like receptor (RLR) pathway had worse overall survival than those without 2 type aberrations. Importantly, patients with IFIH1 and/or DDX3X aberrations in the RLR pathway had higher plasma and NK-cell EBV load. Knockdown of DDX3X in NKYS cells downregulated RLR signaling activities and elevated the expression of EBV-encoded oncogenes such as LMP1 and EBNA1. Genetic defects were prevalent in adult EBV-associated hemophagocytic lymphohistiocytosis patients and patients with chronic active EBV disease of T/NK cell type; these defects were associated with unfavorable prognosis. These findings can help clinicians work out more precise staging of the condition and provide new insights into these EBV-associated diseases.

Impact of Epstein-Barr virus infection in patients with inflammatory bowel disease.

A high prevalence of Epstein-Barr virus (EBV) infection in patients with inflammatory bowel disease (IBD) has been reported in many case reports and studies; thus, the association between EBV and IBD has gained increasing attention. Patients with IBD are at an increased risk of opportunistic EBV infection owing to the common use of immunomodulators. EBV infection in IBD patients can cause various complications, including superimposed viral colitis, which is associated with chronicity, exacerbation, and poor prognosis of refractory IBD, and can induce progression to lymphoproliferative disorders, such as EBV-positive mucocutaneous ulcer (EBVMCU), lymphomatoid granulomatosis (LYG), hemophagocytic lymphohistiocytosis (HLH) and diffuse large B-cell lymphoma (DLBCL). It has been suggested to screen for EBV before initiating immunosuppressive therapy and monitor the status of EBV infection in patients with IBD, especially those who are EBV-seronegative and have a risk of primary EBV infection. Clinicians should also be careful of misdiagnosing IBD and EBV-associated lymphoproliferative diseases due to similarities in both clinical symptoms and endoscopic manifestations. Withdrawal of immunosuppressants has been shown to be an effective strategy to achieve remission of disease at the time of EBV diagnosis, but antiviral therapy remains controversial. The present review aims to describe the characteristics of the complications caused by EBV infection and generalize the recent research progress on and challenges caused by EBV infection in IBD patients. The literature for writing this review was collected from 'PubMed' research engine. The keywords 'inflammatory bowel disease and Epstein-Barr virus' or 'ulcerative colitis and Epstein-Barr virus' or 'Crohn's disease and Epstein-Barr virus' were used to collect the literature and relevant papers were collected to help writing this review.

Acute Epstein‐Barr virus associated haemophagocytosis in an Asian female: What is the diagnosis?

Acute <scp>Epstein‐Barr</scp> virus associated haemophagocytosis in an Asian female: What is the diagnosis?

Characterizing EBV-associated lymphoproliferative diseases and the role of myeloid-derived suppressor cells

AbstractChronic active Epstein-Barr virus (CAEBV) typically presents as persistent infectious mononucleosis-like disease and/or hemophagocytic lymphohistocytosis (HLH), reflecting ectopic Epstein-Barr virus (EBV) infection and lymphoproliferation of T and/or NK cells. Clinical behavior ranges from indolent, stable disease through to rapidly progressive, life-threatening disease. Although it is thought the chronicity and/or progression reflect an escape from immune control, very little is known about the phenotype and function of the infected cells vs coresident noninfected population, nor about the mechanisms that could underpin their evasion of host immune surveillance. To investigate these questions, we developed a multicolor flow cytometry technique combining phenotypic and functional marker staining with in situ hybridization for the EBV-encoded RNAs (EBERs) expressed in every infected cell. This allows the identification, phenotyping, and functional comparison of infected (EBERPOS) and noninfected (EBERNEG) lymphocyte subset(s) in patients' blood samples ex vivo. We have characterized CAEBV and HLH cases with monoclonal populations of discrete EBV-activated T-cell subsets, in some cases accompanied by EBV-activated NK-cell subsets, with longitudinal data on the infected cells' progression despite standard steroid-based therapy. Given that cytotoxic CD8+ T cells with relevant EBV antigen specificity were detectable in the blood of the best studied patient, we searched for means whereby host surveillance might be impaired. This revealed a unique feature in almost every patient with CAEBV studied: the presence of large numbers of myeloid-derived suppressor cells that exhibited robust inhibition of T-cell growth. We suggest that their influence is likely to explain the host's failure to contain EBV-positive T/NK-cell proliferation.

Systemic T-Cell Lymphoproliferative Disease Associated with Epstein-Barr Virus: A Literature Review and a Case Report

Epstein-Barr virus (EBV) is ubiquitous, being identified in 90-95 % of adults. Its reactivation in immunodeficiency conditions often leads to clonal transformation of B-lymphocytes and development of B-cell lymphoproliferative diseases (LPD) and B-cell lymphomas. At the same time, in the countries of North-East and East Asia, as well as Latin America, non-immunocompromised patients sometimes demonstrate the development of EBV-associated T-cell lymphoproliferative diseases. The present paper reports a rare case of EBV-associated systemic T-LPD with lymphadenopathy, splenomegaly as well as acute autoimmune hemolytic anemia in a man of Caucasian race. Complex analysis of anamnestic, pathomorphological, and laboratory data allowed to distinguish this disease from T-cell lymphoma and choose the appropriate patient management strategy.

Reactivation of Epstein-Barr Virus Hepatitis in T/Natural Killer (NK) Cells Mimicking Liver T/NK-Cell Lymphoma

Diagnosis of Epstein-Barr virus (EBV)-associated hepatitis, chronic active EBV infection, and EBV-associated lymphoproliferative diseases, is always challenging due to the overlapping symptoms and lack of diagnostic criteria. We report such a case of a 40-year-old man with unremarkable past medical history. He presented with fever of unknown origin for 1 month with jaundice for 2 days. Physical exams were unremarkable with body temperature at 98.6 °F. His liver function tests were elevated with alanine transaminase (ALT) 559 U/L, aspartate transaminase (AST) 892 U/L, alkaline phosphatase 319 U/L and total bilirubin 4.4 mg/dL. Computed tomography of his chest, abdomen and pelvis did not show lymphadenopathy or hepatosplenomegaly. A liver biopsy showed moderately acute hepatitis with hemophagocytosis, positive Epstein-Barr virus encoding RNA (EBER) in situ hybridization in CD3 and CD4-positive T cells and CD56-positive natural killer (NK) cells. CD20 was negative. The pathology diagnosis was consistent with reactivation of EBV hepatitis but NK-cell lymphoma needs to be excluded. Steatohepatitis with mild activity was present. His blood EBV DNA was 846,000 copies/mL and continued to increase to 2,000,000 copies/mL. Flow cytometric analysis of his bone marrow revealed an increased NK-cell activity but no T/NK-cell lymphoma was identified. Initial treatment with rituxan, etoposide and/or ruxolitinib/acyclovir failed or only had limited effect. However, subsequent valganciclovir greatly improved his conditions. In his 3 months follow-up, the patient was doing well with almost normal liver function tests except mildly elevated ALT (95 U/L) that was due to mild steatohepatitis. EBV DNA PCR was 2,009 copies/mL. To the best of our knowledge, this is the first documented case with reactivation of EBV hepatitis mimicking NK-cell lymphoma in the English literature. With appropriate anti-EBV viral treatments, the patient eventually became asymptomatic and was able to return to his routine life.

EBV-associated lymphoproliferative disorder involving the gastrointestinal tract which mimic IBD in immunocompetent patients: case reports and literature review

IntroductionEpstein-Barr virus (EBV)–associated lymphoproliferative diseases (LPD) with digestive tract involvement in immunocompetent patients is rather rare. Since the symptoms of EBV-associated LPD involving the gastrointestinal tract in immunocompetent patients are similar to those of inflammatory bowel disease (IBD), most patients are initially misdiagnosed.Case presentationIn this paper, we present two cases of EBV-associated T cell LPD involving the colon in immunocompetent patients and review the relevant literature.ConclusionEBV serological testing may help in detecting this disease, and our findings suggest that histopathological evidence of EBV, such as the Epstein-Barr encoding region, is very important to establish the diagnosis.

Systemic Epstein–Barr Virus-Positive T/NK Lymphoproliferative Diseases With SH2D1A/XIAP Hypomorphic Gene Variants

X-linked lymphoproliferative disease (XLP) is one of the X-linked primary immunodeficiency diseases (PIDs) with defective immune response to Epstein–Barr virus (EBV) infection. Chronic active EBV infection (CAEBV) and EBV-hemophagocytic lymphohistiocytosis (HLH) are recognized as systemic EBV-positive T-cell and natural killer (NK)-cell lymphoproliferative diseases (LPDs) arising from the clonal proliferations of EBV-infected T cells and NK cells. A high incidence of CAEBV in East Asia implies the unknown genetic predisposition. In patients with XLP, EBV-infected cells are generally B cells. No mutation of SH2D1A/XIAP genes has ever been identified in patients with systemic EBV-positive T-cell and NK-cell LPD. We report herewith a male case of NK-cell type CAEBV with SH2D1A hypomorphic mutation (c.7G > T, p.Ala3Ser), two male cases of CAEBV/EBV-HLH with XIAP hypomorphic variant (c.1045_1047delGAG, p.Glu349del), and another female case of CD4+CAEBV with the same XIAP variant. The female underwent bone marrow transplantation from an HLA-matched sister with the XIAP variant and obtained a complete donor chimerism and a cure of laryngeal LPD lesion, but then suffered from donor-derived CD4+ T cell EBV-LPD. These observations demonstrated that SH2D1A and XIAP genes are critical for the complete regulation of EBV-positive T/NK cell LPD. X-linked lymphoproliferative disease (XLP) is one of the X-linked primary immunodeficiency diseases (PIDs) reported to have a defective immune response to Epstein–Barr virus (EBV) infection. Mutations in SH2D1A and XIAP genes cause XLP. Systemic EBV-positive T-cell and natural killer (NK)-cell lymphoproliferative diseases (LPDs) consist of three major types: EBV-positive hemophagocytic lymphohistiocytosis (HLH), chronic active EBV infection (CAEBV), and EBV-positive T-cell/NK-cell lymphoma. CAEBV is recognized as a poor prognostic disease of EBV-associated T-cell and NK-cell LPD arising from the clonal proliferation of EBV-infected T cells (CD4+, CD8+, and TCRγδ+) and/or NK cells. The majority of cases with CAEBV were reported from East Asia and South America. In Caucasian patients with CAEBV disease, the target of infection is exclusively B cells. These imply a genetic predisposition to EBV-positive T/NK cell LPD according to ethnicity. In reported cases with XLP, EBV-infected cells are B cells. On the other hand, no mutation of SH2D1A/XIAP genes have been determined in patients with T/NK-cell-type (Asian type) CAEBV. We here describe, for the first time, four case series of CAEBV/EBV-HLH patients who carried the hypomorphic variants of XLP-related genes. These cases included a male patient with CAEBV carrying SH2D1A hypomorphic mutation (c.7G > T, p.Ala3Ser) and two male patients with CAEBV/EBV-HLH carrying the XIAP hypomorphic variant (c.1045_1047delGAG, p.Glu349del), along with another female patient with CAEBV carrying the same XIAP variant. The female case underwent bone marrow transplantation from a healthy HLA-matched sister having the same XIAP variant. Although a complete donor chimerism was achieved with the resolution of laryngeal LPD lesions, systemic donor-derived CD4+ T-cell EBV-LPD developed during the control phase of intractable graft- vs. -host-disease. These observations demonstrated that SH2D1A and XIAP genes are critical for the complete regulation of systemic EBV-positive T/NK-cell LPD.

Epstein Barr Virus Associated B-Cell Lymphomas and Iatrogenic Lymphoproliferative Disorders.

Epstein-Barr virus (EBV) is a ubiquitous herpesvirus, affecting up to 90% of the population. EBV was first identified as an oncogenic virus in a Burkitt lymphoma cell line, though subsequently has been found to drive a variety of malignancies, including diffuse large B-cell lymphoma (DLBCL) and other lymphoma subtypes. EBV has a tropism for B-lymphocytes and has the unique ability to exist in a latent state, evading the host immune response. In cases of impaired cell mediated immunity, as in patients with advanced age or iatrogenic immune suppression, the virus is able to proliferate in an unregulated fashion, expressing viral antigens that predispose to transformation. EBV-positive DLBCL not otherwise specified, which has been included as a revised provisional entity in the 2016 WHO classification of lymphoid malignancies, is thought to commonly occur in older patients with immunosenescence. Similarly, it is well-established that iatrogenic immune suppression, occurring in both transplant and non-transplant settings, can predispose to EBV-driven lymphoproliferative disorders. EBV-positive lymphoproliferative disorders are heterogeneous, with variable clinical features and prognoses depending on the context in which they arise. While DLBCL is the most common subtype, other histologic variants, including Burkitt lymphoma, NK/T-cell lymphoma, and Hodgkin lymphoma can occur. Research aimed at understanding the underlying biology and disease prevention strategies in EBV-associated lymphoproliferative diseases are ongoing. Additionally, personalized treatment approaches, such as immunotherapy and adoptive T-cell therapies, have yielded encouraging results, though randomized trials are needed to further define optimal management.

Viral-Targeted Strategies Against EBV-Associated Lymphoproliferative Diseases.

Epstein-Barr virus (EBV) is strongly associated with a spectrum of EBV-associated lymphoproliferative diseases (EBV-LPDs) ranging from post-transplant lymphoproliferative disorder, B cell lymphomas (e.g., endemic Burkitt lymphoma, Hodgkin lymphoma, and diffuse large B cell lymphoma) to NK or T cell lymphoma (e.g., nasal NK/T-cell lymphoma). The virus expresses a number of latent viral proteins which are able to manipulate cell cycle and cell death processes to promote survival of the tumor cells. Several FDA-approved drugs or novel compounds have been shown to induce killing of some of the EBV-LPDs by inhibiting the function of latent viral proteins or activating the viral lytic cycle from latency. Here, we aim to provide an overview on the mechanisms by which EBV employs to drive the pathogenesis of various EBV-LPDs and to maintain the survival of the tumor cells followed by a discussion on the development of viral-targeted strategies based on the understanding of the patho-mechanisms.

Counteracting survival functions of EBNA3C in Epstein-Barr virus (EBV)-driven lymphoproliferative diseases by combination of SAHA and bortezomib.

Combination of suberoylanilide hydroxamic acid (SAHA) and bortezomib (SAHA/bortezomib) was shown to synergistically induce killing of lymphoblastoid cell lines (LCL) and Burkitt lymphoma (BL) of type III or Wp-restricted latency, both of which express EBNA3A, -3B and -3C proteins. We hypothesize that SAHA/bortezomib can counteract the survival functions conferred by the EBNA3 proteins. We tested the effect of SAHA/bortezomib on the survival of BL cell lines containing EBNA3A, -3B or -3C knockout EBV with or without the respective revertant EBNA3 genes. Isobologram analysis showed that SAHA/bortezomib induced significantly greater synergistic killing of EBNA3C-revertant cells when compared with EBNA3C-knockout cells. Such differential response was not observed in either EBNA3A or -3B revertant versus their knockout pairs. Interestingly, EBNA3C-knockout cells showed significant G2/M arrest whilst EBNA3C-revertant cells and LCLs escaped G2/M arrest induced by SAHA/bortezomib and became more susceptible to the induction of apoptosis. In parallel, SAHA/bortezomib induced stronger expression of p21WAF1 but weaker expression of p-cdc25c, an M-phase inducer phosphatase, in EBNA3C-expressing cells when compared with EBNA3C-knockout cells. SAHA/bortezomib also induced greater growth suppression of EBNA3C-expressing xenografts (EBNA3C-revertant and LCL) than that of EBNA3C-knockout xenografts in SCID mice. In conclusion, our data showed that SAHA/bortezomib could synergistically induce killing of BL and LCL through counteracting the survival functions of EBNA3C, providing a strong basis for clinical testing of this drug combination in patients with EBV-associated lymphoproliferative diseases.

Epstein-Barr Virus Nuclear Antigen Leader Protein Coactivates EP300.

Epstein-Barr virus nuclear antigen (EBNA) leader protein (EBNALP) is one of the first viral genes expressed upon B-cell infection. EBNALP is essential for EBV-mediated B-cell immortalization. EBNALP is thought to function primarily by coactivating EBNA2-mediated transcription. Chromatin immune precipitation followed by deep sequencing (ChIP-seq) studies highlight that EBNALP frequently cooccupies DNA sites with host cell transcription factors (TFs), in particular, EP300, implicating a broader role in transcription regulation. In this study, we investigated the mechanisms of EBNALP transcription coactivation through EP300. EBNALP greatly enhanced EP300 transcription activation when EP300 was tethered to a promoter. EBNALP coimmunoprecipitated endogenous EP300 from lymphoblastoid cell lines (LCLs). EBNALP W repeat serine residues 34, 36, and 63 were required for EP300 association and coactivation. Deletion of the EP300 histone acetyltransferase (HAT) domain greatly reduced EBNALP coactivation and abolished the EBNALP association. An EP300 bromodomain inhibitor also abolished EBNALP coactivation and blocked the EP300 association with EBNALP. EBNALP sites cooccupied by EP300 had significantly higher ChIP-seq signals for sequence-specific TFs, including SPI1, RelA, EBF1, IRF4, BATF, and PAX5. EBNALP- and EP300-cooccurring sites also had much higher H3K4me1 and H3K27ac signals, indicative of activated enhancers. EBNALP-only sites had much higher signals for DNA looping factors, including CTCF and RAD21. EBNALP coactivated reporters under the control of NF-κB or SPI1. EP300 inhibition abolished EBNALP coactivation of these reporters. Clustered regularly interspaced short palindromic repeat interference targeting of EBNALP enhancer sites significantly reduced target gene expression, including that of EP300 itself. These data suggest a previously unrecognized mechanism by which EBNALP coactivates transcription through subverting of EP300 and thus affects the expression of LCL genes regulated by a broad range of host TFs.IMPORTANCE Epstein-Barr virus was the first human DNA tumor virus discovered over 50 years ago. EBV is causally linked to ∼200,000 human malignancies annually. These cancers include endemic Burkitt lymphoma, Hodgkin lymphoma, lymphoma/lymphoproliferative disease in transplant recipients or HIV-infected people, nasopharyngeal carcinoma, and ∼10% of gastric carcinoma cases. EBV-immortalized human B cells faithfully model key aspects of EBV lymphoproliferative diseases and are useful models of EBV oncogenesis. EBNALP is essential for EBV to transform B cells and transcriptionally coactivates EBNA2 by removing repressors from EBNA2-bound DNA sites. Here, we found that EBNALP can also modulate the activity of the key transcription activator EP300, an acetyltransferase that activates a broad range of transcription factors. Our data suggest that EBNALP regulates a much broader range of host genes than was previously appreciated. A small-molecule inhibitor of EP300 abolished EBNALP coactivation of multiple target genes. These findings suggest novel therapeutic approaches to control EBV-associated lymphoproliferative diseases.

EBV-Associated T-Cell Lymphoproliferative Diseases in Young Adults with Unusual Histopathological and Immunophenotypic Features

EBV-associated T-cell lymphoproliferative diseases (EBV T-cell LPDs) in non-immunocompromised hosts are a heterogenous syndrome which is challengeable for both diagnosis and treatment. Here, we report four young patients of EBV T-cell LPDs with unusual histopathological and immunophenotypic features. They presented with intermittent high fever (4/4), multiple lymphadenopathy (4/4), hepatosplenomegaly (4/4), hematochezia (2/4) and high blood EBV antibodies titer (4/4) for seven months to one year. The enteroscopic examination revealed multiple ileocecus ulcers in three of four cases. Histologically, three cases showed similar dense infiltration of polymorphic composition including variable reactive components such as plasma cells and histiocytes as well as atypical lymphocytes and one case was characterized by proliferation of monomorphic atypical lymphocytes. The infiltrating lymphocytes were medium-sized with hyperchromatic nuclei and showed mild cytologic atypia. Abundant eosinophils infiltration and formation of eosinophilic abscess were seen in all cases. Multiple foci of necrosis with granuloma were observed in lymph nodes of all cases, but hemophagocytosis was absent. The immunohistochemical staining showed that infiltrative lymphocytes were CD2 TIA1 CD56﹣, suggesting cytotoxic T-cells origin, but loss of pan-T markers CD3 (2/4), CD5 (4/4) and CD7 (2/4) were frequently observed. Negativity for both CD4 and CD8 (4/4) and silent T-cell receptor (TCR) expression (3/3) were detected. EBV positivity in numerous T-cells was identified by double staining of CD3 and EBER. Three patients died within one year and one patient is alive six months after initial presentation. These unusual pathologic findings prompt us being aware of EBV T-cell LPDs and add to the understanding of this rare disease.

Epstein-Barr Virus (EBV) Reactivation, Its Treatment with Rituximab and Their Impact on Relapse after Allogeneic Hematopoietic Stem Cell Transplantation for Hematological Malignancies

▪Background:Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a potentially curative approach for a wide range of hematologic malignancies. However, the period of prolonged immunodeficiency after this procedure results in significant morbidity and mortality from infectious complications. Epstein-Barr virus (EBV) is a latent γ-herpes virus with B lymphocyte-specific tropism that infects more than 90% of healthy individuals. Following allo-HSCT, EBV reactivation and EBV-related proliferations may be associated with a spectrum of clinical presentations, going from fever to lymphoproliferative diseases (LPD), which arise as a consequence of an outgrowth of B cells latently infected with EBV in the setting of loss or suppression of normal cytotoxic T-cell surveillance. Reduction in immunosuppression and/or pre-emptive therapy with rituximab are strategies nowadays employed to prevent or to treat EBV-associated LPD, which have led to a significant improvement in patient outcome. While the role of T cells and NK cells in graft-versus-leukemia (GVL) immune responses has been established, recent studies have shown a potential contribution of B cells on GVL responses. In this context, the impact of the use of monoclonal antibodies targeting B cells lines on disease control has not been evaluated yet. The aim of this study is to evaluate the incidence of EBV reactivation in patients with hematological malignancies after allo-HSCT, the use of rituximab for its treatment and its impact on the transplantation outcomes.Patients and methods:We evaluated 359 consecutive patients with hematological malignancies who received allo-HSCT and were followed in our center between January 2008 and June 2015; there were 218 (61%) males and 141 (39%) females with a median age of 48 years (range: 18-70), 182 (51%) had acute myeloid leukemia, 44 (12%) multiple myeloma, 34 (9%) myelodysplastic syndrome, 30 (8%) Non-Hodgkin lymphoma, 7 (2%) chronic lymphocytic leukemia, 21 (6%) myeloproliferative syndrome, 14 (4%) Hodgkin disease, 13 (4%) chronic myeloid leukemia and 14 (4%) aplastic anemia. At transplantation, 227 (63%) patients were in complete response (CR) or chronic phase (CP) and 132 (37%) were in less than CR or CP. For conditioning regimen, 171 (48%) were myeloablative and 188 (52%) were reduced intensity. EBV DNA levels in blood were detected by quantitative real-time polymerase chain reaction (RQ-PCR) after weekly monitoring up to 3 months after allo-HSCT. EBV-DNA was considered positive when the copies exceeded 1000 copies/ml. EBV therapeutic intervention, firstly concerned gradual reduction of the doses of immunosuppressive drugs, and then rituximab infusion (375 mg/m², four injections weekly) was administered when the copies exceeded 10 000 copies/ml.Results:Among 359 patients, 222 patients had EBV reactivation after a median time of 1.3 months (0.7-2.5) after allo-HSCT with a cumulative incidence of 48 % (47-50) at 3 months. Among the 222 patients with EBV reactivation, only 35 (15.7%) needed treatment with rituximab. Rituximab was introduced after a median time of 55 days after transplantation with a median number of 5 infusions and a median dose of 2025 mg/m². EBV treatment was successful in all patients, none progressed to LPD. The cumulative incidence of relapse at one and two years for the whole population was 27% (26-28) and 34% (33-35) respectively and the cumulative incidence of transplant-related mortality (TRM) was 22% (21-23) and 25% (24-26) respectively. The multivariate analysis taking into account the type of disease, the type of conditioning, the use of ATG, the disease status at transplantation, the presence of GVHD and the EBV reactivation with/without rituximab, showed that the presence of EBV reactivation was associated with a significant lower relapse rate, HR= 0.52 [0.35-1], p=0.05. The use of rituximab did not compromise the GVL effect and thus had not impact on relapse or overall survival.Conclusion:We showed a positive impact of EBV reactivation on relapse incidence which could be explained as a stimulation of both function and amplification of NK compartment. The strict monitoring of the viral load and therapeutic intervention using rituximab enable a full viral control without any progression into LPD and without any compromise on GVL effect. [Display omitted] DisclosuresMichallet:Bristol-Myers Squibb: Consultancy, Honoraria, Research Funding; Pfizer: Consultancy, Honoraria; Novartis: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; Astellas Pharma: Consultancy, Honoraria; MSD: Consultancy, Honoraria; Genzyme: Consultancy, Honoraria. Nicolini:Ariad: Honoraria, Membership on an entity's Board of Directors or advisory committees; BMS: Consultancy, Honoraria; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding.

Editorial: Memory T Cells: Effectors, Regulators, and Implications for Transplant Tolerance.

Editorial: Memory T Cells: Effectors, Regulators, and Implications for Transplant Tolerance.

Measurement of CD8+ and CD4+ T Cell Frequencies Specific for EBV LMP1 and LMP2a Using mRNA-Transfected DCs.

An EBV-specific cellular immune response is associated with the control of EBV-associated malignancies and lymphoproliferative diseases, some of which have been successfully treated by adoptive T cell therapy. Therefore, many methods have been used to measure EBV-specific cellular immune responses. Previous studies have mainly used autologous EBV-transformed B-lymphoblastoid cell lines (B-LCLs), recombinant viral vectors transfected or peptide pulsed dendritic cells (DCs) as stimulators of CD8+ and CD4+ T lymphocytes. In the present study, we used an interferon-γ (IFN-γ) enzyme-linked immunospot (ELISPOT) assay by using isolated CD8+ and CD4+ T cells stimulated with mRNA-transfected DCs. The frequency of latent membrane protein 1 (LMP1)-specific IFN-γ producing CD4+ T cells was significantly higher than that of LMP2a. The frequency of IFN-γ producing CD4+ T cells was significantly correlated with that of CD8+ T cells in LMP1-specific immune responses (r = 0.7187, Pc < 0.0001). To determine whether there were changes in LMP1- or LMP2a-specific immune responses, subsequent peripheral blood mononuclear cells (PBMCs) samples were analyzed. Significant changes were observed in 5 of the 10 donors examined, and CD4+ T cell responses showed more significant changes than CD8+ T cell responses. CD8+ and CD4+ T cells from EBV-seropositive donors secreted only the Th1 cytokines IFN-γ, TNF-α, and IL-2, while Th2 (IL-4) and Th17 (IL-17a) cytokines were not detected. CD4+ T cells secreted significantly higher cytokine levels than did CD8+ T cells. Analysis of EBV-specific T cell responses using autologous DCs transfected with mRNA might provide a comprehensive tool for monitoring EBV infection and new insights into the pathogenesis of EBV-associated diseases.

Prevention of EBV lymphoma development by oncolytic myxoma virus in a murine xenograft model of post-transplant lymphoproliferative disease

Epstein–Barr virus (EBV) has been associated with a variety of epithelial and hematologic malignancies, including B-, T- and NK cell-lymphomas, Hodgkin's disease (HD), post-transplant lymphoproliferative diseases (LPDs), nasopharyngeal and gastric carcinomas, smooth muscle tumors, and HIV-associated lymphomas. Currently, treatment options for EBV-associated malignancies are limited. We have previously shown that myxoma virus specifically targets various human solid tumors and leukemia cells in a variety of animal models, while sparing normal human or murine tissues. Since transplant recipients of bone marrow or solid organs often develop EBV-associated post-transplant LPDs and lymphoma, myxoma virus may be of utility to prevent EBV-associated malignancies in immunocompromised transplant patients where treatment options are frequently limited. In this report, we demonstrate the safety and efficacy of myxoma virus purging as a prophylactic strategy for preventing post-transplant EBV-transformed human lymphomas, using a highly immunosuppressed mouse xenotransplantation model. This provides support for developing myxoma virus as a potential oncolytic therapy for preventing EBV-associated LPDs following transplantation of bone marrow or solid organ allografts.

The Latent Membrane Protein 1 (LMP1).

Almost exactly twenty years after the discovery of Epstein-Barr virus (EBV), the latent membrane protein 1 (LMP1) entered the EBV stage, and soon thereafter, it was recognized as the primary transforming gene product of the virus. LMP1 is expressed in most EBV-associated lymphoproliferative diseases and malignancies, and it critically contributes to pathogenesis and disease phenotypes. Thirty years of LMP1 research revealed its high potential as a deregulator of cellular signal transduction pathways leading to target cell proliferation and the simultaneous subversion of cell death programs. However, LMP1 has multiple roles beyond cell transformation and immortalization, ranging from cytokine and chemokine induction, immune modulation, the global alteration of gene and microRNA expression patterns to the regulation of tumor angiogenesis, cell-cell contact, cell migration, and invasive growth of tumor cells. By acting like a constitutively active receptor, LMP1 recruits cellular signaling molecules associated with tumor necrosis factor receptors such as tumor necrosis factor receptor-associated factor (TRAF) proteins and TRADD to mimic signals of the costimulatory CD40 receptor in the EBV-infected B lymphocyte. LMP1 activates NF-κB, mitogen-activated protein kinase (MAPK), phosphatidylinositol 3-kinase (PI3-K), IRF7, and STAT pathways. Here, we review LMP1's molecular and biological functions, highlighting the interface between LMP1 and the cellular signal transduction network as an important factor of virus-host interaction and a potential therapeutic target.

Lymphoproliferative Disease in Wiskott-Aldrich Syndrome: Analysis of the French National Registry of Primary Immunodeficiencies

▪Wiskott-Aldrich syndrome (WAS) is an X-linked primary immune deficiency (PID) characterized by persistent thrombocytopenia, eczema and immunodeficiency. It has been suggested that these patients had an increased risk of lymphoproliferative diseases (LPD).We conducted a retrospective study in order to describe LPD occurring in patients with WAS/XLT (X Linked Thrombocytopenia with WAS gene mutation) enrolled in the French national registry for PIDs. Post- hematopoietic stem cell transplantation (HSCT) LPD were excluded. Methodology of the French National Registry for PIDs has been previously described (Clin. Immunol, 2010, ASH 2009 abstract#23950).Eight patients (4%) with LPD were identified among the 197 patients of our cohort. All patients had a WAS and presented first symptoms of WAS before the age of one year. One patient developed a LPD before the clinical diagnosis of WAS.All patients for whom histological charts were available (6/8) developed a B-cell LPD. Three of them had diffuse large B-cell lymphomas and three patients had polymorphic B-cell LPD associated with EBV and without detection of clonal IGHV rearrangement. The median age of cancer diagnosis was 18.3 years (range: 2.0-50.7). All LPD were disseminated diseases, mostly with a central nervous system involvement (4/6 patients). All patients received intensive chemotherapy, except one patient with an EBV-associated LPD who only received rituximab. Three patients obtained a complete remission (CR), two patients had a stable disease (SD) and the last one progressed on therapy. Four patients, of whom two were in CR and one in SD, received an allogeneic HSCT during the year following the diagnosis of LPD. Two patients received peripheral blood-derived stem cells and the two other patients received bone marrow-derived stem cells. There were three matched unrelated donor (MUD) and one haplotype-mismatched family donor, with no ex vivo graft manipulation. Three of these four patients were long-term survivors after LPD (2.7, 7.7 and 9.8 years).The median life expectancy for the entire cohort is 19.1 years (range: 7.8-52.0). Three patients, two of which have received an HSCT, are still alive 2.4, 2.7 and 9.8 years after the cancer onset. The median overall survival among the five remaining patients after diagnosis of LPD was 0.5 years (from 0.3 to 7.7 years).Additional details on genetic, clinical characteristics and a centralized histopathology review are currently being studied and will be presented.In conclusion, the occurrence of LPD is less frequent than expected, may occur only in patients with WAS phenotype rather than XLT. As it impairs the prognosis of patients, HSCT could be a therapeutic option for these patients, even in partial remission of LPD. DisclosuresNo relevant conflicts of interest to declare.