Latent Epstein-Barr Virus Research Articles

Epstein-Barr virus infection-induced inflammasome activation in human monocytes.

Inflammasomes are cytoplasmic sensors that regulate the activity of caspase-1 and the secretion of interleukin-1β (IL-1β) or interleukin-18 (IL-18) in response to foreign molecules, including viral pathogens. They are considered to be an important link between the innate and adaptive immune responses. However, the mechanism by which inflammasome activation occurs during primary Epstein-Barr virus (EBV) infection remains unknown. Human B lymphocytes and epithelial cells are major targets of EBV, although it can also infect a variety of other cell types. In this study, we found that EBV could infect primary human monocytes and the monocyte cell line, THP-1, inducing inflammasome activation. We incubated cell-free EBV with THP-1 cells or primary human monocytes, then confirmed EBV infection using confocal microscopy and flow cytometry. Lytic and latent EBV genes were detected by real-time RT-PCR in EBV-infected monocytes. EBV infection of THP-1 cells and primary human monocytes induced caspase-dependent IL-1β production, while EBV infection of B-cell or T-cell lines did not induce IL-1β production. To identify the sensor molecule responsible for inflammasome activation during EBV infection, we examined the mRNA and the protein levels of NLR family pyrin domain-containing 3 (NLRP3), absent in melanoma 2 (AIM2), and interferon-inducible protein 16 (IFI16). Increased AIM2 levels were observed in EBV-infected THP-1 cells and primary human monocytes, whereas levels of IFI16 and NLRP3 did not show remarkable change. Furthermore, knockdown of AIM2 by small interfering RNA attenuated caspase-1 activation. Taken together, our results suggest that EBV infection of human monocytes induces caspase-1-dependent IL-1β production, and that AIM2, acting as an inflammasome, is involved in this response.

A workflow for in silico design of hIL-10 and ebvIL-10 inhibitors using well-known miniprotein scaffolds.

The over-expression of immune-suppressors such as IL-10 is a crucial landmark in both tumor progression, and latent viral and parasite infection. IL-10 is a multifunctional protein. Besides its immune-cell suppressive function, it also promotes B-cell tumorigenesis of lymphomas and melanoma. Human pathogens like unicellular parasites and viruses that remain latent inside B cells promote the over-expression of hIL-10 upon infection, which inhibits cell-mediated immune surveillance, and at the same time mediates B cell proliferation. The B-cell specific oncogenic latent virus Epstein-Barr virus (EBV) encodes a viral homologue of hIL-10 (ebvIL-10), expressed during lytic viral proliferation. Once expressed, ebvIL-10 inhibits cell-mediated immune surveillance, assuring EBV re-infection. During long-term latency, EBV-infected B cells over-express hIL-10 to assure B-cell proliferation, occasionally inducing EBV-mediated lymphomas. The amino acid sequences of hIL-10 and ebvIL-10 are more than 80% identical and thus have a very similar tridimensional structure. Based on their published crystallographic structures bound to their human receptor IL10R1, we report a structure-based design of hIL-10 and ebvIL-10 inhibitors based on 3 loops from IL10R1 that establish specific hydrogen bonds with the two IL10s. We have grafted these loops onto a permissible loop in three well-known miniprotein scaffolds-the Conus snail toxin MVIIA, the plant-derived trypsin inhibitor EETI, and the human appetite modulator AgRP. Our computational workflow described in detail below was invigorated by the negative and positive controls implemented, and therefore paves the way for future in vitro and in vivo validation assays of the IL-10 inhibitors engineered.

Fibrin-associated EBV-positive Large B-Cell Lymphoma: An Indolent Neoplasm With Features Distinct From Diffuse Large B-Cell Lymphoma Associated With Chronic Inflammation.

Incidental cases of localized fibrin-associated Epstein-Barr virus (EBV)+ large B-cell proliferations have been described at unusual anatomic sites and have been included in the category of diffuse large B-cell lymphoma associated with chronic inflammation (DLBCL-CI) in the WHO Classification. We describe 12 cases and review the literature to define their clinicopathologic spectrum and compare features with typical cases of DLBCL-CI. Median age was 55.5 years with a M:F ratio of 3. In all 12 cases, the lymphoma was an incidental microscopic finding involving atrial myxomas (n=3), thrombi associated with endovascular grafts (n=3), chronic hematomas (n=2), and pseudocysts (n=4). All cases tested were nongerminal center B-cell origin, type III EBV latency, and were negative for MYC rearrangements and alternative lengthening of telomeres by FISH. Most showed high CD30, Ki67, and PD-L1, and low to moderate MYC and p53 expression. Among 11 patients with detailed follow-up, 6 were treated surgically, 3 with cardiac or vascular lesions had persistent/recurrent disease at intravascular sites, and 4 died of causes not directly attributable to lymphoma. Reports of previously published fibrin-associated cases showed similar features, whereas traditional DLBCL-CI cases with a mass lesion had significantly higher lymphoma-associated mortality. Fibrin-associated EBV+ large B-cell lymphoma is clinicopathologically distinct from DLBCL-CI, warranting separate classification. Most cases, particularly those associated with pseudocysts, behave indolently with the potential for cure by surgery alone and may represent a form of EBV+ lymphoproliferative disease rather than lymphoma. However, primary cardiac or vascular disease may have a higher risk of recurrence despite systemic chemotherapy.

Characterization of Epstein-Barr virus-encoded small RNA gene variations in virus associated lymphomas in Northern China.

Epstein-Barr virus (EBV)-encoded small RNAs (EBER1 and EBER2) are highly expressed in all forms of EBV latency in EBV-associated malignancies. EBER gene variations and their association with EBV-associated disease still remain poorly characterized. To investigate the patterns of EBER gene variations and their roles in tumorigenesis, EBER gene sequences were analyzed by nested-PCR and DNA sequencing in 101 lymphomas from Northern China, a non-nasopharyngeal carcinoma (NPC) endemic area. In addition, EBV type 1 and type 2 classifications were made by using nested-PCR assays across type-specific regions in the EBNA2 gene. EB-6m was the dominant subtype (95.0%, 96/101) in lymphoma. The distribution of the EBER subtypes in the four lymphoma groups was not significantly different (p>0.05), neither was that of the EBNA2 type (p>0.05). Compared with previous data in the same area, the distribution of EBER subtypes in lymphoma was similar to that in EBV-associated gastric carcinoma (EBVaGC) and throat washing (TW) from healthy donors (p>0.05), but was significantly different from that of NPC. The EBNA2 type distribution between lymphoma and the other three groups was significantly different (p<0.05). The proportion of type 1 and type 2 dual infections was higher in lymphoma than that in GC, NPC and TW. The mutation 7123nt A→T was identified in 11 of 101 (10.9%, 11/101) lymphomas, significantly more than that in EBV-associated gastric carcinomas (EBVaGC) (0%, 0/50) and throat washings (TWs) from healthy donors (3.3%, 3/92) (p<0.05). These findings indicate that EBER subtypes may not be associated with pathogenesis of lymphoma, but that a point mutation at position 7123nt (A→T) provides a new area for further exploration. Furthermore it is necessary to investigate the role of EBNA2-subtype mixed infections in the establishment of lymphoma.

Epstein-Barr virus gene expression and latency pattern in gastric carcinomas: a systematic review.

A systematic review of literature was conducted to identify all published reports regarding the expression of Epstein-Barr Virus (EBV) proteins/transcripts and EBV latency patterns in EBV-associated gastric carcinomas (EBVaGC). The literature search retrieved 247 papers, of which 25 papers matched the inclusion criteria. The analysis reveals that the most frequently expressed EBV latent proteins are EBNA1 (98.1%) and LMP2A (53.8%), while LMP1 and LMP2B are present in only 10% of cases. Lytic proteins, such as BARF0 and BARF1, and other lytic transcripts are present in almost half of cases. EBVaGC seems to display a unique transcription/latency pattern that does not fit the 'standard' EBV latency patterns and therefore should be further studied to better understand EBVaGC carcinogenesis.

EBV Infection and Glucose Metabolism in Nasopharyngeal Carcinoma.

To establish persistent infection in cells, viruses evolve strategies to alter host cellular pathways to regulate cell proliferation and energy metabolism which support viral infection. Epstein-Barr virus (EBV) undergoes both lytic and latent infection to achieve persistent and lifelong infection in human. EBV readily infects human B cells, driving their transformation to proliferative lymphoblastoid cell lines (LCL), and eventually establishes lifelong latent infection in memory B cells. In contrary, EBV undergoes lytic replication upon infection into normal epithelial cells which is essential for the replication of EBV genome and production of infectious viral particles for transmission through saliva. EBV shuttles between B cells and epithelial cells to complete its infection cycle. EBV infection is closely associated with nasopharyngeal carcinoma (NPC) and is present in practically 100% of undifferentiated NPC. In contrast to undergo lyticinfection of normal pharyngeal epithelium, EBV establishes latent infection in NPC. The switch from lytic infection to latent infection may represent an early and essential step in the development of NPC. Recent studies in both B cells and NPC cells latently infected with EBV reveal alterations in cell metabolism to support persistentand latent EBV infection. Events underlying the switching of lytic to latent EBV infection in NPC cells are largely undefined. Molecular events and alterations of cell metabolism are likely to play crucial roles in switching EBV infection from lytic to latent in NPC cells. Latent EBV infection and expression of viral genes, including LMP1, LMP2, and possibly EBV-encoded micro RNAs, may play essential roles in alterations of cell metabolism to support NPC pathogenesis. Alteration of energy metabolism is an essential hallmark of cancer. The role of altered energy metabolism in host cells in modulating latent and lytic EBV infection in NPC cells is unclear. In this review, we will discuss the impact of genetic alterations in NPC to module cellular metabolism and its influence on latent infection and lytic reactivation of EBV infection in NPC cells. In particular, the role of EBV-encoded genes in driving glucose metabolism and their contribution to NPC pathogenesis will be discussed. This new perspective on the interplay between EBV infection and altered host metabolic pathways in NPC pathogenesis may offer novel and effective therapeutic strategies in the treatment of NPC and other EBV-associated malignancies.

Up-Regulation of Lamin A/C Expression in Epstein-Barr Virus Immortalized B Cells and Burkitt Lymphoma Cell Lines of Activated B Cell Phenotype

Lamin A, B and C, the nuclear intermediate-filament proteins, play a role in epigenetic regulation. Lamin B could be detected in all nucleated cells studied, whereas the lamin A and lamin C isoforms (lamin A/C) encoded by the LMNA gene are co-expressed in most somatic cell types except mature B lymphocytes. Since Epstein-Barr virus (EBV), a human gammaherpesvirus, is associated with tumorigenic processes and is known to alter the epigenotype of its host cells, we studied the expression of the LMNA gene and its epigenetic marks in EBV-carrying human lymphoid cell lines. We observed a high lamin A/C mRNA expression in EBV-immortalized B lymphoblastoid cell lines (LCLs) and in a subset of Burkitt lymphoma (BL) lines characterized by an activated B cell phenotype and a unique latent EBV gene expression pattern (latency III). In these cells the first exon of LMNA was hypomethylated and associated with activating histone marks. In contrast, we observed a low level of lamin A/C mRNA expression in EBV negative BL lines and BL lines with a restricted expression of latent EBV products (latency I). Low LMNA promoter activity was associated with hypermethylation of the LMNA first exon. These data suggest a role for EBV latency products in switching on or upregulating the LMNA promoter (LMNAp) in EBV-infected activated B cells in vitro. Lamin A/C may contribute to the establishment of the activated B cell phenotype. Our data also imply a role of LMNA first exon methylation in the silencing of LMNAp.

Chaetocin reactivates the lytic replication of Epstein-Barr virus from latency via reactive oxygen species.

Oxidative stress, regarded as a negative effect of free radicals in vivo, takes place when organisms suffer from harmful stimuli. Some viruses can induce the release of reactive oxygen species (ROS) in infected cells, which may be closely related with their pathogenicity. In this report, chaetocin, a fungal metabolite reported to have antimicrobial and cytostatic activity, was studied for its effect on the activation of latent Epstein-Barr virus (EBV) in B95-8 cells. We found that chaetocin remarkably up-regulated EBV lytic transcription and DNA replication at a low concentration (50 nmol L-1). The activation of latent EBV was accompanied by an increased cellular ROS level. N-acetyl-L-cysteine (NAC), an ROS inhibitor, suppressed chaetocin-induced EBV activation. Chaetocin had little effect on histone H3K9 methylation, while NAC also significantly reduced H3K9 methylation. These results suggested that chaetocin reactivates latent EBV primarily via ROS pathways.

Association of Epstein–Barr Virus (EBV) with malignancy of the nasopharynx in Lagos, Nigeria

Background: Nasopharyngeal cancer is a rare cancer worldwide. It is now found to be increasing in incidence in Nigeria, though the incidence is lower when compared to countries in the Mediterranean basin, North Africa and Southeast Asia. This cancer is frequently associated with Epstein-Barr virus (EBV), but no previous study was done in this centre to document the association. Aim and Objectives: To assess the EBV status in Nigerian patients with nasopharyngeal cancer by using Latent membrane protein – 1 immunohistochemistry. ( LMP-1IHC). Materials and Methods: Thirty-five (35) out of Seventy-five (75) histologically-confirmed cases of nasopharyngeal malignancy (NPM) seen in the Department of Anatomic and Molecular Pathology of Lagos University Teaching Hospital, Lagos between January 2003 and December 2012 were available for EBV study using LMP-1 IHC. Results: EBV LMP-1 IHC was positive in thirty (86%) out of the thirty-five cases studied and these consisted of, 14 cases of undifferentiated carcinoma (WHO type 3), 5 cases of non-Hodgkin lymphoma and 2 cases of adenocarcinoma. Also seven out of eleven cases of differentiated non-keratinizing squamous cell carcinoma (WHO type 2) were EBV positive and two of the three cases of keratinizing squamous cell carcinoma (WHO type 1) were EBV positive. Conclusion: This study has shown that nasopharyngeal malignancy in our centre is highly associated with EBV latency as detected by LMP-1 immunohistochemistry.

Identification of Epstein-Barr Virus in the Human Placenta and Its Pathologic Characteristics.

Epstein-Barr virus (EBV), a common pathogen in humans, is suspected as the cause of multiple pregnancy-related pathologies including depression, preeclampsia, and stillbirth. Moreover, transmission of EBV through the placenta has been reported. However, the focus of EBV infection within the placenta has remained unknown to date. In this study, we proved the expression of latent EBV genes in the endometrial glandular epithelial cells of the placenta and investigated the cytological characteristics of these cells. Sixty-eight placentas were obtained from pregnant women. Tissue microarray was constructed. EBV latent genes including EBV-encoding RNA-1 (EBER1), Epstein-Barr virus nuclear antigen 1 (EBNA1), late membrane antigen (LMP1), and RPMS1 were detected with silver in situ hybridization and/or mRNA in situ hybridization. Nuclear features of EBV-positive cells in EBV-infected placenta were compared with those of EBV-negative cells via image analysis. Sixteen placentas (23.5%) showed positive expression of all 4 EBV latent genes; only the glandular epithelial cells of the decidua showed EBV gene expression. EBV infection status was not significantly correlated with maternal, fetal, or placental factors. The nuclei of EBV-positive cells were significantly larger, longer, and round-shaped than those of EBV-negative cells regardless of EBV-infection status of the placenta. For the first time, evidence of EBV gene expression has been shown in placental tissues. Furthermore, we have characterized its cytological features, allowing screening of EBV infection through microscopic examination.

Quantitative multi-target RNA profiling in Epstein-Barr virus infected tumor cells

Epstein-Barr virus (EBV) is etiologically linked to multiple acute, chronic and malignant diseases. Detection of EBV-RNA transcripts in tissues or biofluids besides EBV-DNA can help in diagnosing EBV related syndromes. Sensitive EBV transcription profiling yields new insights on its pathogenic role and may be useful for monitoring virus targeted therapy. Here we describe a multi-gene quantitative RT-PCR profiling method that simultaneously detects a broad spectrum (n=16) of crucial latent and lytic EBV transcripts. These transcripts include (but are not restricted to), EBNA1, EBNA2, LMP1, LMP2, BARTs, EBER1, BARF1 and ZEBRA, Rta, BGLF4 (PK), BXLF1 (TK) and BFRF3 (VCAp18) all of which have been implicated in EBV-driven oncogenesis and viral replication. With this method we determine the amount of RNA copies per infected (tumor) cell in bulk populations of various origin. While we confirm the expected RNA profiles within classic EBV latency programs, this sensitive quantitative approach revealed the presence of rare cells undergoing lytic replication. Inducing lytic replication in EBV tumor cells supports apoptosis and is considered as therapeutic approach to treat EBV-driven malignancies. This sensitive multi-primed quantitative RT-PCR approach can provide broader understanding of transcriptional activity in latent and lytic EBV infection and is suitable for monitoring virus-specific therapy responses in patients with EBV associated cancers.

NKG2A-Expressing Natural Killer Cells Dominate the Response to Autologous Lymphoblastoid Cells Infected with Epstein-Barr Virus.

Epstein–Barr virus (EBV) is a human γ-herpesvirus that establishes latency and lifelong infection in host B cells while achieving a balance with the host immune response. When the immune system is perturbed through immunosuppression or immunodeficiency, however, these latently infected B cells can give rise to aggressive B cell lymphomas. Natural killer (NK) cells are regarded as critical in the early immune response to viral infection, but their role in controlling expansion of infected B cells is not understood. Here, we report that NK cells from healthy human donors display increased killing of autologous B lymphoblastoid cell lines (LCLs) harboring latent EBV compared to primary B cells. Coculture of NK cells with autologous EBV+ LCL identifies an NK cell population that produces IFNγ and mobilizes the cytotoxic granule protein CD107a. Multi-parameter flow cytometry and Boolean analysis reveal that these functional cells are enriched for expression of the NK cell receptor NKG2A. Further, NKG2A+ NK cells more efficiently lyse autologous LCL than do NKG2A− NK cells. More specifically, NKG2A+2B4+CD16−CD57−NKG2C−NKG2D+ cells constitute the predominant NK cell population that responds to latently infected autologous EBV+ B cells. Thus, a subset of NK cells is enhanced for the ability to recognize and eliminate autologous, EBV-infected transformed cells, laying the groundwork for harnessing this subset for therapeutic use in EBV+ malignancies.

Lymphoepithelioma-like gastric carcinoma: clinicopathological characteristics and infection status

BackgroundLymphoepithelioma-like gastric carcinoma (LLGC) is a rare subtype of gastric carcinoma (GC) characterized by prominent lymphocytic infiltration. LLGC may be associated with latent Epstein–Barr virus (EBV) infection or microsatellite instability (MSI). This study aims to assess the clinicopathological characteristics, EBV infection, and MSI status in LLGC. MethodsA retrospective analysis of GC patients submitted to potentially curative resection between 2009 and 2014 was performed. The LLGC subtype specimens were examined for EBV by in situ hybridization and MSI by immunohistochemical analysis. The LLGC profile was analyzed accordingly to clinicopathological parameters. ResultsFrom 255 patients, seven were identified on the pathological report as LLGC. Six cases were EBV-positive and one had MSI, showing loss of MLH1 and PMS2 expression. LLGC was more frequently seen in men, and the mean age was 69 years. When compared to non-LLGC, LLGC cases were larger (∼5.8 cm) poorly differentiated tumors and had lower incidence of lymph node metastasis (P = 0.045). Mean number of lymph nodes dissected in the LLGC group was 39.5, and only one patient had a single positive lymph node. In addition, two patients presented associated lesions. LLGC was not associated with HER-2, chromogranin and synaptophysin positivity or Helicobacter pylori infection. ConclusionsDistinct pathological aspects and clinical behavior of LLGC reinforce the need for proper recognition of this histological subtype to choose better therapeutic approaches.

LMP1 signaling pathway activates IRF4 in latent EBV infection and a positive circuit between PI3K and Src is required.

Interferon (IFN) regulatory factors (IRFs) have crucial roles in immune regulation and oncogenesis. We have recently shown that IRF4 is activated through c-Src-mediated tyrosine phosphorylation in virus-transformed cells. However, the intracellular signaling pathway triggering Src activation of IRF4 remains unknown. In this study, we provide evidence that Epstein-Barr virus (EBV) latent membrane protein 1 (LMP1) promotes IRF4 phosphorylation and markedly stimulates IRF4 transcriptional activity, and that Src mediates LMP1 activation of IRF4. As to more precise mechanism, we show that LMP1 physically interacts with c-Src, and the phosphatidylinositol 3 kinase (PI3K) subunit P85 mediates their interaction. Depletion of P85 by P85-specific short hairpin RNAs disrupts their interaction and diminishes IRF4 phosphorylation in EBV-transformed cells. Furthermore, we show that Src is upstream of PI3K for activation of both IRF4 and Akt. In turn, inhibition of PI3K kinase activity by the PI3K-speicfic inhibitor LY294002 impairs Src activity. Our results show that LMP1 signaling is responsible for IRF4 activation, and further characterize the IRF4 regulatory network that is a promising therapeutic target for specific hematological malignancies.

Epstein–Barr virus microRNAs reduce immune surveillance by virus-specific CD8+ T cells

Infection with Epstein-Barr virus (EBV) affects most humans worldwide and persists life-long in the presence of robust virus-specific T-cell responses. In both immunocompromised and some immunocompetent people, EBV causes several cancers and lymphoproliferative diseases. EBV transforms B cells in vitro and encodes at least 44 microRNAs (miRNAs), most of which are expressed in EBV-transformed B cells, but their functions are largely unknown. Recently, we showed that EBV miRNAs inhibit CD4+ T-cell responses to infected B cells by targeting IL-12, MHC class II, and lysosomal proteases. Here we investigated whether EBV miRNAs also counteract surveillance by CD8+ T cells. We have found that EBV miRNAs strongly inhibit recognition and killing of infected B cells by EBV-specific CD8+ T cells through multiple mechanisms. EBV miRNAs directly target the peptide transporter subunit TAP2 and reduce levels of the TAP1 subunit, MHC class I molecules, and EBNA1, a protein expressed in most forms of EBV latency and a target of EBV-specific CD8+ T cells. Moreover, miRNA-mediated down-regulation of the cytokine IL-12 decreases the recognition of infected cells by EBV-specific CD8+ T cells. Thus, EBV miRNAs use multiple, distinct pathways, allowing the virus to evade surveillance not only by CD4+ but also by antiviral CD8+ T cells.

Lenalidomide, Thalidomide, and Pomalidomide Reactivate the Epstein-Barr Virus Lytic Cycle through Phosphoinositide 3-Kinase Signaling and Ikaros Expression.

Lenalidomide, thalidomide, and pomalidomide (LTP) are immunomodulatory agents approved for use in multiple myeloma, but in some settings, especially with alkylating agents, an increase in Hodgkin lymphoma and other secondary primary malignancies (SPM) has been noted. Some of these malignancies have been linked to Epstein-Barr virus (EBV), raising the possibility that immunomodulatory drugs disrupt latent EBV infection. We studied the ability of LTP to reactivate latently infected EBV-positive cell lines in vitro and in vivo, and evaluated the EBV viral load in archived serum samples from patients who received a lenalidomide, thalidomide, and dexamethasone (LTD) combination. Treatment of EBV-infected B-cell lines with LTP at physiologically relevant concentrations induced the immediate early gene BZLF1, the early gene BMRF1, and the late proteins VCA and BCFR1. This occurred in the potency order pomalidomide > lenalidomide > thalidomide, and the nucleoside analogue ganciclovir enhanced the cytotoxic effects of lenalidomide and pomalidomide in Burkitt lymphoma cells in vitro and in vivo EBV reactivation was related to PI3K stimulation and Ikaros suppression, and blocked by the PI3Kδ inhibitor idelalisib. Combinations of lenalidomide with dexamethasone or rituximab increased EBV reactivation compared with lenalidomide alone and, importantly, lenalidomide with melphalan produced even greater reactivation. We conclude LTP may reactivate EBV-positive resting memory B cells thereby enhancing EBV lytic cycle and host immune suppression. Clin Cancer Res; 22(19); 4901-12. ©2016 AACR.

Anti-Cancer Effect of Quercetin in Xenograft Models with EBV-Associated Human Gastric Carcinoma.

Licorice extracts have been widely used in herbal and folk medications. Glycyrrhiza contains diverse range of biological compounds including triterpenes (glycyrrhizin, glycyrrhizic acid) and flavonoids (quercetin, liquiritin, liquiritigenin, glabridin, licoricidin, isoliquiritigenin). The flavonoids in licorice are known to have strong anti-cancer activities. Quercetin, the most abundant flavonoid, has been shown to have anti-ulcer, anti-cancer, antioxidant, and anti-inflammatory properties. Latent Epstein-Barr virus (EBV) infection can lead to serious malignancies, such as, Burkitt’s lymphoma, Hodgkin’s disease and gastric carcinoma(GC), and (Epstein-Barr virus associated gastric carcinoma) EBVaGC is one of the most common EBV-associated cancers. In this study, the authors first examined the anti-cancer effects of quercetin and isoliquiritigenin in vivo xenograft animal models implanted with EBV(+) human gastric carcinoma (SNU719) or EBV(−) human gastric carcinoma (MKN74), and then explored the molecular mechanisms responsible for their anti-cancer activities. The results obtained showed that anti-cancer effect of quercetin was greater than isoliquiritigenin in mice injected with EBV(+) human gastric carcinoma (SNU719) cells. On the other hand, quercetin and isoliquiritigenin had similar anti-cancer effects in mice injected with EBV(−) human gastric carcinoma (MKN74) cells. Interestingly, quercetin inhibited EBV viral protein expressions, including EBNA-1 and LMP-2 proteins in tumor tissues from mice injected with EBV(+) human gastric carcinoma. Quercetin more effectively induced p53-dependent apoptosis than isoliquiritigenin in EBV(+) human gastric carcinoma, and this induction was correlated with increased expressions of the cleaved forms of caspase-3, -9, and Parp. In EBV(−)human gastric carcinoma (MKN74), both quercetin and isoliquiritigenin induced the expressions of p53, Bax, and Puma and the cleaved forms of caspase-3 and -9 and Parp at similar levels.

Active CMV and EBV infections in renal transplant recipients with unexplained fever and elevated serum creatinine

Proper identification of active cytomegalovirus (CMV) and Epstein-Barr virus (EBV) infections are helpful for monitoring antiviral treatment in transplant recipients. Qualitative and quantitative CMV, EBV DNA PCR techniques in the context of serological tests are performed for early detection and differentiation of active and latent CMV and EBV infections in renal transplantation. Basically, 129 renal transplanted recipients monitored carefully and hospitalized for unexplained elevated creatinine levels or high fever and 21 of their donors were studied. CMV DNA was detected in 63.5% of the febrile episodes following transplantation and in 46.42% of readmitted patients using qualitative PCR method. In the first group, 15% of the patients and in the second group 42.85% of the patients had copy numbers more than cutoff point (900 copies/mL). Cutoff point had 100% sensitivity and 82.5% specificity for active and symptomatic CMV infection. Only 15.5% of the subjects were positive for EBV infection by qualitative PCR method. Among them 5% had >2000 copies/mL and were symptomatic. One subject with a history of three times hospitalization had higher EBV viral load and developed post-transplant lymphoproliferative disorder. CMV load was significantly correlated with elevated creatinine levels (OR = 3.1, p = 0.006), abnormal heart sounds (OR = 4.7; p = 0.02) and hypertension (OR = 3.6; p = 0.03). Only qRT-PCR could differentiate between latent and active infections and might be clinically useful for monitoring symptomatic CMV and EBV infections and initiation of the antiviral therapy. Elevated creatinine levels, hypertension, and abnormal heart sounds could be considered as main manifestations of HCMV infection in kidney recipients.

Epstein–Barr virus and regulatory T cells in Egyptian paediatric patients with acute B lymphoblastic leukaemia

ObjectiveAcute B lymphoblastic leukaemia (B-ALL) is the most common type of childhood malignancy worldwide but little is known of its origin. Recently, many studies showed both a high incidence of...

Getting Back to the Basics of Translational Research.

How do viruses cause cancer? As a naive graduate student in the years before kits and personal computers, I found myself fascinated by viral “immortalization,” the process by which a virus induces a cell to live forever. The immortalizing virus of interest was named Epstein–Barr Virus (EBV) after its co-discoverers and was isolated from tumors commonly found in African children. EBV was also known to be a major cause of infectious mononucleosis, commonly referred to as the “kissing disease,” and a potential agent in chronic fatigue syndromes. For these reasons, I suspect, EBV was slow to get the respect it deserved. We now know that EBV is found in diverse tumors and is responsible for almost ~200,000 cancer cases each year. At the time, my interests were to understand the viral mechanisms that transform a resting mortal cell to replicate uncontrollably. It was also tempting to speculate over a beer whether viral immortalization could reveal fresh insights into human mortality and life span. On more sober occasions, the question of how a virus could cause cancer seemed a big enough challenge. Over the years, my interest in viral immortalization morphed into an investigation of viral gene regulation and latency. How does EBV establish long-term latent infection and express only a few essential viral genes necessary to keep the host cell dividing and alive? To get at these questions, we needed to combine many different methods ranging from functional genomics to structural biology, requiring expertise in diverse disciplines far beyond my capabilities. Building a team of collaborators with the appropriate expertise and identifying the most appropriate technologies is essential. Selecting the right tools and team to answer the bigger questions is not always straightforward, but establishing multi-disciplinary collaborations can open many otherwise closed doors. Among the most important and challenging steps in a research career is identifying a significant long-term research problem. This sounds easy, but it has many snags. Highly ambitious long-term goals, like curing EBV cancers, may not be achievable without more mundane short-term groundwork. And short-term goals, like the need to get the next paper published or grant funded, can sometimes distract from the long-term plan. Working on tangential problems is necessary to overcome obstacles and can lead to new and unanticipated discoveries. But keeping sight of the long-term goal can be a great strength for a research program. If all goes well, the research focus will be aligned with reviewers and funding agencies. Fortunately for EBV research, foundations like Cancer Research UK have recently recognized the eradication of EBV-associated cancers as one of the decade’s grand challenges. So how best to respond to this grand challenge? From my perspective, studying the basic mechanisms of EBV latency is the best path to identify viral-specific targets for therapeutic intervention. At least half of my group is committed to developing small molecules to either inhibit EBV latency or stimulate viral reactivation, two possible strategies to treat latent viruses in cancer cells. Viral reactivation from latency is attractive because it is likely to trigger the host immune system to eliminate EBV-infected cancer cells. Inhibiting latency should lead to the loss of viral DNA from cancer cells that depend on EBV for growth and survival. In principle, inhibitors of EBV latency or activators of viral lytic antigens should be valuable therapies to treat EBV-associated cancer and related disease. Of course these more ambitious projects are risky and require vast sums of funding. The pharmaceutical industry and venture capitalists have no tolerance for financing the early stages of long-term, high-risk projects, so public funding of academic investigators will be the only possible path forward. Although there are many academics who consider translational research a corruption of academic freedom, the urgency of real-world unmet medical needs becomes very apparent if one only takes a look at the economic and human cost of disease. It will be essential for academic investigators to fulfill these obligations to society. Ironically, an editorial opposing the funding of basic academic research was recently written by Matt Ridley in The Wall Street Journal arguing that public sponsorship of basic research blocks private investment and derails innovation. In fact, the only chance for innovative solutions to long-term challenges will be through public funding of basic academic research. Encouraging scientists to focus on unmet medical needs as well as conceptual advances in basic science should be a high priority for public and private investment in a healthy future. Image 1 Paul M. Lieberman, Hilary Koprowski Professor, The Wistar Institute.