Systemic Epstein-Barr virus-positive T-cell lymphoma of childhood combined with hemophagocytic lymphohistiocytosis: a case report.

Background: Systemic Epstein-Barr virus (EBV)-positive T-cell lymphoma of childhood (STCLC) is a rare disease with few clinical reports and high mortality. By exploring the clinical manifestations of a child with STCLC in our hospital auxiliary examination and diagnostic and therapeutic process, to deepen pediatricians' understanding of this disease. Case Description: This paper describes a 5-year-old Chinese girl who presented with acute fever and epistaxis. After admission, relevant ancillary tests indicated the presence of hemophagocytic lymphohistiocytosis (HLH) and the combination of EBV infection in this patient. Pathology of the cervical lymph node biopsy and bone marrow flow cytology examination indicated STCLC, and a diagnosis of STCLC combined with HLH was clear. Although the girl was clearly diagnosed within a few days and treated with chemotherapy and symptomatic support, she eventually died on the 6th day after admission due to progressive worsening of her disease. Conclusions: STCLC is a rare T-cell lymphoproliferative disorder that occurs primarily in the setting of acute EBV infection, usually presenting as HLH. It is a rapidly progressive and fatal disease of children and young adults characterized by monoclonal expansions of EBV-positive T-cells with an activated cytotoxic phenotype and by malignant proliferation. The mortality rate is close to 100%.

Myelin Oligodendrocyte Glycoprotein Antibody Disease (MOGAD)-Monophasic Optic Neuritis and Epstein-Barr Virus (EBV): A Case Report of Rare Comorbid Diagnoses in an Adolescent From a Remote Greek Island.

A unique case of a female adolescent diagnosed with myelin oligodendrocyte glycoprotein (MOG) monophasic optic neuritis with Epstein-Barr virus (EBV) reactivation antibody profile on a remote Greek island is presented, highlighting the challenges of diagnosing rare conditions in rural settings and the importance of connecting centers of expertise with regional hospitals. The 16-year-old patient presented with progressive vision loss, headache, and retrobulbar pain in the right eye. Initial ophthalmological examinations showed decreased visual acuity and color vision deterioration. Magnetic resonance imaging (MRI) revealed optic perineuritis and edema. Cerebrospinal fluid (CSF) analysis excluded oligoclonal bands, and blood analysis was positive for both anti-MOG antibodies and EBV reactivation. Expert opinion and blood immunophenotyping confirmed the neuroimmunological condition. This case not only underscores the value of telemedicine in overcoming diagnostic challenges in rural settings but also contributes to the scientific discussion on neuroimmunological aspects and the potential role of EBV as an underlying factor in acquired demyelinating syndromes (ADS), beyond multiple sclerosis (MS).

TianTan Vaccinia Virus-based EBV vaccines targeting both latent and lytic antigens elicits potent immunity against lethal EBV challenge in humanized mice.

AbstractEpstein-Barr virus (EBV) infection has been related to multiple epithelial cancers and lymphomas. Current efforts in developing a prophylactic EBV vaccine have focused on inducing neutralizing antibodies. However, given the lifelong and persistent nature of EBV infection following primary infection, it is rationalized that an ideal vaccine should elicit both humoral and cellular immune responses targeting multiple stages of the EBV lifecycle. This study used a DNA vector and a TianTan vaccinia virus to express key EBV antigens, including BZLF1, EBNA1, EBNA3B, and gH/gL, to generate multi-antigen vaccines. The multi-antigen vaccine expressing all four antigens and the multi-antigen vaccine expressing BZLF1, EBNA1, and EBNA3B showed comparable protection effects and prevented 100% and 80% of humanized mice, respectively, from EBV-induced fatal B cell lymphoma by activating BZLF1, EBNA1, and EBNA3B specific T cell. The vaccine expressing lytic protein BZLF1 elicited stronger T cell responses and conferred superior protection compared to vaccines targeting single latent EBNA1 or EBNA3B. The vaccine solely expressing gH/gL exhibited no T cell protective effects in our humanized mice model. Our study implicates the potential of EBV vaccines that induce potent cellular responses targeting both latent and lytic phases of the EBV life cycle in the prevention of EBV-induced B cell lymphoma.

High Somatization Rates, Frequent Spontaneous Recovery, and a Lack of Organic Biomarkers in Post-Covid-19 Condition.

INTRODUCTION: Many patients report neuropsychiatric symptoms after SARS-CoV-2 infection. Data on prevalence of post-COVID-19 condition (PCC) vary due to the lack of specific diagnostic criteria, the report of unspecific symptoms, and reliable biomarkers. METHODS: PCC patients seen in a neurological outpatient department were followed for up to 18 months. Neurological examination, SARS-CoV-2 antibodies, Epstein-Barr virus antibodies, and cortisol levels as possible biomarkers, questionnaires to evaluate neuropsychiatric symptoms and somatization (Patient Health Questionnaires D [PHQ-D]), cognition deficits (Montreal Cognitive Assessment [MoCA]), sleep disorders (ISS, Epworth Sleepiness Scale [ESS]), and fatigue (FSS) were included. RESULTS: A total of 175 consecutive patients (78% females, median age 42 years) were seen between May 2021 and February 2023. Fatigue, subjective stress intolerance, and subjective cognitive deficits were the most common symptoms. Specific scores were positive for fatigue, insomnia, and sleepiness and were present in 95%, 62.1%, and 44.0%, respectively. Cognitive deficits were found in 2.3%. Signs of somatization were identified in 61%, who also had an average of two symptoms more than patients without somatization. Overall, 28% had a psychiatric disorder, including depression and anxiety. At the second visit (n = 92), fatigue (67.3%) and insomnia (45.5%) had decreased. At visit three (n = 43), symptom load had decreased in 76.8%; overall, 51.2% of patients were symptom-free. Biomarker testing did not confirm an anti-EBV response. SARS-CoV-2-specific immune reactions increased over time, and cortisol levels were within the physiological range. CONCLUSION: Despite high initial symptom load, 76.8% improved over time. The prevalence of somatization and psychiatric disorders was high. Our data do not confirm the role of previously suggested biomarkers in PCC patients.

Prevention of post-transplant lymphoproliferative disorder in pediatric kidney transplant recipients.

BACKGROUND: Post-transplant lymphoproliferative disorder (PTLD) is a devastating complication of immunosuppressive treatment in both solid organ transplantations (SOT) and hematopoietic stem cell transplantations (HSCT). Epstein-Barr virus (EBV) infection precedes PTLD in 90% of patients. Rituximab, a monoclonal anti-CD20 antibody, depletes B-lymphocytes, which are the ultimate reservoir for EBV. Although rituximab therapy is commonly used as a preventive measure for PTLD in high-risk HSCT, it is not established in SOT. METHODS: Pediatric kidney transplant recipients (PKTR) underwent routine EBV-PCR surveillance. Patients with increasing viral loads, despite immunosuppressive dose reduction, were managed with preventive rituximab therapy. RESULTS: Between 2012 and 2023, we identified eight episodes of asymptomatic EBV-PCR-positive blood tests in seven out of 65 PKTR (11%) under our care. EBV DNAemia emerged 120-720 days post-transplantation. Five of seven patients with EBV DNAemia (71%) were EBV-seronegative prior to transplantation. All five patients did not respond to MMF dose reduction and were therefore treated with preventive rituximab therapy. Following this treatment, EBV PCR clearance was observed in all patients with only minimal complications. CONCLUSIONS: PKTR who are EBV-naïve prior to transplantation are expected to have a higher prevalence of EBV DNAemia. We found that PKTR who were EBV seronegative prior to transplantation were less likely to achieve EBV clearance in response to immunosuppression dose reduction. We suggest that rituximab therapy in PKTR may be safe and effective in EBV clearance and PTLD prevention.

Multifocal Epstein-Barr Virus-Positive Mucocutaneous Ulcers in a Patient With Crohn's Disease.

Epstein-Barr Virus-positive mucocutaneous ulcer (EBVMCU) is a rare and new category of mature B-cell neoplasms commonly linked to immunosuppression. It often has a benign course and regresses spontaneously after discontinuation or dose reduction of immunosuppressive agents. We report the case of a 48-year-old woman on long-term azathioprine therapy for rectosigmoid Crohn's disease. In contrast to the prevalent sites typically associated with EBVMCU, such as the oral mucosa and skin, this patient was found to have locations in the gastrointestinal tract and upper neck. These areas tested positive for histopathology consistent with EBVMCU and were excised due to bowel perforation and concern for malignancy.

Epstein-Barr Virus-Associated Aggressive Natural Killer Cell Leukemia: Challenges and Emerging Therapies.

A 24-year-old Ecuadorian female, previously diagnosed with acute fatty liver (AFL) during pregnancy, developed constitutional symptoms, jaundice, and abdominal pain in a subsequent pregnancy, prompting investigations that suggested a recurrence of AFL. She underwent an elective abortion, which resulted in the resolution of her abdominal pain, and a liver biopsy, which showed granulomatous inflammation and lymphocytic infiltration. She later presented with abdominal distention, productive cough, and persistent constitutional symptoms and jaundice. Extensive laboratory and imaging studies indicated sepsis, acute liver injury, and disseminated intravascular coagulopathy. Her serum Epstein-Barr virus (EBV) level was elevated. Special staining of her previous liver biopsy revealed EBV-positive natural killer (NK) cells. A bone marrow biopsy also revealed EBV-positive NK cells. She was diagnosed with aggressive NK cell leukemia (ANKL) with or without chronic active EBV (CAEBV). Treatment included dexamethasone, atovaquone, bortezomib, and ganciclovir, with plans for a stem cell transplant. However, her course was complicated by infections and multi-organ failure, resulting in her passing. This case highlights the rarity and challenges in managing EBV-associated ANKL, emphasizing the need for early detection and improved treatment options, with stem cell transplantation offering the best prognosis.

The simultaneous presence of active BK, Epstein Barr, and human cytomegalovirus infection and their correlation by host factors in patients suspected of kidney transplant rejection.

AIMS: This study aims to evaluate the presence of EBV, HCMV, and BKV genomic sequences in the plasma samples (active infection/viremia) of kidney transplant recipients suspected of rejection and to investigate host and risk factors related to the activation of these viruses in these patients. METHODS: In this cross-sectional single-center study, plasma samples were collected from 98 suspected kidney transplant rejection patients at Labafinejad Hospital, Tehran, Iran, between December 2022 and June 2023. Quantitative real-time PCR assays for HCMV, EBV, and BK were performed using GeneProof Real-time PCR kits. ROC curve analysis was used to determine the viral load cutoff point for each virus. FINDINGS: HCMV active viremia was detected in 18 (18.36%) recipients, EBV active viremia in 7 (7.14%), and BKV active viremia in 5 (5.10%). ROC results indicated viral load cutoff points of 778, 661, and 457 points for HCMV, EBV, and BKV, respectively. The duration of time after transplantation significantly differed between active viremia and no viremia groups (120.5 vs. 46 months, P = 0.014). In the BKV active viremia group, the increase in creatinine compared to baseline creatinine was significantly higher than in the no viremia group (2.7 vs. 0.8, P = 0.017). The odds ratio of HCMV active viremia in patients taking tacrolimus was 2.84 times higher, and the odds of HCMV active viremia in patients taking antithymocyte globulin was 3.01 times higher than in patients not taking these drugs. CONCLUSION: Rapid and timely diagnosis of viral active infections in kidney transplant patients is crucial for effective disease management and implementation of appropriate treatment strategies. Identifying potential risk factors, including host and treatment-related factors that influence transplantation, can facilitate the development of suitable preventive strategies.

The Epstein-Barr virus small capsid protein BFRF3 disrupts the NF-кB signaling pathway by inhibiting p65 activity.

Epstein-Barr virus (EBV), a common gamma herpesvirus, establishes a life-long latent infection in the host to defend against innate immune recognition, which is closely related to a variety of malignant tumors, but its specific mechanism is unclear. BFRF3, an EBV-encoded small capsid protein, is mainly involved in the assembly of the viral capsid structure and the maintenance of its stability. Here, we showed that BFRF3 can inhibit TNF-α-mediated NF-кB promoter activation. Moreover, BFRF3 downregulates NF-кB-mediated promoter activation and transcription of inflammatory cytokines, including IL-6 and IL-8. Dual-luciferase reporter assay demonstrated that BFRF3 restrains NF-кB promoter activity at or below the p65 level, and coimmunoprecipitation analysis revealed that BFRF3 not only interacts with p65 but also binds to its critical truncated Rel homology domain (RHD) and transcriptional activation domain (TAD). However, BFRF3 does not affect the dimerization of p65-p50, but overexpression of BFRF3 reduces the nuclear accumulation of p65, and the phosphorylation of p65 (Ser536) is repressed during BFRF3 transfection and EBV lytic infection, which promotes the proliferation of EBV. Overall, our study suggested that BFRF3 may play a crucial role in antiviral immunity to defend against EBV infection by inhibiting NF-κB activity.

Viral Deregulation of Apoptotic Pathways and Its Correlation With Adverse Pregnancy Outcomes.

OBJECTIVE: Our objective was to correlate parvovirus-B19 and Epstein-Barr virus (EBV) infections with apoptotic biomarker levels in tissues from placentas from spontaneous abortions and cases of elective termination of pregnancy. We also explored if viral presence could cause spontaneous abortions by trying to associate the levels of pro-apoptotic markers with adverse pregnancy outcomes. MATERIALS AND METHODS: We used 194 placental samples, of which 152 came from spontaneous abortions and were the study group and 42 controls came from cases of elective pregnancy termination. Hematoxylin and eosin (H&E) staining was performed to investigate morphological changes in the tissues, and then indirect immunohistochemistry to evaluate the expression of B19, EBV, M30, terminal deoxynucleotidyl transferase assay (TUNEL), and nuclear factor kappa B (NF-kB). Statistical analysis was performed using SPSS v. 19.0 (IBM). RESULTS: Higher levels of apoptosis were observed in the spontaneous abortion group (p<0.001) with statistical significance and their presence was also correlated with statistical significance with viral infection (p<0.001). Also, viral infections were observed only in cases of spontaneous abortion. When simple and multivariate logistic regression was performed we confirmed that viral presence remained an independent prognostic factor for high expression of all apoptotic biomarkers with statistical significance (p<0.001). CONCLUSIONS: Our results indicate that viral presence can lead to deregulation of apoptotic pathways within the maternal-fetal environment and thus work as a trigger event for spontaneous abortions.

Significance of longitudinal Epstein-Barr virus DNA combined with multipoint tumor response for dynamic risk stratification and treatment adaptation in nasopharyngeal carcinoma.

Dynamic therapy response is strongly associated with cancer outcomes. This study aimed to evaluate the significance of longitudinal Epstein-Barr virus (EBV) DNA and radiological tumor regression in risk stratification and response-adaptive treatment in locally-advanced nasopharyngeal carcinoma (LA-NPC). In total, 1,312 patients from two centers were assigned to the training and validation cohorts. Based on the multipoint examination of EBV-DNA and tumor response, four post-induction chemotherapy, four mid-radiotherapy, and four post-radiotherapy subgroups were established. Then seven phenotypes were further generated according to different permutations and combinations. These phenotypes were subsequently congregated into four response clusters, which reflect distinct biological treatment responses. The four response clusters correlated with an evident 5-year progression-free survival in both the training and external validation cohorts (5-year: training cohort 91.1%, 82.8%, 30.6%, and 10.0%; external validation 94.4%, 55.6%, 40.0%, and 12.7%) had superior prognostic performance compared to TNM staging and nomogram model (concordance index: training cohort-0.825 vs. 0.603 vs. 0.756 and external validation-0.834 vs. 0.606 vs. 0.789). Importantly, the response clusters exhibited an excellent capability in selecting candidates who can benefit from adjuvant chemotherapy. In conclusion, risk stratification based on the dynamic assessment of both radiological and biological responses can significantly enhance prognostic insights and shed light on individualized treatment modifications in LA-NPCs.

Gammaherpesvirus Infection Stimulates Lung Tumor-Promoting Inflammation.

Lung tumor-promoting environmental exposures and γherpesvirus infections are associated with Type 17 inflammation. To test the effect of γherpesvirus infection in promoting lung tumorigenesis, we infected mutant K-Ras-expressing (K-RasLA1) mice with the murine γherpesvirus MHV68 via oropharyngeal aspiration. After 7 weeks, the infected mice displayed a more than 2-fold increase in lung tumors relative to their K-RasLA1 uninfected littermates. Assessment of cytokines in the lung revealed that expression of Type 17 cytokines (Il-6, Cxcl1, Csf3) peaked at day 7 post-infection. These observations correlated with the post-infection appearance of known immune mediators of tumor promotion via IL-17A in the lungs of tumor-bearing mice. Surprisingly, Cd84, an immune cell marker mRNA, did not increase in MHV68-infected wild-type mice lacking lung tumors. Csf3 and Cxcl1 protein levels increased more in the lungs of infected K-RasLA1 mice relative to infected wild-type littermates. Flow cytometric and transcriptomic analyses indicated that the infected K-RasLA1 mice had increased Ly6Gdim/Ly6Chi immune cells in the lung relative to levels seen in uninfected control K-RasLA1 mice. Selective methylation of adenosines (m6A modification) in immune-cell-enriched mRNAs appeared to correlate with inflammatory infiltrates in the lung. These observations implicate γherpesvirus infection in lung tumor promotion and selective accumulation of immune cells in the lung that appears to be associated with m6A modification of mRNAs in those cells.

Rash decisions: Unmasking a risk phenotype in adults with persistent delayed penicillin allergy sensitized during historic infection with Epstein-Barr virus.

Background: Penicillin-associated exanthems in the setting of infectious mononucleosis caused by Epstein-Barr virus (EBV) are often viewed as a transient event, not a true allergy. Recent evidence challenges this and suggests that a notable subset of patients retain penicillin hypersensitivity. Objective: We investigated the occurrence and predictors of persistent adulthood hypersensitivity in those with penicillin-associated rash occurring in the setting of EBV infection. Methods: Retrospective analysis of data of patients referred for penicillin allergy testing to an Australian tertiary-care hospital captured from 2015 to 2023 was carried out. Results: Of 2066 patients, 23 (1%) had penicillin-associated rash during an historic EBV infection; 16 (70%) were female; and median (interquartile range) age was 18 (16-20) years at index reaction and 38 (33.5-57) years at allergy testing. Skin prick testing and delayed intradermal testing to a penicillin panel were performed, followed by oral provocation challenge in those testing negative. Persistent sensitization was shown in 6 (26%) of 23; 4 (67%) of 6 positive delayed intradermal testing; and 3 (50%) of 6 had positive oral challenge test. Notably, 5 (83%) of 6 had a severe maculopapular exanthem with facial swelling, including 2 (33%) of 6 with probable drug reaction with eosinophilia and systemic symptoms (aka DRESS) during the index reaction, compared to 0 of 17 in patients tolerating penicillin on reexposure. Conclusion: This study highlights the requirement of allergy testing in adult patients reporting a penicillin-associated severe maculopapular exanthem in the setting of EBV, even if it occurred during childhood or adolescence.

EBNA-1 antibody and autoimmune rheumatic diseases: A Mendelian Randomization Study.

Background: Numerous studies have investigated a possible correlation between Epstein-Barr virus (EBV) and autoimmune rheumatic diseases (ARDs). However, establishing a cause-and-effect relationship remains a challenging endeavor. This study employs Mendelian randomization to examine the impact of EBV nuclear antigen-1 antibody (EBNA-1) antibody levels on the susceptibility to nine distinct ARDs, including rheumatoid arthritis (RA), primary Sjogren's syndrome (PSS), systemic lupus erythematosus (SLE), undifferentiated reactive arthritis (UA), systemic sclerosis (SSc), adult-onset Still's disease (AOSD), psoriatic arthritis (PsA), dermatomyositis (DM), and ankylosing spondylitis (AS). Methods: The researchers applied a two-sample Mendelian randomization approach, utilizing online data from separate cohorts of European descent. We drew upon data from GWAS related to EBNA-1 antibody levels and the nine autoimmune-related disorders. Our primary analyses predominantly relied on the Inverse Variance Weighted methodology, complemented by a range of sensitivity assessments. Results: Our analysis revealed significant direct associations between EBNA-1 antibody levels and the risk of developing PSS (95 % CI: 0.44 to 0.85, p = 0.003), PsA (95 % CI: 0.36 to 0.99, p = 0.044), AS (95 % CI: 0.07 to 0.88, p = 0.031), and UA (95 % CI: 0.56 to 0.96, p = 0.025). These results remained consistent through comprehensive sensitivity analyses. However, no clear associations were found for the other specified conditions. Conclusions: Our findings provide compelling evidence that EBNA-1 antibody levels play a role in developing ARDs. These findings enhance our understanding of ARD pathogenesis and hold substantial promise for developing potential treatment strategies.

Epstein-Barr Virus BARF1 Is Expressed in Lung Cancer and Is Associated with Cancer Progression.

BACKGROUND: Epstein-Barr virus (EBV) is involved in the development of lymphomas, nasopharyngeal carcinomas (NPC), and a subgroup of gastric carcinomas (GC), and has also been detected in lung carcinomas, even though the role of the virus in this malignancy has not yet been established. BamH1-A Rightward Frame 1 (BARF1), a suggested exclusive epithelial EBV oncoprotein, is detected in both EBV-associated GCs (EBVaGC) and NPC. The expression and role of BARF1 in lung cancer is unknown. METHODS: A total of 158 lung carcinomas including 80 adenocarcinomas (AdCs) and 78 squamous cell carcinomas (SQCs) from Chilean patients were analyzed for EBV presence via polymerase chain reaction (PCR), Immunohistochemistry (IHC), or chromogenic in situ hybridization (CISH). The expression of BARF1 was evaluated using Reverse Transcription Real-Time PCR (RT-qPCR). Additionally, A549 and BEAS-2B lung epithelial cells were transfected with a construct for ectopic BARF1 expression. Cell proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) were evaluated. RESULTS: We found that EBV was present in 37 out of 158 (23%) lung carcinomas using PCR. Considering EBV-positive specimens using PCR, IHC for Epstein-Barr nuclear antigen 1 (EBNA1) detected EBV in 24 out of 30 (80%) cases, while EBERs were detected using CISH in 13 out of 16 (81%) cases. Overall, 13 out of 158 (8%) lung carcinomas were shown to be EBV-positive using PCR/IHC/CISH. BARF1 transcripts were detected in 6 out of 13 (46%) EBV-positive lung carcinomas using RT qPCR. Finally, lung cells ectopically expressing BARF1 showed increased migration, invasion, and EMT. CONCLUSIONS: EBV is frequently found in lung carcinomas from Chile with the expression of BARF1 in a significant subset of cases, suggesting that this viral protein may be involved in EBV-associated lung cancer progression.

Epstein-Barr virus induces host shutoff extensively via BGLF5-independent mechanisms.

Epstein-Barr virus (EBV) is a ubiquitous oncogenic virus associated with multiple cancers and autoimmune diseases. Unlike most herpesviruses, EBV reactivation from latency occurs asymptomatically, allowing it to spread efficiently to other hosts. However, available models are limited by the inefficient and asynchronous reactivation from latency into lytic replication. To address this problem, we develop a dual-fluorescent lytic reporter (DFLR) EBV that specifically labels cells in the early and late stages of replication. Using lymphoblastoid cell lines transformed by DFLR EBV as a model for EBV reactivation in B cells, we observe extensive reprogramming of the host cell transcriptome during lytic cycle progression. This includes widespread shutoff of host gene expression and disruption of mRNA processing. Unexpectedly, host shutoff remains extensive even in cells infected with DFLR EBV deleted for the BGLF5 nuclease. These findings implicate BGLF5-independent mechanisms as the primary drivers of host transcriptome remodeling during EBV lytic replication.

Epstein-Barr virus-positive, primary cutaneous marginal zone lymphoma, with transformation: Case report and review of the literature.

Epstein-Barr Virus (EBV) positive primary cutaneous marginal zone lymphoma (PCMZL) is uncommon and subsequent transformation is rare. METHODS: We report a patient with EBV positive PCMZL with subsequent transformation to plasmablastic lymphoma and review the literature for transformed PCMZL to assess clinical and pathologic characteristics. In the case we describe, the patient presented with multifocal PCMZL, developed large B cell transformation with plasmacytic differentiation, followed by plasmablastic transformation (PBL), and ultimately died of disease progression despite multiple lines of therapy. Past history was significant for psoriatic arthritis (multiple prior lines of immunomodulatory therapy). The lymphomas and non-involved bone marrow share the same somatic DNMT3A and TET2 mutations, suggesting clonal relatedness and an association with clonal hematopoiesis (CH). RESULTS: Eighteen cases complied the cohort (seventeen cases from the literature and the case reported herein). Nearly half of the eighteen cases of PCMZL with transformation died of progressive disease (44%). Transformed cases were more commonly seen in patients with >2 sites at initial diagnosis. EBV was assessed in 5 patients, 3 were positive (all died of disease). Two patients with NGS studies demonstrated TET2 and DNMT3A mutations. CONCLUSIONS: Transformation of EBV positive PCMZL appears to be a poor prognostic indicator, with our reported case being the first well defined case transformed to PBL, suspected to arise from myeloid-CH.

Metabolic regulation of the tumour and its microenvironment: The role of Epstein-Barr virus.

The Epstein-Barr virus (EBV), the first identified human tumour virus, infects over 95% of the individuals globally and has the potential to induce different types of cancers. It is increasingly recognised that EBV infection not only alters cellular metabolism, contributing to neoplastic transformation, but also utilises several non-cell autonomous mechanisms to shape the metabolic milieu in the tumour microenvironment (TME) and its constituent stromal and immune cells. In this review, we explore how EBV modulates metabolism to shape the interactions between cancer cells, stromal cells, and immune cells within a hypoxic and acidic TME. We highlight how metabolites resulting from EBV infection act as paracrine factors to regulate the TME, and how targeting them can disrupt barriers to immunotherapy.

Epstein-Barr virus replication within differentiated epithelia requires pRb sequestration of activator E2F transcription factors.

Epstein-Barr virus (EBV) co-infections with human papillomavirus (HPV) have been observed in oropharyngeal squamous cell carcinoma. Modeling EBV/HPV co-infection in organotypic epithelial raft cultures revealed that HPV16 E7 inhibited EBV productive replication through the facilitated degradation of the retinoblastoma protein pRb/p105. To further understand how pRb is required for EBV productive replication, we generated CRISPR-Cas9 pRb knockout (KO) normal oral keratinocytes (NOKs) in the context of wild-type and mutant K120E p53. EBV replication was examined in organotypic rafts as a physiological correlate for epithelial differentiation. In pRb KO rafts, EBV DNA copy number was statistically decreased compared to vector controls, regardless of p53 context. Loss of pRb did not affect EBV binding or internalization of calcium-treated NOKs or early infection of rafts. Rather, the block in EBV replication correlated with impaired immediate early gene expression. An EBV infection time course in rafts with mutant p53 demonstrated that pRb-positive basal cells were initially infected with delayed replication occurring in differentiated layers. Loss of pRb showed increased S-phase progression makers and elevated activator E2F activity in raft tissues. Complementation with a panel of pRb/E2F binding mutants showed that wild type or pRb∆685 mutant capable of E2F binding reduced S-phase marker gene expression, rescued EBV DNA replication, and restored BZLF1 expression in pRb KO rafts. However, pRb KO complemented with pRb661W mutant, unable to bind E2Fs, failed to rescue EBV replication in raft culture. These findings suggest that EBV productive replication in differentiated epithelium requires pRb inhibition of activator E2Fs to restrict S-phase progression.IMPORTANCEA subset of human papillomavirus (HPV)-positive oropharyngeal squamous cell carcinoma is co-positive for Epstein-Barr virus (EBV). Potential oncogenic viral interactions revealed that HPV16 E7 inhibited productive EBV replication within the differentiated epithelium. As E7 mediates the degradation of pRb, we aimed to establish how pRb is involved in EBV replication. In the context of differentiated epithelium using organotypic raft culture, we evaluated how the loss of pRb affects EBV lytic replication to better comprehend EBV contributions to carcinogenesis. In this study, ablation of pRb interfered with EBV replication at the level of immediate early gene expression. Loss of pRb increased activator E2Fs and associated S-phase gene expression throughout the differentiated epithelium. Complementation studies showed that wild type and pRb mutant capable of binding to E2F rescued EBV replication, while pRb mutant lacking E2F binding did not. Altogether, these studies support that in differentiated tissues, HPV16 E7-mediated degradation of pRb inhibits EBV replication through unregulated E2F activity.