Epstein-Barr Virus-associated Hemophagocytic Lymphohistiocytosis Research Articles

Highly sensitive detection of Epstein-Barr virus-infected cells by EBER flow FISH.

When Epstein-Barr virus (EBV) infection is suspected, identification of infected cells is important to understand the pathogenesis, determinine the treatment strategy, and predict the prognosis. We used the PrimeFlow™ RNA Assay Kit with a probe to detect EBV-encoded small RNAs (EBERs) and multiple surface markers, to identify EBV-infected cells by flow cytometry. We analyzed a total of 24 patients [11 with chronic active EBV disease (CAEBV), 3 with hydroa vacciniforme lymphoproliferative disorder, 2 with X-linked lymphoproliferative disease type 1 (XLP1), 2 with EBV-associated hemophagocytic lymphohistiocytosis, and 6 with posttransplant lymphoproliferative disorder (PTLD)]. We compared infected cells using conventional quantitative PCR methods and confirmed that infected cell types were identical in most patients. Patients with CAEBV had widespread infection in T and NK cells, but a small amount of B cells were also infected, and infection in patients with XLP1 and PTLD was not limited to B cells. EBV-associated diseases are believed to be complex pathologies caused by EBV infecting a variety of cells other than B cells. We also demonstrated that infected cells were positive for HLA-DR in patients with CAEBV. EBER flow FISH can identify EBV-infected cells with high sensitivity and is useful for elucidating the pathogenesis.

Characteristics of Virology and Immune Inflammation of Epstein-Barr Virus Infection Related Non-Neoplastic Diseases in Children.

The goal was to study the difference of virological, immunologic, and inflammatory indicators between Epstein-Barr associated infectious mononucleosis (EBV-IM) and EBV associated hemophagocytic lymphohistiocytosis (EBV-HLH) and to explore the evaluation indicators for monitoring the therapeutic efficacy of EBV-HLH. Twenty children with EBV-IM (IM group) and 10 children with EBV-HLH (HLH group) were selected. Virology indicators were detected; the absolute count of lymphocyte, and lymphocyte subsets were detected; the levels of immunoglobulin and ferritin were assayed. Compared to the IM group, the HLH group showed a decrease in EBV-specific VCA-IgM antibody levels (U = 29.0, p = 0.006) and an increase in EBV-specific NA-IgG antibody levels (U = 17.0, p = 0.001), while there was no significant difference in EB-DNA loads (t = 0.417, p = 0.680). The counts of lymphocytes, and various lymphocyte subsets in the HLH group were lower than those in the IM group. Inflammatory markers in the HLH group were significantly higher than those in IM group. Dynamic monitoring of virological, immunological, and inflammatory indicators in HLH patients during treatment showed that EBV DNA gradually decreased in patients with good prognosis. Inflammatory indicators significantly decreased and returned to normal, lymphocyte count significantly increased and returned to normal during treatment. However, patients with poor prognosis showed rebound increase in EBV DNA and inflammatory indicators in the later stage of treatment, while lymphocyte count further decreased with the recurrence of the disease. Exhausted and damaged immune function in host by persistent stimulation of EB viral antigen is one of the main pathogeneses of EB-HLH. Lymphocyte count and serum ferritin level are effective indicators to monitor the therapeutic efficacy during the treatment to HLH.

Combined Use of Emapalumab With Ruxolitinib and Dexamethasone as an Effective Treatment for Epstein-Barr Virus-Associated Hemophagocytic Lymphohistiocytosis Complicated With Multiorgan Damage and Severe Infection.

Anti-interferon-γ monoclonal antibody emapalumab and JAK1/2 inhibitors ruxolitinib have been widely reported for the treatment of hemophagocytic lymphohistiocytosis (HLH) recently. These targeted drugs have fewer side effects and may provide new options for patients with HLH who are refractory to previous treatment or intolerant to chemotherapy. Herein, we reported a case of Epstein-Barr virus-related HLH, which did not respond well to HLH-94 plus ruxolitinib and developed severe fungal infection. The disease was successfully controlled after a combination therapy of emapalumab, ruxolitinib, and dexamethasone.

Single-Cell Transcriptomic Analysis of Epstein-Barr Virus-Associated Hemophagocytic Lymphohistiocytosis.

Epstein-Barr virus (EBV) infection can lead to infectious mononucleosis (EBV-IM) and, more rarely, EBV-associated hemophagocytic lymphohistiocytosis (EBV-HLH), which is characterized by a life-threatening hyperinflammatory cytokine storm with immune dysregulation. Interferon-gamma (IFNγ) has been identified as a critical mediator for primary HLH; however, the detailed role of IFNγ and other cytokines in EBV-HLH is not fully understood. In this study, we used single-cell RNA sequencing to characterize the immune landscape of EBV-HLH and compared it with EBV-IM. Three pediatric patients with EBV-HLH with different backgrounds,one with X-linked lymphoproliferative syndrome type 1 (XLP1), two with chronic active EBV disease (CAEBV), and two patients with EBV-IM were enrolled. The TUBA1B + STMN1 + CD8 + T cell cluster, a responsive proliferating cluster with rich mRNA detection, was explicitly observed in EBV-IM, and the upregulation of SH2D1A-the gene responsible for XLP1-was localized in this cluster. This proliferative cluster was scarcely observed in EBV-HLH cases. In EBV-HLH cases with CAEBV, upregulation of LAG3 was observed in EBV-infected cells, which may be associated with an impaired response by CD8 + T cells. Additionally, genes involved in type I interferon (IFN) signaling were commonly upregulated in each cell fraction of EBV-HLH, and activation of type II IFN signaling was observed in CD4 + T cells, natural killer cells, and monocytes but not in CD8 + T cells in EBV-HLH. In conclusion, impaired responsive proliferation of CD8 + T cells and upregulation of type I IFN signaling were commonly observed in EBV-HLH cases, regardless of the patients' background, indicating the key features of EBV-HLH.

Secondary hemophagocytic lymphohistiocytosis in pediatric patients with visceral leishmaniasis and Epstein-Barr virus infection.

Visceral leishmaniasis-associated hemophagocytic lymphohistiocytosis (VL-HLH) is indistinguishable from those of HLH of other etiologies due to the overlap symptoms, posing a serious threat to life. In this study, we aimed to provide insights for early diagnosis and improve outcomes in pediatric patients with VL-HLH. We retrospectively analyzed the clinical and laboratory data of 10 pediatric patients with VL-HLH and 58 pediatric patients with Epstein-Barr virus-associated hemophagocytic lymphohistiocytosis (EBV-HLH). The median time from symptom onset to cytopenia in patients with VL-HLH and EBV-HLH was 11 days (interquartile range, 7-15 days) and five days (interquartile range, 3.75-9.25 days) (P = 0.005). Both groups showed liver injury and increased lactate dehydrogenase levels; however the levels of aspartate aminotransferase, alanine aminotransferase, direct bilirubin, and lactate dehydrogenase in patients with VL-HLH were significantly lower than those in patients with EBV-HLH (P < 0.05). The fibrinogen and triglyceride levels were almost normal in VL-HLH patients but were significantly altered in EBV-HLH cases ( P < 0.05). The positive rate of first bone marrow microscopy examination, anti-rK39IgG detection, and blood metagenomic next-generation sequencing was 50%, 100%, and 100%, respectively. After VL diagnosis, eight patients were treated with sodium stibogluconate and two were treated with liposomal amphotericin B. All the patients with VL-HLH recovered. Our study demonstrates that regular triglyceride and fibrinogen levels in pediatric patients with VL-HLH may help in differential diagnosis from EBV-HLH. VL-HLH is milder than EBV-HLH, with less severe liver injury and inflammatory responses, and timely treatment with antileishmanial agents is essential to improve the outcomes of pediatric patients with VL-HLH.

Novel plasma microRNA expression features in diagnostic use for Epstein-Barr virus-associated febrile diseases

BackgroundEpstein-Barr virus (EBV) is widely infected in humans and causes various diseases. Among them, microRNAs of EBV play a key role in the progression of EBV-associated febrile diseases. There're few specific indicators for rapid differential diagnosis of various febrile diseases associated with EBV, and the lack of more reliable screening methods with high diagnostic utility has led to spaces for improvement in the accurate diagnosis and efficient treatment of relevant patients, making EBV infection a complicated clinical problem. With recent advances in plasma microRNA testing, the apparent presence of EBV microRNAs in plasma can help screen for EBV infection. The gene networks targeted by these microRNAs can also indicate potential biomarkers of EBV-associated febrile diseases. This study aimed to identify some novel miRNAs as potential biomarkers for early diagnosis of respectively EBV-associated febrile diseases. Materials and methodsA total of 110 participants were recruited for this task. First, we performed high-throughput sequencing and preliminary PCR validation of differentially expressed miRNAs in 15 participants with EBV-associated fever (divided into 5 groups, common EBV carriers, infectious mononucleosis (IM) and chronic active EBV infection (CAEBV), EBV-associated Hemophagocytic Lymphohistiocytosis (EBV-HLH), and 3 healthy individuals). After a comprehensive analysis, 10 miRNAs with abnormal expression were screened, and then qRT-PCR was performed in the rest of 95 participants to detect the validation of miRNAs expression in plasma samples. Thereafter, we further investigated their potential for clinical application in EBV-related febrile diseases by using a combination of Gene Ontology analysis, Kyoto Encyclopedia of Genes and Genomes pathway analysis, and Protein-protein interaction network analysis. ResultsThrough identification and detailed analysis of the obtained data, we found significant differences in the expression of Hsa-miR-320d, EBV-miR-BART22, and EBV-miR-BART2-3p in blood samples from patients with different EBV-related febrile diseases. We found that the expression levels of Hsa-miR-320d, EBV-miR-BART22, and EBV-miR-BART2-3p in plasma are indicative of determining different disease types of EBV-related febrile diseases, while EBV-miR-BART22 and EBV-miR-BART2-3p may be potential therapeutic targets. ConclusionThe expression levels of Hsa-miR-320d, EBV-miR-BART22, and EBV-miR-BART2-3p suggest that they may be used as transcriptional features for early differential diagnosis of EBV-related febrile diseases, and EBV-miR-BART22 and EBV-miR-BART2-3p may be potential therapeutic targets.

Comparison of plasma proteomic profiles of patients with Epstein-Barr virus-associated hemophagocytic lymphohistiocytosis and infectious mononucleosis.

Primary Epstein-Barr virus (EBV) infection occasionally causes EBV-infectious mononucleosis (EBV-IM) and EBV-hemophagocytic lymphohistiocytosis (EBV-HLH). Although EBV-IM is mostly mild and self-limiting, EBV-HLH is a life-threatening disease characterized by excessive immune activation. However, the pathogenesis of EBV-HLH is yet to be fully elucidated. A diagnostic biomarker for EBV-HLH is desirable because early diagnosis and treatment are critical for the effective management of patients. In this study, the proteomic profiling of plasma was performed using liquid chromatography-mass spectrometry to identify proteins specific to EBV-IM and EBV-HLH. Furthermore, pathway analysis was performed for the proteins upregulated in patients with EBV-IM and EBV-HLH. Compared to healthy controls, 63 and 18 proteins were upregulated in patients with EBV-IM and EBV-HLH, respectively. Pathway and process enrichment analyses revealed that the complement system was the most enriched category of upregulated proteins in EBV-IM, whereas proteins related to immune effector processes were the most enriched in EBV-HLH. Among the 18 proteins upregulated in EBV-HLH, seven were exclusive to EBV-HLH. These specific proteins were associated with three pathways, and apolipoprotein E was commonly found in all the pathways. Proteomic analysis may provide new insights into the host response to EBV infection and the pathogenesis of EBV-related diseases.

Caebv Disease Originates from Infected Hematopoietic Stem Cells

Background Chronic active Epstein-Barr Virus (EBV) disease (CAEBV) is lethal syndrome due to persistent EBV infection. When diagnosed as CAEBV, EBV infection was observed in multiple hematopoietic lineages, but the etiology of CAEBV is still elusive. Methods Bone marrow and peripheral cells derived from five CAEBV patients, one EBV-associated hemophagocytic lymphohistiocytosis (EBV-HLH) patient, and two healthy controls were analyzed. Multiple assays were applied to identify and characterize EBV-infected cells, including quantitative PCR (qPCR), PrimeFlow and single-cell RNA-sequencing (scRNA-seq). Based on scRNA-seq data, alterations in gene expression of particular cell types were analyzed between CAEBV patients and controls, and between infected and uninfected cells. One CAEBV patient was treated with allogeneic hematopoietic stem cell transplantation (HSCT), and the samples derived from this patient were analyzed again six month after HSCT. Results EBV infected the full spectrum of the hematopoietic system including both lymphoid and myeloid lineages, so as hematopoietic stem cells (HSCs) in the CAEBV patients. EBV-infected HSCs exhibited a higher differentiation rate towards downstream lineages, and the EBV infection had an impact on both the innate and adaptive immunity, resulting in inflammatory symptoms. EBV infected cells were thoroughly removed from the hematopoietic system after HSCT. Conclusions Token together,multiple lines of evidence presented in this study suggest that CAEBV disease originates from the infected hematopoietic stem cells. Figure 1. EBV-infected HSCs were detected in CAEBV patients with multiple assays.

Single-Cell RNA Sequencing Reveals Contradictory Immune Microenvironment in Hemophagocytic Syndrome

Hemophagocytic lymphohistiocytosis (HLH), a formidable and rare clinical syndrome impacting multiple organs, manifests as a perilous condition. However, the intricate pathophysiological mechanisms underlying HLH remain yet to be fully elucidated. In this study, we obtained peripheral blood samples from 9 HLH patients spanning the years 2018 to 2022. Employing single-cell RNA sequencing (scRNA-seq), we conducted a comprehensive analysis of the alterations in the immune microenvironment, delving into the role of EBV infection in HLH and its influence on pathogenesis. By comparing with health donor peripheral blood , we identified a total of 11 significant immune cell populations among 72,779 single-cell transcriptomes, comprising 20,337 cells from healthy controls and 52,442 cells from HLH patients. Notably, we found that the proportion of T cells increased significantly, and other immune cells decreased in HLH patients. In terms of function, we found that the overall T cell killing ability was weakened, and the factor secretion ability was enhanced. Concerning myeloid cells, the impact of CD16+ monocytes on T cells exhibited a pronounced increase, along with an enhanced ability to secrete IL-18 factors, potentially serving as the main source of cells promoting T cell function. Different subpopulations of T cells have significant changes in the pathogenesis of HLH. Remarkably, both CD4+ T cells and CD8+ T cells exhibited HLA-DR+GZMK+ cell subgroups with upregulated IFN-γ production and response, suggesting their association with the hyperinflammatory state in HLH. A correlation analysis between the proportion of HLA-DRA+ GZMK+ T cells and the severity score revealed a significant correlation (p&amp;lt;0.05), with the ROC curve further affirming their predictability in assessing high inflammatory activity. Among CD4+ T cells, the proportion of Treg cells and the secretion of inhibitory factors such as TGFb1 decreased. Additionally, we noted the presence of a functionally impaired subset of CBLB+CD8+ T cells, indicating the considerable variability in T cell subset functionality in HLH. Next, a comprehensive analysis of the alterations in myeloid cells, B cells, and NK cells in HLH patients was conducted. We observed a substantial reduction in the number of DC cells and immune-related functions within various myeloid cell subsets. Regarding B cells, an increase in anergy B cells was observed, whereas the population of plasma cells, an effective B cell type, exhibited a decrease. In the context of NK cells, there was a significant increase in the proportion of NK cell subsets associated with IFN-γ secretion. Moreover, EBV-infected T cells in EBV-associated HLH were identified as the principal source of IFN-γ. Through single-cell EBV-CALL analysis, we found that IFN-γ secretion was primarily attributed to EBV-infected T cells, subsequently influencing CD16+ monocytes. Additionally, IL-16 production was predominantly observed in EBV-infected CD16+ monocytes, thereby impacting lymphocytes. Moreover, Monocytes were stimulated by EBV-infected CD4+ T cells and Atypical T cells, leading to the secretion of a pro-inflammatory factors such as MIF. Furthermore, our investigation unveiled that EBNA2 could promote MYC into the nucleus and enhance the proliferation and IFN-γ secretion of T cells. In conclusion, our comprehensive analysis of the immune microenvironment changes following the onset of HLH has yielded valuable insights and novel perspectives for understanding the underlying mechanisms and potential clinical treatments of this condition.

Clinical and Pathological Characteristics of Severe EBV Infection in Japanese Children; a Disease Spectrum from Acute Liver Failure to Hemophagocytic Lymphohistiocytosis

Abstract Background Epstein-Barr virus (EBV) is a ubiquitous herpesvirus known as a cause of infectious mononucleosis (IM). While most EBV infection is mild and self-limited, severe EBV diseases such as severe IM, EBV-associated hemophagocytic lymphohistiocytosis (HLH) and EBV-associated acute liver failure (ALF) may occur on rare occasions. Interestingly, the severe EBV disease with ALF may occasionally present with overlapping features of HLH; however, it remains unknown if the presence or absence of HLH reflects a distinctive underlying pathophysiology of EBV infection. To investigate the clinical and pathological characteristics of EBV-associated ALF among children, we conducted a single-center, retrospective study at the largest tertiary children’s hospital in Japan. Methods We enrolled children under 18 years old who required medical management in the intensive care unit due to primary EBV infection between January 2010 and April 2021. Patients with underlying immunodeficiencies and other medical conditions were excluded. Clinical data, type of EBV-infected cell lineage, and pathological exam were reviewed. Fourteen patients with EBV-associated ALF were identified in the initial screening, and two were excluded (due to underlying medical conditions). Thus, the details of 12 patients were further analyzed. Five patients fulfilled the criteria for HLH, while seven presented as isolated ALF only. Results The most common symptoms of these patients include fever (92%), vomiting (58%), and consciousness disturbance (100%). EBV loads were higher when HLH co-existed (median 2,700 copies/μg of DNA vs. 130,000 copies/μg of DNA, p = 0.01). Identification of EBV-infected cells revealed that the most common cell type was CD8+T cells (8/12, 80%), followed by CD3+T cells (1/12, 10%), and CD19+ cells (2/12, 20%). EBV infected in both CD8+T and CD19+ cells in one case and EBV-infected cells were not identified in two cases. In addition, an increase of activated CD8+ T cells (CD3+CD8+HLA-DR+) was observed in both HLH and ALF cases. Among 12 patients with ALF, seven patients underwent life-saving liver transplantation (LT) due to ALF. Of five patients with ALF+HLH, all patients received corticosteroid, four received cyclosporin or etoposide, and one underwent hematopoietic stem cell transplantation (HSCT). Overall mortality is 17% (2/12 patients), and both patients were complicated with HLH. Conclusion This study suggests that HLH and ALF can be an indistinguishable spectrum of severe EBV disease, and the underlying pathogenesis may be associated with the unusual type of EBV-infected cells. Hence, the identification of EBV-infected cells should be considered to guide the diagnosis and treatment. Our study also revealed that LT and cytotoxic anti-inflammatory therapy effectively treated patients with EBV-associated ALF with or without HLH, given the high mortality (&amp;gt;70%) of this disease spectrum. Further studies investigating EBV-associated ALF pathology are warranted to determine optimal management.

Comparative study of biomarkers for the early identification of Epstein–Barr virus-associated hemophagocytic lymphohistiocytosis in infectious mononucleosis

Background and aimEpstein-Barr virus-associated hemophagocytic lymphohistiocytosis (EBV-HLH) and infectious mononucleosis (EBV-IM) share mimic symptoms in the early stages of childhood development. We aimed to examine the clinical features and laboratory indices of these two diseases in children and uncover unique indicators to assist pediatricians in identifying these diseases early.MethodsWe collected clinical data from 791 pediatric patients diagnosed with EBV-IM or EBV-HLH, compared the clinical traits and laboratory biomarkers presented in the two groups, and constructed predictive models based on them.ResultsPatients with EBV-IM had greater ratios of cervical lymphadenopathy, eyelid edema, and tonsillitis, whereas individuals with EBV-HLH were more likely to have hepatomegaly and splenomegaly. When using the criteria of interleukin (IL)-10 > 89.6 pg/mL, interferon (IFN)-γ > 45.6 pg/mL, ferritin > 429 μg/L, D-dimer > 3.15 mg/L and triglycerides > 2.1 mmol/L, the sensitivity was 87.9%, 90.7%, 98.1%, 91.1% and 81.5% to predict EBV-HLH, while the specificity was 98.4%, 96.3%, 96.5%, 94.1% and 80.6%, respectively. A logistic regression model based on four parameters (IL-10, ferritin, D-dimer, and triglycerides) was established to distinguish EBV-HLH patients from EBV-IM patients, with a sensitivity of 98.0% and a specificity of 98.2%.ConclusionsIL-10, IFN-γ, ferritin and D-dimer levels are significantly different between EBV-HLH and EBV-IM. Predictive models based on clinical signs and laboratory findings provide simple tools to distinguish the two situations.

Chronic active Epstein-Barr virus disease originates from infected hematopoietic stem cells

AbstractChronic active Epstein-Barr virus (EBV) disease (CAEBV) is a lethal syndrome because of persistent EBV infection. When diagnosed as CAEBV, EBV infection was observed in multiple hematopoietic lineages, but the etiology of CAEBV is still elusive. Bone marrow and peripheral cells derived from 5 patients with CAEBV, 1 patient with EBV-associated hemophagocytic lymphohistiocytosis, and 2 healthy controls were analyzed. Multiple assays were applied to identify and characterize EBV-infected cells, including quantitative polymerase chain reaction, PrimeFlow, and single-cell RNA-sequencing (scRNA-seq). Based on scRNA-seq data, alterations in gene expression of particular cell types were analyzed between patients with CAEBV and controls, and between infected and uninfected cells. One patient with CAEBV was treated with allogeneic hematopoietic stem cell transplantation (HSCT), and the samples derived from this patient were analyzed again 6 months after HSCT. EBV infected the full spectrum of the hematopoietic system including both lymphoid and myeloid lineages, as well as the hematopoietic stem cells (HSCs) of the patients with CAEBV. EBV-infected HSCs exhibited a higher differentiation rate toward downstream lineages, and the EBV infection had an impact on both the innate and adaptive immunity, resulting in inflammatory symptoms. EBV-infected cells were thoroughly removed from the hematopoietic system after HSCT. Taken together, multiple lines of evidence presented in this study suggest that CAEBV disease originates from the infected HSCs, which might potentially lead to innovative therapy strategies for CAEBV.

An analysis of sintilimab combined with ruxolitinib as compassionate therapy for 12 adults with EBV-associated hemophagocytic lymphohistiocytosis.

Epstein-Barr virus-associated hemophagocytic lymphohistiocytosis (EBV-HLH) is a severe hyperinflammatory illness that affects adults and is caused by an EBV infection. Without allogeneic hematopoietic stem cell transplantation (allo-HSCT), the overall survival of adult patients with EBV-HLH is unsatisfactory, necessitating the development of innovative therapeutic approaches. The clinical records of twelve EBV-HLH patients who received sintilimab therapy combined with ruxolitinib on a compassionate basis at the First Affiliated Hospital of Soochow University were retrospectively examined in this investigation. All the patients responded without fever, but three patients relapsed within a week. Among the nine patients achieving complete response (CR), 55.6% (5/9) maintained CR for >4.5 months, and 33.3% (3/9) relapsed following CR. Neither patients with no response (NR) nor relapsed patients were fit for allo-HSCT, and all died soon after discharge. Six patients had clinical CR with a median follow-up of 5 (4.4-14.7) months. There were no documented severe negative effects. Additional information on this innovative treatment for adult EBV-HLH is provided in our report.

Sintilimab treatment for chronic active Epstein–Barr virus infection and Epstein–Barr virus-associated hemophagocytic lymphohistiocytosis in children

BackgroundChronic active Epstein–Barr virus infection (CAEBV) and Epstein–Barr virus-associated hemophagocytic lymphohistiocytosis (EBV-HLH) are rare but life-threatening progressive diseases triggered by EBV infection. Glucocorticoid/immunosuppressants treatment is temporarily effective; however, most patients relapse and/or progress. Hematopoietic stem cell transplantation (HSCT) is a potentially curative therapy; however, there are risks of transplantation-associated complications. Currently there is no standard treatment for CAEBV and EBV-HLH. Programmed death protein 1 (PD-1) inhibitors have achieved a high response in many EBV-related diseases. Sintilimab (a recombinant human IgG4 monoclonal antibody against PD-1) disrupts the interaction between PD-1 and its ligand, leading to T cell reinvigoration.MethodsA retrospective analysis was performed on three children with CAEBV or EBV-HLH in the Children’s Hospital of Soochow University between 12 December 2020 and 28 November 2022. The efficacy of sintilimab was evaluated.ResultsThree patients, including two males and one female, were analyzed. Among them, two children were diagnosed with CAEBV with intermittent fever for more than four years, and one child was diagnosed with EBV-HLH. After sintilimab treatment and a mean follow-up of 17.1 months (range 10.0–23.3 months), patients 1 and 3 achieved a complete clinical response and patient 2 achieved a partial clinical response. All three children showed a > 50% decrease in EBV-DNA load in both blood and plasma. EBV-DNA copies in sorted T, B, and NK cells were also markedly decreased after sintilimab treatment.ConclusionOur data supported the efficacy of PD-1 targeted therapy in certain patients with CAEBV and EBV-HLH, and suggested that sintilimab could provide a cure for these diseases, without HSCT. More prospective studies and longer follow-up are needed to confirm these conclusions.

T cell-mediated immunity during Epstein-Barr virus infections in children.

Epstein-Barr virus (EBV) infection is extremely common worldwide, with approximately 90% of adults testing positive for EBV antibodies. Human are susceptible to EBV infection, and primary EBV infection typically occurs early in life. EBV infection can cause infectious mononucleosis (IM) as well as some severe non-neoplastic diseases, such as chronic active EBV infection (CAEBV) and EBV-associated hemophagocytic lymphohistiocytosis (EBV-HLH), which can have a heavy disease burden. After primary EBV infection, individuals develop robust EBV-specific T cell immune responses, with EBV-specific CD8+ and part of CD4+ T cells functioning as cytotoxic T cells, defending against virus. Different proteins expressed during EBV's lytic replication and latent proliferation can cause varying degrees of cellular immune responses. Strong T cell immunity plays a key role in controlling infection by decreasing viral load and eliminating infected cells. However, the virus persists as latent infection in EBV healthy carriers even with robust T cell immune response. When reactivated, it undergoes lytic replication and then transmits virions to a new host. Currently, the relationship between the pathogenesis of lymphoproliferative diseases and the adaptive immune system is still not fully clarified and needs to be explored in the future. Investigating the T cell immune responses evoked by EBV and utilizing this knowledge to design promising prophylactic vaccines are urgent issues for future research due to the importance of T cell immunity.

Diagnostic and disease severity determination criteria for hydroa vacciniforme lymphoproliferative disorders and severe mosquito bite allergy.

Hydroa vacciniforme lymphoproliferative disorder (HV-LPD) and severe mosquito bite allergy (SMBA) are both cutaneous forms of Epstein-Barr virus (EBV)-associated T/natural killer (NK) cell LPDs and are closely related to chronic active EBV disease (CAEBV) and EBV-associated hemophagocytic lymphohistiocytosis (EBV-HLH). HV-LPD is further divided into classic HV, a benign subtype mediated by EBV-positive γδT cells, and systemic HV, another life-threatening subtype mainly associated with EBV-positive αβT or γδT cells. The vast majority of patients with SMBA have increased numbers of EBV-infected NK cells in the blood. Clinical symptoms of HV-LPD and SMBA often overlap in the same patient and may progress to more serious disease conditions equivalent to the systemic form of CAEBV. To define the disease spectrum of HV-LPD and SMBA, we propose the diagnostic criteria and the determination criteria for disease severity. The proposed diagnostic criteria are consistent with those for CAEBV and EBV-HLH in the guidelines for the management for CAEBV and related disorders 2023.

Stevens-Johnson Syndrome complicated by Epstein-Barr virus-associated hemophagocytic lymphohistiocytosis in a child.

Stevens-Johnson Syndrome complicated by Epstein-Barr virus-associated hemophagocytic lymphohistiocytosis in a child.

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.

Combination of gemcitabine, pegaspargase, etoposide, and dexamethasone (GPED) in treatment of advanced extranodal NK/T-cell lymphoma.

Combination of gemcitabine, pegaspargase, etoposide, and dexamethasone (GPED) in treatment of advanced extranodal NK/T-cell lymphoma.

Hemophagocytic Lymphohistiocytosis: an Under-recognized and Life-threatening Condition.

Diagnosis of hemophagocytic lymphohistiocytosis is a challenge in Nepal because of limited resources and the high prevalence of tropical febrile illness mimicking hemophagocytic lymphohistiocytosis. We retrospectively reviewed medical records of 21 patients who were diagnosed with hemophagocytic lymphohistiocytosis from 2010 to 2015 at a single center in Nepal. Two patients had a mutation in their perforin gene and underwent successful haploidentical stem cell transplantation. Marrow hemophagocytosis was found only in 57% of the patients. Five patients had hematological malignancy and were treated with disease-specific chemotherapy. Seven patients developed hemophagocytic lymphohistiocytosis secondary to an infection, including visceral leishmaniasis, scrub typhus, and Epstein Barr virus. EBV-associated hemophagocytic lymphohistiocytosis was refractory to hemophagocytic lymphohistiocytosis 94 protocol, including the addition of rituximab. Malignancy and infection-associated hemophagocytic lymphohistiocytosis was more common. The most common clinical presentations included fever, splenomegaly, hyponatremia, liver function derangement, hyperfibrinogenemia, hyperferritinemia, and cytopenia. With a mortality of 29% in our study cohort, hemophagocytic lymphohistiocytosis should be considered a lethal disease, and clinicians should maintain a high index of suspicion to diagnose this disease. Keywords: Hemophagocytic lymphohistiocytosis; infection; malignancy.