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Background: Hemophagocytic Lymphohistiocytosis (HLH) carries a high mortality rate. Current existing risk-evaluation methodologies fall short and improved predictive methods are needed. This study aimed to forecast 30-day mortality in adult HLH patients using 11 distinct machine learning (ML) algorithms., Methods: A retrospective analysis on 431 adult HLH patients from January 2015 to September 2021 was conducted. Feature selection was executed using the least absolute shrinkage and selection operator. We employed 11 ML algorithms to create prediction models. The area under the curve (AUC), sensitivity, specificity, positive predictive value, negative predictive value, F1 score, calibration curve and decision curve analysis were used to evaluate these models. We assessed feature importance using the SHapley Additive exPlanation (SHAP) approach., Results: Seven independent predictors emerged as the most valuable features. An AUC between 0.65 and 1.00 was noted among the eleven ML algorithms. The gradient boosting decision tree (GBDT) algorithms demonstrated the most optimal performance (1.00 in the training cohort and 0.80 in the validation cohort). By employing the SHAP method, we identified the variables that contributed to the model and their correlation with 30-day mortality. The AUC of the GBDT algorithms was the highest when using the top 4 (ferritin, UREA, age and thrombin time (TT)) features, reaching 0.99 in the training cohort and 0.83 in the validation cohort. Additionally, we developed a web-based calculator to estimate the risk of 30-day mortality., Conclusions: With GBDT algorithms applied to laboratory data, accurate prediction of 30-day mortality is achievable. Integrating these algorithms into clinical practice could potentially improve 30-day outcomes., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

High ferritin is an important and sensitive biomarker for the various forms of hemophagocytic lymphohistiocytosis (HLH), a diverse and deadly group of cytokine storm syndromes. Early action to prevent immunopathology in HLH often includes empiric immunomodulation, which can complicate etiologic work-up and prevent collection of early/pre-treatment research samples. To address this, we instituted an alert system at UPMC Children's Hospital where serum ferritin > 1000 ng/mL triggered real-time chart review, assessment of whether the value reflected "inflammatory hyperferritnemia (IHF)", and biobanking of remnant samples from consenting IHF patients. We extracted relevant clinical data; periodically measured serum total IL-18, IL-18 binding protein (IL-18BP), and CXCL9; retrospectively classified patients by etiology into infectious, rheumatic, or immune dysregulation; and subjected a subgroup of samples to a 96-analyte biomarker screen. 180 patients were identified, 30.5% of which had IHF. Maximum ferritin levels were significantly higher in patients with IHF than with either hemoglobinopathy or transplant, and highly elevated total IL-18 levels were distinctive to patients with Stills Disease and/or Macrophage Activation Syndrome (MAS). Multi-analyte analysis showed elevation in proteins associated with cytotoxic lymphocytes in all IHF samples when compared to healthy controls and depression of proteins such as ANGPT1 and VEGFR2 in samples from hyperferritinemic sepsis patients relative to non-sepsis controls. This real-time IFH screen proved feasible and efficient, validated prior observations about the specificity of IL-18, enabled early sample collection from a complex population, suggested a unique vascular biomarker signature in hyperferritinemic sepsis, and expanded our understanding of IHF heterogeneity., (© 2024. The Author(s).)

Magnesium transporter 1 (MAGT1) gene loss-of-function variants lead to X-linked MAGT1 deficiency with increased susceptibility to EBV infection and N-glycosylation defect (XMEN), a condition with a variety of clinical and immunological effects. In addition, MAGT1 deficiency has been classified as a congenital disorder of glycosylation (CDG) due to its unique role in glycosylation of multiple substrates including NKG2D, necessary for viral protection. Due to the predisposition for EBV, this etiology has been linked with hemophagocytic lymphohistiocytosis (HLH), however only limited literature exists. Here we present a complex case with HLH and EBV-driven classic Hodgkin lymphoma (cHL) as the presenting manifestation of underlying immune defect. However, the patient's underlying immunodeficiency was not identified until his second recurrence of Hodgkin disease, recurrent episodes of Herpes Zoster, and after he had undergone autologous hematopoietic stem cell transplant (HSCT) for refractory Hodgkin lymphoma. This rare presentation of HLH and recurrent lymphomas without some of the classical immune deficiency manifestations of MAGT1 deficiency led us to review the literature for similar presentations and to report the evolving spectrum of disease in published literature. Our systematic review showcased that MAGT1 predisposes to multiple viruses (including EBV) and adds risk of viral-driven neoplasia. The roles of MAGT1 in the immune system and glycosylation were highlighted through the multiple organ dysfunction showcased by the previously validated Immune Deficiency and Dysregulation Activity (IDDA2.1) score and CDG-specific Nijmegen Pediatric CDG Rating Scale (NPCRS) score for the patient cohort in the systematic review., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

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., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Varicella zoster virus (VZV) is a neurotropic alphaherpesvirus exclusively infecting humans, causing two distinct pathologies: varicella (chickenpox) upon primary infection and herpes zoster (shingles) following reactivation. In susceptible individuals, VZV can give rise to more severe clinical manifestations, including disseminated infection, pneumonitis, encephalitis, and vasculopathy with stroke. Here, we describe a 3-year-old boy in whom varicella followed a complicated course with thrombocytopenia, hemorrhagic and necrotic lesions, pneumonitis, and intermittent encephalopathy. Hemophagocytic lymphohistiocytosis (HLH) was strongly suspected and as the condition deteriorated, HLH therapy was initiated. Although the clinical condition improved, longstanding hemophagocytosis followed despite therapy. We found that the patient carries a rare monoallelic variant in autocrine motility factor receptor (AMFR), encoding a ubiquitin ligase involved in innate cytosolic DNA sensing and interferon (IFN) production through the cyclic GMP-AMP synthase-stimulator of IFN genes (cGAS-STING) pathway. Peripheral blood mononuclear cells (PBMCs) from the patient exhibited impaired signaling downstream of STING in response dsDNA and 2'3'-cGAMP, agonists of cGAS and STING, respectively, and fibroblasts from the patient showed impaired type I IFN responses and significantly increased VZV replication. Overexpression of the variant AMFR R594C resulted in decreased K27-linked STING ubiquitination compared to WT AMFR. Moreover, ImageStream technology revealed reduced STING trafficking from ER to Golgi in cells expressing the patient AMFR R594C variant. This was supported by a dose-dependent dominant negative effect of expression of the patient AMFR variant as measured by IFN-β reporter gene assay. Finally, lentiviral transduction with WT AMFR partially reconstituted 2'3'-cGAMP-induced STING-mediated signaling and ISG expression in patient PBMCs. This work links defective AMFR-STING signaling to severe VZV disease and hyperinflammation and suggests a direct role for cGAS-STING in the control of viral infections in humans. In conclusion, we describe a novel genetic etiology of severe VZV disease in childhood, also representing the first inborn error of immunity related to a defect in the cGAS-STING pathway., (© 2024. The Author(s).)

Hemophagocytic lymphohistiocytosis (HLH) is a life-threatening hyperinflammatory syndrome characterized by excessive activation of the immune system, along with uncontrolled proliferation of activated macrophages and lymphocytes. The clinical features of HLH often overlap with the clinical features of other severe inflammatory conditions such as sepsis, hindering accurate and timely diagnosis. In this study, we performed a data-independent acquisition mass spectrometry-based plasma proteomic analysis of 33 pediatric patients with HLH compared with four control groups: 39 healthy children, 43 children with sepsis, 39 children hospitalized in the pediatric intensive care unit without confirmed infections, and 21 children with acute Epstein-Barr virus infection. Proteomic comparisons between the HLH group and each of the control groups showed that HLH was characterized by alterations in complement and coagulation cascades, neutrophil extracellular trap formation, and platelet activation pathways. We identified eight differentially expressed proteins in patients with HLH, including plastin-2 (LCP1), vascular cell adhesion protein 1, fibrinogen beta chain, fibrinogen gamma chain, serum amyloid A-4 protein, extracellular matrix protein 1, apolipoprotein A-I, and albumin. LCP1 emerged as a candidate diagnostic marker for HLH with an area under the curve (AUC) of 0.97 in the original cohort and an AUC of 0.90 (sensitivity = 0.83 and specificity = 1.0) in the validation cohort. Complement C1q subcomponent subunit B was associated with disease severity in patients with HLH. Based on comparisons with multiple control groups, this study provides a proteomic profile and candidate biomarkers of HLH, offering researchers novel information to improve the understanding of this condition., (© 2023. The Author(s).)

Purpose: Allogeneic hematopoietic stem cell transplantation (HSCT) is an established therapy for many inborn errors of immunity (IEI). The indications for HSCT have expanded over the last decade. The study aimed to collect and analyze the data on HSCT activity in IEI in Russia., Methods: The data were collected from the Russian Primary Immunodeficiency Registry and complemented with information from five Russian pediatric transplant centers. Patients diagnosed with IEI by the age of 18 years and who received allogeneic HSCT by the end of 2020 were included., Results: From 1997 to 2020, 454 patients with IEI received 514 allogeneic HSCT. The median number of HSCTs per year has risen from 3 in 1997-2009 to 60 in 2015-2020. The most common groups of IEI were immunodeficiency affecting cellular and humoral immunity (26%), combined immunodeficiency with associated/syndromic features (28%), phagocyte defects (21%), and diseases of immune dysregulation (17%). The distribution of IEI diagnosis has changed: before 2012, the majority (65%) had severe combined immunodeficiency (SCID) and hemophagocytic lymphohistiocytosis (HLH), and after 2012, only 24% had SCID and HLH. Of 513 HSCTs, 48.5% were performed from matched-unrelated, 36.5% from mismatched-related (MMRD), and 15% from matched-related donors. In 349 transplants T-cell depletion was used: 325 TCRαβ/CD19+ depletion, 39 post-transplant cyclophosphamide, and 27 other. The proportion of MMRD has risen over the recent years., Conclusion: The practice of HSCT in IEI has been changing in Russia. Expanding indications to HSCT and SCID newborn screening implementation may necessitate additional transplant beds for IEI in Russia., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Human inborn errors of immunity (IEI) affecting the type I interferon (IFN-I) induction pathway have been associated with predisposition to severe viral infections. Hemophagocytic lymphohistiocytosis (HLH) is a life-threatening systemic hyperinflammatory syndrome that has been increasingly associated with inborn errors of IFN-I-mediated innate immunity. Here is reported a novel case of complete deficiency of STAT2 in a 3-year-old child that presented with typical features of HLH after mumps, measles, and rubella vaccination at the age of 12 months. Due to the life-threatening risk of viral infection, she received SARS-CoV-2 mRNA vaccination. Unfortunately, she developed multisystem inflammatory syndrome in children (MIS-C) after SARS-CoV-2 infection, 4 months after the last dose. Functional studies showed an impaired IFN-I-induced response and a defective IFNα expression at later stages of STAT2 pathway induction. These results suggest a possible more complex mechanism for hyperinflammatory reactions in this type of patients involving a possible defect in the IFN-I production. Understanding the cellular and molecular links between IFN-I-induced signaling and hyperinflammatory syndromes can be critical for the diagnosis and tailored management of these patients with predisposition to severe viral infection., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Purpose: To evaluate the role of circulating Epstein-Barr virus (EBV) DNA in lymphoma-associated hemophagocytic lymphohistiocytosis (HLH)., Methods: We conducted a retrospective cohort study to explore the clinical and prognostic significance of EBV DNA in lymphoma-associated HLH. We included adult patients with combined diagnoses of lymphoma and HLH from January 2010 and November 2022 by retrieving the medical record system., Results: A total of 281 patients with lymphoma-associated HLH were identified. Elevated whole-blood EBV DNA was observed in 54.4% (153/281) of patients, and the median copy number was significantly higher in the T/NK-cell malignancies (199,500, interquartile range, 30,000-1,390,000) than that in the B-cell non-Hodgkin lymphoma (5520, interquartile range, 1240-28,400, P < 0.001). The optimum cutoff for predicting survival was 16,100 copies/mL. Compared to the patients with EBV DNA ≤ 16,100 copies/mL, those with EBV DNA > 16,100 copies/mL were younger and had more T/NK-cell malignancies, lower levels of neutrophils and fibrinogen, and higher levels of hemoglobin, alanine aminotransferase, aspartate aminotransferase, lactic dehydrogenase, and β 2 -microglobulin. A higher load of EBV DNA (> 16,100 copies/mL), thrombocytopenia (< 100 × 10 9 /L), neutropenia (< 1 × 10 9 /L), hypofibrinogenemia (≤ 1.5 g/L), and elevated levels of creatinine (> 133 μmol/L) were independent adverse predictors of 60-day overall survival and overall survival. A prognostic index based on EBV DNA and the other four factors was established to categorize the patients into four groups with significantly different outcomes., Conclusion: Our study identified high EBV load as a risk factor for lymphoma-associated HLH and established a prognostic index to predict outcomes., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Purpose: The first step in diagnosing hemophagocytic lymphohistiocytosis (HLH) is to suspect its presence and then order the appropriate diagnostic tests. The development of screening procedures for HLH could facilitate early diagnosis. In this study, we evaluated the utility of fever, splenomegaly, and cytopenias as screening criteria for identifying pediatric HLH at an early stage, built a screening model using commonly measured laboratory parameters, and developed a step-wise screening procedure for pediatric HLH., Methods: The medical records of 83,965 pediatric inpatients, including 160 patients with HLH, were collected retrospectively. The utility of fever, splenomegaly, hemoglobin level, and platelet and neutrophil counts at hospital admission as screening criteria for HLH was evaluated. For HLH patients who might be missed by screening based on the presence of fever, splenomegaly, and cytopenias, a screening model using common laboratory parameters was developed. Following that, a three-step screening procedure was then developed., Results: The criteria of cytopenias affecting two or more lineages plus fever or splenomegaly had a sensitivity of 51.9% and a specificity of 98.4% for identifying HLH in pediatric inpatients. Our screening score model comprises six parameters: splenomegaly, platelet count, neutrophil count, albumin level, total bile acid level, and lactate dehydrogenase level. The use of the validation set had a sensitivity of 87.0% and a specificity of 90.6%. A three-step screening procedure has been developed: Step 1: Is fever or splenomegaly present? (Yes: risk for HLH should be considered, go to Step 2; No: less likely HLH); Step 2: Are cytopenias affecting at least two lineages? (Yes: consider HLH; No: go to Step 3); Step 3: Calculate the screening score. Is the sum of the score greater than 37? (Yes: consider HLH; No: less likely HLH). The overall sensitivity and specificity of the three-step screening procedure were 91.9% and 94.4%, respectively., Conclusion: A significant proportion of pediatric HLH patients present at the hospital without having all three symptoms: fever, splenomegaly, and cytopenias. Our three-step screening procedure, utilizing commonly available clinical and laboratory parameters, can effectively identify pediatric patients who may be at high risk for HLH., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Autosomal recessive mutations in RAB27A are associated with Griscelli syndrome type 2 (GS2), characterized by hypopigmentation and development of early-onset, potentially fatal hemophagocytic lymphohistiocytosis (HLH). We describe a 35-year old male who presented with recurrent fever, was diagnosed with Epstein-Barr virus-driven chronic lymphoproliferation, fulfilled clinical HLH criteria, and who carried a novel homozygous RAB27A c.551G > A p.(R184Q) variant. We aimed to evaluate the contribution of the identified RAB27A variant in regard to the clinical phenotype as well as cellular and biochemical function. The patient displayed normal pigmentation as well as RAB27A expression in blood-derived cells. However, patient NK and CD8 + T cell exocytosis was low. Ectopic expression of the RAB27A p.R184Q variant rescued melanosome distribution in mouse Rab27a-deficient melanocytes, but failed to increase exocytosis upon reconstitution of human RAB27A-deficient CD8 + T cells. Mechanistically, the RAB27A p.R184Q variant displayed reduced binding to SLP2A but augmented binding to MUNC13-4, two key effector proteins in immune cells. MUNC13-4 binding was particularly strong to an inactive RAB27A p.T23N/p.R184Q double mutant. RAB27A p.R184Q was expressed and could facilitate melanosome trafficking, but did not support lymphocyte exocytosis. The HLH-associated RAB27A variant increased Munc13-4 binding, potentially representing a novel mode of impairing RAB27A function selectively in hematopoietic cells., (© 2022. The Author(s).)

Hemophagocytic lymphohistiocytosis (HLH) is a life-threatening immune dysregulation syndrome characterized by uncontrolled immune cell activation. Timely diagnosis is important, since early treatment can improve survival rates. However, completing all assessments needed to reach ≥5 positive criteria out of the 8 HLH-2004 criteria can be time consuming and may delay timely initiation of treatment. Hence, we applied a data-driven approach to identify a minimal parameter set for early decision-making towards the initiation of HLH-specific treatment. We retrospectively evaluated 165 patients from five Dutch tertiary hospitals with suspected HLH. Sixteen pHLH (median age 0.5 years) and 70 sHLH patients (median age 8.7 years) were identified using the HLH-2004 criteria. Clustering analysis and multi-receiver operator characteristics were used to identify parameters distinctive of HLH. The presence of either increased ferritin, cytopenia in ≥2 lineages, or splenomegaly distinguished HLH from non-HLH cases with a negative predictive value of 100%. A minimal parameter set consisting of 2 major criteria (phagocytosis and splenomegaly) and 3 minor criteria (cytopenia, increased ferritin, and increased triglycerides/low fibrinogen) predicted HLH with 95% (88-99) sensitivity and 94% (86-98) specificity. This finding was replicated in an independent retrospective validation cohort of 109 US patients (n = 109). By dividing a subset of the HLH-2004 criteria into major and minor criteria, this strategy uses the evaluation of less than 5 criteria to quickly identify patients with HLH. When confirmed in a prospective setting, this approach could be of value for timely diagnosis and treatment of HLH., (© 2021. The Author(s).)

Hemophagocytic lymphohistiocytosis (HLH) is a syndrome of severe immune dysregulation that encompasses a broad range of underlying genetic diseases and infectious triggers. Monogenic conditions, autoimmune diseases, and infections can all drive the phenotype of HLH and associated immune hyperactivation with hypercytokinemia. A diagnosis of HLH usually requires a combination of clinical and laboratory findings; there is no single sensitive and specific diagnostic test, which often leads to "diagnostic dilemmas" and delays in treatment initiation. Ferritin levels, one of the most commonly used screening tests, were collected across a large tertiary care pediatric hospital to identify the positive predictive value for HLH. Herein, we present several cases that illustrate the clinical challenges of confirming an HLH diagnosis. Additionally, we report on the utility of establishing a formal multi-disciplinary group to aid the prompt diagnosis and treatment of patients presenting with HLH-like pathophysiologies., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Familial hemophagocytic lymphohistiocytosis (HLH) is a life-threatening hyperinflammatory syndrome resulting from defective cytotoxicity. A previously healthy 3-month-old female presented with fever, irritability, abdominal distention, and tachypnea. She ultimately met all eight HLH-2004 diagnostic criteria, accompanied by elevated CXCL9. Initial empiric anti-inflammatory treatment included anakinra and IVIg, which stabilized ferritin and cytopenias. She had molecular and genetic confirmation of perforin deficiency and was started on dexamethasone and etoposide per HLH-94. She clinically improved, though CXCL9 and sIL-2Ra remained elevated. She was readmitted at week 8 for relapsed HLH without clear trigger and HLH-94 induction therapy was reinitiated. Her systemic HLH symptoms failed to respond and she soon developed symptomatic CNS HLH. She was incidentally found to have multifocal lung and kidney nodules, which were sterile and consisted largely of histiocytes and activated, oligoclonal CD8 T cells. The patient had a laboratory response to salvage therapy with alemtuzumab and emapalumab, but progressive neurologic decline led to withdrawal of care. This report highlights HLH foci manifest as pulmonary/renal nodules, demonstrates the utility of monitoring an array of HLH biomarkers, and suggests possible benefit of earlier salvage therapy.

Many biomarkers have been proposed for the diagnosis of secondary hemophagocytic lymphohistiocytosis (HLH) in adults, but comparative studies are lacking. We analyzed ferritin, glycosylated ferritin, soluble CD25, CD163 and CD14, IL-6, IFN-γ, IL-18, IL-10, IL-1ß, IL-12p70, IL-17α, IP-10, and CXCL9 levels to differentiate HLH from sepsis in critically ill patients. Of 120 patients, HLH was confirmed for 14 patients. Among the biomarkers tested, ferritin, IL-18, and glycosylated ferritin were the most efficient parameters for early diagnosis of HLH. With a sensitivity set at 85%, ferritin, IL-18, and glycosylated ferritin were the biomarkers with the highest specificity: 84, 79, and 71% respectively. Combining IL-18 with the HScore provided a new score with an increased specificity compared to the HScore alone, 86% compared to 70% with a sensitivity set at 100%. A distinct cytokine pattern was highlighted in patients with malignancy-triggered HLH, with highly increased levels of INF-ɣ and CXCL9, compared to HLH secondary to infection. This is the largest study available to date, comparing diagnostic biomarkers for HLH on a cohort of critically ill adult patients. Serum ferritin was the most discriminating parameter for early diagnosis of secondary HLH. IL18 * HScore was identified as a highly potential score.

Isolated neuroinflammatory disease has been described in case reports of familial hemophagocytic lymphohistiocytosis (FHL), but the clinical spectrum of disease manifestations, response to therapy and prognosis remain poorly defined. We combined an international survey with a literature search to identify FHL patients with (i) initial presentation with isolated neurological symptoms; (ii) absence of cytopenia and splenomegaly at presentation; and (iii) systemic HLH features no earlier than 3 months after neurological presentation. Thirty-eight (20 unreported) patients were identified with initial diagnoses including acute demyelinating encephalopathy, leukoencephalopathy, CNS vasculitis, multiple sclerosis, and encephalitis. Median age at presentation was 6.5 years, most commonly with ataxia/gait disturbance (75%) and seizures (53%). Diffuse multifocal white matter changes (79%) and cerebellar involvement (61%) were common MRI findings. CSF cell count and protein were increased in 22/29 and 15/29 patients, respectively. Fourteen patients progressed to systemic inflammatory disease fulfilling HLH-2004 criteria at a mean of 36.9 months after initial neurological presentation. Mutations were detected in PRF1 in 23 patients (61%), RAB27A in 10 (26%), UNC13D in 3 (8%), LYST in 1 (3%), and STXBP2 in 1 (3%) with a mean interval to diagnosis of 28.3 months. Among 19 patients who underwent HSCT, 11 neurologically improved, 4 were stable, one relapsed, and 3 died. Among 14 non-transplanted patients, only 3 improved or had stable disease, one relapsed, and 10 died. Isolated CNS-HLH is a rare and often overlooked cause of inflammatory brain disease. HLH-directed therapy followed by HSCT seems to improve survival and outcome.

Purpose: Hemophagocytic lymphohistiocytosis (HLH) is a rare systematic immune disease manifested with excessive activation of lymphocytes and macrophages. This study was designed to explore the feasible prognostic factors of secondary HLH (sHLH)., Method: We retrospectively analyzed 179 patients with newly diagnosed sHLH from January 2016 to May 2019 according to the HLH-2004 protocol. Baseline characteristics and laboratory results were reviewed., Results: The median age of all patients was 53 years, with a male/female ratio of 1.45. The commonest cause of HLH was malignancy. Of the 179 patients, 48.6% presented with Epstein-Barr virus (EBV) infection, 92.8% with hemocytopenia (at least 2 lineages), 60.3% with hypofibrinogenemia, 43.0% with hypertriglyceridemia (≥ 3 mmol/L), 99.4% with high ferritin, 97.8% with fever, 72.1% with splenomegaly, and 72.6% with hemophagocytosis. As to their prognosis, 122 patients died; the median survival was 88 days, with a 2-year survival rate of 26.72%. Univariate analysis confirmed neutrophil-to-lymphocyte ratio ˃ 2.53, lymphocyte-to-monocyte ratio (LMR) ≤ 4.43, platelet-to-lymphocyte ratio ˃ 227.27, red blood cell distribution width ˃ 14.6, red blood cell distribution width-to-platelet ratio (RPR) > 0.33, EBV infection, platelet ≤ 34 × 10 9 /L, fibrinogen ≤ 1.34 g/L, alkaline phosphatase ˃ 182.4 U/L, adenosine deaminase ˃ 69.2 U/L, and ferritin ˃ 2318 ng/mL were associated with an inferior survival. In a multivariate model, LMR, RPR, and ferritin were considered as three independent factors., Conclusion: Some blood-based inflammatory markers, which can be easily and cheaply detected, are significantly associated with the OS of HLH patients. LMR and RPR, superior to NLR, PLR, RDW, can be taken to predict the OS of patients with HLH.

Epstein-Barr virus (EBV) reactivation causes serious diseases in immunocompromised hosts, such as acquired immunodeficiency syndrome (AIDS). We report on a case of plasmablastic lymphoma (PBL) with hemophagocytic lymphohistiocytosis (HLH).A-53-year-old Japanese man was diagnosed with PBL and AIDS. In addition to combined antiretroviral therapy, HyperCVAD (cyclophosphamide, doxorubicin, vincristine, prednisone)/high-dose methotrexate + cytarabine was initiated immediately. Partial remission was attained with chemotherapy. However, the patient developed HLH and died despite intensive therapy. Autopsy findings suggested that PBL was controlled, and immunosuppression appeared to cause fatal infection. The patient showed high titers of EBV viral-capsid antigen (VCA)-IgG (1:2560) on PBL diagnosis and high EBV-DNA levels throughout the clinical course. Moreover, EBV-DNA was detected in the fraction of CD8-positive cells, which strongly supports the pathogenesis of EBV-associated HLH.Our report highlights the importance of EBV control in patients with EBV-positive AIDS lymphoma. EBV not only behaves as the etiologic pathogen of PBL but also can be a trigger of HLH, the fatal complication. Careful follow-up of the EBV status should be performed, and if needed, preemptive anti-EBV therapy should also be considered to prevent EBV-associated complications such as HLH.

We report our experience in using flow cytometry-based immunological screening prospectively as a decision tool for the use of genetic studies in the diagnostic approach to patients with hemophagocytic lymphohistiocytosis (HLH). We restricted genetic analysis largely to patients with abnormal immunological screening, but included whole exome sequencing (WES) for those with normal findings upon Sanger sequencing. Among 290 children with suspected HLH analyzed between 2010 and 2014 (including 17 affected, but asymptomatic siblings), 87/162 patients with "full" HLH and 79/111 patients with "incomplete/atypical" HLH had normal immunological screening results. In 10 patients, degranulation could not be tested. Among the 166 patients with normal screening, genetic analysis was not performed in 107 (all with uneventful follow-up), while 154 single gene tests by Sanger sequencing in the remaining 59 patients only identified a single atypical CHS patient. Flow cytometry correctly predicted all 29 patients with FHL-2, XLP1 or 2. Among 85 patients with defective NK degranulation (including 13 asymptomatic siblings), 70 were Sanger sequenced resulting in a genetic diagnosis in 55 (79%). Eight patients underwent WES, revealing mutations in two known and one unknown cytotoxicity genes and one metabolic disease. FHL3 was the most frequent genetic diagnosis. Immunological screening provided an excellent decision tool for the need and depth of genetic analysis of HLH patients and provided functionally relevant information for rapid patient classification, contributing to a significant reduction in the time from diagnosis to transplantation in recent years.

Objective: We assessed the clinical features and outcomes based on therapeutic options adopted during hospital stay for adult patients with macrophage activation syndrome and secondary hemophagocytic lymphohistiocytosis (MAS/sHLH)., Methods: We conducted a retrospective chart review of all adult patients (age ≥ 18 years) diagnosed with MAS/sHLH at our center between 2010 and 2015. Inclusion criteria for patients were diagnosis of MAS/sHLH during admission and patients meeting at least 5 out of 8 of Henter's criteria or at least 4 out of 6 of the criteria that were tested., Results: Nineteen adult patients with MAS/sHLH met the inclusion criteria from January 2010 to October 2015 (median age 48 years; female 68.4%). Treatment had been personalized, depending on the clinical presentation and course of disease. Majority of the patients received anakinra, cyclosporine, intravenous immunoglobulins (IVIG), and steroids. Fourteen (74%) patients survived, with clinical improvement by the time of discharge. After excluding the three patients with underlying leukemia/lymphoma who opted for palliative care and subsequently died, the survival rate was 88%., Conclusion: A modified diagnostic and treatment protocol for adult patients with MAS/sHLH that incorporated graded introduction of medications based on clinical presentation and cytokine profile resulted in the best adult survival rate reported in literature.

Background: X-linked inhibitor of apoptosis protein (XIAP) deficiency is a rare immunodeficiency that is characterized by recurrent hemophagocytic lymphohistiocytosis (HLH) and splenomegaly and sometimes associated with refractory inflammatory bowel disease (IBD). Although hematopoietic stem cell transplantation (HSCT) is the only curative therapy, the outcomes of HSCT for XIAP deficiency remain unsatisfactory compared with those for SLAM-associated protein deficiency and familial HLH., Aim: To investigate the outcomes and adverse events of HSCT for patients with XIAP deficiency, a national survey was conducted., Methods: A spreadsheet questionnaire was sent to physicians who had provided HSCT treatment for patients with XIAP deficiency in Japan., Results: Up to the end of September 2016, 10 patients with XIAP deficiency had undergone HSCT in Japan, 9 of whom (90%) had survived. All surviving patients had received a fludarabine-based reduced intensity conditioning (RIC) regimen. Although 5 patients developed post-HSCT HLH, 4 of them survived after etoposide administration. In addition, the IBD associated with XIAP deficiency improved remarkably after HSCT in all affected cases., Conclusion: The RIC regimen and HLH control might be important factors for successful HSCT outcomes, with improved IBD, in patients with XIAP deficiency.

Purpose: Familial hemophagocytic lymphohistiocytosis type 3 (FHL3) is a genetic disorder that results in immune dysregulation. It requires prompt and accurate diagnosis. A natural killer (NK) cell degranulation assay is often used to screen for FHL3 patients. However, we recently encountered two cases of late-onset FHL3 carrying novel UNC13D missense mutations: in these cases, the degranulation assays using freshly isolated and interleukin (IL)-2-activated NK cells yielded contradictory results. Since the defective degranulation of CD57 + cytotoxic T lymphocytes (CTLs) in these cases was helpful for making the diagnosis, we assessed whether the CD57 + CTL degranulation assay more effectively identified FHL3 patients than the NK cell assays., Methods: Forty additional patients with hemophagocytic lymphohistiocytosis were prospectively screened for FHL3 by measuring the perforin expression in NK cells and the expression of Munc13-4, syntaxin-11, and Munc18-2 in platelets and by performing NK cell and CTL degranulation assays. The results were confirmed by genetic analysis., Results: The freshly isolated NK cell degranulation assay detected FHL3 patients with high sensitivity (100%) but low specificity (71%). The IL-2-stimulated NK cell assay had improved specificity, but 3 out of the 31 non-FHL3 patients still showed degranulation below the threshold level. The CD57 + CTL degranulation assay identified FHL3 patients with high sensitivity and specificity (both 100%)., Conclusions: The CD57 + CTL degranulation assay more effectively identified FHL3 patients than the NK cell-based assays.

The X-linked inhibitor of apoptosis (XIAP) deficiency is a primary immunodeficiency characterized by Epstein-Barr virus (EBV)-driven hemophagocytic lymphohistiocytosis (HLH), splenomegaly, and colitis. Here, we present, for the first time, granulomatous hepatitis and granulomatous and lymphocytic interstitial lung disease (GLILD) as manifestations of XIAP deficiency. We report successful treatment of GLILD in XIAP deficiency with rituximab and azathioprine and discuss the role of XIAP deficiency in immune dysregulation.

Familial Hemophagocytic Lymphohistiocytosis type 2 (FHL2) results from mutations in PRF1. We described two unrelated individuals who presented with FHL, in whom severely impaired NK cytotoxicity and decrease perforin expression was observed. DNA sequencing of PRF1 demonstrated that both were not only heterozygous for the p.54R > C/91A > V haplotype but also presented with the novel variant p.47G > V at the perforin protein. Perforin mRNA was found to be increased in a individual with that genotype. A carrier of the novel variant also demonstrated altered perforin mRNA and protein expression. Phylogenetic analysis and multiple alignments with perforin orthologous demonstrated a high level of conservation at Gly47. PolyPhen-2 and PROVEAN predicted p.47G > V to be "probably damaging" and "deleterious", respectively. A thermodynamic analysis showed that this variant was highly stabilizing, decreasing the protein internal energy. The ab initio perforin molecular modeling indicated that Gly47 is buried inside the hydrophobic core of the MACPF domain, which is crucial for the lytic pore formation and protein oligomerization. After the in silico induction of the p.47G > V mutation, Val47 increased the interactions with the surrounding amino acids due to its size and physical properties, avoiding a proper conformational change of the domain. To our knowledge, this is the first description supporting that p.47G > V is a pathogenic variant that in conjunction with p.54R > C/91A > V might result in the clinical phenotype of FHL2.

Familial hemophagocytic lymphohistiocytosis (FHL) is a genetically heterogeneous hyperinflammatory syndrome, caused by an uncontrolled and ineffective proliferation and activation of T-lymphocytes, NK-cells, and macrophages that infiltrate multiple organs. Herein, a patient is presented who suffered from hepatitis and atypical brain lesions. Genetic studies revealed a homozygous mutation in the STXP2 gene; and thus, the diagnosis of FHL5 was confirmed.

Familial hemophagocytic lymphohistiocytosis (FHL), is a rare autosomal recessive disorder characterized by an impairment of cytotoxic cells and uncontrolled activation of macrophages. This study presents the first description of four patients with FHL type 2 in Latin America. Patient 1 fulfilled the disease diagnostic criteria since 2 months of age, whereas patients 2, 3 and 4 exhibited the typical manifestations of the disease only later in their childhood. The PRF1 genetic analysis in these patients revealed two previously reported mutations: L17fsx50 and R54C. Interestingly, seven out of the 8 alleles evaluated here in patients carried the haplotype R54C/A91V, suggesting that this is a highly frequent FHL type 2 allele in Colombia. This haplotype confers residual cytotoxic function leading to late onset disease. Therefore, this report highlights the remarkable complexity of FHL diagnostic, emphasizing the importance of the genetic characterization of the disease.

Deficiency of X-linked inhibitor of apoptosis (XIAP) caused by XIAP/BIRC4 gene mutations is an inherited immune defect recognized as X-linked lymphoproliferative syndrome type 2. This disease is mainly observed in patients with hemophagocytic lymphohistiocytosis (HLH) often associated with Epstein-Barr virus infection. We described nine Japanese patients from six unrelated families with XIAP deficiency and studied XIAP protein expression, XIAP gene analysis, invariant natural killer T (iNKT) cell counts, and the cytotoxic activity of CD8(+) alloantigen-specific cytotoxic T lymphocytes. Of the nine patients, eight patients presented with symptoms in infancy or early childhood. Five patients presented with recurrent HLH, one of whom had severe HLH and died after cord blood transplantation. One patient presented with colitis, as did another patient's maternal uncle, who died of colitis at 4 years of age prior to diagnosis with XIAP deficiency. Interestingly, a 17-year-old patient was asymptomatic, while his younger brother suffered from recurrent HLH and EBV infection. Seven out of eight patients showed decreased XIAP protein expression. iNKT cells from patients with XIAP deficiency were significantly decreased as compared with age-matched healthy controls. These results in our Japanese cohort are compatible with previous studies, confirming the clinical characteristics of XIAP deficiency.

Introduction: Hemophagocytic lymphohistiocytosis (HLH) is a life-threatening disease with major diagnostic and therapeutic difficulties, basically comprising two different conditions: primary and secondary forms. Recent advances regarding molecular diagnosis may be useful to distinguish from one another, especially in sporadic cases starting in early infancy., Materials and Methods: In this report, we evaluated three Argentinean patients with clinical suspicion of HLH, but without family history. We excluded mutations in the perforin gene but identified in the three patients a novel homozygous deletion (c. 581&#95;584delTGCC; p.Leu194ProfsX2) in the gene-encoding syntaxin 11 (STX11), causing a premature termination codon., Results and Conclusion: Each parent from the three unrelated families resulted heterozygous for this deletion confirming the diagnosis of familial hemophagocytic lymphohistiocytosis type 4. Patients shared the same single-nucleotide polymorphism profile in STX11 gene, and genotyping at ten microsatellites surrounding this gene support the presence of a single-haplotype block carrying the novel mutation.