RAB27A Mutations Research Articles

Case Report on Griscelli Syndrome with Hemophagocytic Lymphohistiocytosis

Objective: This case report was written with the intention of educating pediatric caregivers regarding the early detection and management of Griscelli Syndrome type 2 (GS2), thereby preventing fatal consequences, and to draw light on the necessity for premarital genetic counselling and education in public. Case Report: A two months and 7 days old male baby presented with tachycardia, fever, abdominal distension, cough, loose stools, and vomiting. Head to toe examination showed hypo-pigmented hair with silvery grey eyebrows and eyelashes. Bone marrow aspiration showed increased histiocytes exhibiting haemophagocytosis. Suspecting Griscelli Syndrome, genetic test was done which indicated RAB27A gene mutations, thereby confirming the GS2 diagnosis. The child was treated with 2 litres of oxygen with nasal prong, meropenem, fluconazole, dexamethasone and packed red blood cell transfusion. In view of worsening sepsis baby was given with vancomycin, tigecycline and cyclosporine and later intubated. Conclusion: Consanguineous marriages are common in India and are one of the major reason for autosomal recessive disorders like GS2 in newborns. This highlights the need of both premarital genetic counseling and education among public. Keywords: Hypopigmented hair, haemophagocytosis, abdominal distension, RAB27A mutation

Hemophagocytic lymphohistiocytosis: pediatric hepatic perspective

BackgroundHepatic manifestations of hemophagocytic lymphohistiocytosis (HLH), an underrecognized primary presentation in pediatric age group, mandate high levels of suspicion for early diagnosis.AimThis is to study the frequencies of clinical and laboratory hepatic involvement in patients with familial/primary or secondary/acquired HLH in relation to disease reactivation and outcome.MethodsA 6-month retrospective cohort study recruited 35 patients with HLH. Detailed clinical, laboratory, and genetic characteristics of HLH were collected. Hepatic transaminases and synthetic liver functions were collected at presentation, weeks 2 and 8 after starting treatment, and at time of reactivation. Biochemical liver involvement was considered when alanine aminotransferase (ALT) lived three-times more than the upper normal level. Overall (OS) and reactivation free survival were analyzed according to liver involvement.ResultsTwenty patients (57%) had genetically confirmed HLH, 12 (34.3%) had MUNC13D mutations, 3 (8.5%) had STXBP2 mutations, and 5 (14.3%) had RAB27A mutations, while 9 (25.7%) had no genetic mutations with 4 of them had secondary HLH. Six patients (17.2%) patients had unknown genetics status. Median (IQR) age of the whole group was 18 months (6–36) with an age range of 2–108 months. Liver enlargement was detected at diagnosis in 29 (82.9%) and at reactivation in 18 (51.4%) patients. Eight (22.86%) patients had biochemical hepatic involvement at presentation with no significant difference in their demographic, initial clinical presentation, survival, or the type of mutant gene according to liver involvement.ConclusionVariable hepatic biochemical involvement might be the presenting manifestation of HLH at diagnosis and upon reactivation, yet it did not impact disease outcome.

Clinical Presentations, Diagnosis, and Genetic Features of Hemophagocytic Lymphohistiocytosis: A Single Institutional Experience With the Saudi Population.

Background Hemophagocytic lymphohistiocytosis (HLH) is an uncommon, potentially fatal condition caused by high immune activation. The present study aimed to identify the clinical manifestations, geographic distribution, and associated pathogenic genetic mutations of HLH in Saudi Arabia. Method A retrospective cross-sectional study was conducted at King Fahad Medical City (KFMC), with a total of 59 patients diagnosed with HLH in the period between 2006 and 2018. All genetic results and clinical and biochemical data were retrieved and statistically analyzed using IBM SPSS Statistics for Windows, Version 25 (Released 2017; IBM Corp., Armonk, New York, United States). Results The results revealed that 48 patients (81.4%) had 15 pathogenic mutations of primary HLH whereas 8 (13.6%) patients had no genetic mutation. The most common variant mutation identified was c.1430C>T of the STXBP2 gene (42.4% of total patients), followed by c.1122G>A of the PRF1 gene (10.2% of patients), which demonstrated a distinctive geographic and tribal association. Patients with RAB27A mutation tend to present at an older age than the others with a median age of presentation of 5.5 months vs 2 months for patients with PRF1 mutations. No significant differences in clinical features were observed among the various groups. Conclusion This study highlights the incidence of genetic mutations among the Saudi population with HLH. The STXBP2 is the most common mutation followed by PRF1 mutations, many mutation variants are associated with a distinctive tribal and geographic association.

Hepatic Manifestations of Pediatric Hemophagocytic Lymphohistocytosis

Background Hemophagocytic lymphohistiocytosis (HLH) is a rare hyper-inflammatory disorder.it is caused by benign systemic overgrowth of macrophages in the reticulo-endothelial system, leading to cytokine storm. The main features of HLH are; fever, splenomegaly, bi/pancytopenia; hyperferritinemia, hyper-triglyceridemia and hypofibrinogenemia; which if not diagnosed and early treated progresses to disseminated intravascular coagulopathy (DIC), multi-organ dysfunction with dismal outcome. Hepatic manifestations are not well recognized primary presentation in pediatric patients with HLH. This presentation mandates high levels of suspicious for early diagnosis. Aim of the Work To study the hepatic involvement clinical, laboratory, and pathological in patients with Familial/primary (1ry) HLH and in patients with secondary (2ry) acquired HLH. Patients and Methods A 6 month retrospective cohort study included 35 patients with genetically confirmed HLH divided into familial HLH by its types, X linked lymphoproliferative syndrome, HLH with partial albinism and secondary HLH; following at pediatric hematology/oncology clinic, Ain Shams University. By detailed review of clinical and laboratory data that include hepatic transaminases and synthetic liver function done at the time of presentation, at week 2, 8 from treatment start and at time of reactivation; Liver biopsy results and genetic analysis were recorded. Biochemical liver involvement was considered when alanine aminotransferase was three folds the upper level of normal at presentation. Overall and reactivation free survival were analyzed according to liver involvement Results Thirty five patients with HLH were recruited with age range of 2-108 months, 20(57.1%) of patients with HLH were genetically confirmed and 12(34.3%) of them had MUNC13D mutations, 3(8.6%) had STXBP2 mutation and 5(14.3%) had RAB27A mutation while 4(11.4%) had secondary HLH. 29(82.9%) had liver enlargement at diagnosis with hepatic reactivation in 18(51.4%). 8(22.8%) of patients had biochemical liver involvement; there was no significant difference in their demographic data or their clinical presentation, their final outcome or the type of mutant gene according to liver involvement. Conclusion HLH is a frequently lethal disease with a high variability in clinical presentation .it must be considered a differential in patients presenting with clinical signs of hepatic dysfunction especially when it is accompanied by cytopenias. Significant biochemical liver involvement is an under recognized presentation of HLH. Hepatic involvement did not impact response to treatment and disease outcome.

Novel RAB27A Variant Associated with Late-Onset Hemophagocytic Lymphohistiocytosis Alters Effector Protein Binding

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.

Griscelli Syndrome Type 2 Sine Albinism: Unraveling Differential RAB27A Effector Engagement.

Griscelli syndrome type 2 (GS-2) is an inborn error of immunity characterized by partial albinism and episodes of hemophagocytic lymphohistiocytosis (HLH). It is caused by RAB27A mutations that encode RAB27A, a member of the Rab GTPase family. RAB27A is expressed in many tissues and regulates vesicular transport and organelle dynamics. Occasionally, GS-2 patients with RAB27A mutation display normal pigmentation. The study of such variants provides the opportunity to map distinct binding sites for tissue-specific effectors on RAB27A. Here we present a new case of GS-2 without albinism (GS-2 sine albinism) caused by a novel missense mutation (Val143Ala) in the RAB27A and characterize its functional cellular consequences. Using pertinent animal cell lines, the Val143Ala mutation impairs both the RAB27A–SLP2-A interaction and RAB27A–MUNC13-4 interaction, but it does not affect the RAB27A–melanophilin (MLPH)/SLAC2-A interaction that is crucial for skin and hair pigmentation. We conclude that disruption of the RAB27A–MUNC13-4 interaction in cytotoxic lymphocytes leads to the HLH predisposition of the GS-2 patient with the Val143Ala mutation. Finally, we include a review of GS-2 sine albinism cases reported in the literature, summarizing their genetic and clinical characteristics.

RAB27A mutation in a patient with griscelli syndrome type 2, successfully cured by hematopoietic stem cell transplantation: Sustained remission

Griscelli syndrome (GS) is a rare autosomal recessive disorder. It features with clinical constellation of skin hypopigmentation, silvery-gray hair, associated with neurological involvement (type I), or defective cell-mediated immunity with hemophagocytosis (type II) that leads to recurrent infection, and patients succumb to death in early childhood or isolated “quiet” GS (type III). To our knowledge, more than sixty patients have been reported mainly from the Mediterranean region with the first reported Saudi cases, reported by Harfi et al. in 1992. The outcome of GS is guarded if not treated early due to recurrent fulminant infections, disease recurrence, and the impending severity of central nervous system disease. Hematopoietic stem cell transplantation (HSCT) is the single current therapeutic treatment for GS II. HSCT, therefore, should be performed as early as possible to prevent potential complications. The reported patient had sustained remission and normal hematopoietic function after allogeneic bone marrow transplant from a human leukocyte antigen-identical sibling donor. GS II syndrome should be considered in any child with hypopigmented skin, silvery-gray hair, and primary immunodeficiency. Early recognition and early HSCT intervention are associated with favorable outcome.

Next Generation Proteomics of Human Neutrophil Granulocytes in Monogenic Disease

Background and objectiveNeutrophil granulocytes (NG) are critical mediators of innate immunity and tissue regeneration. Few monogenic diseases affecting their differentiation and/or function have been described, yet a systems-biology perspective has not yet been opened. We set out to define the proteome and transcriptome of NG from healthy individuals and exome-sequenced patients aiming to develop a multi-dimensional perspective for diagnostic purposes.Patients and methodsWe analyzed highly purified peripheral blood neutrophil granulocytes from 68 healthy individuals, 6 children with severe congenital neutropenia (SCN) associated with ELANE mutations, 5 children with chronic granulomatous disease (CGD) with CYBA (2) or CYBB (3) mutations and 3 children with leukocyte adhesion deficiency (LAD, 2x ITGB2 mutation, 1x SLC35C1 mutation). In addition we analyzed proteomes of 2 children with genetically undetermined neutrophil defects. Cells were isolated from fresh venous blood using magnetic bead-based negative selection (purity >99%). Whole cell proteome analysis was done by data-independent acquisition using a Thermo Fisher QExactive HF mass spectrometer.ResultsWe quantified a total of ~4300 proteins in each sample. This is the first study to analyze highly purified neutrophil granulocytes from a large cohort of healthy individuals as well as from patients with rare diseases affecting differentiation and/or function of NG. Intraindividual reproducibility was shown for one donor sampled at 3 different time points with a correlation coefficient of ~0.89. Principal component analysis demonstrated separation of the proteome of healthy and diseased cells. Interestingly, only 6 proteins constituted for 50% of the neutrophils proteome with the antimicrobial proteins DEF3A and S100A8 being the most abundant.Stringent differential expression analysis (p<0.01) showed specific differences between the neutrophil proteome of healthy individuals and patients with monogenic diseases. In patients with LAD, only few proteins were found to be present in decreased (7 proteins) or increased (5 proteins) abundance. Of note, proteins with lowest expression levels in LAD1 (ITGB2 mutation) were the integrins ITGB2, ITGAM and ITGAX while in LAD2 (SLC35C1 mutation) integrin expression was not affected. In contrast we noticed lack of PRCP, HDHD2 and XRCC4 protein. In patients with CGD, CYBA and CYBB were decreased in expression, independent of the underlying genotype. In contrast, the other members of the NADPH oxidase complex (NCF1/2/4, Rac1/2) were not found to be differentially expressed. This suggests that both membrane-bound proteins CYBB and CYBA stabilize each other. In total, CGD neutrophils showed limited aberrations with 23 overexpressed and 11 underexpressed proteins. In striking contrast to LAD and CGD, SCN patients with ELANE mutations had a much higher number of proteins with increased (93 proteins) or decreased (47 proteins) expression levels. It is currently unknown whether these imbalances are due to the immaturity of peripheral blood neutrophils in SCN or rather secondary to unfolded protein response and cellular stress mechanisms.In ongoing studies, we analyze the transcriptome of healthy neutrophil granulocytes in correlation to the proteome. Preliminary data indicate that there is poor correlation (PCC 0.11) between protein and mRNA levels, indicating that granule proteins are synthesized during earlier steps of differentiation.In two patients where exome sequencing did not reveal any underlying mutation, proteome analysis showed decreased expression of NCF1 and RAB27A, respectively. In both patients we were able to use targeted sequencing of amplified DNA to show the causative mutations in NCF1 and RAB27A. Multi-omic analysis may therefore be superior to unidimensional genomic sequencing to establish a molecular diagnosis.ConclusionIn-depth mass spectrometry-based proteomics provides new insights into disease-specific alterations of neutrophil granulocytes and may help to refine molecular diagnostics. DisclosuresNo relevant conflicts of interest to declare.

Ferritin is secreted via 2 distinct nonclassical vesicular pathways

AbstractFerritin turnover plays a major role in tissue iron homeostasis, and ferritin malfunction is associated with impaired iron homeostasis and neurodegenerative diseases. In most eukaryotes, ferritin is considered an intracellular protein that stores iron in a nontoxic and bioavailable form. In insects, ferritin is a classically secreted protein and plays a major role in systemic iron distribution. Mammalian ferritin lacks the signal peptide for classical endoplasmic reticulum–Golgi secretion but is found in serum and is secreted via a nonclassical lysosomal secretion pathway. This study applied bioinformatics and biochemical tools, alongside a protein trafficking mouse models, to characterize the mechanisms of ferritin secretion. Ferritin trafficking via the classical secretion pathway was ruled out, and a 2:1 distribution of intracellular ferritin between membrane-bound compartments and the cytosol was observed, suggesting a role for ferritin in the vesicular compartments of the cell. Focusing on nonclassical secretion, we analyzed mouse models of impaired endolysosomal trafficking and found that ferritin secretion was decreased by a BLOC-1 mutation but increased by BLOC-2, BLOC-3, and Rab27A mutations of the cellular trafficking machinery, suggesting multiple export routes. A 13-amino-acid motif unique to ferritins that lack the secretion signal peptide was identified on the BC-loop of both subunits and plays a role in the regulation of ferritin secretion. Finally, we provide evidence that secretion of iron-rich ferritin was mediated via the multivesicular body–exosome pathway. These results enhance our understanding of the mechanism of ferritin secretion, which is an important piece in the puzzle of tissue iron homeostasis.

Perforin and CD107a testing is superior to NK cell function testing for screening patients for genetic HLH

AbstractPrimary hemophagocytic lymphohistiocytosis (HLH) can be caused by biallelic mutations in PRF1, encoding perforin, or UNC13D, STXBP2, STX11, RAB27A, LYST, and AP3B1, encoding proteins involved in cytotoxic lymphocyte degranulation. Natural killer (NK)–cell cytotoxicity assays can quickly screen for all of these genetic diseases, facilitating treatment, but combining NK-cell perforin expression and CD107a upregulation tests can as well. To determine the relative diagnostic accuracies for each approach, we retrospectively reviewed screening test performance in 1614 patients referred for HLH evaluation. For each test, we generated a receiver operating characteristic (ROC) curve, and calculated area under the curve (AUC) and diagnostic parameters at optimal threshold. We generated an AUC for combining perforin and CD107a tests by creating a logistic regression model and applying model-generated coefficients to patient values. Sensitivities of NK-cell function, perforin mean channel fluorescence (MCF), and CD107a MCF to detect biallelic mutations were 59.5%, 96.6%, and 93.8%, with specificities of 72.0%, 99.5%, and 73%. AUCs for NK-cell cytotoxicity, perforin MCF, CD107a MCF, and combined perforin and CD107a MCFs were 0.690, 0.971, 0.860, and 0.838. Perforin and CD107a tests are more sensitive and no less specific compared with NK cytotoxicity testing for screening for genetic HLH and should be considered for addition to current HLH criteria.

Dynamic modulation of cortical actin at the immunological synapse controls lytic granule secretion in cytotoxic T lymphocytes

Abstract CD8+ cytotoxic T lymphocytes (CTLs) are critical for eliminating virally infected cells. Upon target recognition, CTLs reorganize their actin cytoskeleton to facilitate the polarized secretion of specialized granules containing lytic proteins that kill cells. Because a single CTL can kill multiple targets, granule secretion must be tightly regulated, ensuring only appropriate cells are killed, and preserving granules for serial killing. However, how CTLs regulate secretion remains unclear. Here, we used live confocal and TIRF microscopy to evaluate how cortical actin affects granule secretion at the membrane. While previous work has shown that clearance of actin at the synapse precedes granule secretion, we find that after granule fusion, cortical actin recovers and no further secretion is observed. Depolymerization of recovered actin led to resumed granule fusion, suggesting that recovered actin acts as a barrier to prevent further lytic granule secretion. We further show that CTLs from mice unable to secrete granules due to a Rab27a mutation that prevents granule fusion, failed to recover cortical actin at the synapse. This suggests that granule fusion itself triggers actin recovery in CTLs. Thus, cortical actin both regulates, and is regulated by, secretion. Finally, using fluorescent reporters, we correlated the clearance and recovery of actin with PIP2. PIP2 binds numerous actin regulatory proteins, suggesting phosphatidylinositol distribution in the membrane as a potential mechanism through which CTLs regulate the density of cortical actin during killing. Our work provides insight into actin-related mechanisms regulating CTL secretion that may serial killing capacity during immune responses.

Exome sequencing for simultaneous mutation screening in children with hemophagocytic lymphohistiocytosis.

In the present study, we used exome sequencing to analyze PRF1, UNC13D, STX11, and STXBP2, as well as genes associated with primary immunodeficiency disease (RAB27A, LYST, AP3B1, SH2D1A, ITK, CD27, XIAP, and MAGT1) in Thai children with hemophagocytic lymphohistiocytosis (HLH). We performed mutation analysis of HLH-associated genes in 25 Thai children using an exome sequencing method. Genetic variations found within these target genes were compared to exome sequencing data from 133 healthy individuals. Variants identified with minor allele frequencies <5% and novel mutations were confirmed using Sanger sequencing. Exome sequencing data revealed 101 non-synonymous single nucleotide polymorphisms (SNPs) in all subjects. These SNPs were classified as pathogenic (n=1), likely pathogenic (n=16), variant of unknown significance (n=12), or benign variant (n=72). Homozygous, compound heterozygous, and double-gene heterozygous variants, involving mutations in PRF1 (n=3), UNC13D (n=2), STXBP2 (n=3), LYST (n=3), XIAP (n=2), AP3B1 (n=1), RAB27A (n=1), and MAGT1 (n=1), were demonstrated in 12 patients. Novel mutations were found in most patients in this study. In conclusion, exome sequencing demonstrated the ability to identify rare genetic variants in HLH patients. This method is useful in the detection of mutations in multi-gene associated diseases.

Pedigree Gene Investigation and Parameters of NK Cell Activity, CD107a Degranulation Amd HLH Related Defective Protein Play Significant Role in the Diagnosis of Primary HLH

Abstract This study aims to evaluate the significance of pedigree genetic screening and rapid immunological parameters in the diagnosis of primary HLH and explore their correlation. Twelve Chinese families of primary HLH with mutations in PRF1, UNC13D, LYST, RAB27A, SH2D1A, BIRC4 were recruited and conducted pedigree investigation, including family genetic screening, NK cell activity, CD107a degranulation and expression of HLH related defective protein. Ten patients were identified with homozygous, compound heterozygous, or hemizygous mutations in PRF1, UNC13D, RAB27A, SH2D1A and BIRC4, including one case of homozygous mutation (c.1349C>T:p.T450M) and four cases of compound heterozygous mutations (frameshift deletion c.65delC:p.P22fs and c.G503A:p.S168N; c.1168C>Tp.R390X and c.1349C>T:p.T450M; frameshift mutation c.1090_1091delCT:p.T364fsX93 and c.1349C>T:p.T450M; non-frameshift deletions c.1083_1094del:p.361_365del and c.T172C :p.S58P) in PRF1, one case of homozygous mutation (c.G2588A:p.G863D) and one compound heterozygous (c.C640T:p.R214X and c.G407A:p.C136Y) in UNC13D, one case of homozygous mutation (c.C244T:p.R82C) in RAB27A, and two male cases of hemizygous mutations in SH2D1A(c.32T>G:p.I11S) and BIRC4(c.G592A:p.V198M). Meanwhile, two patients were identified mutations in two or three different gene, including a male with a hemizygous mutation in SH2D1A(c.7G>T:p.A3S) and a heterozygous mutation(c.127C>A: p.L43M) in PRF1, and a female with three hemizygous mutations respectively in UNC13D(c.G680A:p.R227H), LYST(c.A8368C:p.K2790Q) and BIRC4(c.C962G:p.A321G).The primary HLH patients and their family members presented different levels of decreased NK cell activity. Individuals who were found mutations in PRF1, SH2D1A and BIRC4 showed low expression of perforin, SAP and XIAP. And the patients with homozygous and compound heterozygous mutations in UNC13D and RAB27A showed significant reducing of cytotoxic degranulation function and their family members presented normal or different levels of decreased CD107a. The patient with combination defects involving two genes(UNC13D and LYST) in the degranulation pathway showed decreased of CD107a degranulation which implied a digenic inheritance and diagnosed primary HLH. Pedigree genetic screening and rapid detection of immunological parameters play an important role in the diagnosis of primary HLH and demonstrate good consistency. DisclosuresNo relevant conflicts of interest to declare.

A heterozygous RAB27A mutation causes delayed cytolytic granule polarization and hemophagocytic lymphohistiocytosis in 2 unrelated teenagers

Abstract Frequently fatal, primary hemophagocytic lymphohistiocytosis (HLH) occurs in infancy resulting from homozygous mutations in natural killer (NK) and CD8 T cell cytolytic pathway genes. Secondary HLH presents after infancy and may be associated with heterozygous mutations in HLH genes. We report 2 unrelated teenagers with HLH and an identical heterozygous RAB27A mutation (259 G&amp;gt;C). The contribution of this Rab27A missense (A87P) mutation on NK cell cytotoxicity was studied by cloning it into a lentiviral expression vector prior to introduction into the human NK-92 cell line. NK cell degranulation (CD107a expression), target cell conjugation, and K562 target cell lysis was compared between mutant and wild-type (WT) transduced NK-92 cells. Polarization of granzyme B to the immunologic synapse and interaction of mutant Rab27A (A87P) with Munc13-4 were explored by confocal microscopy and proximity ligation assay (PLA), respectively. Over-expression of the RAB27A mutation had no effect on cell conjugate formation between the NK and target cells but decreased NK cell cytolytic activity and degranulation. Moreover, the mutant Rab27A protein was predicted to disrupt binding to WT Munc13-4 by crystal structure modelling, and decreased interaction of Rab27A (A87P) with WT Munc13-4 was shown by PLA in situ and by co-immunoprecipiation in vitro. Finally, Rab27A (A87P) over-expression in NK-92 cells delayed granzyme B polarization toward the immunologic synapse, as noted by confocal microscopy, with a resulting increase in interferon-γ production, a cytokine responsible for HLH. This heterozygous RAB27A mutation blurs the genetic distinction between primary and secondary HLH by contributing to HLH via a partial dominant-negative effect.

A novel Rab27a mutation binds melanophilin, but not Munc13-4, causing immunodeficiency without albinism

A novel Rab27a mutation binds melanophilin, but not Munc13-4, causing immunodeficiency without albinism

Prognostic Factors and Long-Term Outcome in 52 Turkish Children With Hemophagocytic Lymphohistiocytosis.

Hemophagocytic lymphohistiocytosis is a syndrome of pathologic immune activation that shares similar clinical and laboratory phenotypes with severe sepsis. Recent studies led to better recognition of hemophagocytic lymphohistiocytosis by clinicians, but no consensus exists on the criteria for high-risk patients. We retrospectively reviewed the medical records of patients diagnosed with hemophagocytic lymphohistiocytosis to analyze the risk factors associated with poor outcome. Pediatric intensive care and hematology units of three tertiary hospitals in Turkey. Fifty-two children with hemophagocytic lymphohistiocytosis. None. There were a total of 52 children meeting the diagnostic criteria of Histiocytic Society. Of them, 28 (54%) had a primary hemophagocytic lymphohistiocytosis. Mutation studies were performed in 18 of 28 patients (65%). Fourteen of them had PRF1, STX11, STXBP2, and UNC13D mutations, and four had Rab27a and LYST mutations. The remaining 24 patients (46%) were defined as having secondary hemophagocytic lymphohistiocytosis. Twenty-one of them had infection-associated hemophagocytic lymphohistiocytosis, and three had lysinuric protein intolerance. The mortality rate was significantly higher in primary hemophagocytic lymphohistiocytosis (64%) than in secondary hemophagocytic lymphohistiocytosis (16%) (p < 0.05). There were no significant differences for survival rate between hemophagocytic lymphohistiocytosis 94 (44%) and hemophagocytic lymphohistiocytosis 2004 (64%) protocols (p > 0.05). Age below 2 years, hyperferritinemia, thrombocytopenia, high disseminated intravascular coagulation score at diagnosis, and no clinical response at 2 weeks of treatment were independent prognostic factors for poor prognosis. Our data suggest that disseminated intravascular coagulation score greater than or equal to 5 can be used in the definition of high-risk patients. Early recognition of poor risk factors has important prognostic and therapeutic implications.

Patients with Griscelli syndrome and normal pigmentation identify RAB27A mutations that selectively disrupt MUNC13-4 binding

BackgroundFamilial hemophagocytic lymphohistiocytosis (FHL) is a rare and often fatal disorder characterized by defective cellular cytotoxicity and hyperinflammation, and the only cure known to date is hematopoietic stem cell transplantation. Mutations in RAB27A, LYST, and AP3B1 give rise to FHL associated with oculocutaneous albinism, and patients with FHL are usually only screened for mutations in these genes when albinism is observed. A number of patients with FHL and normal pigmentation remain without a genetic diagnosis.ObjectiveWe asked whether patients with FHL with immunodeficiency but with normal pigmentation might sometimes have mutations that affected cellular cytotoxicity without affecting pigmentation.MethodsWe carried out mutation analysis of RAB27A, LYST, and AP3B1 in patients with FHL with pigment dilution, as well as a cohort with no clinical evidence of pigment dilution but no mutations in the other known FHL-related genes (PRF1, STXBP2, and UNC13D).ResultsWe identify patients with Griscelli syndrome type 2 with biallelic mutations in RAB27A in the absence of albinism. All 6 patients carried mutations at amino acids R141, Y159, or S163 of Rab27a that disrupt the interaction of Rab27a with Munc13-4, without impairing the interaction between melanophilin and Rab27a.ConclusionThese studies highlight the need for RAB27A sequencing in patients with FHL with normal pigmentation and identify a critical binding site for Munc13-4 on Rab27a, revealing the molecular basis of this interaction.

A novel RAB-27A mutation causing Griscelli syndrome type 2 with severe central nervous system involvement: Case report and review of literature

Griscelli syndrome (GS) is one of the rare autosomal recessive disease characterized by hypopigmentation of the hair, hepatosplenomegally, primary immunodeficiency and neurological manifestations. It was described by Griscelli et al. (1978) in France, who reported two girls who were presented with silver gray hair, several episodes of fever, hepatosplenomegally and pancytopenia. There are three types of Griscelli syndrome based on clinical features and genetic mutations. We report a five month old girl diagnosed as Griscelli syndrome type 2 (OMIM #607624) presenting with significant central nervous system involvement. The molecular studies of the RAB27A gene were performed. Coding sequence revealed a novel homozygous deletion at (c.138delC). In communities with high incidence of consanguineous marriages, we should keep in mind the rare primary immunodeficiency diseases with autosomal recessive inheritance. Early diagnosis and treatment will help in improvement of the outcome. Key words: Griscelli syndrome type 2, RAB27A, CNS involvement.

The risk of hemophagocytic lymphohistiocytosis in Hermansky-Pudlak syndrome type 2

AbstractGenetic disorders of lymphocyte cytotoxicity predispose patients to hemophagocytic lymphohistiocytosis (HLH). Reduced lymphocyte cytotoxicity has been demonstrated in Hermansky-Pudlak syndrome type 2 (HPS2), but only a single patient was reported who developed HLH. Because that patient also carried a potentially contributing heterozygous RAB27A mutation, the risk for HLH in HPS2 remains unclear. We analyzed susceptibility to HLH in the pearl mouse model of HPS2. After infection with lymphocytic choriomeningitis virus, pearl mice developed all key features of HLH, linked to impaired virus control caused by a moderate defect in CTL cytotoxicity in vivo. However, in contrast to perforin-deficient mice, the disease was transient, and all mice fully recovered and controlled the infection. An additional heterozygous Rab27a mutation did not aggravate the cytotoxicity defect or disease parameters. In the largest survey of 22 HPS2 patients covering 234 patient years, we identified only 1 additional patient with HLH and 2 with incomplete transient HLH-like episodes, although cytotoxicity or degranulation was impaired in all 16 patients tested. HPS2 confers a risk for HLH that is lower than in Griscelli or Chediak-Higashi syndrome, probably because of a milder defect in cytotoxicity. Preemptive hematopoietic stem cell transplantation does not appear justified in HPS2.

Novel RAB27A Mutation Distinguishes Hypopigmentation From Cytotoxic Dysfunction in Griscelli Syndrome Type 2

Griscelli syndrome type 2 is an autosomal recessive primary immunodeficiency caused by RAB27A mutation. It is characterized by albinism, pigmentary clumping in hair shafts leading to silvery-grey hair and propensity towards hemophagocytic lymphohistiocytosis (HLH).