Gaucher Disease Phenotype Research Articles

The spectrum of neurological manifestations and genotype-phenotype correlation in Indian children with Gaucher disease.

Gaucher disease (GD), one of the most frequent autosomal recessive lysosomal storage disorders, occurs due to bi-allelic pathogenic variants in the GBA1. Worldwide, the c.1448T>C (L483P) homozygous pathogenic variant is reported to be associated with neurological GD phenotype. Clinical distinction between GD1 and GD3 may be challenging due to subtle neurological features. Objective methods to evaluate neurological signs and saccades may help in early diagnosis. This study was conducted to assess the neurological phenotype, and its severity using a modified severity scoring tool (mSST), and the genotype-phenotype correlation. A total of 45 children aged 2 years 6months to 15 years with a confirmed enzymatic and molecular diagnosis of GD with or without therapy were recruited. mSST tool was used to assess the severity of the neurological phenotype. A digital eye movement tracker (View Point Tracker) was used to assess eye movements. Clinical and genetic findings were analyzed. Out of 45 patients, 39 (86.7%) had at least one neurological phenotype detected using the mSST tool, with impairment of cognitive function (68.8%, 31/45) being the commonest feature. Thirty-two of 45 (71%) were assessed for saccadic eye movements using the eye tracker. Of these, 62.5% (20/32) had absent saccades. Four children (8.9%, 4/32) without clinical oculomotor apraxia had absent saccades on the viewpoint eye tracker. Overall, 77.7% (35/45), had homozygosity for c.1448T>C in GBA1 of which 91.4% (32/35) had neurological manifestations. Other alleles associated with neurological phenotype included c.1603C>T(p.R535C), c.1184C>T (p.S395F), c.115+1G>A (g.4234G>A), c.260G>A (p.R87Q) and c.1352A>G (p.Y451C). To conclude, in India, the c.1448T>C pathogenic variant in GBA1 is the commonest and is associated with neurological phenotype of GD. Therefore, every patient of GD should be assessed using the mSST scoring tool for an early pick up of neurological features. The routine use of a viewpoint eye tracker in children with GD would be useful for early recognition of saccadic abnormalities.

PGRN deficiency exacerbates, whereas a brain penetrant PGRN derivative protects, GBA1 mutation-associated pathologies and diseases

Mutations in GBA1, encoding glucocerebrosidase (GCase), cause Gaucher disease (GD) and are also genetic risks in developing Parkinson's disease (PD). Currently, the approved therapies are only effective for directly treating visceral symptoms, but not for primary neuronopathic involvement in GD (nGD). Progranulin (PGRN), encoded by GRN, is a novel modifier of GCase, but the impact of PGRN in GBA1 mutation-associated pathologies invivo remains unknown. Herein, Grn-/- mice crossed into Gba9v/9v mice, a Gba1 mutant line homozygous for the Gba1 D409V mutation, generating Grn-/-Gba9v/9v (PG9V) mice. PG9V mice exhibited neurobehavioral deficits, early onset, and more severe GD phenotypes compared to Grn-/- and Gba9v/9v mice. Moreover, PG9V mice also displayed PD-like phenotype. Mechanistic analysis revealed that PGRN deficiency caused severe neuroinflammation with microgliosis and astrogliosis, along with impaired autophagy associated with the Gba1 mutation. A PGRN-derived peptide, termed ND7, ameliorated the disease phenotype in GD patient fibroblasts exvivo. Unexpectedly, ND7 penetrated the blood-brain barrier (BBB) and effectively ameliorated the nGD manifestations and PD pathology in Gba9v/null and PG9V mice. Collectively, this study not only provides the first line of invivo but also exvivo evidence demonstrating the crucial role of PGRN in GBA1/Gba1 mutation-related pathologies, as well as a clinically relevant mouse model for mechanistic and potential therapeutics studies for nGD and PD. Importantly, a BBB penetrant PGRN-derived biologic was developed that may provide treatment for rare lysosomal storage diseases and common neurodegenerative disorders, particularly nGD and PD.

Progranulin deficiency markedly exacerbates Gaucher disease phenotypes in Gba1 mutant mice

Progranulin deficiency markedly exacerbates Gaucher disease phenotypes in Gba1 mutant mice

Gaucher disease: achievements and prospects

Gaucher disease (GD) is the most common lysosomal storage disorder, resulting from a deficiency in the activity of a lysosomal enzyme glucocerebrosidase, which is involved in the catabolism of sphingolipids. The phenomenal progress in understanding the pathogenesis and development of specific therapy of this disease over the past 60 years dramatically changed the clinical phenotype of GD, turning a severe progressive disorder into an asymptomatic metabolic defect. The evolution of the understanding of GD associated with fundamental discoveries in the field of cell biology, biochemistry and genetics may be of interest to a wide audience as a model of the effective work of the scientific community in the treatment of rare metabolic pathology.

Clinical Characteristics, Molecular Background, and Survival of Egyptian Patients With Gaucher Disease Over a 20-Year Follow-up.

This study analyzes the general disease characteristics, impact of enzyme replacement therapy (ERT), and overall survival (OS) of 156 Egyptian patients with Gaucher disease (GD) enrolled on hormone replacement from 1998 to 2017. The mean age at diagnosis was 32.46±12.68 months. Anemia was noted at diagnosis in 50%, thrombocytopenia in 30.7%, severe splenomegaly in 58.7%, severe hepatomegaly in 11.9%, and skeletal findings were detected in 24.3% of the patients. The most prevalent GD type was type 3 (54.5%). Twenty-two of type 3 patients had no neurological manifestations at diagnosis, and 12 developed variable central nervous system manifestations during follow-up. The most common neurological features were limited eye movements, oculomotor apraxia, and squint. Of the 60 patients for whom genotypes were obtained, homozygous L444P was the most common (n=35/60, 58.3%). Treatment with ERT (imiglucerase) revealed significant improvements in blood indices, organ volumes, and growth parameters (P<0.05). Ten (11.7%) type 3 patients did not develop any neurological manifestations under ERT over 20 years. Mortality was 16%, and the 20-year OS was 73.3%. We conclude that in Egypt, type 3 is the most prevalent phenotype of GD, and homozygous L444P is the predominant GBA genotype of GD. Early age at diagnosis and treatment with ERT over 20 years revealed significant improvements in disease manifestations, with an OS of 73.3%.

PR001 gene therapy increased GCase activity and improved neuronopathic Gaucher disease phenotypes

PR001 gene therapy increased GCase activity and improved neuronopathic Gaucher disease phenotypes

Genetic characterization of the Albanian Gaucher disease patient population.

Gaucher disease (GD) is a recessive metabolic disorder caused by a deficiency of the GBA gene‐encoded enzyme β‐glucocerebrosidase. We characterized a cohort of 36 Albanian GD patients, 31 with GD type 1 and 5 affected by GD types 2, 3, and an intermediate GD phenotype between type 2 and type 3. Of the 12 different GBA alleles that we detected, the most frequently observed was p.Asn409Ser, followed by p.[Asp448His;His294Gln]. The prevalence of the p.Leu483Pro allele was approximately 10‐fold lower than reported in other populations. We identified a novel pathogenic missense variant (c.1129G>A; p.Ala377Thr). All five of our non‐type 1 patients had genotypes consisting of the p.[Asp448His;His294Gln] allele in combination with another severe GBA allele. The median Lyso‐Gb1 level of treated patients carrying the p.[Asp448His;His294Gln] and no p.Asn409Ser allele was significantly higher than that of treated individuals homozygous or compound heterozygous for the p.Asn409Ser allele. In conclusion, the most important distinguishing features of the Albanian GD patient population are the underrepresentation of the p.Leu483Pro allele and an unusually high number of p.[Asp448His;His294Gln] alleles originating from a common Balkan founder event. The presence of at least one p.Asn409Ser allele is associated with mild disease and low Lyso‐Gb1 biomarker levels, while compound heterozygosity involving p.[Asp448His;His294Gln] and no p.Asn409Ser entails severe phenotypes and high Lyso‐Gb1 levels.

Biophysical Analysis of Lipid Domains by Fluorescence Microscopy.

The study of the structure and dynamics of membrane domains in vivo is a challenging task. However, major advances could be achieved through the application of microscopic and spectroscopic techniques coupled with the use of model membranes, where the relations between lipid composition and the type, amount and properties of the domains present can be quantitatively studied.This chapter provides protocols to study membrane organization and visualize membrane domains by fluorescence microscopy both in artificial membrane and living cell models of Gaucher Disease (GD ). We describe a bottom-up multiprobe methodology, which enables understanding how the specific lipid interactions established by glucosylceramide, the lipid that accumulates in GD , affect the biophysical properties of model and cell membranes, focusing on its ability to influence the formation, properties and organization of lipid raft domains. In this context, we address the preparation of (1) raft-mimicking giant unilamellar vesicles labeled with a combination of fluorophores that allow for the visualization and comprehensive characterization of those membrane domains and (2) human fibroblasts exhibiting GD phenotype to assess the biophysical properties of biological membrane in living cells using fluorescence microscopy.

Hyperimmunoglobulinemia and IgG Subclass Abnormalities in Children With Gaucher Disease.

Gaucher disease (GD) is the most common lysosomal storage disorder, the aim of the current study was to investigate hyperimmunoglobulinemia and abnormalities of serum immunoglobulin G (IgG) subclasses in children with GD and the relation of those findings to the GD phenotype and genotype, duration of enzyme replacement therapy (ERT), and infection frequency. The study included 20 Egyptian children with GD receiving ERT and 20 age-matched and sex-matched healthy children as controls. Serum Ig and serum IgG subclass levels were measured in the children with GD. Serum IgG subclass levels were measured in the control subjects. Hyperimmunoglobulinemia was present in 15 of the 20 (75%) children with GD. In addition, it is found significantly lower IgG2 levels and significantly higher IgG3 levels in the GD group than in the control group (P<0.001 and <0.006, respectively). Patients with 12 infections per year had significantly higher IgG3 levels compared with patients with 6 infections per year (P=0.022). In conclusion, hyperimmunoglobulinemia and IgG subclass abnormalities occur in children with GD who are on ERT and may be related to recurrent infections.

Enzyme replacement therapy in adult patients with type I Gaucher disease

Enzyme replacement therapy (ERT) is the standard for the treatment of Gaucher disease (GD). A lifelong intravenous administration of a recombinant analogue of human glucocerebrosidase compensates for the functional deficiency of its own enzyme. The use of ERT has changed the clinical phenotype of GD, a severe progressive disease has been turned into the status of an asymptomatic metabolic defect. At the same time, a reduced dosing ERT regimen applied in Gaucher patients who had achieved therapeutic goals has not yet been developed.

Gaucher Disease: New Expanded Classification Emphasizing Neurological Features.

Gaucher disease (GD) is a rare inherited metabolic disorder and the most common lysosomal storage disorder, caused by a deficiency in glucocerebrosidase enzyme activity. It has been classified according to the neurological manifestations into three types: type 1, without neuropathic findings, type 2 with acute infantile neuropathic signs and type 3 or chronic neuropathic form. However, report of new variants has led to the expansion of phenotype as a clinical phenotype of GD considered as a continuum of phenotypes. Therefore, it seems that a new classification is needed to cover new forms of the disease.

Brain-penetrant heat shock protein amplifier arimoclomol enhances GCase activity in in vitro Gaucher disease models

Brain-penetrant heat shock protein amplifier arimoclomol enhances GCase activity in in vitro Gaucher disease models

Progressive myoclonus epilepsy in Gaucher Disease due to a new Gly\u2013Gly mutation causing loss of an Exonic Splicing Enhancer

BackgroundPatients with Gaucher Disease (GD) exhibit three phenotypes, including type 1 (non-neuronopathic), type 2 (acute neuronopathic), and type 3 (subacute neuronopathic).AimIdentifying which GBA changes represent benign polymorphisms and which may result in disease-causing mutations is essential for diagnosis and genotype/phenotype correlations but is often challenging.ResultsHere, we describe a patient with type 3 GD, presenting with drug-resistant epilepsy, who bears a set of GBA polymorphic variants including the novel c.363A > G (Gly82Gly) synonymous mutation. In silico predictions, mRNA and functional studies revealed that the new Gly82Gly mutation causes skipping of GBA exon 4, leading to a severe reduction of the wild type GBA mRNA. This is the first report of a synonymous change causing GD through loss of an exonic splicing enhancer sequence.The synonymous mutation is in trans with the Asn188Ser missense mutation, thus making the Asn188Ser responsible for the patient’s phenotype and strengthening the association of Asn188Ser with the particular neurological phenotype of type 3 GD.ConclusionWe strengthen the association of Asn188Ser with the type 3 GD phenotype and progressive myoclonus epilepsy. Our data confirm that in silico predictions and mRNA analysis are mandatory in discriminating pathological mutations from the background of harmless polymorphisms, especially synonymous changes.

Gaucher disease iPSC-derived osteoblasts have developmental and lysosomal defects that impair bone matrix deposition

Gaucher disease (GD) is caused by bi-allelic mutations in GBA1, the gene that encodes acid β-glucocerebrosidase (GCase). Individuals affected by GD have hematologic, visceral and bone abnormalities, and in severe cases there is also neurodegeneration. To shed light on the mechanisms by which mutant GBA1 causes bone disease, we examined the ability of human induced pluripotent stem cells (iPSC) derived from patients with Types 1, 2 and 3 GD, to differentiate to osteoblasts and carry out bone deposition. Differentiation of GD iPSC to osteoblasts revealed that these cells had developmental defects and lysosomal abnormalities that interfered with bone matrix deposition. Compared with controls, GD iPSC-derived osteoblasts exhibited reduced expression of osteoblast differentiation markers, and bone matrix protein and mineral deposition were defective. Concomitantly, canonical Wnt/β catenin signaling in the mutant osteoblasts was downregulated, whereas pharmacological Wnt activation with the GSK3β inhibitor CHIR99021 rescued GD osteoblast differentiation and bone matrix deposition. Importantly, incubation with recombinant GCase (rGCase) rescued the differentiation and bone-forming ability of GD osteoblasts, demonstrating that the abnormal GD phenotype was caused by GCase deficiency. GD osteoblasts were also defective in their ability to carry out Ca2+-dependent exocytosis, a lysosomal function that is necessary for bone matrix deposition. We conclude that normal GCase enzymatic activity is required for the differentiation and bone-forming activity of osteoblasts. Furthermore, the rescue of bone matrix deposition by pharmacological activation of Wnt/β catenin in GD osteoblasts uncovers a new therapeutic target for the treatment of bone abnormalities in GD.

A Global Delphi Consensus Initiative for Early Diagnosis of Gaucher Disease: Key Presenting Signs and Patient Co-Variables in Type 1 Disease

Background: Gaucher disease (GD) is associated with a broad spectrum of signs and symptoms, with phenotypes ranging from fatal perinatal to asymptomatic adult forms. Given the heterogeneous presentation and rarity of the disease, misdiagnosis is common and diagnosis is often delayed, which can lead to avoidable morbidities and potentially irreversible tissue damage. Among the three phenotypes of GD, type 1 is the most prevalent in the Western hemisphere. As part of the global Gaucher earlier diagnosis consensus (GED-C) initiative, we report here the signs and patient co-variables that are regarded by expert physicians as most indicative of type 1 GD in its early stages. The overarching goal of the GED-C initiative is to generate a web-based point-scoring system that can be used by clinicians across specialties to facilitate identification of patients who may benefit from diagnostic testing for GD.Methods: In an anonymous iterative Delphi process, a panel of expert physicians was asked to provide free-text answers to a series of open questions, including: "Which unexplained signs and co-variables may be important to consider in early type 1 GD?" An independent facilitator categorized responses from round 1 into themes, which were checked and consolidated by the two non-voting co-chairs of the initiative to generate a set of summary factors. In round 2, panel members independently rated the importance of each factor using a 5-point Likert scale (1 = not important, 5 = extremely important). Factors assigned an importance score of at least 3 by more than 75% of respondents were provisionally classified as major; other factors were classified as minor. In round 3, panel members rated their level of agreement with the provisional classification of factors as major using a 5-point pivoted Likert scale (1 = strongly disagree; 3 = neither agree nor disagree; 5 = strongly agree). Consensus was defined as more than 67% of respondents agreeing or strongly agreeing (a score of ≥ 4) with the classification; if consensus was not reached, factors were classified as minor.Results: In total, 22 physicians with expertise in type 1 GD were recruited to the GED-C panel from 16 countries. Round 1 (100% response, n = 22) yielded 104 phrases, which were grouped into 38 themes, then consolidated as 21 factors. In round 2 (100% response, n = 22), 10 factors were provisionally classified as major. In round 3 (100% response, n = 22), consensus was reached on 9 major factors in early type 1 GD, including 7 presenting signs and 2 patient co-variables. The mean importance scores (round 2) and agreement scores (round 3) awarded to these 9 major factors are given in Table 1. Minor factors included asthenia, bleeding or bruising, dyslipidaemia, fatigue, gallstones, growth retardation, low bone mineral density and a family history of Parkinson disease.Table 1. Mean scores of importance and agreement for 9 major factors in rounds 2 and 3, respectively.Discussion: Definitive diagnostic tests for GD have been available for several years, but patient referral for testing is impeded by several issues, including a general lack of knowledge among clinicians of the signs and co-variables that should arouse a suspicion of GD. The presenting signs and patient co-variables identified by this multidisciplinary consensus initiative will help clinicians to identify those patients who may benefit from diagnostic testing for GD. Several algorithms have been devised with the aim of facilitating GD diagnosis, but these may be perceived as complex by the non-specialist. The next stage of the GED-C initiative will be to use the factors identified here to create a point-scoring system that clinicians of any specialty can use to obtain clear direction regarding the need to test a patient for GD.Acknowledgment: Submitted on behalf of the GED-C panel members and the European Hematology Association Scientific Working Group 'Quality of Life and Symptoms'. Administration of the GED-C initiative was funded by an unrestricted educational grant from Shire. [Display omitted] DisclosuresKuter:Amgen: Consultancy; Eisai: Consultancy; Genzyme: Consultancy; GlaxoSmithKline: Consultancy; ONO: Consultancy; Pfizer: Consultancy; Shionogi: Consultancy; Shire: Consultancy; 3SBios: Consultancy; Bristol-Myers Squibb: Research Funding; Protalix: Research Funding; Rigel: Research Funding. Salek:Agios: Consultancy; Servier: Consultancy; Shire: Consultancy; Bristol-Myers Squibb: Research Funding; Novartis: Research Funding; Sanofi: Research Funding. Mehta:Actelion: Honoraria, Other: Travel grants, Research Funding; Genzyme: Honoraria, Other: Travel grants, Research Funding; Protalix/Pfizer: Honoraria, Other: Travel grants, Research Funding; Shire: Honoraria, Other: Travel grants, Research Funding.

Key Presenting Signs and Patient Co-Variables in Early Diagnosis of Type 3 Gaucher Disease: A Global Delphi Consensus Initiative

Background: Among the three phenotypes of Gaucher disease (GD), type 1 is the most prevalent in the Western hemisphere, but types 2 and 3 are increasingly seen, and occur with similar prevalence to type 1 in parts of Asia. There is a spectrum of signs and symptoms among these phenotypes, which range from fatal perinatal to asymptomatic adult disease, and this heterogeneity contributes to relatively high levels of misdiagnosis and delays in diagnosis of GD. As part of the global Gaucher earlier diagnosis consensus (GED-C) initiative, we report here the signs and patient co-variables that are regarded by expert physicians as most indicative of type 3 GD in its early stages. The overarching goal of the GED-C initiative was to generate a web-based point-scoring system for use across clinical specialties to facilitate identification of patients who may benefit from diagnostic testing for GD.Methods: In an anonymous, iterative Delphi process, a panel of expert physicians was asked to provide free-text answers to a series of open questions, including: "Which unexplained signs and co-variables may be important to consider in early type 3 GD?" An independent facilitator categorized responses from round 1 into themes, which were checked and consolidated by the two non-voting co-chairs of the initiative to generate a set of summary factors. In round 2, panel members independently rated the importance of each factor using a 5-point Likert scale (1 = not important, 5 = extremely important). Factors that were assigned an importance score of at least 3 by more than 75% of respondents were provisionally classified as major; other factors were classified as minor. In round 3, panel members rated their level of agreement with the provisional classification of factors as major using a 5-point pivoted Likert scale (1 = strongly disagree; 3 = neither agree nor disagree; 5 = strongly agree). Consensus was defined as more than 67% of respondents agreeing or strongly agreeing (a score of ≥ 4) with the classification; if consensus was not reached, factors were classified as minor.Results: In total, 19 physicians with expertise in type 3 GD were recruited to the GED-C panel from 14 countries. Round 1 (100% response, n = 19) yielded 70 phrases, which were grouped into 34 themes, then consolidated as 23 factors. In round 2 (100% response, n = 19), 16 factors were provisionally classified as major. In round 3 (100% response, n = 19), consensus was reached on 10 major factors in early type 3 GD, including 9 presenting signs and 1 patient co-variable. The mean importance scores (round 2) and agreement scores (round 3) awarded to these 10 major factors are given in Table 1. Minor factors included bleeding or bruising, cardiovascular calcification, cognitive deficit, growth retardation, hyperferritinaemia, aged 18 years or younger, Jewish ancestry and a family history of Parkinson disease.Discussion: Definitive diagnostic tests for GD have been available for several years, but patient referral for testing is impeded by several issues, including a lack of knowledge among clinicians of the signs and co-variables that should arouse suspicion of GD. In type 3 GD, the problem is exacerbated by the relative rarity of the phenotype. Presenting signs and patient co-variables identified by this multidisciplinary consensus initiative will help clinicians identify those patients who may benefit from diagnostic testing for GD. Several algorithms have been devised to facilitate GD diagnosis, but non-specialists may perceive these as complex. The GED-C initiative will use the factors identified here to create a point-scoring system that clinicians of any specialty can use to obtain clear direction regarding the need to test a patient for GD.Acknowledgment: Submitted on behalf of the GED-C panel members and the European Hematology Association Scientific Working Group 'Quality of Life and Symptoms'. Administration of the GED-C initiative was funded by an unrestricted educational grant from Shire. [Display omitted] DisclosuresKuter:Eisai: Consultancy; Genzyme: Consultancy; MedImmune: Consultancy; Rigel: Consultancy, Research Funding; CRICO: Other: Paid expert testimony; Pfizer: Consultancy; Protalex: Research Funding; GlaxoSmithKline: Consultancy; Amgen: Consultancy, Paid expert testimony; Bristol-Myers Squibb: Research Funding; ONO: Consultancy; Shionogi: Consultancy; Shire: Consultancy; Syntimmune: Consultancy; 3SBios: Consultancy. Salek:Novartis: Research Funding; Bristol-Myers Squibb: Research Funding; Shire: Consultancy; Servier: Consultancy; Sanofi: Research Funding; Agios: Consultancy. Mehta:Protalix/Pfizer: Honoraria, Other: travel grant, Research Funding; Genzyme: Honoraria, Other: travel grant, Research Funding; Actelion: Honoraria, Other: travel grant, Research Funding; Shire: Honoraria, Other: travel grant, Research Funding.

GBA p.T369M substitution in Parkinson disease: Polymorphism or association? A meta-analysis

The lysosomal enzyme glucocerebrosidase (GCase), encoded by GBA, has an important role in Parkinson disease (PD). GBA mutation carriers have an increased risk for PD, earlier age at onset, faster progression, and various nonmotor symptoms including cognitive decline, REM sleep behavior disorder, hyposmia, and autonomic dysfunction.(1) Furthermore, GCase enzymatic activity is reduced in the peripheral blood(2) and brain(3) of noncarrier, sporadic PD patients. Biallelic GBA mutations, which have been classified as "severe" or "mild," may cause Gaucher disease (GD), a lysosomal storage disorder. Mild mutations may lead to GD type 1, and 2 severe mutations result in neuronopathic GD (type 2 and type 3).(4) There are 2 GBA variants, p.E326K and p.T369M, which do not cause GD in homozygous carriers, but may modify GCase activity and GD phenotype. It is now clear that p.E326K is a risk factor for PD,(5) but whether p.T369M is associated with PD is still controversial. In some studies, the p.T369M substitution was associated with PD,(6) while in others it had similar or increased frequency in controls. Of interest, it was recently demonstrated that the GBA p.T369M substitution was associated with reduced enzymatic activity in patients with PD and controls compared with that in noncarriers (7.64 vs 11.93 μmol/L/h, p < 0.001).(2) Of interest, it was even lower than the average enzymatic activity of the p.E326K substitution, which was 9.81 μmol/L/h. Because clinical trials on GBA-associated PD are ongoing, and because treatment specifically targeting GBA is likely to be available in the future, it is important to determine whether the GBA p.T369M substitution is associated with PD.

Properties of neurons derived from induced pluripotent stem cells of Gaucher disease type 2 patient fibroblasts: potential role in neuropathology.

Gaucher disease (GD) is caused by insufficient activity of acid β-glucosidase (GCase) resulting from mutations in GBA1. To understand the pathogenesis of the neuronopathic GD, induced pluripotent stem cells (iPSCs) were generated from fibroblasts isolated from three GD type 2 (GD2) and 2 unaffected (normal and GD carrier) individuals. The iPSCs were converted to neural precursor cells (NPCs) which were further differentiated into neurons. Parental GD2 fibroblasts as well as iPSCs, NPCs, and neurons had similar degrees of GCase deficiency. Lipid analyses showed increases of glucosylsphingosine and glucosylceramide in the GD2 cells. In addition, GD2 neurons showed increased α-synuclein protein compared to control neurons. Whole cell patch-clamping of the GD2 and control iPSCs-derived neurons demonstrated excitation characteristics of neurons, but intriguingly, those from GD2 exhibited consistently less negative resting membrane potentials with various degree of reduction in action potential amplitudes, sodium and potassium currents. Culture of control neurons in the presence of the GCase inhibitor (conduritol B epoxide) recapitulated these findings, providing a functional link between decreased GCase activity in GD and abnormal neuronal electrophysiological properties. To our knowledge, this study is first to report abnormal electrophysiological properties in GD2 iPSC-derived neurons that may underlie the neuropathic phenotype in Gaucher disease.

Characterization of Variants in the Glucosylceramide Synthase Gene and their Association with Type 1 Gaucher Disease Severity

The extreme phenotypic variability of patients with Gaucher disease (GD) is not completely explained by glucocerebrosidase gene mutations. It has been proposed that genetic modifiers might influence GD phenotype. We examined seven polymorphisms of the glucosylceramide synthase gene (UGCG) and their correlation with severity of GD. Five UGCG variants were significantly associated with disease severity, according to the DS3 scoring system: c.-295C>T, c.-232_-241ins10, c.98+50A>G, c.98+68A>T, and c.861A>G. Heterozygous [N370S]+[L444P] patients with c.[-232_-241ins10;98+50G] haplotype had a significantly lower DS3 score in relation to patients carrying only one of these polymorphisms. Electrophoretic mobility shift assay analysis showed an increased nuclear protein binding ability for the G allele at the cDNA position c.98+50, as well as an altered pattern for the c.-232_-241ins10 allele. The promoter activity of the haplotypes decreased significantly with respect to wild type activity in HepG2 and COS-7 cells (-14% and -16% for the c.[-232_-241ins10;98+50A] haplotype, -44% and -25% for c.[-222nonins;98+50G] haplotypes, and -64% and -75% for c.[-232_-241ins10;98+50G] haplotype, respectively). These data indicate that the c.-232_-241ins10 and c.98+50A>G variants are modifying factors of GD severity, which can partly explain the variability in severity of the disease.

Selective reduction of bis(monoacylglycero)phosphate ameliorates the storage burden in a THP-1 macrophage model of Gaucher disease

Bis(monoacylglycero)phosphate (BMP) assists lysosomal function by facilitating interaction of hydrolases and activator proteins with sphingolipid substrates. Impaired lysosomal degradation of the sphingolipid glucosylceramide (GC) occurs in Gaucher disease due to an inherited deficiency of acid β-glucosidase, with secondary BMP alterations. We investigated the nature of BMP accumulation and whether its correction reduced the storage burden in a THP-1 macrophage model of Gaucher disease. Using sucrose gradients and detergent solubility, 98% of BMP resided in the detergent-soluble membranes (DSM) rather than in the detergent-resistant membranes (DRM) where 73% of GC predominated. There was a 2-fold widespread elevation in BMP, including the saturated, mono- and polyunsaturated species. Linoleic acid in the culture media selectively reduced BMP from 4.2 nmol/mg to 0.49 nmol/mg (except 18:1/18:2) and prevented up to one third of GC, dihexosylceramide (DHC), and trihexosylceramide (THC) from accumulating. The 2-fold reduction in these sphingolipids occurred only in the DRM and did not reduce 18:1/16:0. However, once GC had accumulated, linoleic acid could not reverse it, DHC, or THC, despite effectively reducing BMP. These results imply a causative link for BMP in the pathobiology of Gaucher disease and demonstrate that linoleic acid can shield the cell from excessive substrate accumulation.