55-Base pair deletion in certain patients with Gaucher disease complicates screening for common Gaucher alleles

Mutations in the glucocerebrosidase gene which result in Gaucher disease can originate from the highly homologous glucocerebrosidase pseudogene. A 55-bp deletion in exon 9, which corresponds to a 55-bp segment absent from the pseudogene, has been identified in patients with Gaucher disease. We have developed a simple polymerase chain reaction (PCR)-based method to detect this 55-bp deletion, and have found this mutation in 3 of 75 DNA samples (4%) collected from patients with Gaucher disease. Commonly used PCR-based screening methods for specific Gaucher mutations frequently make use of primers either within or surrounding the 55-bp gap to selectively distinguish the glucocerebrosidase gene from the pseudogene. However, if the 55-bp deletion in exon 9 occurs, primers will either fail to produce an amplification product or will produce a shortened product which will be falsely attributed to the pseudogene. This could lead to inaccurate genotyping and genetic counseling for some Gaucher patients and their families. We therefore recommend that laboratories using PCR-based screening techniques involving primers in this region initially determine whether this 55-bp sequence is present. © 1996 Wiley-Liss, Inc.     

Movement and mood disorder in two brothers with Gaucher disease

Gaucher disease (GD) is a lysosomal storage disorder with a wide spectrum of phenotypic presentations. We report the case histories of two adult brothers with GD who developed both parkinsonism and psychiatric symptoms. Direct sequencing and real-time polymerase chain reaction were used to establish that the patients were homozygous for mutation L444P. While parkinsonism has been described previously in GD, these patients had atypical features, including a complicated mood disorder. The comorbidity of GD and a mood disorder is a new finding, as psychiatric manifestations of GD have been described rarely. The etiology of the mental illness could be related to the processes contributing to the development of parkinsonism.     

DNA mutation analysis of Gaucher patients

We evaluated 62 Gaucher patients to determine whether patients with similar phenotypes had the same DNA point mutations. Genomic DNA from these Gaucher patients was screened for the 3 most frequent single-point mutations, occurring in 69% of the 124 patient alleles, and resulting in changes in amino acids 370, 444, and 463. Many different genotypes were observed, at least one of which is present in all 3 types of Gaucher disease. No specific symptom complex could be correlated with a unique genotype. Even the more clinically homogeneous subgroups of Gaucher patients contained several genotypes. This study further emphasizes the need for caution in making clinical predictions on the basis of current genotype analysis, especially since one might not discern a fetus affected with type 2 disease by current DNA studies. The severity of involvement in type 1 disease could also not be predicted. Thus, even limiting our focus to 3 isolated common point mutations, a given genotype cannot be uniquely correlated with a specific prognosis.     

Novel mutations in type 2 Gaucher disease in Chinese and their functional characterization by heterologous expression

We investigated 10 unrelated Chinese patients with type 2 Gaucher disease and performed ex vivo expression for the novel mutations to characterize their functional defects. These patients were diagnosed by enzymatic assays and clinicopathologic features over the past five years in a national centre in China. Genomic DNA was sequenced by a two-stage PCR approach for mutations in the functional GBA gene. Novel mutations were expressed with baculovirus-transfected Sf21 cells. Six novel mutations were found (in traditional nomenclature): P122L, Y363C, N382K, L383R, L385P, and M416V. Review of reported mutations indicated clustering of type 2 mutations in three regions of the GBA gene. Expression of novel mutations revealed that the enzyme defect could arise from one of two mechanisms: loss of catalytic activity (Y363C and M416V) or enzyme instability (P122L and N382K).     

Two new mild homozygous mutations in Gaucher disease patients: Clinical signs and biochemical analyses

Gaucher disease (GD) is a lysosomal storage disorder resulting from impaired activity of lysosomal β-glucocerebrosidase. More than 60 mutations have been described in the GBA gene. They have been classified as lethal, severe, and mild on the basis of the corresponding phenotype. The fact that most GD patients are compound heterozygous and that most type 1 patients bear the N370S allele, which by itself causes a mild phenotype, make it difficult to correlate the clinical signs with the mutations. Besides N370S, about 10 mild mutations have been described, but only one undoubtedly classified as mild was found at homozygosity. Here we report 2 novel mutations, I402T and V375L, at homozygosity in 2 adult Italian type 1 GD patients. Some properties of the I402T fibroblast enzyme have been compared to those of the enzyme from cells of several N370S/N370S patients. Analysis of the catalytic properties and heat stability as well as the response to phosphatidylserine and sphingolipid activator protein indicate a marked similarity between the 2 enzymes. The finding of another, unrelated patient bearing the I402T mutation (in this case as a compound heterozygote with mutation N370S) suggests that this allele might be quite frequent in the area of Sicily from where both patients originated. In conclusion, the phenotypic expression in the 2 homozygous patients presented here and the biochemical data for one of them allowed the classification of these mutations as mild thus extending the group of mild mutations found at homozygosity. Am. J. Med. Genet. 70:437–443, 1997. © 1997 Wiley-Liss, Inc.     

Gaucher disease paradigm: From ERAD to comorbidity

Mutations in the GBA gene, encoding the lysosomal acid beta‐glucocerebrosidase (GCase), lead to deficient activity of the enzyme in the lysosomes, to glucosylceramide accumulation and to development of Gaucher disease (GD). More than 280 mutations in the GBA gene have been directly associated with GD. Mutant GCase variants present variable levels of endoplasmic reticulum (ER) retention, due to their inability to correctly fold, and undergo ER‐associated degradation (ERAD) in the proteasomes. The degree of ER retention and proteasomal degradation is one of the factors that determine GD severity. In the present review, we discuss ERAD of mutant GCase variants and its possible consequences in GD patients and in carriers of GD mutations. Hum Mutat 33:1398–1407, 2012. © 2012 Wiley Periodicals, Inc.     

Type I Gaucher disease due to homozygosity for the 259T mutation in a Bedouin patient

A 26-year-old Bedouin with moderate thrombocytopenia and enlarged spleen and liver was diagnosed as having type I Gaucher disease based on the presence of Gaucher cells in the bone marrow biopsy and enzymatic determination of glucocerebrosidase activity. Molecular analysis excluded 10 common mutations in the glucocerebrosidase gene. Homozygosity for the C → T mutation in nucleotide 259 of the cDNA (1763 genomic) was detected by digestion with restriction enzyme StyI after an amplification of a portion of exon 3 by mismatched primers. This is the first known case of homozygosity for this mutation. The fact that it produces a very mild phenotype, confirms a previous suggestion that 259T can be classified as a “mild” mutation. Association of the 259T mutation with the “Pv 1.1 +” haplotype is consistent with a common origin of the mutated alleles. Am. J. Med. Genet. 72:77–78, 1997. © 1997 Wiley-Liss, Inc.     

A mutation in SCARB2 is a modifier in Gaucher disease

Lysosomal integral membrane protein type 2 (LIMP-2) is responsible for proper sorting and lysosomal targeting of glucocerebrosidase, the enzyme deficient in Gaucher disease (GD). Mutations in the gene for LIMP-2, SCARB2, are implicated in inherited forms of myoclonic epilepsy, and myoclonic epilepsy is part of the phenotypic spectrum associated with GD. We investigated whether SCARB2 mutations impact the Gaucher phenotype focusing on patients with myoclonic epilepsy, including a pair of siblings with GD who were discordant for myoclonic seizures. Sequencing of SCARB2 genomic and cDNA identified a heterozygous, maternally inherited novel mutation, c.1412A>G (p.Glu471Gly), in the brother with GD and myoclonic epilepsy, absent from his sibling and controls. Glucocerebrosidase activity, Western blots, real-time PCR, and immunofluorescence studies demonstrated markedly decreased LIMP-2 and glucocerebrosidase in cells from the sibling with (p.Glu471Gly) LIMP-2, and diminished glucocerebrosidase in lysosomes. The cells secreted highly glycosylated enzyme and showed mistrafficking of glucocerebrosidase. Sequencing of SCARB2 in 13 other subjects with GD and myoclonic epilepsy and 40 controls failed to identify additional mutations. The study provides further evidence for the association of LIMP-2 and myoclonic epilepsy, explains the drastically different phenotypes encountered in the siblings, and demonstrates that LIMP-2 can serve as a modifier in GD.     

The D409H variant in GBA1: Challenges in predicting the Gaucher phenotype in the newborn screening era

Gaucher disease (GD) is an autosomal recessive disorder resulting from glucocerebrosidase deficiency due to pathologic variants in GBA1. While clinically heterogeneous, GD encompasses three types, non-neuronopathic (GD1), acute neuronopathic (GD2), and chronic neuronopathic (GD3). Newborn screening (NBS), which has made remarkable inroads in detecting certain diseases before detrimental health consequences and fatality ensues, is now being piloted for GD in several states and countries. Early on, clinical features of GD2 can overlap with GD3; hence, predicting outcome is challenging. As NBS for GD becomes more available, the increased detection of GD in neonates is inevitable. As a result, health care providers and families will be faced with uncertainty with respect to clinical management. Since more severe GBA1 variants are generally associated with neuronopathic GD, there has been an increased dependence on genotypic information. We present an infant detected by NBS with genotype D409H(p.Asp448His)/RecNciI (p.Leu483Pro; p.Ala495Pro;p.Val499=). To assist in genetic counseling, we performed a retrospective review of other patients in our cohort carrying D409H and reviewed the literature. The study illustrates the challenges faced in counseling for infants with neuronopathic GD, even with known GBA1 variants, and the tough management decisions that can ensue from detection in newborns.     

Gaucher disease and parkinsonism, a molecular link theory

Mutant GBA was found recently to be the most prevalent risk factor for familial parkinsonism. The two diseases do not share common symptoms and there is no direct pathway to explain the mechanism by which GBA mutations can confer the risk. Increased burden on the degradative pathway caused by defective glucocerebrosidase, or toxic side effects of glycosylated lipids accumulation were proposed to explain brain damage. Both hypotheses are not sufficient to explain the linkage. In order to develop a more inclusive theory we introduced into the model the prion theory and the second hit. Other possibilities are also brought into consideration.     

Glucocerebrosidase genotype of Gaucher patients in The Netherlands: Limitations in prognostic value

Gaucher disease is a recessively inherited lysosomal storage disorder that is caused by a deficiency in glucocerebrosidase activity. The clinical expression is markedly heterogeneous with respect to age of onset, progression, severity, and neurological involvement. The relative incidence of glucocerebrosidase (GC) mutations has been studied extensively for Jewish but not for non-Jewish Caucasian patient populations. The present survey on mutant GC genotypes prevalent in Gaucher disease in The Netherlands was taken of 72 patients from different genetic backgrounds. This number is more than half the total number of affected Gaucher patients to be expected on the basis of the incidence of the disorder in this country. Analysis of nine GC mutations led to the identification of 74% of the mutant GC alleles in patients from 44 unrelated Dutch families (i.e., families that have lived in The Netherlands for at least several generations) and of 44% of the mutant GC alleles in patients from nine unrelated families that recently immigrated from both European and non-European countries. The N370S (cDNA 1226G) GC mutation proved to occur most frequently (41%) in the unrelated Dutch patients and less frequently (6%) in the unrelated immigrant patients and was always associated with the nonneuronopathic (Type 1) form of the disease. Apart from the association of the N370S mutation with Type 1 Gaucher disease, the prognostic value of GC genotyping was limited, since a particular GC genotype did not correlate closely to a specific clinical course, or to a specific relative responsiveness to enzyme-supplementation therapy. Hum Mutat 10:348–358, 1997. © 1997 Wiley-Liss, Inc.     

Mutation analysis of 28 Gaucher disease patients: The Australasian experience

Gaucher disease is the most common lysosomal storage disease. It is an autosomal recessive disorder that results from a deficiency of β-glucocerebrosidase. Three clinical phenotypes have been described: non-neuronopathic, acute neuronopathic, and subacute neuronopathic. Genomic DNA from 28 Australasian patients of diverse ethnic origin with Gaucher disease was screened for 3 common mutations (1226G, 1448C and 84GG) using the amplification refractory mutation system (ARMS), and one uncommon mutation (1504T) by restriction enzyme digestion. Thirty-eight of the 56 independent alleles in these patients were characterized, with 1448C present in 42% and 1226G in 28% of the alleles. The 1226G mutation was associated only with the non-neuronopathic phenotype and 7 of the 15 patients who carried the 1448C mutation developed neuronopathic disease. Three infants who died in the neonatal period following a rapidly progressive neurodegenerative course carried no identifiable mutations. The 84GG mutation was carried by 2 Jewish patients and 1504T was present in one patient. It is now possible to rapidly identify the common Gaucher mutations using ARMS and restriction enzyme digestion, and our findings confirm the heterogeneity of mutations in Gaucher disease. It is also possible to predict in part the phenotypic outcome when screening patients for these mutations. We consider mutation analysis to be of most use in prenatal diagnosis and for carrier detection within affected families. © 1994 Wiley-Liss, Inc.     

Identification of two novel and four uncommon missense mutations among Chinese Gaucher disease patients

Gaucher disease is the most prevalent lysosomal storage disease. It is panethnic and results from an inherited deficiency of glucocerebrosidase. Most mutations to date have been identified among Jewish and non-Jewish Caucasian patients; mutations in Chinese patients are largely unknown. We have performed nucleotide sequence analysis of PCR-amplified glucocerebrosidase genomic DNA from five unrelated Chinese patients affected with type 1 (non-neuropathic) Gaucher disease. A novel heterozygous C → T mutation at cDNA nucleotide position 475 (R120W) was detected in a patient who is also heterozygous for a C → T transition at cDNA nucleotide position 259 (R48W). In a second patient, a novel, heterozygous T → G transversion at cDNA 226 (F37V) was detected. Mutation 1448 (L444P), the most prevalent mutation among non-Jewish Caucasian Gaucher patients, was found in the heterozygous form in four patients. The mutations in the second Gaucher allele in the other three patients are mutations 254 (G46E), 680 (N188S), and 754 (F213I), which were recently reported in Korean, Arab, and Chinese (Taiwanese) patients. We have developed screening methods that utilize PCR amplification of glucocerebrosidase genomic DNA and Eco571, Nci1, Hinc11, BsaJ1, and Bsr1 restriction endonuclease analyses for the detection of each of these mutations. The population genetics of some of these Gaucher alleles and their implications in genotype/phenotype correlation are discussed. Am. J. Med. Genet. 71:172–178, 1997. © 1997 Wiley-Liss, Inc.     

Linkage disequilibrium of common Gaucher disease mutations with a polymorphic site in the pyruvate kinase (PKLR) gene

Gaucher disease (GD), caused by a deficiency of the lysosomal enzyme glucocerebrosidase (GBA), is the most common human glycolipid storage disease. The incidence of the disease is particularly high in the Ashkenazi Jewish population, with a carrier frequency of 0.068. The 1226A→G and 84GG mutations are the two predominant disease-causing alleles. We investigated the association of various mutations in the GBA gene with different alleles of a highly polymorphic site in the adjacent pyruvate kinase (PKLR) gene. Ninety-seven unrelated type I GD patients of various genotypes were studied to determine their genotype for the PKLR gene trinucleotide repeat polymorphism. One hundred out of 104 (96%) alleles carrying the 1226G mutation also carried the A1 allele of the PKLR gene, which is present in only 6.7% of the control population. The calculated linkage disequilibrium between 1226G and the A1 allele of the PKLR gene is 0.957. Mutation 84GG was found to be uniquely associated with the PKLR A6 allele, with a linkage disequilibrium of 1.00. The association of several less frequent GD mutations with PKLR alleles was also studied. These results support the hypothesis that the 1226G and 84GG mutations in the Ashkenazi Jewish population each originated in a single founder. Further studies of the association of the 1226G and 84GG mutations with PKLR alleles in European non-Jewish GD patients could help in the study of the chronological order of these mutations and may shed light on the history of the Ashkenazi Jews in the past two millennia. Am. J. Med. Genet. 78:233–236, 1998. © 1998 Wiley-Liss, Inc.     

Activity of glucocerebrosidase in extracts of different cell types from type 1 Gaucher disease patients

Glucocerebrosidase activity in extracts of leukocytes, Epstein-Barr virus transformed lymphocytes and fibroblasts from Portuguese Type 1 Gaucher disease patients was studied. The residual glucocerebrosidase activity in all extracts from patients was less than 25% if measured in the presence of bile salt taurocholate. However, if measured in the absence of bile salt the residual enzyme activity in extracts from patients was cell type specific: it was severely reduced in the case of fibroblasts, mildly reduced in the case of lymphoblasts and not significantly reduced in the case of leukocytes. The glucocerebrosidase activity in extracts from all control cell types was stimulated by taurocholate. In the patients the enzyme activity in fibroblasts extracts was also stimulated but that in lymphoblasts and leukocytes was inhibited by the bile salt. The differences in glucocerebrosidase activity (in the absence of taurocholate) in extracts from different cell types from Gaucher disease patients are attributable to differences in the proportion of glucocerebrosidase present as a monomer with low activity (form I) and as a highly active aggregate (form II) that may also contain sphingolipid activator protein 2 (SAP-2). In extracts from leukocytes and lymphocytes from Type 1 Gaucher disease patients, but not in those from fibroblasts, a relatively high proportion of enzyme is present in aggregated form with near normal specific activity.     

Gaucher disease: N370S glucocerebrosidase gene frequency in the Portuguese population

In the Portuguese population the most frequent form of Gaucher disease is type 1. The N370S glucocerebrosidase gene mutation accounts for 63% of mutated alleles. The frequency of this mutation was accurately determined in the Portuguese population, which does not present an Ashkenazi Jewish genetic background. A gene frequency of 0.0043, with 95% confidence limits between 0.0023 and 0.0063, was obtained studying the genomic DNA of 2000 blood cards randomly sampled from the national neonatal screening program. On the basis of this frequency a significantly high number of homozygotes for the N370S mutation should be expected in the Portuguese population. This finding supports the idea that the majority of homozygotes for this mutation present a very mild clinical phenotype and remain undiagnosed.     

The identification of eight novel glucocerebrosidase (GBA) mutations in patients with Gaucher disease

Mutations in the gene encoding for the lysosomal enzyme glucocerebrosidase (GBA) result in Gaucher disease. In this study, seven novel missense mutations in the glucocerebrosidase gene (A136E, H162P, K198E, Y205C, F251L, Q350X and I402F) and a splice site mutation (IVS10+2T-->A) were identified by direct sequencing of three amplified segments of the glucocerebrosidase gene. Five of the novel mutations were found in patients with neuronopathic forms of Gaucher disease, two of which, K198E and F251L, appear to be associated with type 2 Gaucher disease.