Aberrant splicing at catalytic site as cause of infantile onset glycogen storage disease type II (GSDII): Molecular identification of a novel IVS9 (+2GT→GC) in combination with rare IVS10 (+1GT→CT)

Glycogen storage disease type II (GSDII) results from deleterious mutations in acid α‐glucosidase gene. To date several mutant alleles have been studied including missense and nonsense mutations, insertions, small and large deletions as well as splice site mutations. Apart from IVS1 (? 13→G), 525delT, and Δ18, the other mutations are rare and often unique to single patients. Moreover, the molecular findings also observed in the different ethnic groups makes it difficult to attempt to correlate genotype and phenotype to explain the origin of clinical variability. Even though there are no conclusive genotype phenotype correlations, the in frame splice site mutations identified up until now have been found associated with the juvenile/adult onset of GSDII. In this study we describe a novel in frame splicing defect, IVS9 (+2GT→GC), identified in combination with the rare IVS10 (+1GT→CT) mutation in a patient with classic infantile GSDII disease. Because both mutations occur at the catalytic site region, it is likely that the alteration of both catalytic function and steric conformation of the enzyme may be responsible for the most severe form of the disease. © 2001 Wiley‐Liss, Inc.     

Biochemical genetics of the Lapland dog model of glycogen storage disease type II (acid α-glucosidase deficiency)

A recently described canine model (Lapland dog) of glycogen storage disease type II (GSD II, Pompe disease, acid α-glucosidase deficiency) was identified with several biochemical genetic methods. Complementation studies in which fibroblasts from a GSD II dog were fused with fibroblasts derived from control dogs and from human patients with different clinical forms of the disease did not lead to restoration of acid α-glucosidase activity in the heterokaryon cell populations. These results indicate that acid α-glucosidase deficiency is the primary defect in canine GSD II and that there is a close genetic parallelism with human GSD II. Immunotitration analysis of the residual acid α-glucosidase activity in the canine GSD II fibroblasts and liver demonstrated that this residual activity was not due to acid α-glucosidase enzyme, in which respect canine GSD II was similar to the infantile form of the human disease. Double immunodiffusion studies showed the presence of catalytically inactive acid α-glucosidase enzyme protein in canine GSD II. This is consistent with a structural gene mutation. It is concluded that canine GSD II in the Lapland dog is a homologous model of the infantile form of human GSD II, a conclusion in concordance with clinical and pathological studies.     

Twenty-two novel mutations in the lysosomal alpha-glucosidase gene (GAA) underscore the genotype-phenotype correlation in glycogen storage disease type II

Patients with glycogen storage disease type II (GSDII, Pompe disease) suffer from progressive muscle weakness due to acid alpha-glucosidase deficiency. The disease is inherited as an autosomal recessive trait with a spectrum of clinical phenotypes. We have investigated 29 cases of GSDII and thereby identified 55 pathogenic mutations of the acid alpha-glucosidase gene (GAA) encoding acid maltase. There were 34 different mutations identified, 22 of which were novel. All of the missense mutations and two other mutations with an unpredictable effect on acid alpha-glucosidase synthesis and function were transiently expressed in COS cells. The effect of a novel splice-site mutation was investigated by real-time PCR analysis. The outcome of our analysis underscores the notion that the clinical phenotype of GSDII is largely dictated by the nature of the mutations in the GAA alleles. This genotype-phenotype correlation makes DNA analysis a valuable tool to help predict the clinical course of the disease.     

Development of a clinical assay for detection of GAA mutations and characterization of the GAA mutation spectrum in a Canadian cohort of individuals with glycogen storage disease, type II

Glycogen storage disease, type II (GSDII; Pompe disease; acid maltase deficiency) is an autosomal recessive disease caused by mutations of the GAA gene that lead to deficient acid alpha-glucosidase enzyme activity and accumulation of lysosomal glycogen. Although measurement of acid alpha-glucosidase enzyme activity in fibroblasts remains the gold standard for the diagnosis of GSDII, analysis of the GAA gene allows confirmation of clinical or biochemical diagnoses and permits predictive and prenatal testing of individuals at risk of developing GSDII. We have developed a clinical molecular test for the detection of GAA mutations based on cycle sequencing of the complete coding region. GAA exons 2-20 are amplified in six independent PCR using intronic primers. The resulting products were purified and sequenced. Preliminary studies using this protocol were conducted with DNA from 21 GSDII-affected individuals from five centers across Canada. In total, 41 of 42 mutations were detected (96.7% detection rate). Mutations spanned intron 1 through exon 19 and included nine novel mutations. Haplotype analysis of recurrent mutations further suggested that three of these mutations are likely to have occurred independently at least twice. Additionally, we report the identification of the c.-32-13T>G GAA mutation in an individual with infantile variant GSDII, despite reports of this mutation being associated almost exclusively with late-onset forms of the disease. The development of a clinical molecular test provides an important tool for the management and counseling of families and individuals with GSDII, and has provided useful information about the GAA mutation spectrum in Canada.     

Mutation profile of the GAA gene in 40 Italian patients with late onset glycogen storage disease type II

Glycogen storage disease type II (GSDII) is a recessively inherited disorder due to the deficiency of acid alpha-glucosidase (GAA) that results in impaired glycogen degradation and its accumulation in the lysosomes. We report here the complete molecular analysis of the GAA gene performed on 40 Italian patients with late onset GSDII. Twelve novel alleles have been identified: missense mutations were functionally characterized by enzyme activity and protein processing in a human GAA-deficient cell line while splicing mutations were studied by RT-PCR and in silico analysis. A complex allele was also identified carrying three different alterations in cis. The c.-32-13T > G was the most frequent mutation, present as compound heterozygote in 85% of the patients (allele frequency 42.3%), as described in other late onset GSDII Caucasian populations. Interestingly, the c.-32-13T > G was associated with the c.2237G > A (p.W746X) in nine of the 40 patients. Genotype-phenotype correlations are discussed with particular emphasis on the subgroup carrying the c.-32-13T > G/c.2237G > A genotype.     

The identification of five novel mutations in the lysosomal acid a-(1,4) glucosidase gene from patients with glycogen storage disease type II

The autosomal recessive disorder Glycogen Storage Disease Type II (GSDII) is caused by a deficiency in the lysosomal enzyme acid α-glucosidase. We have optimised a procedure to use fluorescent DNA sequencing technology to screen for mutations within the α-glucosidase gene from UK patients with GSDII. Five previously unknown mutations in six patients (4 early onset infantile and 2 late onset adult) have been found. The mutations are an insertion of a C residue in exon 2 (InsC258), an insertion of a G residue in exon 16 (InsG2242), a deletion of 20 nucleotides in exon 4 Δ, and a nonsense mutation in exon 16 (G2237A - Trp746Stop). All will result in the introduction of a premature stop codon in the coding region, predicting a truncated and non-functional protein. The final mutation is a duplication of 18 nucleotides in exon 19 (Ins18nt2776) and will result in the insertion of an additional six amino acids into the protein chain after Asn925 (Gly-Val-Pro-Val-Ser-Asn). Hum Mutat 11:413, 1998. © 1998 Wiley-Liss, Inc.     

Frequent mutation in Chinese patients with infantile type of GSD II in Taiwan: Evidence for a founder effect

Glycogen storage disease type II (GSD II, Pompe's disease), an autosomal recessive inherited disease, is caused by the deficiency of acid α-D-glucosidase, which results in the impaired glycogen degradation in lysosome and causes excess glycogen accumulation in lysosome. In Taiwan, the infantile form of GSD II is the most common type of glycogen storage diseases. The frequency of C1935A mutant allele is 0.8 in these Chinese patients. In this study, we analyzed four single point polymorphic markers (324, 1203, 2065, 2338) by ACRS-based RFLP. We observed that the alleles possessing the C1935A mutation in 19 of 25 Chinese patients who were heterozygous or homozygous have conserved polymorphic markers, and all of C1935A mutant alleles in these patients are linked to a specific haplotype. The allele frequency of this specific haplotype in 19 Chinese patients and in 42 normal individuals is 0.95 and 0.17, respectively (P<0.005,χ² = 66.018). This result suggests that the C1935A mutation in Chinese patients with infantile form of GSD II is due to the founder effect. Hum Mutat 11:306–312, 1998. © 1998 Wiley-Liss, Inc.     

Nosology of lysosomal glycogen storage diseases without in vitro acid maltase deficiency. Delineation of a neonatal form

We describe a boy with an early lethal hypertrophic vacuolar cardiomyopathy of neonatal onset. Abnormal intra- and extralysosomal glycogen storage disease was demonstrated in heart and skeletal muscles. Glycogen content was twice the normal in muscles and over 3-fold the normal in the heart. In this organ, over 50% of the intracellular space was occupied by glycogen and possibly oligosaccharides, as demonstrated by the quantitative morphometric analysis of electron micrographs. The activity of acid α-glucosidase was increased in the heart, skeletal muscles, and liver, but was normal in leukocytes. A review of the 11 previously published pedigrees of lysosomal glycogen storage disease with normal in vitro α-glucosidase activity allows the delineation of three clinical entities: juvenile and neonatal pseudo-Pompe diseases and partial Pompe disease. Partial Pompe disease, due to the tissue-specific absence of acid α-glucosidase, was observed in a single patient. The most common form is the late-onset pseudo-Pompe disease, which is characterized by severe cardiomyopathy and mild myopathy appearing in the second or third decade, prominent arrhythmia with Wolf-Parkinson-White syndrome, and sometimes mental retardation. Patients reported as suffering from Antopol disease probably belong to this group. Dominant inheritance (autosomal or X linked) is likely in most families. The present report appears to be the first one to describe a rapidly fatal neonatal form of lysosomal glycogenosis without acid maltase deficiency. The mode of inheritance of this form is not known. Differential diagosis includes Pompe disease (similar histology) and cardiac phosphorylase b kinase deficiency (similar clinical course). The delineation of neonatal pseudo-Pompe disease makes enzymatic confirmation mandatory in each case suspected of Pompe disease. Am. J. Med. Genet. 72:135–142, 1997. © 1997 Wiley-Liss, Inc.     

A nonsense mutation due to a single base insertion in the 3′-coding region of glycogen debranching enzyme gene associated with a severe phenotype in a patient with glycogen storage disease type IIIa

Glycogen storage disease type III (GSD-III) is an autosomal recessive disease resulting from deficient glycogen debranching enzyme (GDE) activity. A child with GDE deficient in both liver and muscle (GSD-IIIa) had recurrent hypoglycemia, seizures, severe cardiomegaly, and hepatomegaly and died at 4 years of age. Analysis of the GDE gene in this child by single-strand conformation polymorphism, followed by direct DNA sequencing and restriction analysis, revealed an insertion of a nucleotide A into position 4529 of the GDE cDNA (4529insA). This insertion resulted in substitution of a tyrosine to a stop codon at amino acid 1510 (Y1510X). The 4529insA mutation appeared to be homozygous in this patient and was not found in 20 unrelated controls or 18 other GSD-III patients (14 GSD-IIIa and 4 GSD-IIIb). This is the first identification of a disease mutation in this gene, and the data suggest that homozygous 4529insA may be associated with a severe phenotype in GSD-IIIa. © 1997 Wiley-Liss, Inc.     

Compound heterozygous patient with glycogen storage disease type III: Identification of two novel AGL mutations,a donor splice site mutation of Chinese origin and a 1‐bp deletion of Japanese origin

Glycogen storage disease type III (GSD III) is an autosomal recessive disorder caused by deficiency of glycogen‐debranching enzyme (AGL). We studied a 2‐year‐old GSD III patient whose parents were from different ethnic groups. Nucleotide sequence analysis of the patient showed two novel mutations: a single cytosine deletion at nucleotide 2399 (2399delC) in exon 16, and a G‐to‐A transition at the +5 position at the donor splice site of intron 33 (IVS33+5G>A). Analysis of the mRNA produced by IVS33+5G>A showed aberrant splicing: skipping of exon 33 and activation of a cryptic splice site in exon 34. Mutational analysis of the family revealed that the 2399delC was inherited from her father, who is of Japanese origin, and the IVS33+5G>A from her mother, who is of Chinese descent, establishing that the patient was a compound heterozygote. To our knowledge, this is the first report of a mutation identified in a GSD III patient from the Chinese population. Am. J. Med. Genet. 93:211–214, 2000. © 2000 Wiley‐Liss, Inc.     

Pompe disease: design, methodology, and early findings from the Pompe Registry

Pompe disease is an autosomal recessive, progressive, debilitating, and often fatal neuromuscular disorder caused by deficiency of lysosomal acid α-glucosidase (GAA). It is characterized by the accumulation of glycogen in muscle tissue that leads to progressive muscle weakness and loss of function. It presents as a broad spectrum of clinical phenotypes, with varying rates of progression, symptom onset, degree of organ involvement, and severity. The Pompe Registry represents worldwide data collection on this rare and clinically heterogeneous disease. This report describes the design, methodology, and early findings from the Registry and presents an overview of the Registry population over a five-year period from its inception in September 2004 through September 2009. Among the 742 patients from 28 countries in the Registry, 70% (517/742) reported symptom onset >12 months of age and 23% (170/742) reported symptom onset ≤12 months of age. Seventy-eight percent (582/742) of registry patients have received enzyme replacement therapy. Overall, Registry data appear to be consistent with smaller natural history studies in terms of symptoms and disease course in classical infantile Pompe disease (≤12 months of age with cardiomyopathy) and late-onset Pompe disease (>12 months of age). In addition, a subset of patients with symptom onset ≤12 months of age do not have cardiomyopathy (14.7%); these patients appear to have a later age at first symptoms and diagnosis than their peers with cardiomyopathy. As the largest dataset on Pompe disease to date, the Pompe Registry will serve to improve recognition of the disease, enhance understanding of the variable disease course, and offer insights into treated and untreated disease course.     

Six novel β‐galactosidase gene mutations in Brazilian patients with GM1‐gangliosidosis

GM1‐gangliosidosis is a lysosomal storage disease caused by a deficiency of acid β‐galactosidase. Three clinical forms are recognized—infantile, juvenile, and adult—based on age of onset and severity of the symptoms. We have performed molecular analysis of a large cohort of GM1 patients (19 Brazilian and one Uruguayan), using nonradioactive single‐strand conformation polymorphism (SSCP) and restriction enzyme analysis of genomic DNA. Six novel mutations (R121S, V240M, D491N, 638–641insT, 895–896insC, 1622–1627insG) and two previously described point mutations (R59H, R208C) were identified. Together they accounted for 90% of the disease alleles of the patients. Two mutations, 1622–1627insG and R59H, were present in 18 of 20 patients. In addition, four polymorphisms (L10P, L12L, R521C, S532G) were identified. All cases reported are infantile GM1 gangliosidosis. This report constitutes the most comprehensive molecular study to date of this disorder in infantile patients. Since GM1‐gangliosidosis is the most common lysosomal storage disorder in Southern Brazil, molecular diagnosis will be important for genetic counseling, carrier detection and prenatal diagnosis in index families. Hum Mutat 13:401–409, 1999. © 1999 Wiley‐Liss, Inc.