Mucopolysaccharidosis type II: identification of 30 novel mutations among Latin American patients.

In this study, 103 unrelated South-American patients with mucopolysaccharidosis type II (MPS II) were investigated aiming at the identification of iduronate-2-sulfatase (IDS) disease causing mutations and the possibility of some insights on the genotype-phenotype correlation The strategy used for genotyping involved the identification of the previously reported inversion/disruption of the IDS gene by PCR and screening for other mutations by PCR/SSCP. The exons with altered mobility on SSCP were sequenced, as well as all the exons of patients with no SSCP alteration. By using this strategy, we were able to find the pathogenic mutation in all patients. Alterations such as inversion/disruption and partial/total deletions of the IDS gene were found in 20/103 (19%) patients. Small insertions/deletions/indels (<22 bp) and point mutations were identified in 83/103 (88%) patients, including 30 novel mutations; except for a higher frequency of small duplications in relation to small deletions, the frequencies of major and minor alterations found in our sample are in accordance with those described in the literature.

Bloodspot acylcarnitine and amino acid analysis in cord blood samples: efficacy and reference data from a large cohort study.

BACKGROUND: In order to test the feasibility of cord blood screening for inherited metabolic disease, a two-year cohort study of births in six obstetric units from five towns in the north of England was undertaken. These towns have a high prevalence of consanguineous marriages, largely among the immigrant Asian community. The purpose of the study was to determine whether early detection of metabolic disease was possible and whether early intervention would improve prognosis. METHODS: Following parental consent, cord blood samples were collected at birth and analysed for acylcarnitine and amino acid profiles by tandem mass spectrometry in one of two laboratories. One laboratory used butylated derivatives, the other used underivatized samples. The same laboratories performed routine blood spot neonatal screening at 5-7 days of age on these babies. Patients with positive results were investigated and treated by a metabolic paediatrician as soon as possible. RESULTS: 24,983 births were examined. 12,952 samples were analysed as butyl derivatives, 12,031 samples were analysed underivatized. The following disorders were detected: medium-chain acyl-CoA dehydrogenase (MCAD) deficiency (1 case), 3-methylcrotonyl-CoA carboxylase (MCC) deficiency (2 cases), maternal carnitine transporter defect (2 cases), maternal MCC (1 case). The following disorders were diagnosed subsequently but were not detected by the cord blood screening: phenylketonuria (PKU) (1 case), maple syrup urine disease (MSUD) (2 cases), argininosuccinic aciduria (1 case), methylmalonic acidaemia (MMA) (1 case), glutaric aciduria type 2 (1 case), MCAD deficiency (2 cases), 3-hydroxy-3-methylglutaryl-CoA lyase deficiency (1 case). Comprehensive reference data for all analytes by both methods were obtained. CONCLUSIONS: Cord blood testing is of limited value in detecting inherited metabolic disease. The metabolites associated with most disorders examined were not elevated in cord blood. Some maternal disorders, carnitine transporter defect and 3-methlycrotonyl-CoA carboxylase deficiency, are detected. These remain of uncertain clinical significance. Comprehensive reference data have been obtained that will facilitate future interpretation of studies in cord blood.

Ethnicity of children with homozygous c.985A>G medium-chain acyl-CoA dehydrogenase deficiency: findings from screening approximately 1.1 million newborn infants.

OBJECTIVES: It has been suggested that homozygous c.985A>G medium-chain acyl-CoA dehydrogenase deficiency (MCADD) is a disease of White ethnic origin but little is known regarding its ethnic distribution. We estimated ethnic-specific homozygous c.985A>G MCADD birth prevalence from a large-scale UK newborn screening study. METHODS: Homozygous c.985A>G MCADD cases were ascertained in six English newborn screening centres between 1 March 2004 and 28 February 2007 by screening approximately 1.1 million newborns using tandem mass spectrometry analysis of underivatised blood spot samples to quantitate octanoylcarnitine (C8). Follow-up biochemistry and mutation analyses for cases (mean triplicate C8 value >/=0.5 micromol/L) were reviewed to confirm diagnosis. Ethnicity was ascertained from clinician report and denominators from 2001 UK Census estimates of ethnic group of children less than one year. RESULTS: Sixty-four infants were c.985A>G MCADD homozygotes (overall prevalence 5.8 per 100,000 live births; 95% CI 4.4-7.2). Sixty (93%) were White, two (3%) were mixed/other and two were of unknown ethnic origin. No Asian or Black homozygotes were identified. Proportions of White, mixed/other, Asian and Black births in screening regions were estimated, yielding homozygous c.985A>G MCADD birth prevalence of 6.9 per 100,000 (95% CI 5.2-8.8) in White, and 95% CI estimates of 0-2.7 per 100,000 in Asian and 0-5.8 in Black populations. The c.985A>G carrier frequency in the White group was estimated at one in 65 (95% CI 1/74, 1/61) under Hardy-Weinberg conditions. CONCLUSION: c.985A>G homozygous MCADD is not found in Black and Asian ethnic groups that have been screened at birth in England. This is consistent with the earlier published observations suggesting that MCADD due to the c.985A>G mutation is a disease of White ethnic origin.

Skewed X inactivation is associated with phenotype in a female with adrenal hypoplasia congenita.

Adrenal hypoplasia congenita (AHC) can occur due to deletions or mutations in the DAX 1 (NR0B1) gene on the X chromosome (OMIM 300200). This form of AHC is therefore predominantly seen in boys. Deletion of the DAX 1 gene can also be part of a larger contiguous deletion including the centromeric dystrophin and glycerol kinase (GK) genes. We report a girl with a de novo deletion at Xp21.2 on the maternal chromosome, including DAX1, the GK gene and 3' end of the dystrophin gene, who presented with salt losing adrenal insufficiency and moderate developmental delay, but relatively mild features of muscular dystrophy. Investigation using the androgen receptor as a marker gene identified skewed inactivation of the X chromosome. In the patient's leucocytes, the paternal X chromosome was completely inactive, but in muscle 20% of the active chromosomes were of paternal origin. Thus skewed X inactivation (deletion on the active maternal X chromosome with an inactive paternal X chromosome) is associated with AHC in a female. Variability in X inactivation between tissues may account for the pronounced salt loss and adrenal insufficiency but mild muscular dystrophy.

The natural history of Niemann-Pick disease type C in the UK.

Niemann-Pick disease type C (NPC) is an autosomal recessive, neurovisceral lipid storage disorder. Mutations in two genes (NPC1 and NPC2) produce indistinguishable clinical phenotypes by biochemical mechanisms that have not yet been entirely clarified. The wide spectrum of clinical presentations of NPC includes hepatic and pulmonary disease as well as a range of neuropsychiatric disorders. Late-onset disease has been increasingly recognized as the biochemical diagnosis of NPC has been more widely applied in adult neurology clinics. The clinical presentation and follow-up of 94 patients with NPC is described, 58 of whom were still alive at the time this report was prepared. The age at diagnosis ranged from the prenatal period (with hydrops fetalis) up to 51 years. This review of NPC patients in the UK confirms the phenotypic variability of this inherited lipid storage disorder reported elsewhere. Although a non-neuronopathic variant has been described, most patients in this series who survived childhood inevitably suffered neurological and in some cases neuropsychiatric deterioration. While symptomatic treatment, such as anticholinergic and antiepileptic drugs, can alleviate some aspects of the disease, there is a clear need to develop a specific treatment for this progressively debilitating neurodegenerative disorder.

Ornithine aminotransferase deficiency: diagnostic difficulties in neonatal presentation.

We describe two unrelated cases of ornithine aminotransferase (OAT) deficiency with rare neonatal presentation of hyperammonaemia. The diagnosis in the neonatal presentation of OAT deficiency is hampered as hyperornithinaemia is absent. Enzyme and mutation studies confirmed the diagnosis. OAT deficiency should be included in differential diagnosis of neonatal hyperammonaemia.

Methylmalonic aciduria: follow-up and enzymology on the original case after 36 years.

A 36-year follow-up on the original patient described with methylmalonic aciduria has shown that she has methylmalonyl-CoA apomutase deficiency. The main clinical problem associated with her methylmalonic aciduria is progressive renal impairment requiring commencement of haemodialysis at 42 years of age.

Lactic acidosis and developmental delay due to deficiency of E3 binding protein (protein X) of the pyruvate dehydrogenase complex.

Pyruvate dehydrogenase deficiency is an important cause of primary lactic acidosis. Most cases occur as a result of mutations in the gene for the E1 alpha subunit of the complex, with a small number resulting from mutations in genes for other components, most commonly the E3 and E3-binding protein subunits. We describe pyruvate dehydrogenase E3-binding protein deficiency in two siblings in each of two unrelated families from Kuwait. The index patient in each family had reduced pyruvate dehydrogenase activity in cultured fibroblasts and no detectable immunoreactive E3-binding protein. Both were homozygous for nonsense mutations in the E3-binding protein gene, one involving the codon for glutamine 266, the other the codon for tryptophan 5.

Juvenile Sandhoff disease--nine new cases and a review of the literature.

Juvenile Sandhoff disease (McKusick 268800) is a rare lysosomal storage disorder with only 12 cases recorded in the literature. This condition is also referred to as the subacute form of hexosaminidase deficiency. We describe 9 new cases of Pakistani origin and compare these with the other published cases. Ataxia and speech abnormalities were the commonest presentation. Constipation and urinary incontinence were frequent and may be due to autonomic neuropathy. Cherry-red spot was not noted in any of our cases. Increased lower limb reflexes were the commonest physical finding. Significant delay in diagnosis may be due to the nonspecific presentation of this condition. Diagnosis was on the basis of hexosaminidase deficiency. Residual enzyme activity did not correlate with the clinical picture. Emerging therapies make early diagnosis of this disorder important.

Straight-chain acyl-CoA oxidase deficiency presenting with dysmorphia, neurodevelopmental autistic-type regression and a selective pattern of leukodystrophy.

We report a rare case of straight-chain acyl-CoA oxidase deficiency (McKusick 264470) presenting with dysmorphism, neurodevelopmental regression and leukodystrophy.

Niemann-Pick disease: sixteen-year follow-up of allogeneic bone marrow transplantation in a type B variant.

Allogenic bone marrow transplantation (BMT) was carried out on a 3-year-old white caucasian girl with Niemann-Pick disease (NPD) type B. The donor was her unaffected brother. The patient was neurologically normal at the time of transplantation. Engraftment was based on cytogenetic studies and increased leukocyte acid sphingomyelinase (ASM) activity. However, liver biopsies taken up to 33 months post transplantation showed only a moderate reduction in stored sphingomyelin and no significant increase in ASM activity. The post-transplantation period was complicated by severe graft-versus-host disease and a respiratory arrest. By 6 years of age, neurological involvement was observed, including bilateral cherry red spots. The proband is now severely mentally and physically disabled. Liver cirrhosis has continued to progress despite the BMT, and haematemesis due to portal hypertension occurred at 17 years of age. However, pulmonary infiltration regressed after BMT and there has been no clinical evidence of pulmonary insufficiency.

2-methyl-3-hydroxybutyryl-CoA dehydrogenase deficiency in a 23-year-old man.

2-Methyl-3-hydroxybutyryl-CoA dehydrogenase (EC 1.1.1.178) deficiency is a recently described defect of isoleucine catabolism. The disorder is characterized by normal early development followed by a progressive loss of mental and motor skills. Deterioration may be rapid or may follow a slower decline with a possible stabilization of the disorder on a low-protein diet and appropriate medication. We report a 23-year-old man with 2-methyl-3-hydroxybutyryl-CoA dehydrogenase deficiency with a very mild clinical course. He had apparently normal early development and remained relatively well until the age of 6 years, when he contracted measles. Following this illness, his motor skills and school progress deteriorated. At 15 years he had significant dysarthria, and generalized rigidity with some dystonic and unusual posturing. He was then treated with a low-protein high-carbohydrate diet with a good response in terms of balance and gait. At 18 years he was given benzhexol (Artane), increased slowly from 2 mg to 6 mg daily, resulting in improvement in tremor and dystonia. At 23 years he can dress himself and works in sheltered employment but remains severely dysarthric.

Niemann-Pick disease type C in adults.

Although it is often perceived as a paediatric disorder, significant numbers of patients with Niemann-Pick disease type C present for the first time in adult life or survive into adult life. The presentation in these patients differs from that seen in the classical juvenile form of the disease. Adult patients are often referred to clinicians with psychosis or other major psychiatric problems. The dystonia with preserved intellectual functioning can be mistaken for other basal ganglia disorders such as Wilson disease. The presence of vertical gaze palsy is an important clinical clue and, in the presence of a modest increase in plasma chitotriosidase activity, can be very helpful in the differential diagnosis. The diagnosis should be confirmed in suspected cases by filipin staining of cultured fibroblasts, as well as cholesterol esterification studies and DNA mutation analysis.

Cochlear implantation of a patient with a previously undescribed mitochondrial DNA defect.

Mitochondrial DNA (mtDNA) defects are responsible for symptom complexes that are characteristically heterogeneous but are typically represented by muscle weakness and neurological deficits. One common feature of mitochondrial disease is deafness. This report details the assessment and outcome of a patient with a previously undescribed mtDNA rearrangement who underwent cochlear implantation. The patient shows a marked improvement in sentence recognition tests and recognition of environmental sounds. Patients with profound sensorineural hearing loss due to mtDNA defects should be considered as candidates for cochlear implantation when they no longer benefit from conventional hearing aids.

Succinyl-CoA:3-ketoacid CoA transferase (SCOT): cloning of the human SCOT gene, tertiary structural modeling of the human SCOT monomer, and characterization of three pathogenic mutations.

The activity of succinyl-CoA:3-ketoacid CoA transferase (SCOT; locus symbol OXCT; EC 2.8.3.5) is the main determinant of the ketolytic capacity of tissues. Hereditary SCOT deficiency causes episodic ketoacidosis. Here we describe the human SCOT gene, which spans more than 100 kb and contains 17 exons, on chromosome 5p13. We report pathogenic missense mutations in three SCOT-deficient patients designated GS04, 05, and 06. GS04 is a G219E/G324E compound; GS05 is a V221M homozygote, and GS06 is a G324E homozygote. We constructed a tertiary structural model of human SCOT by homology modeling based on the known structure of Acidaminococcus fermentans glutaconate CoA transferase. The model predicts that V221 and G219 are on the dimerizing surface, whereas G324 is near the active site. SCOT activity was reduced to a comparable degree in all three patients, but in a transient expression assay in SCOT-deficient fibroblasts, cDNAs containing G219E and G324E produced no detectable activity, whereas V221M constructs yielded approximately 10% of the control peptide level and detectable specific activity. Interestingly, GS05 had the mildest clinical course reported to date and detectable levels of SCOT protein in fibroblasts.

Five cases of isolated glycerol kinase deficiency, including two families: failure to find genotype:phenotype correlation.

Little is understood of the genotype/phenotype correlations in X linked glycerol kinase deficiency (GKD) where most cases are caused by extensive deletions of Xp21, which often include genes flanking the GK locus. Few cases of isolated GKD have been investigated where the phenotype is not influenced by neighbouring genes. In this paper, we present the mutation data from four confirmed and one suspected case of non-deletion, isolated, X linked GKD and therefore extend the base of patients that can allow an assessment of genotype/phenotype correlations for this disease. The mutations found were two terminations leading to premature truncation of the GK polypeptide chain, one insertion, and an amino acid substitution. Phenotypic variation was observed in two families, where there was more than one affected subject carrying the same mutation, confirming previous studies that suggest there is no correlation between disease severity and genotype. Furthermore, the nature of the mutation in different families does not appear to influence the spectrum of phenotypic variation. In addition, one coding polymorphism in exon 3 has been found. The characterisation of the gene structure has been completed and shows that instead of 19 there are 21 exons.

Compound heterozygosity for a Wolman mutation is frequent among patients with cholesteryl ester storage disease.

Cholesteryl ester storage disease and Wolman disease are rare autosomal recessive lipoprotein-processing disorders caused by mutations in the gene encoding human lysosomal acid lipase. Thus far we have elucidated the genetic defects in 15 unrelated CESD patients. Seven were homozygotes for the prevalent hLAL exon 8 splice junction mutation which results in incomplete exon skipping, while eight probands were compound heterozygotes for E8SJM and a rare mutation on the second chromosome. In this report, we describe the molecular basis of CESD in three compound heterozygous subjects of Czech and Irish origin. RFLP and DNA sequence analysis revealed that they were heteroallelic for the common G(934)-->A substitution in exon 8 of the hLAL gene and a mutation which, if inherited on both alleles, would be expected to result in complete loss of enzyme activity and to cause Wolman disease. In patients A. M. and J. J., two nucleotide deletions in exons 7 and 10 were detected, involving a T at position 722, 723, or 724 and a G in a stretch of five guanosines at positions 1064;-1068 of the hLAL cDNA. Both mutations result in premature termination of protein translation at residues 219 and 336, respectively, and in the production of truncated, inactive enzymes. Subject D. H., in contrast, is a compound heterozygote for the Arg(44)-->Stop mutation previously described in a French CESD proband. Combined with data in the literature, our results demonstrate that compound heterozygosity for a mutation causing Wolman disease is common among cholesteryl ester storage disease patients.

Trifunctional protein deficiency: three families with significant maternal hepatic dysfunction in pregnancy not associated with E474Q mutation.

We report five families with trifunctional protein deficiency in which, during pregnancy, three mothers experienced significant hepatic disease when carrying an affected fetus. Diagnoses were based on increased levels of long-chain hydroxyacylcarnitines and deficiencies of 3-hydroxyacyl-CoA dehydrogenase (LCHAD) and 3-ketoacyl-CoA thiolase activity in fibroblasts. All affected infants lacked the common E474Q mutation associated with isolated LCHAD deficiency. This mutation is thought to be a predisposing factor for maternal hepatic disease in pregnancy. Our findings suggest that other defects in this enzyme complex might be responsible for maternal hepatic complications in pregnancy.

Defective PEX gene products correlate with the protein import, biochemical abnormalities, and phenotypic heterogeneity in peroxisome biogenesis disorders.

Peroxisome biogenesis disorders (PBD) comprise three phenotypes including Zellweger syndrome (ZS) (the most severe), neonatal adrenoleucodystrophy, and infantile Refsum disease (IRD) (the most mild), and can be classified into at least 12 genetic complementation groups, which are not predictive of the phenotypes. Several pathogenic genes for PBD groups have been identified, but the relationship between the defective gene products and phenotypic heterogeneity has remained unclear. We identified a mutation in the PEX2 gene in an IRD patient with compound heterozygosity for a missense mutation and the known nonsense mutation detected in ZS patients. In transfection experiments using the peroxisome deficient CHO mutant, Z65 with a nonsense mutation in the PEX2 gene, we noted the E55K mutation had mosaic activities of peroxisomal protein import machinery and residual activities of peroxisomal functions, including dihydroxyacetone phosphate acyltransferase and beta oxidation of very long chain fatty acids. The nonsense mutation severely affects these peroxisomal functions as well as the protein import. These data suggest that allelic heterogeneity of the PEX gene affects the peroxisomal protein import and functions and regulates the clinical severity in PBD.

Genomic structure and identification of 11 novel mutations of the PEX6 (peroxisome assembly factor-2) gene in patients with peroxisome biogenesis disorders.

The PEX6 (peroxisome assembly factor-2, PAF-2) gene encodes a member of the AAA protein (ATPases associated with diverse cellular activities) family and restores peroxisome assembly in fibroblasts from peroxisome biogenesis disorder patients belonging to complementation group C (group 4 in the United States). We have now clarified the genomic DNA structure of human PEX6 and identified mutations in patients from various ethnic groups. The human PEX6 gene consists of 17 exons and 16 introns, spanning about 14kb. The largest exon, exon 1, has at least 952 bp nucleotides. Eleven novel mutations (18 alleles) were identified by direct sequencing of the PEX6 cDNA from 10 patients. All these mutations have been confirmed in the corresponding genomic DNA. There was no common mutation, but an exon skip was identified in two unrelated Japanese patients. Most of the mutations led to premature termination or large deletions of the PEX6 protein and resulted in the most severe peroxisome biogenesis disorder phenotype of Zellweger syndrome. A patient with an atypical Zellweger syndrome had a missense mutation that was shown to disrupt the cell's ability to form peroxisomes. This mutation analysis will aid in understanding the functions of the PEX6 protein in peroxisomal biogenesis. Hum Mutat 13:487-496, 1999.