Late Infantile Metachromatic Leukodystrophy Research Articles

Occurrence, distribution, and phenotype of arylsulfatase A mutations in patients with metachromatic leukodystrophy

Occurrence, distribution, and phenotype of arylsulfatase A (ASA) mutations were investigated in 27 patients with metachromatic leukodystrophy (MLD) from Central Europe, mainly from Austria (n = 15) and Poland (n = 9). Genomic DNA from leukocytes, fibroblasts, or paraffin-embedded, formalin-fixed brain or nerve tissue, respectively, was tested by natural or mutated primer-modulated PCR restriction, fragment length polymorphism for the eight most common European mutations: R84Q, S96F, 459+1G > A, I179S, A212V, 1204+1G > A, P426L, and 1401del11bp. The overall identification rate of unrelated MLD alleles was the highest, in adult (90%), medium in juvenile (50%), and lowest in late infantile (36%) MLD patients. The two common alleles, 459+1G > A and P426L, together accounted for 42% of all 50 unrelated MLD alleles investigated; I179S was observed in 6 of 50 MLD alleles (12%). Thus, I179S was far more frequent than hitherto thought and appears to be a third common mutation in Europe. Moreover, a different allelic distribution between Austrian and Polish juvenile patients was disclosed, indicating genetic heterogeneity of MLD even within Central Europe. The genotype-phenotype correlation suggested by Polten et al. [N Engl J Med 324:18-22, 1991] was not followed by all of our MLD patients. Moreover, some MLD patients with identical ASA mutations presented with different phenotypes. This may be due, at least in some cases, to the presence of an additional mutation on individual mutant alleles. Therefore, prediction of the clinical course from single mutation analysis is not possible.

Three-dimensional brain visualization for metachromatic leukodystrophy

The basic understanding of many neurogenetic diseases requires study of the clinical, biochemical, and pathological aspects. To study the pathological aspects, the organs affected by the disease must be observed. We have used volume visualization techniques to create three-dimensional (3D) brain images of a patient with late infantile metachromatic leukodystrophy (MLD). The 3D brain images showed clearly, stereographically, and non-invasively the intracerebral lesion. This lesion, which indicated hyperintensity in magnetic resonance (MR) images, extended throughout the periventricular white matter. The 3D brain images are provided to integrate information. Volumetric ray-casting was useful in obtaining directly images of the entire brain and in allowing an intuitive understanding of the extension of the lesion in three dimensions and of the extent of the defects in the MLD brain. Isosurfacing facilitated a clear extraction of the lesion located by volumetric ray-casting. Each technique used in this study played a role in visualization and their use was complementary. 3D brain images will promote morphological investigation of neurogenetic diseases.

Characterization of two arylsulfatase A missense mutations D335V and T274M causing late infantile metachromatic leukodystrophy

Characterization of two arylsulfatase A missense mutations D335V and T274M causing late infantile metachromatic leukodystrophy

Characterization of two arylsulfatase A missense mutations D335V and T274M causing late infantile metachromatic leukodystrophy.

Metachromatic leukodystrophy is a lysosomal storage disorder caused by the deficiency of arylsulfatase A. We describe a novel missense mutation in exon 6 causing the substitution of Asp335 by Val. In transient transfections no enzyme activity could be expressed from the arylsulfatase A cDNA carrying this mutation. Examination of the effects of the mutation in cells stably overexpressing the mutant enzyme revealed, that the mutant enzyme is catalytically inactive and degraded in an early biosynthetic compartment. We have also investigated the effects of a previously identified mutation (T274M). The specific catalytic activity of the Met274 substituted arylsulfatase is reduced to about 35% of the normal enzyme when measured with an artificial substrate. Most of this enzyme is also degraded in an early biosynthetic compartment.

Discrimination Between Metachromatic Leukodystrophy and Pseudo-Deficiency of Arylsulfatase A by Restriction Digest of Amplified Gene Fragments

Mutations causing metachromatic leukodystrophy and pseudo-deficiency were detected in the arylsulfatase A gene by methods based on different wild-type and mutant restriction sites. After polymerase chain reaction amplification of fragments of the arylsulfatase A gene and digestion by the appropriate endonuclease, the mixtures were separated by polyacrylamide gel electrophoresis and visualized by ethidium bromide staining. The common splice mutation in intron 2 (459 + 1G-->A) causing, in homozygosity, late-infantile metachromatic leukodystrophy and the common missense mutation in exon 8 (P426L) causing, in homozygosity, adult or juvenile metachromatic leukodystrophy were found to abolish Bst NI and Aci I sites, respectively. The polyadenylation pseudo-deficiency mutation (1619A-->G) was found to create a Mae III restriction site. The N-glycosylation pseudo-deficiency mutation (N350S) does not produce or destroy any known restriction site, and in this case, introduction of a single nucleotide mismatch in one of the primers enabled the authors to create a Bfa I site in the mutant allele.

Molecular basis of late infantile metachromatic leukodystrophy in the Habbanite Jews.

Late infantile metachromatic leukodystrophy (MLD) is a neurodegenerative disease, most commonly caused by the deficiency of the lysosomal enzyme arylsulfatase A (ARSA). Late infantile MLD is frequent (1/75 live birth) in a small Jewish community which lived in Habban, isolated from the other Jewish populations. The gene coding for ARSA was sequenced in one of the Habbanite patients, who was found to be homozygous for an allele having three mutations. Two mutations are A to G transitions in the ARSA gene at positions 1788 and 2723, causing the loss of an N-glycosylation site and a polyadenylation signal, respectively. These mutations are characteristics for the ARSA pseudodeficiency (PD) allele, which in homozygozity is associated with low enzymatic activity, but does not cause disease. The third mutation, which occurred on the background of the PD allele, is a C to T transition at position 2119, predicting a substitution of proline-377 by leucine (P377L). Biosynthesis studies performed with cells expressing the ARSA cDNA into which this mutation was introduced demonstrated a severely reduced half-life of the mutant enzyme. Five of 10 patients from the Habbanite community could be studied and were homozygous for the P377L allele. These observations confirm the genealogical data which pointed to a common ancestor for all the carriers of MLD among the Habbanite Jews. In addition, the same mutation was demonstrated to be relatively frequent among the Yemenite Jews. The origin and the means by which the mutation spread between the two communities remain unknown.

Presymptomatic late-infantile metachromatic leukodystrophy treated with bone marrow transplantation

At 8 months of age, before clinical neurologic deterioration, the younger of two sisters with metachromatic leukodystrophy received a transplant of bone marrow from her haploidentical, heterozygote mother. Compared with the course in the older, affected, untreated sibling, the onset of neurologic regression was delayed 1 year and progressed at a slower rate. (J PEDIATR 1994;125:755-8)

Metachromatic leukodystrophy among southern Alaskan Eskimos: molecular and genetic studies.

Metachromatic leukodystrophy (MLD) is an autosomal recessive disorder resulting from the inability to metabolize sulphatide, an important component of myelin. Although there is significant clinical variability between patients, most have the late-infantile form. It is one of the most common lysosomal disorders involving mental deterioration and is found throughout the world. The great majority of the cases have a deficiency of arylsulphatase A activity. Accurate diagnosis of MLD is complicated by the presence of so-called pseudodeficiency alleles and the need to receive specimens for biochemical testing within 24-48 h of collection. We report the identification of the mutation (a g-to-a transition in the first nucleotide of intron 4) in the arylsulphatase A gene causing late-infantile MLD among the Eskimo population of southern Alaska. As all patients and family members from living and deceased patients had the same mutation, a mutation-based test was developed to identify patients and carriers that can be done on dried blood spots sent via regular mail service. A possible genetic link between this population and the Navajo Indians of the southwestern United States is proposed.

Late infantile metachromatic leukodystrophy in Israel

Metachromatic Leukodystrophy (MLD) is a neurodegenerative disease in which the lysosomal enzyme, Aryl sulfatase A (ARSA) is deficient. The disease is inherited as an autosomal recessive trait and its frequency is estimated to be 1 40,000 live births. The gene of ARSA has been cloned and up to now eight mutations causing MLD have been reported. Another mutation, PD, leads to the deficiency of the enzyme in vitro (pseudodeficiency) without any known clinical effect. The PD mutation is frequent in all populations. In Israel, late infantile MLD was found to be very frequent in a small Jewish isolate, the Habbanite Jews ( 1 75 live births). The molecular analysis demonstrated that in the Habbanite population, the mutation occurred on an allele with the PD mutation. The loss of ARSA activity is due to a point mutation C > T leading to a change of proline to leucine. MLD is also frequent among Moslem Arabs in Jerusalem. The mutation is a transition G > A destroying the splice donor site of exon 2. This mutation has been reported in patients with the late infantile MLD from different ethnic groups. The Christian Arabs in Israel also have a high incidence of the disease ( 1 10,000 live births); the mutation in this population is still unknown. Knowledge of the different mutations causing MLD in these defined populations will allow a carrier screening program to be carried out and prevent the birth of additional affected children.

Metachromatic leukodystrophy: a nonsense mutation (Q486X) in the arylsulphatase A (ARSA) gene

Metachromatic Leukodystrophy (MLD) is an autosomal recessive lysosomal storage disorder characterised by defective catabolism of myelin and the accumulation of cerebroside sulphate in neural tissue. The disorder is caused by a deficiency of the lysosomal hydrolase, arylsulphatase A (ARSA). MLD patients present with a broad spectrum of clinical phenotypes, the disorder has therefore been divided into three broad subtypes based on the age of onset of clinical symptoms. Late-infantile MLD is typically diagnosed before the age of 2 years, juvenile between ages 3 and 16 and adult MLD which is diagnosed after puberty (1). Biochemical diagnosis of MLD is complicated by the existence of an ARSA pseudodeficiency allele, which causes the loss of the major mRNA species (2.1 kb) of ARSA and a concomitant 90% reduction in the level of ARSA activity. Approximately 1% of the normal population is homozygous for this allele with no apparent clinical sequelae (2). The 507 amino acid ARSA polypeptide is encoded by a compact 3.2 kb gene containing 8 exons (3). Each of the ARSA exons and intron boundaries were PCR amplified and subjected to single-stranded conformation polymorphism analysis (SSCP) to detect sequence alterations (4). Exons exhibiting alterations in electrophoretic mobility on SSCP gels were sequenced using a modified linear based PCR sequencing protocol (5). SSCP analysis of the ARSA gene from a late-infantile MLD patient revealed a shift in the mobility of a band derived from exon v m (Figure 1). Sequence analysis of exon 8 revealed a heterozygous C to T transition at nucleotide number 1458 of the ARSA cDNA resulting in the alteration of a glutamine codon (CAG) at position 486 to a stop codon (TAG), (Q486X). No other sequence changes have yet been observed in this patient. Allele specific oligonucleotides were designed to detect the normal (5'-CGCCCTGCAGATCTGCTG-3') and mutant (5'-CGCCCTGTAGATCTGCTG-3') sequences Of Q486X (Figure 2) on Southern blotted filters containing exon 8 PCR products. After hybridisation filters were washed twice in 4xSSC for 10 minutes and once at 60°C in 2 xSSC, 0.1 % SDS (4). The patient's mother was found to be heterozygous for Q ggX (her father was unavailable for testing), however the mutation was not found in 27 other MLD patients, 21 ARSA pseudodeficient or 42 normal individuals.

Bone marrow transplantation for late infantile metachromatic leukodystrophy: Pathogenic investigation for graft rejection

A Japanese boy aged 2 years 11 months with late infantile metachromatic leukodystrophy underwent bone marrow transplantation (BMT) from his human leukocyte antigen (HLA) identical but mixed lymphocyte culture reactive father. Chimerism and increased arylsulfatase A activities of leukocytes had been observed with retarded progression of neurological deterioration during the first 3 months post-BMT. Graft rejection gradually occurred and donor cells were almost completely eliminated from the patient at 1 year after BMT. The process of neurodegeneration progressed clinically and neuroradiologically. Three possible reasons for the pathogenesis of graft rejection are: (i) T cell depletion of donor marrow cells as graft-versus-host disease (GVHD) prophylaxis; (ii) a slightly weak conditioning regimen: and (iii) a small number of marrow cells transplanted. It is stressed that as BMT is still a preliminary therapy for metachromatic leukodystrophy indications, conditioning, and GVHD prophylaxis for BMT should be considered individually.

Mutations in the arylsulfatase A gene of Japanese patients with metachromatic leukodystrophy.

To understand the molecular basis of metachromatic leukodystrophy (MLD) in Japanese patients, we analyzed the presence of three known mutant arylsulfatase A (ASA) alleles in 9 Japanese patients with MLD. Two of these mutant alleles (designated 609A and 2381T) were reported to be relatively frequent in a sample of predominantly Caucasian MLD. The other allele, with a substitution of Gly-99 by Asp (allele 445A), had been identified in a Japanese adult form of MLD in a heterozygous combination. We have found that allele 445A has a moderately high incidence among Japanese patients with MLD, and that homozygosity results in the late-infantile form. Neither allele 609A nor 2381T was found in Japanese patients examined in this study. Analysis on the nucleotide sequence of the ASA genes from another late-infantile MLD patient revealed the presence of a previously unreported G-to-A mutation at the 1,070th nucleotide of the ASA gene (designated 1070A). This results in a substitution of Gly-245 by Arg. This 1070A mutation was also found heterozygously in a juvenile MLD patient. When the 1070A mutation was introduced into the ASA cDNA and evaluated by transient expression studies, no enzyme activity was induced. These results suggest that Japanese MLD patients have a different distribution of ASA mutations from that found in a predominantly Caucasian population.

New sites of ocular involvement in late-infantile metachromatic leukodystrophy revealed by histopathologic studies.

New sites of ocular involvement in late-infantile metachromatic leukodystrophy revealed by histopathologic studies.

An arylsulfatase A (ARSA) missense mutation (T274M) causing late-infantile metachromatic leukodystrophy

Metachromatic leukodystrophy (MLD) is an autosomal recessive lysosomal storage disorder caused by a deficiency of arylsulfatase A (ARSA; EC 3.1.6.8). The 8 ARSA exons and adjacent intron boundaries from a patient with late-infantile metachromatic leukodystrophy were polymerase chain reaction (PCR) amplified in seven discrete reactions. Amplified ARSA exons were analysed for the presence of sequence alterations by single-strand conformation polymorphism analysis, followed by direct sequencing of PCR products. The patient was found to be homozygous for a C-->T transition in exon IV that results in the substitution of a highly conserved threonine residue at amino acid 274 with a methionine (T274M). Analysis of a further 29 MLD patients revealed the presence of five additional homozygotes for T274M. All 6 T274M homozygotes (representing four families) were of Lebanese descent, and all were known to be the result of consanguineous marriages. The altered amino acid is rigidly conserved among 10 sulfatases from Escherichia coli to humans; therefore, it is most likely that the resultant mutant protein will have little or no enzyme activity. This is consistent with the very low ARSA activity measured in these patients and their uniformly severe clinical presentation.

Missense mutations in the arylsulphatase A genes of metachromatic leukodystrophy patients.

Novel predicted disease-causing mutations have been defined in three patients with metachromatic leukodystrophy (MLD). The first new mutation is a C-->A change at base 884 in exon 5 of the arylsulphatase A (ASA) gene causing a serine to tyrosine substitution at position 295 of the protein (S295Y). A late-infantile MLD patient was found to be homozygous for this mutation. The second mutation is a G-->A substitution at nucleotide 1144 in exon 7, that causes a glutamic acid to lysine substitution at amino acid 382 (E382K). A juvenile MLD patients was found to be homozygous for this mutation. Finally an adult MLD patient has been shown to be heterozygous for two novel point mutations in exon 3. These are both C-->T changes at position 635 and 671 that result in alanine to valine substitutions at amino acids 212 (A212V) and 224 (A224V) of the ASA protein.

An 11-bp deletion in the arylsulfatase A gene of a patient with late infantile metachromatic leukodystrophy

Metachromatic leukodystrophy is a lysosomal storage disorder caused by the deficiency of arylsulfatase A. Examination of the arylsulfatase A gene in a patient suffering from late infantile metachromatic leukodystrophy revealed an 11-bp deletion in exon 8. Although this allele produces normal amounts of ASA mRNA, no arylsulfatase A cross-reacting material could be detected in cultured fibroblasts from the patient. The patient was found to be a compound heterozygote, the other allele is also known to generate no ASA polypeptides. This patient is another example where absence of ASA polypeptides correlates with the severe late infantile form of metachromatic leukodystrophy.

Treatment of Late Infantile Metachromatic Leukodystrophy by Bone Marrow Transplantation

METACHROMATIC leukodystrophy of the late infantile type, an autosomal recessive inherited disorder caused by a deficiency of arylsulfatase A activity, is characterized by progressive mental regression, loss of speech, quadriparesis, peripheral neuropathy, and death within a few years of onset.1 We describe a 10-year-old girl with metachromatic leukodystrophy in whom neurophysiologic function and sulfatide metabolism had improved after she received a bone marrow transplant five years before. These results may encourage the use of bone marrow transplantation in other patients with metachromatic leukodystrophy early in the course of the disease, when the degree of central nervous system impairment and the . . .

Lysosulfatide (Galactosylsphingosine‐3‐O‐Sulfate) from Metachromatic Leukodystrophy and Normal Human Brain

The glycosphingolipid pattern was examined in three cases of late infantile metachromatic leukodystrophy (MLD): one with a relatively short (2.5 years), one with a long (7.8 years), and one with a very long (13.2 years) survival time. All values were compared with those of age-matched normal controls. The cerebroside concentration was reduced to 25, 12, and 4%, respectively, in the MLD white matter, whereas the sulfatide concentration was increased up to 200% of the control value. The yield of myelin was reduced to less than 15% in the early case and to less than 3 and 1%, respectively, in the two later cases. There was no sign of increased sulfatide proportion in the myelin. The ganglioside pattern was normal in cerebral gray matter, but in the white matter, contents of gangliosides of the lacto series were significantly increased, in particular, the ganglioside suggested by us as being characteristic of reactive astrocytosis. For the first time, lysosulfatide was identified in MLD and normal human brains by mass spectrometry and radioimmunoaffinity TLC using specific monoclonal antibody. Its quantity was found to be similar in normal and MLD brains. These findings support our postulation that the lysoglycosphingolipids are synthesized de novo from sphingosine and that they do not play a key role in pathogenetic mechanisms.

Accumulation of lysosulfatide (sulfogalactosylsphingosine) in tissues of a boy with metachromatic leukodystrophy

Abnormal accumulation of lysosulfatide (sulfogalactosylsphingosine) was evident in autopsied tissues from a boy with late-infantile metachromatic leukodystrophy. The concentration was high in the cerebral white matter, spinal cord and sciatic nerve (116–787 pmol/mg protein) and low in the cerebral gray matter, kidney and liver (4–40 pmol/mg protein). As is the case with galactosylsphingosine, lysosulfatide inhibited cytochrome c oxidase activity, in a dose-dependent manner. Judging from the tissue distribution of the accumulated lysosulfatide and because of the cytotoxity, the lysosulfatide presumably explains the demyelination seen in the nervous tissues of patients with metachromatic leukodystrophy.

Multiple Sulfatase Deficiency

A 9-year-old girl with a phenotype similar to a mucopolysaccharidosis (MPS) and a clinical history characteristic of late infantile metachromatic leukodystrophy (MLD) is reported from the Department of Neurology, National Defense Medical Center, Taipei, Taiwan, Republic of China; the Developmental and Metabolic Neurology Branch, NIH, Bethesda, Maryland; and Department of Pediatrics (Dr. Horwitz), University of Chicago, Chicago, Illinois.