PURPOSE: To investigate the MR findings of childhood metachromaticleukodystrophy (MLD). METH- ODS: Nine MR imaging studies in seven children (five girls and two boys, 10 to 32 months old) with MLD were evaluated retrospectively for the extent and progression of white matter abnormalities and the presence of contrast enhancement. RESULTS: All seven cases showed symmetric, confluent high signal intensity on T2-weighted images in the periventricular white matter and centrum semiovale. A posterior predominance of white matter abnormalities was noted in all cases. Although initially spared from demyelination in all cases, in one case, the subcortical U fibers were later involved in demyeli- nation on follow-up MR studies. Other sites of involvement were the genu (n 5 5) and splenium (n 5 6) of the corpus callosum, the posterior limbs of the internal capsule (n 5 5), the descending pyramidal tracts (n 5 4), the claustrum (n 5 4), and the cerebellar white matter (n 5 2); diffuse brain atrophy was seen in two cases. No enhancement of the lesion was seen on any of the five postcontrast examinations. A “tigroid” pattern, previously described in cases of Pelizaeus-Merzbacher disease, was noted in the centrum semiovale in six cases. CONCLUSION: In late-infantile MLD, demyelination is more promi- nent in the occipital region. In addition to demyelination of the periventricular white matter, common manifestations include a “tigroid” pattern and involvement of the corpus callosum, the internal capsule, and the corticospinal tract.
Gene-based delivery can establish a sustained supply of therapeutic proteins within the nervous system. For diseases characterized by extensive CNS and peripheral nervous system (PNS) involvement, widespread dis- tribution of the exogenous gene may be required, a challenge to in vivo gene transfer strategies. Here, using lentiviral vectors (LVs), we efficiently transduced hematopoietic stem cells (HSCs) ex vivo and evaluated the potential of their progeny to target therapeutic genes to the CNS and PNS of transplanted mice and correct a neurodegenerative disorder, metachromaticleukodystrophy (MLD). We proved extensive repopulation of CNS microglia and PNS endoneurial macrophages by transgene-expressing cells. Intriguingly, recruitment of these HSC-derived cells was faster and more robust in MLD mice. By transplanting HSCs transduced with the aryl- sulfatase A gene, we fully reconstituted enzyme activity in the hematopoietic system of MLD mice and pre- vented the development of motor conduction impairment, learning and coordination deficits, and neu- ropathological abnormalities typical of the disease. Remarkably, ex vivo gene therapy had a significantly higher therapeutic impact than WT HSC transplantation, indicating a critical role for enzyme overexpression in the HSC progeny. These results indicate that transplantation of LV-transduced autologous HSCs represents a potentially efficacious therapeutic strategy for MLD and possibly other neurodegenerative disorders.
Metachromaticleukodystrophy (MLD) is a demyelinating lysosomal storage disorder for which new treat- ments are urgently needed. We previously showed that transplantation of gene-corrected hematopoietic stem progenitor cells (HSPCs) in presymptomatic myeloablated MLD mice prevented disease manifestations. Here we show that HSC gene therapy can reverse neurological deficits and neuropathological damage in affected mice, thus correcting an overt neurological disease. The efficacy of gene therapy was dependent on and pro- portional to arylsulfatase A (ARSA) overexpression in the microglia progeny of transplanted HSPCs. We dem- onstrate a widespread enzyme distribution from these cells through the CNS and a robust cross-correction of neurons and glia in vivo. Conversely, a peripheral source of enzyme, established by transplanting ARSA- overexpressing hepatocytes from transgenic donors, failed to effectively deliver the enzyme to the CNS. These results indicate that the recruitment of gene-modified, enzyme-overexpressing microglia makes the enzyme bioavailable to the brain and makes therapeutic efficacy and disease correction attainable. Overall, our data provide a strong rationale for implementing HSPC gene therapy in MLD patients.
With pulse-chase study of 1-[14C]stearic acid-labeled cerebroside sulfate (14C-CS) and subsequent subcellular fractionation by Percoll gradient, the metabolism of CS and translocation of its metabolites in human skin fibroblasts from controls, metachromaticleukodystrophy (MLD), and globoid cell leukodystrophy (GLD) were studied. In control skin fibroblasts, CS was transported to lysosome and metabolized there to galactosylceramide (GalCer) and ceramide (Cer) within 1 h. During the chase period, radioactivity was
Abstract: Metachromaticleukodystrophy (MLD) is an autosomal recessive lysosomal storage disorder mainly caused by the arysulfatase A (ARSA) gene mutations, which results in ARSA activity deficient to accumulate sulfatide in the oligodendrocytes and in the Schwann cells. On the basis of the age of onset, MLD is characterized by three clinical subtypes: late infantile, juvenile, and adult. In this manuscript we report a novel ARSA gene mutation c.302delG in a Chinese late infantile form MLD patient. The frameshift mutation c.302delG changes translated amino acid sequence and creates a premature stop codon in exon 2 at residue 107 (G101Afs*7) according to Mutation Taster Database analysis. Moreover, the mutation c.302delG also damages the protein structure in comparison to that of wild type ARSA protein through SWISS-MODEL. Combined with the patient’s typical late infantile presentation, we speculate it may be the cause of MLD.
Results: The median age at disease onset was 1 year and 3 months with the first clinical symptom being gait disturbance. All five patients became bed-ridden at a median age of 2 years and 5 months. Nerve conduction velocity revealed demyelinating polyneuropathy and brain MRI disclosed tigroid and leopard skin pattern of dysmyelination in all 5 patients. All patients had decreased ARSA activities in leukocytes accounting for 15.88 % to 30.75 % of controls. Five novel mutations, p.A316D, p.G303R, p.Q176X, p.R293X, and c.749 insGCGGGCCA, were identified in our case series. Eighteen patients, including our 5 patients, were reported in Asian countries. A total of 22 different disease-causing alleles were found, in which p.W320X was identified in Taiwan and China, and p.G101V was found in Taiwan and Korea. Conclusions: Patients with late infantile metachromaticleukodystrophy exhibited a rapid and devastating clinical course. The pattern of dysmyelination on brain MRI together with peripheral demyelination polyneuropathy indicates that evaluation of ARSA activity in leukocytes is warranted. A wide diversity of ARSA gene mutations was noted in Asia.
Metachromaticleukodystrophy (MLD) is an autosomal recessively inherited metabolic disease characterized by deficient activity of the lysosomal enzyme arylsulfatase A. Its deficiency results in accumulation of sulfatides in neural and visceral tissues, and causes demyelination of the central and peripheral nervous system. This leads to a broad range of neurological symptoms and eventually premature death. In asymptomatic patients with juvenile and adult MLD, treatment with allogeneic hematopoietic stem cell transplantation (HCT) provides a symptomatic and survival benefit. However, this treatment mainly impacts brain white matter, whereas the peripheral neuropathy shows no or only limited response. Data about the impact of peripheral neuropathy in MLD patients are currently lacking, although in our experience peripheral neuropathy causes significant morbidity due to neuropathic pain, foot deformities and neurogenic bladder disturbances. Besides, the reasons for residual and often progressive peripheral neuropathy after HCT are not fully understood. Preliminary studies suggest that peripheral neuropathy might respond better to gene therapy due to higher enzyme levels achieved than with HCT. However,
outlined in the 2009 US Food and Drug Administration (FDA) patient-reported outcomes (PROs) guidance and expectations of European regulators [16, 17]. This study was conducted in accordance with the ethical principles outlined in the Declaration of Helsinki 2008 and reviewed and approved by the RTI International Institutional Re- view Board. Informed consent was obtained before partici- pation. Figure 1 shows a schematic of the instrument methodological process followed in the development of the content for the new COA measure: Impact of Juvenile MetachromaticLeukodystrophy on Physical Activities (IMPA) scale. The methodological details of each stage of the development process follow.
A deficiency of arylsulfatase A (ASA) causes metachromaticleukodystrophy (MLD), a lysosomal storage disorder character- ized by accumulation of sulfatide, a severe neurological phenotype and early death. The efficacy of enzyme replacement ther- apy (ERT) has previously been determined in ASA knockout (ASA-/-) mice representing the only available animal model for MLD. Repeated intravenous injection of human ASA (hASA) improved the nervous system pathology and function, but also elicited a progressive humoral immune response leading to treatment resistance, anaphylactic reactions, and high mortality. In contrast to ASA-/- mice, most MLD patients express mutant hASA which may entail immunological tolerance to substituted wildtype hASA and thus protect from immunological complications. To test this notion, a cysteine-to-serine substitution was introduced into the active site of the hASA and the resulting inactive hASA-C69S variant was constitutively expressed in ASA-/- mice. Mice with sub- to supranormal levels of mutant hASA expression were analyzed. All mice, including those showing transgene expression below the limit of detection, were immunologically unresponsive to injected hASA. More than 100-fold overexpression did not induce an overt new phenotype except occasional intralysosomal deposition of minor amounts of glycogen in hepatocytes. Furthermore, long-term, low-dose ERT reduced sulfatide storage in peripheral tissues and the central nervous system indicating that high levels of extracellular mutant hASA do not prevent cellular uptake and lysosomal targeting of substituted wildtype hASA. Due to the tol- erance to hASA and maintenance of the MLD-like phenotype, the novel transgenic strain may be particularly advantageous to assess the benefit and risk of long-term ERT.
Abstract: Metachromaticleukodystrophy (MLD) disorder is a rare lysosomal storage disorder that leads to severe neurological symptoms and an early death. MLD occurs due to the deficiency of enzyme arylsulfatase A (ARSA) in leukocytes, and patients with MLD excrete sulfatide in their urine. In this study, the ARSA gene in 12 non-consanguineous MLD patients and 40 healthy individuals was examined using polymerase chain reaction sequencing. Furthermore, the structural and functional effects of new mutations on ARSA were analyzed using SIFT (sort- ing intolerant from tolerant), I-Mutant 2, and PolyPhen bioinformatics software. Here, 4 new pathogenic homozygous mutations c.585G.T, c.661T.A, c.849C.G, and c.911A.G were detected. The consequence of this study has extended the genotypic spectrum of MLD patients, paving way to a more effective method for carrier detection and genetic counseling.
Notes: The figure shows the biosynthetic pathways of sulfatide, the sphingolipid that accumulates during metachromaticleukodystrophy MLD. The specific enzymes described in each step of the pathway are in italics. The rounded boxes depict the products or substrates of the reactions. The green shaded square shapes show different therapeutic approaches for MLD treatment. Two current substrate reduction therapies for another lysosomal disease (Gaucher disease) are also shown: N-Butyldeoxynojirimycin (NB-DNJ) miglustat (Zavesca ® , Actelion Pharmaceuticals Ltd, Allschwil, Switzerland), and eliglustat tartrate. Both of these small molecules competitively inhibit ceramide glucosyltransferase.
galactosylceramide, ceramide, and stearic acid, which was reutilized in the synthesis of the major lipids found in cultured fibroblasts. Uptake and metabolism of the [ 14 C]CS into cells from typical and atypical patients and carriers of metachromaticleukodystrophy (MLD), Krabbe disease, and Farber disease were observed. Cells from patients with late infantile MLD could not metabolize the CS at all, while cells from an adult MLD patient and from a variant MLD patient could metabolize ~40 and 15%, respectively, of the CS taken up. These results are in contrast to the in vitro results that demonstrated a severe deficiency of
(BMT Database). Those whose MLD diagnosis could be retrospectively confirmed were included for this analysis. MLD diagnosis was considered confirmed if (1) the pa- tient had low ARSA with elevated urine sulfatides (n = 39); or (2) the patient had low ARSA, radiographic and clinical evidence of leukodystrophy and an affected sibling with low ARSA and elevated urine sulfatides (n = 1). For each patient, data were collected from the BMT Data- base, medical record review, and/or parental telephone surveys (approved by the Institutional Review Board [IRB] and following the provision of informed consent). All efforts were made to extract the following information per patient: age at clinical MLD onset, reason for diagno- sis, family history of MLD, age at HSCT, transplant condi- tioning intensity, allograft source, donor-recipient human leukocyte antigen (HLA) compatibility, occurrence and time to neutrophil and platelet recovery, occurrence and severity of acute and chronic graft-versus-host disease (aGvHD, cGvHD) [18, 19], most recent post- transplant donor hematopoietic engraftment, most recent leukocyte ARSA activity, survival status, cause of death (where applicable), and time to most recent follow-up or death.
Certain limitations of this study should, however, be noted. Because the study is retrospective, there may be some variabil- ity in the MRI technique, though this might have been miti- gated, in part, by the use of a standardized leukodystrophy protocol. Moreover, the small sample size—readily explained by the rarity of the disorder—limits its statistical power. An- other limitation is that DWI and ADC values were not avail- able for our own control population.
We found that MLD has a fairly characteristic pattern that does not appear to differ among the late infantile, juvenile, and adult-onset groups. A sheet-like area of abnormal T2 signal- intensity hyperintensity initially envelops the frontal and pa- rietal periventricular and central white matter regions, faint in mild disease and denser in moderate-to-severe disease. As se- vere disease develops, the sheet of white matter signal-inten- sity abnormality also involves the inner half of the subcortical white matter and a tigroid pattern then emerges. The corpus callosum is involved in mild disease, but not as prominently as in ALD or globoid cell leukodystrophy (GLD). Projection fi- ber involvement, cerebellar white matter involvement, and basal ganglia/thalamic involvement are often present in severe disease.
Mendelian adult-onset leukodystrophies are a spectrum of rare inherited progressive neurodegenerative disorders affecting the white matter of the central nervous system. Among these, cerebral autosomal dominant and recessive arteriopathy with subcortical infarcts and leukoencephalopathy, cerebroretinal vasculopathy, metachromaticleukodystrophy, hereditary diffuse leukoencephalopathy with spheroids, and vanishing white matter disease present with rapidly progressive dementia as dominant feature and are caused by mutations in NOTCH3, HTRA1, TREX1, ARSA, CSF1R, EIF2B1, EIF2B2, EIF2B3, EIF2B4, and EIF2B5, respectively. Given the rare incidence of these disorders and the lack of unequivocally diagnostic features, leukodystrophies are frequently misdiagnosed with common sporadic dementing diseases such as Alzheimer’s disease (AD), raising the question of whether these overlapping phenotypes may be explained by shared genetic risk factors. To investigate this intriguing hypothesis, we have combined gene expression analysis (1) in 6 different AD mouse strains (APPPS1, HOTASTPM, HETASTPM, TPM, TAS10, and TAU) at 5 different developmental stages (embryo [E15], 2, 4, 8, and 18 months), (2) in APPPS1 primary cortical neurons under stress conditions (oxygen-glucose deprivation) and single-variantebased and single-gene e based (c-alpha test and sequence kernel association test (SKAT)) genetic screening in a cohort composed of 332 Caucasian late-onset AD patients and 676 Caucasian elderly controls. Csf1r was signiﬁcantly overexpressed (log2FC > 1, adj. p-value < 0.05) in the cortex and hippocampus of aged HOTASTPM mice with extensive A b dense-core plaque pathology. We identi ﬁ ed 3 likely pathogenic mutations in CSF1R TK domain (p.L868R, p.Q691H, and p.H703Y) in our discovery and validation cohort, composed of 465 AD and mild cognitive impairment (MCI) Caucasian patients from the United Kingdom.