Search Results (12)

Newborn screening laboratories are increasingly adding lysosomal storage disorders (LSDs), such as Mucopolysaccharidosis I (MPS I) and Pompe disease, to their screening panels. Without newborn screening, LSDs are frequently diagnosed only after the onset of symptoms; late detection can lead to profound and irreversible organ damage and mortality. While screening of these disorders has accelerated over the past five years, there is little published information regarding the potential correlation of demographic variables (age at sample collection, birthweight, gestational age, gender, etc.) with lysosomal enzyme activity. The Missouri State Public Health Laboratory prospectively screened more than 475,000 newborns for MPS I, Pompe disease, Gaucher disease, and Fabry disease between 15 January 2013 and 15 May 2018. This report investigates trends between several demographic variables and activities of four lysosomal enzymes: α-L-iduronidase (IDUA), acid α-glucosidase (GAA), acid β-glucocerebrosidase (GBA), and acid α-galactosidase (GLA). This information provides a valuable resource to newborn screening laboratories for the implementation of screening for lysosomal storage disorders and the establishment of screening cutoffs. Full article

The mucopolysaccharidoses (MPS), Pompe Disease (PD), and Krabbe disease (KD) are inherited conditions known as lysosomal storage disorders (LSDs) The resulting enzyme deficiencies give rise to progressive symptoms. The United States Department of Health and Human Services’ Recommended Uniform Screening Panel (RUSP) suggests LSDs for inclusion in state universal newborn screening (NBS) programs and has identified screening deficiencies in MPS I, KD, and PD NBS programs. MPS I NBS programs utilize newborn dried blood spots and assay alpha L-iduronidase (IDUA) enzyme to screen for potential cases. Glycosaminoglycans (GAGs) offer potential as a confirmatory test. KD NBS programs utilize galactocerebrosidase (GaLC) as an initial test, with psychosine (PSY) activity increasingly used as a confirmatory test for predicting onset of Krabbe disease, though with an excessive false positive rate. PD is marked by a deficiency in acid α-glucosidase (GAA), causing increased glycogen, creatine (CRE), and other biomarkers. Bivariate normal limit (BVNL) methods have been applied to GaLC and PSY activity to produce a NBS tool for KD, and more recently, to IDUA and GAG activity to develop a NBS tool for MPS I. A BVNL tool based on GAA and CRE is in development for infantile PD diagnosis. Early infantile KD, MPS I, and PD cases were pre-symptomatically identified by BVNL-based NBS tools. This article reviews these developments, discusses how they address screening deficiencies identified by the RUSP and may improve NBS more generally. Full article

Mucopolysaccharidosis type I (MPS I) is an inherited autosomal recessive disease resulting from mutation of the α-l-Iduronidase (IDUA) gene. New unknown mutated nucleotides of idua have increasingly been discovered in newborn screening, and remain to be elucidated. In this study, we found that the z-Idua enzymatic activity of zebrafish idua-knockdown embryos was reduced, resulting in the accumulation of undegradable metabolite of heparin sulfate, as well as increased mortality and defective phenotypes similar to some symptoms of human MPS I. After microinjecting mutated z-idua-L346R, -T364M, -E398-deleted, and -E540-frameshifted mRNAs, corresponding to mutated human IDUA associated with MPS I, into zebrafish embryos, no increase in z-Idua enzymatic activity, except of z-idua-E540-frameshift-injected embryos, was noted compared with endogenous z-Idua of untreated embryos. Defective phenotypes were observed in the z-idua-L346R-injected embryos, suggesting that failed enzymatic activity of mutated z-Idua-L346R might have a dominant negative effect on endogenous z-Idua function. However, defective phenotypes were not observed in the z-idua-E540-frameshifted-mRNA-injected embryos, which provided partial enzymatic activity. Based on these results, we suggest that the z-Idua enzyme activity assay combined with phenotypic observation of mutated-idua-injected zebrafish embryos could serve as an alternative platform for a preliminary assessment of mutated idua not yet characterized for their role in MPS I. Full article

Deficit of the IDUA (α-L-iduronidase) enzyme causes the lysosomal storage disorder mucopolysaccharidosis type I (MPS I), a rare pediatric neurometabolic disease, due to pathological variants in the IDUA gene and is characterized by the accumulation of the undegraded mucopolysaccharides heparan sulfate and dermatan sulfate into lysosomes, with secondary cellular consequences that are still mostly unclarified. Here, we report a new fruit fly RNAi-mediated knockdown model of a IDUA homolog (D-idua) displaying a phenotype mimicking some typical molecular features of Lysosomal Storage Disorders (LSD). In this study, we showed that D-idua is a vital gene in Drosophila and that ubiquitous reduction of its expression leads to lethality during the pupal stage, when the precise degradation/synthesis of macromolecules, together with a functional autophagic pathway, are indispensable for the correct development to the adult stage. Tissue-specific analysis of the D-idua model showed an increase in the number and size of lysosomes in the brain and muscle. Moreover, the incorrect acidification of lysosomes led to dysfunctional lysosome-autophagosome fusion and the consequent block of autophagy flux. A concomitant metabolic drift of glycolysis and lipogenesis pathways was observed. After starvation, D-idua larvae showed a quite complete rescue of both autophagy/lysosome phenotypes and metabolic alterations. Metabolism and autophagy are strictly interconnected vital processes that contribute to maintain homeostatic control of energy balance, and little is known about this regulation in LSDs. Our results provide new starting points for future investigations on the disease’s pathogenic mechanisms and possible pharmacological manipulations. Full article

Mucopolysaccharidosis type I (MPS I) is a rare autosomal recessive disorder caused by deleterious mutations in the α-L-iduronidase (IDUA) gene. Until now, MPS I in Vietnamese has been poorly addressed. Five MPS I patients were studied with direct DNA sequencing using Illumina technology confirming pathogenic variants in the IDUA gene. Clinical characteristics, additional laboratory results, and family history were collected. All patients have presented with the classical characteristic of MPS I, and α-L-iduronidase activity was low with the accumulation of glycosaminoglycans. Three variants in the IDUA gene (c.1190-10C>A (Intronic), c.1046A>G (p.Asp349Gly), c.1862G>C (p.Arg621Pro) were identified. The c.1190-10C>A variant represents six of the ten disease alleles, indicating a founder effect for MPS I in the Vietnamese population. Using biochemical and genetic analyses, the precise incidence of MPS I in this population should accelerate early diagnosis, newborn screening, prognosis, and optimal treatment. Full article

The lysosomal storage disorder, mucopolysaccharidosis I (MPSI), results from mutations in IDUA, the gene that encodes the glycosaminoglycan-degrading enzyme α-L-iduronidase. Newborn screening efforts for MPSI have greatly increased the number of novel IDUA variants identified, but with insufficient experimental evidence regarding their pathogenicity, many of these variants remain classified as variants of uncertain significance (VUS). Defining pathogenicity for novel IDUA variants is critical for decisions regarding medical management and early intervention. Here, we describe a biochemical platform for the characterization of IDUA variants that relies on viral delivery of IDUA DNA into IDUA-deficient HAP1 cells and isolation of single cell expression clones. The relative specific activity of wild-type and variant α-iduronidase was determined using a combination of Western blot analysis and α-iduronidase activity assays. The specific activity of each variant enzyme was consistent across different single cell clones despite variable IDUA expression and could be accurately determined down to 0.05–0.01% of WT α-iduronidase activity. With this strategy we compared the specific activities of known pseudodeficiency variants (p.His82Gln, p.Ala79Thr, p.Val322Glu, p.Asp223Asn) or pathogenic variants (p.Ser633Leu, p.His240Arg) with variants of uncertain significance (p.Ser586Phe, p.Ile272Leu). The p.Ser633Leu and p.His240Arg variants both show very low activities consistent with their association with Scheie syndrome. In our experiments, however, p.His240Arg exhibited a specific activity five times higher than p.Ser633Leu in contrast to other reports showing equivalent activity. Cell clones expressing the p.Ser586Phe and p.Ile272Leu variants had specific activities in the range of other pseudodeficiency variants tested. Our findings show that pseudodeficiency and pathogenic variants can be distinguished from each other with regard to specific activity, and confirms that all the pseudodeficiency variants variably reduce α-iduronidase activity. We envision this platform will be a valuable resource for the rigorous assessment of the novel IDUA variants emerging from the expansion of newborn screening efforts. Full article

Mucopolysaccharidosis type I (MPS I) is a progressive lysosomal storage disease, with neurological and visceral involvement, in which early diagnosis through newborn screening (NBS) and early treatment can improve outcomes. We present our first 5 years of experience with laboratory and clinical management of NBS for MPS I. Since 2015, we have screened 160,011 newborns by measuring α-L-iduronidase (IDUA) activity and, since 2019, glycosaminoglycans (GAGs) in dried blood spot (DBS) as a second-tier test. Positive screening patients were referred to our clinic for confirmatory clinical and molecular testing. We found two patients affected by MPS I (incidence of 1:80,005). Before the introduction of second-tier testing, we found a high rate of false-positives due to pseudodeficiency. With GAG analysis in DBS as a second-tier test, no false-positive newborns were referred to our clinic. The confirmed patients were early treated with enzyme replacement therapy and bone-marrow transplantation. For both, the clinical outcome of the disease is in the normal range. Our experience confirms that NBS for MPS I is feasible and effective, along with the need to include GAG assay as a second-tier test. Follow-up of the two positive cases identified confirms the importance of early diagnosis through NBS and early treatment to improve the outcome of these patients. Full article

Mucopolysaccharidosis type I (MPS I) is caused by genetic deficiency of α-l-iduronidase and impairment of lysosomal catabolism of heparan sulfate and dermatan sulfate. In the brain, these substrates accumulate in the lysosomes of neurons and glial cells, leading to neuroinflammation and neurodegeneration. Their storage also affects lysosomal homeostasis-inducing activity of several lysosomal proteases including cathepsin B (CATB). In the central nervous system, increased CATB activity has been associated with the deposition of amyloid plaques due to an alternative pro-amyloidogenic processing of the amyloid precursor protein (APP), suggesting a potential role of this enzyme in the neuropathology of MPS I. In this study, we report elevated levels of protein expression and activity of CATB in cortex tissues of 6-month-old MPS I (Idua -/- mice. Besides, increased CATB leakage from lysosomes to the cytoplasm of Idua -/- cortical pyramidal neurons was indicative of damaged lysosomal membranes. The increased CATB activity coincided with an elevated level of the 16-kDa C-terminal APP fragment, which together with unchanged levels of β-secretase 1 was suggestive for the role of this enzyme in the amyloidogenic APP processing. Neuronal accumulation of Thioflavin-S-positive misfolded protein aggregates and drastically increased levels of neuroinflammatory glial fibrillary acidic protein (GFAP)-positive astrocytes and CD11b-positive activated microglia were observed in Idua -/- cortex by confocal fluorescent microscopy. Together, our results point to the existence of a novel CATB-associated alternative amyloidogenic pathway in MPS I brain induced by lysosomal storage and potentially leading to neurodegeneration. Full article

Enzyme-based newborn screening for Mucopolysaccharidosis type I (MPS I) has a high false-positive rate due to the prevalence of pseudodeficiency alleles, often resulting in unnecessary and costly follow up. The glycosaminoglycans (GAGs), dermatan sulfate (DS) and heparan sulfate (HS) are both substrates for α-l-iduronidase (IDUA). These GAGs are elevated in patients with MPS I and have been shown to be promising biomarkers for both primary and second-tier testing. Since February 2016, we have measured DS and HS in 1213 specimens submitted on infants at risk for MPS I based on newborn screening. Molecular correlation was available for 157 of the tested cases. Samples from infants with MPS I confirmed by IDUA molecular analysis all had significantly elevated levels of DS and HS compared to those with confirmed pseudodeficiency and/or heterozygosity. Analysis of our testing population and correlation with molecular results identified few discrepant outcomes and uncovered no evidence of false-negative cases. We have demonstrated that blood spot GAGs analysis accurately discriminates between patients with confirmed MPS I and false-positive cases due to pseudodeficiency or heterozygosity and increases the specificity of newborn screening for MPS I. Full article

Mucopolysaccharidosis (MPS) I is a severe lysosomal storage disease caused by α-L-iduronidase (IDUA) deficiency, which results in accumulation of non-degraded glycosaminoglycans in lysosomes. Costly enzyme replacement therapy (ERT) is the conventional treatment for MPS I. Toward producing a more cost-effective and safe alternative to the commercial mammalian cell-based production systems, we have produced recombinant human IDUA in seeds of an Arabidopsis mutant to generate the enzyme in a biologically active and non-immunogenic form containing predominantly high mannose N-linked glycans. Recombinant enzyme in ERT is generally thought to require a mannose 6-phosphate (M6P) targeting signal for endocytosis into patient cells and for intracellular delivery to the lysosome. Toward effecting in planta phosphorylation, the human M6P elaboration machinery was successfully co-expressed along with the recombinant human IDUA using a single multi-gene construct. Uptake studies using purified putative M6P-IDUA generated in planta on cultured MPS I primary fibroblasts indicated that the endocytosed recombinant lysosomal enzyme led to substantial reduction of glycosaminoglycans. However, the efficiency of the putative M6P-IDUA in reducing glycosaminoglycan storage was comparable with the efficiency of the purified plant mannose-terminated IDUA, suggesting a poor in planta M6P-elaboration by the expressed machinery. Although the in planta M6P-tagging process efficiency would need to be improved, an exciting outcome of our work was that the plant-derived mannose-terminated IDUA yielded results comparable to those obtained with the commercial IDUA (Aldurazyme^® (Sanofi, Paris, France)), and a significant amount of the plant-IDUA is trafficked by a M6P receptor-independent pathway. Thus, a plant-based platform for generating lysosomal hydrolases may represent an alternative and cost-effective strategy to the conventional ERT, without the requirement for additional processing to create the M6P motif. Full article

Early diagnosis of lysosomal storage diseases (LSDs) through newborn screening (NBS) has been adapted widely. The National Taiwan University Hospital Newborn Screening Center launched the four-plex tandem mass spectrometry LSD newborn screening test in 2015. The test determined activities of acid α-glucosidase (GAA; Pompe), acid α-galactosidase (GLA; Fabry), acid β-glucocerebrosidase (ABG; Gaucher), and acid α-l-iduronidase (IDUA; MPS-I) in dried blood spots (DBS). Through 2017, 64,148 newborns were screened for these four LSDs. The screening algorithm includes enzyme activity/ratio as the cutoffs for the first screening test and a second-tier test for Pompe disease screening. The second-tier Pompe disease screening test measured activity inhibition by acarbose. Twenty-nine newborns required a confirmatory test; six were confirmed to have Pompe disease, and nine were confirmed to have Fabry disease. The screen-positive rate for Pompe disease was 0.031%. Therefore, in Pompe disease newborn screening, a validated 2nd tier test is necessary to decrease false positives. Full article

Mucopolysaccharidoses (MPS’s) represent a subgroup of lysosomal storage diseases related to a deficiency of enzymes that catalyze glycosaminoglycans degradation. Mucopolysaccharidosis type I (MPS I) is a rare autosomal recessive disorder caused by a deficiency of α-l-iduronidase encoded by the IDUA gene. Partially degraded heparan sulfate and dermatan sulfate accumulate progressively and lead to multiorgan dysfunction and damage. The aim of this study is to describe the clinical, biochemical, and molecular characteristics of 13 Algerian patients from 11 distinct families. MPS I diagnosis was confirmed by molecular study of the patients’ IDUA gene. Clinical features at the diagnosis and during the follow-up are reported. Eighty-four percent of the studied patients presented with a mild clinical phenotype. Molecular study of the IDUA gene allowed the characterization of four pathological variations at the homozygous or compound heterozygote status: IDUA NM_00203.4:c.1598C>G-p.(Pro533Arg) in 21/26 alleles, IDUA NM_00203.4:c.532G>A-p.(Glu178Lys) in 2/26 alleles, IDUA NM_00203.4:c.501C>G-p.(Tyr167*) in 2/26 alleles, and IDUA NM_00203. 4: c.1743C>G-p.(Tyr581*) in 1/26 alleles. This molecular study unveils the predominance of p.(Pro533Arg) variation in our MPS I patients. In this series, the occurrence of some clinical features linked to the Scheie syndrome is consistent with the literature, such as systematic valvulopathies, corneal opacity, and umbilical hernia; however, storage signs, facial dysmorphic features, and hepatomegaly were more frequent in our series. Screening measures for these debilitating diseases in highly consanguineous at-risk populations must be considered a priority health problem. Full article