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Hurler-Scheie syndrome (MPS IH/S), also known as mucopolysaccharidosis type I-H/S (MPS IH/S), is a lysosomal storage disorder caused by deficiency of the enzyme alpha-L-iduronidase (IDUA) leading to the accumulation of glycosaminoglycans (GAGs) in various tissues, resulting in a wide range of symptoms affecting different organ systems. Postgenomic omics technologies offer the promise to understand the changes in proteome, phosphoproteome, and phosphorylation-based signaling in MPS IH/S. Accordingly, we report here a large dataset and the proteomic and phosphoproteomic analyses of fibroblasts derived from patients with MPS IH/S ( n = 8) and healthy individuals ( n = 8). We found that protein levels of key lysosomal enzymes such as cathepsin D, prosaposin, arylsulfatases (arylsulfatase A and arylsulfatase B), and IDUA were downregulated. We identified 16,693 unique phosphopeptides, corresponding to 4,605 proteins, in patients with MPS IH/S. We found that proteins related to the cell cycle, mitotic spindle assembly, apoptosis, and cytoskeletal organization were differentially phosphorylated in MPS IH/S. We identified 12 kinases that were differentially phosphorylated, including hyperphosphorylation of cyclin-dependent kinases 1 and 2, hypophosphorylation of myosin light chain kinase, and calcium/calmodulin-dependent protein kinases. Taken together, the findings of the present study indicate significant alterations in proteins involved in cytoskeletal changes, cellular dysfunction, and apoptosis. These new observations significantly contribute to the current understanding of the pathophysiology of MPS IH/S specifically, and the molecular mechanisms involved in the storage of GAGs in MPS more generally. Further translational clinical omics studies are called for to pave the way for diagnostics and therapeutics innovation for patients with MPS IH/S.

Mucopolysaccharidosis type I (MPS I) is a rare genetic disorder caused by mutations in the IDUA gene, leading to alpha-L-iduronidase enzyme deficiency and resulting in the accumulation of glycosaminoglycans (GAG; heparan and dermatan sulfate) in lysosomes. The consequent GAG accumulation within cells leads to organ dysfunction and a range of debilitating symptoms. Enzyme replacement therapy (ERT) is the prevailing treatment, but its limitations (including high cost, time requirements, inefficiency in treatment of central nervous system (CNS), and immunogenicity) necessitate exploration of alternative therapeutic strategies. This research propose a novel approach leveraging the synergistic effects of ERT and resveratrol-induced autophagy. Resveratrol, with its immunomodulatory and GAG degradation-stimulating properties, holds a promise in mitigating immune responses triggered by ERT. Moreover, its ability to penetrate the blood-brain barrier presents a potential solution for addressing CNS manifestations. This study employed cells from MPS I patients to investigate the combined effects of resveratrol and the enzyme. Evaluation of the therapeutic impact involved assessing GAG accumulation, enzyme testing, and examining lysosome functionality and the autophagy process through fluorescence microscopy and Western blotting. The combined therapy stimulated the lysosomal mannose-6-phosphate receptor (M6PR) and lysosome biogenesis through the transcription factor EB (TFEB). Additionally, initial block of autophagy in autophagosome formation was relieved after the combined therapy and resveratrol alone. Together with increased enzyme activity through stimulation of the receptor, this synergistic therapy can be considered a new potential treatment for MPS I patients, improving their overall quality of life., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Genetic deficiency of alpha-L-iduronidase causes mucopolysaccharidosis type I (MPS-I) disease, due to accumulation of glycosaminoglycans (GAGs) including chondroitin/dermatan sulfate (CS/DS) and heparan sulfate (HS) in cells. Currently, patients are treated by infusion of recombinant iduronidase or by hematopoietic stem cell transplantation. An alternative approach is to reduce the L-iduronidase substrate, through limiting the biosynthesis of iduronic acid. Our earlier study demonstrated that ebselen attenuated GAGs accumulation in MPS-I cells, through inhibiting iduronic acid producing enzymes. However, ebselen has multiple pharmacological effects, which prevents its application for MPS-I. Thus, we continued the study by looking for novel inhibitors of dermatan sulfate epimerase 1 (DS-epi1), the main responsible enzyme for production of iduronic acid in CS/DS chains. Based on virtual screening of chemicals towards chondroitinase AC, we constructed a library with 1,064 compounds that were tested for DS-epi1 inhibition. Seventeen compounds were identified to be able to inhibit 27%-86% of DS-epi1 activity at 10 μM. Two compounds were selected for further investigation based on the structure properties. The results show that both inhibitors had a comparable level in inhibition of DS-epi1while they had negligible effect on HS epimerase. The two inhibitors were able to reduce iduronic acid biosynthesis in CS/DS and GAG accumulation in WT and MPS-I fibroblasts. Docking of the inhibitors into DS-epi1 structure shows high affinity binding of both compounds to the active site. The collected data indicate that these hit compounds may be further elaborated to a potential lead drug used for attenuation of GAGs accumulation in MPS-I patients., (© The Author(s) 2024. Published by Oxford University Press.)

The main approach used in the current therapy of mucopolysaccharidosis (MPS) is to reduce the levels of glycosaminoglycans (GAGs) in cells, the deposits considered to be the main cause of the disease. Previous studies have revealed significant differences in the expression of genes encoding proteins involved in many processes, like those related to actin filaments, in MPS cells. Since the regulation of actin filaments is essential for the intracellular transport of specific molecules, the process which may affect the course of MPSs, the aim of this study was to evaluate the changes that occur in the actin cytoskeleton and focal adhesion in cells derived from patients with this disease, as well as in the MPS I mouse model, and to assess whether they could be potential therapeutic targets for different MPS types. Western-blotting, flow cytometry and transcriptomic analyses were employed to address these issues. The levels of the key proteins involved in the studied processes, before and after specific treatment, were assessed. We have also analyzed transcripts whose levels were significantly altered in MPS cells. We identified genes whose expressions were changed in the majority of MPS types and those with particularly highly altered expression. For the first time, significant changes in the expression of genes involved in the actin cytoskeleton structure/functions were revealed which may be considered as an additional element in the pathogenesis of MPSs. Our results suggest the possibility of using the actin cytoskeleton as a potential target in therapeutic approaches for this disease.

Synthesis of a series of l-iduronic acid (IdoA)- and imino-IdoA-typed C-glycosides for modulating α-l-iduronidase (IDUA) activity is described. In an enzyme inhibition study, IdoA-typed C-glycosides were more potent than imino-IdoA analogs, with the most potent IdoA-typed C-glycoside 27c showing an IC 50 value of 1 μM. On the other hand, co-treatment of 12 with rh-α-IDUA in mucopolysaccharidosis type I (MPS I) fibroblasts exhibited a nearly 3-fold increase of the IDUA activity, resulting in a clear reduction of the accumulated heparan sulfate (HS) compared to the exogenous enzyme treatment alone. This is the first report of small molecules facilitating IDUA stabilization, enhancing enzyme activity, and reducing accumulated HS in MPS I cell-based assays, which reveals that small molecules as rh-α-IDUA stabilizers to improve enzyme replacement therapy (ERT) efficacy toward MPS I is feasible and promising., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Wei-Chieh Cheng reports financial support was provided by Ministry of Science and Technology, R.O.C. Wei-Chieh Cheng has patent IDURONIDASE STABILIZERS AND USES THEREOF pending to Wei-Chieh Cheng., (Copyright © 2022 Elsevier Masson SAS. All rights reserved.)

Mucopolysaccharidosis type I Hurler syndrome (MPS IH) is a severe lysosomal storage disorder caused by alpha-l-iduronidase (IDUA) deficiency. Premature truncation mutations (PTC) are the most common (50%-70%) type of IDUA mutations and correlate with MPS IH. Nonsense suppression therapy is a therapeutic approach that aims to induce stop codon readthrough. The different ability of gentamicin to bind mutant mRNA in readthrough is determined by nucleotide sequence (PTC context: UGA > UAG > UAA) and inserted amino acid including the nucleotide position +4 of the PTC, as well as the mRNA secondary structure. We used COS-7 cells to investigate the functional characteristics of p.Q500X and p.R619X, IDUA variants and the effects of gentamicin in inducing stop codon readthrough of seven IDUA variants including p.Q500X, p.R619X, p.Q70X, p.E299X, p.W312X, p.Q380X, and p.W402X. Moreover, we performed prediction of RNA secondary structure using the online tool RNAfold. We found that cells treated with gentamicin showed significantly enhanced full-length IDUA expression and restored IDUA activity, in a dose-dependent manner, only in cells expressing cDNA with W312X, Q380X, W402X, and R619X. Among the readthrough-responsive variants, we observed UGA PTC in W312X, W402X and R619X; and UAG PTC with C at nucleotide +4 in Q380X. Changes of RNA secondary structure were noted only in mutants with readthrough-responsive variants including W312X, Q380X, W402X, and R619X. Additional preclinical studies of selected PTCs with potential readthrough, using drugs with less oto-nephrotoxicity, in patient's skin fibroblasts and animal model are necessary for the premise of personalized medicine., Competing Interests: Declaration of competing interest The authors declare that they have no conflicts of interest in relation to the manuscript., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Abstarct: Suppressing translation termination at premature termination codons (PTCs), termed readthrough, is a potential therapy for genetic diseases caused by nonsense mutations. Ataluren is a compound that has shown promise for clinical use as a readthrough agent. However, some reports suggest that ataluren is ineffective at suppressing PTCs. To further evaluate the effectiveness of ataluren as a readthrough agent, we examined its ability to suppress PTCs in a variety of previously untested models. Using NanoLuc readthrough reporters expressed in two different cell types, we found that ataluren stimulated a significant level of readthrough. We also explored the ability of ataluren to suppress a nonsense mutation associated with Mucopolysaccharidosis I-Hurler (MPS I-H), a genetic disease that is caused by a deficiency of α-L-iduronidase that leads to lysosomal accumulation of glycosaminoglycans (GAGs). Using mouse embryonic fibroblasts (MEFs) derived from Idua-W402X mice, we found that ataluren partially rescued α-L-iduronidase function and significantly reduced GAG accumulation relative to controls. Two-week oral administration of ataluren to Idua-W402X mice led to significant GAG reductions in most tissues compared to controls. Together, these data reveal important details concerning the efficiency of ataluren as a readthrough agent and the mechanisms that govern its ability to suppress PTCs., Key Messages: Ataluren promotes readthrough of PTCs in a wide variety of contexts. Ataluren reduces glycosaminoglyan storage in MPS I-H cell and mouse models. Ataluren has a bell-shaped dose-response curve and a narrow effective range., (© 2022. The Author(s).)

Mucopolysaccharidoses (MPS) are inherited metabolic diseases caused by storage of glycosaminoglycans (GAGs), however, various modulations of the course of these diseases were identified recently due to impairment of different cellular processes. Here, using transcriptomic analyses in cells derived from patients suffering from eleven types of MPS, we demonstrated that expression of dozens to hundreds of genes coding for proteins involved in signal transduction processes is significantly changed in MPS cell relative to controls. When studying membrane estrogen receptor 1 (GPER1) and oxytocin receptor (OXTR) in more detail, we unexpectedly found formation of aggregates of GPER1 in MPS I, and those of OXTR in both MPS I and MPS II cells. The presence of these aggregates did not correlate with levels of expression of GPER1 and OXTR genes and levels of corresponding gene products. On the other hand, the aggregates disappeared in cells treated with enzymes which are otherwise deficient in MPS I and MPS II, causing efficient degradation of GAGs. We demonstrated that GPER1 and OXTR aggregates might be formed due to interactions with GAGs rather than arising from changes of levels of these proteins in cells., (Copyright © 2022 The Authors. Published by Elsevier GmbH.. All rights reserved.)

Mucopolysaccharidosis type I (MPS-I) is a rare lysosomal storage disorder caused by deficiency of the enzyme alpha-L-iduronidase, which removes iduronic acid in both chondroitin/dermatan sulfate (CS/DS) and heparan sulfate (HS) and thereby contributes to the catabolism of glycosaminoglycans (GAGs). To ameliorate this genetic defect, the patients are currently treated by enzyme replacement and bone marrow transplantation, which have a number of drawbacks. This study was designed to develop an alternative treatment by inhibition of iduronic acid formation. By screening the Prestwick drug library, we identified ebselen as a potent inhibitor of enzymes that produce iduronic acid in CS/DS and HS. Ebselen efficiently inhibited iduronic acid formation during CS/DS synthesis in cultured fibroblasts. Treatment of MPS-I fibroblasts with ebselen not only reduced accumulation of CS/DS but also promoted GAG degradation. In early Xenopus embryos, this drug phenocopied the effect of downregulation of DS-epimerase 1, the main enzyme responsible for iduronic production in CS/DS, suggesting that ebselen inhibits iduronic acid production in vivo. However, ebselen failed to ameliorate the CS/DS and GAG burden in MPS-I mice. Nevertheless, the results propose a potential of iduronic acid substrate reduction therapy for MPS-I patients., (© The Author(s) 2021. Published by Oxford University Press.)

Background: Allogeneic hematopoietic stem-cell transplantation is the standard of care for Hurler syndrome (mucopolysaccharidosis type I, Hurler variant [MPSIH]). However, this treatment is only partially curative and is associated with complications., Methods: We are conducting an ongoing study involving eight children with MPSIH. At enrollment, the children lacked a suitable allogeneic donor and had a Developmental Quotient or Intelligence Quotient score above 70 (i.e., none had moderate or severe cognitive impairment). The children received autologous hematopoietic stem and progenitor cells (HSPCs) transduced ex vivo with an α-L-iduronidase (IDUA)-encoding lentiviral vector after myeloablative conditioning. Safety and correction of blood IDUA activity up to supraphysiologic levels were the primary end points. Clearance of lysosomal storage material as well as skeletal and neurophysiological development were assessed as secondary and exploratory end points. The planned duration of the study is 5 years., Results: We now report interim results. The children's mean (±SD) age at the time of HSPC gene therapy was 1.9±0.5 years. At a median follow-up of 2.10 years, the procedure had a safety profile similar to that known for autologous hematopoietic stem-cell transplantation. All the patients showed prompt and sustained engraftment of gene-corrected cells and had supraphysiologic blood IDUA activity within a month, which was maintained up to the latest follow-up. Urinary glycosaminoglycan (GAG) excretion decreased steeply, reaching normal levels at 12 months in four of five patients who could be evaluated. Previously undetectable levels of IDUA activity in the cerebrospinal fluid became detectable after gene therapy and were associated with local clearance of GAGs. Patients showed stable cognitive performance, stable motor skills corresponding to continued motor development, improved or stable findings on magnetic resonance imaging of the brain and spine, reduced joint stiffness, and normal growth in line with World Health Organization growth charts., Conclusions: The delivery of HSPC gene therapy in patients with MPSIH resulted in extensive metabolic correction in peripheral tissues and the central nervous system. (Funded by Fondazione Telethon and others; ClinicalTrials.gov number, NCT03488394; EudraCT number, 2017-002430-23.)., (Copyright © 2021 Massachusetts Medical Society.)

Mucopolysaccharidosis type I (MPS I) is a lysosomal storage disorder characterized by alpha-L-iduronidase (IDUA) deficiency, an enzyme responsible for glycosaminoglycan degradation. Musculoskeletal impairment is an important component of the morbidity related to the disease, as it has a major impact on patients' quality of life. To understand how this disease affects bone structure, morphological, biomechanical and histological analyses of femurs from 3- and 6-month-old wild type (Idua +/+) and MPS I knockout mice (Idua -/-) were performed. Femurs from 3-month-old Idua -/- mice were found to be smaller and less resistant to fracture when compared to their age matched controls. In addition, at this age, the femurs presented important alterations in articular cartilage, trabecular bone architecture, and deposition of type I and III collagen. At 6 months of age, femurs from Idua -/- mice were more resistant to fracture than those from Idua +/+. Our results suggest that the abnormalities observed in bone matrix and articular cartilage in 3-month-old Idua -/- animals caused bone tissue to be less flexible and more likely to fracture, whereas in 6-month-old Idua -/- group the ability to withstand more load before fracturing than wild type animals is possibly due to changes in the bone matrix., (Copyright © 2020 Elsevier GmbH. All rights reserved.)

Mucopolysaccharidosis type I (MPS I) is a lysosomal storage disease caused by a mutation in the IDUA gene, which codes α-L-iduronidase (IDUA), a lysosomal hydrolase that degrades two glycosaminoglycans (GAGs): heparan sulfate (HS) and dermatan sulfate (DS). GAGs are macromolecules found mainly in the extracellular matrix and have important signaling and structural roles which are essential to the maintenance of cell and tissue physiology. Nondegraded GAGs accumulate in various cell types, which characterizes MPS I as a multisystemic progressive disease. Many tissues and vital organs have been described in MPS I models, but there is a lack of studies focused on their effects on the reproductive tract. Our previous studies indicated lower sperm production and morphological damage in the epididymis and accessory glands in male MPS I mice, despite their ability to copulate and to impregnate females. Our aim was to improve the testicular characterization of the MPS I model, with a specific focus on ultrastructural observation of the different cell types that compose the seminiferous tubules and interstitium. We investigated the testicular morphology of 6-month-old male C57BL/6 wild-type (Idua+/+) and MPS I (Idua-/-) mice. We found vacuolated cells widely present in the interstitium and important signs of damage in myoid, Sertoli and Leydig cells. In the cytoplasmic region of Sertoli cells, we found an increased number of vesicles with substrates under digestion and a decreased number of electron-dense vesicles similar to lysosomes, suggesting an impaired flux of substrate degradation. Conclusions: Idua exerts an important role in the morphological maintenance of the seminiferous tubules and the testicular interstitium, which may influence the quality of spermatogenesis, having a greater effect with the progression of the disease.

Background : Mucopolysaccharidosis type I-Hurler (MPS1-H) is a severe genetic lysosomal storage disorder due to loss-of-function mutations in the IDUA gene. The subsequent complete deficiency of alpha l-iduronidase enzyme is directly responsible of a progressive accumulation of glycosaminoglycans (GAG) in lysosomes which affects the functions of many tissues. Consequently, MPS1 is characterized by systemic symptoms (multiorgan dysfunction) including respiratory and cardiac dysfunctions, skeletal abnormalities and early fatal neurodegeneration. Methods : To understand mechanisms underlying MPS1 neuropathology, we generated induced pluripotent stem cells (iPSC) from a MPS1-H patient with loss-of-function mutations in both IDUA alleles. To avoid variability due to different genetic background of iPSC, we established an isogenic control iPSC line by rescuing IDUA expression by a lentivectoral approach. Results : Marked differences between MPS1-H and IDUA -corrected isogenic controls were observed upon neural differentiation. A scratch assay revealed a strong migration defect of MPS1-H cells. Also, there was a massive impact of IDUA deficiency on gene expression (340 genes with an FDR <0.05). Conclusions : Our results demonstrate a hitherto unknown connection between lysosomal degradation, gene expression and neural motility, which might account at least in part for the phenotype of MPS1-H patients.

Mucopolysaccharidosis type I (MPS I) is a lysosomal disease with progressive central nervous system involvement. This study examined the lipid, cholesterol, and myelin basic protein composition of white matter in the corpus callosum of MPS I mice. We studied 50 week-old, male MPS I mice and littermate, heterozygote controls (n = 12 per group). Male MPS I mice showed lower phosphatidylcholine and ether-linked phosphatidylcholine quantities than controls (p < 0.05). Twenty-two phospholipid or ceramide species showed significant differences in percent of total. Regarding specific lipid species, MPS I mice exhibited lower quantities of sphingomyelin 18:1, phosphatidylserine 38:3, and hexosylceramide d18:1(22:1) mH 2 O than controls. Principal components analyses of polar, ceramide, and hexosylceramide lipids, respectively, showed some separation of MPS I and control mice. We found no significant differences in myelin gene expression, myelin basic protein, or total cholesterol in the MPS I mice versus heterozygous controls. There was a trend toward lower proteolipid protein-1 levels in MPS I mice (p = 0.06). MPS I mice show subtle changes in white matter composition, with an unknown impact on pathogenesis in this model., (© 2020 AOCS.)

During brain maturation, cation-independent mannose-6-phosphate receptor (CI-MPR), a key transporter for lysosomal hydrolases, decreases significantly on the blood-brain barrier (BBB). Such a phenomenon leads to poor brain penetration of therapeutic enzymes and subsequent failure in reversing neurological complications in patients with neuropathic lysosomal storage diseases (nLSDs), such as Hurler syndrome (severe form of mucopolysaccharidosis type I [MPS I]). In this study, we discover that upregulation of microRNA-143 (miR-143) contributes to the decline of CI-MPR on the BBB during development. Gain- and loss-of-function studies showed that miR-143 inhibits CI-MPR expression and its transport function in human endothelial cells in vitro. Genetic removal of miR-143 in MPS I mice enhances CI-MPR expression and improves enzyme transport across the BBB, leading to brain metabolic correction, pathology normalization, and correction of neurological functional deficits 5 months after peripheral protein delivery at clinically relevant levels that derived from erythroid/megakaryocytic cells via hematopoietic stem cell-mediated gene therapy, when otherwise no improvement was observed in MPS I mice at a parallel setting. These studies not only uncover a novel role of miR-143 as an important modulator for the developmental decline of CI-MPR on the BBB, but they also demonstrate the functional significance of depleting miR-143 for "rescuing" BBB-anchored CI-MPR on advancing CNS treatment for nLSDs., (Copyright © 2020 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.)

The cause of neurodegeneration in MPS mouse models is the focus of much debate and what the underlying cause of disease pathology in MPS mice is. The timing of development of pathology and when this can be reversed or impacted is the key to developing suitable therapies in MPS. This study is the first of its kind to correlate the biochemical changes with the functional outcome as assessed using non-invasive behaviour testing across multiple mucopolysaccharidosis (MPS) mouse models. In the MPS brain, the primary lysosomal enzyme dysfunction leads to accumulation of primary glycosaminoglycans (GAGs) with gangliosides (G M2 and G M3 ) being the major secondary storage products. With a focus on the neuropathology, a time course experiment was conducted in MPS I, MPS IIIA, MPS VII (severe and attenuated models) in order to understand the relative timing and level of GAG and ganglioside accumulation and how this correlates to behaviour deficits. Time course analysis from 1 to 6 months of age was conducted on brain samples to assess primary GAG (uronic acid), β-hexosaminidase enzyme activity and levels of G M2 and G M3 gangliosides. This was compared to a battery of non-invasive behaviour tests including open field, inverted grid, rotarod and water cross maze were assessed to determine effects on motor function, activity and learning ability. The results show that the GAG and ganglioside accumulation begins prior to the onset of detectable changes in learning ability and behaviour. Interestingly, the highest levels of GAG and ganglioside accumulation was observed in the MPS IIIA mouse despite having 3% residual enzyme activity. Deficits in motor function were clearly observed in the severe Gus mps/mps , which were significantly delayed in the attenuated Gus tm(L175F)Sly model despite their minimal increase in detectable enzyme activity. This suggests that genotype and residual enzyme activity are not indicative of severity of disease pathology in MPS disease and there exists a window when there are considerable storage products without detectable functional deficits which may allow an alteration to occur with therapy., (Crown Copyright © 2020. Published by Elsevier Inc. All rights reserved.)

Mucopolysaccharidosis type I is a genetic disorder with impaired glycosaminoglycan degradation. Cardiac pathologic involvement in this subset of patients is predominantly valvular heart disease. Valvular heart disease seen in these patients will most likely require surgical intervention in their lifetime. Only a limited amount of reports are dedicated to the cardiac surgical management of mucopolysaccharidoses. We present the case of a 32-year-old female with Hurler-Scheie syndrome who required multiple valve replacements due to progression of valvular dysfunction and decline in the quality of life. Multidisciplinary evaluation and discussion early are crucial for quality of life optimization in this cohort of patients.

Our previous study delivered zinc finger nucleases to treat mice with mucopolysaccharidosis type I (MPS I), resulting in a phase I/II clinical trial (ClinicalTrials.gov: NCT02702115). However, in the clinical trial, the efficacy needs to be improved due to the low transgene expression level. To this end, we designed a proprietary system (PS) gene editing approach with CRISPR to insert a promoterless α-l-iduronidase (IDUA) cDNA sequence into the albumin locus of hepatocytes. In this study, adeno-associated virus 8 (AAV8) vectors delivering the PS gene editing system were injected into neonatal and adult MPS I mice. IDUA enzyme activity in the brain significantly increased, while storage levels were normalized. Neurobehavioral tests showed that treated mice had better memory and learning ability. Also, histological analysis showed efficacy reflected by the absence of foam cells in the liver and vacuolation in neuronal cells. No vector-associated toxicity or increased tumorigenesis risk was observed. Moreover, no off-target effects were detected through the unbiased genome-wide unbiased identification of double-stranded breaks enabled by sequencing (GUIDE-seq) analysis. In summary, these results showed the safety and efficacy of the PS in treating MPS I and paved the way for clinical studies. Additionally, as a therapeutic platform, the PS has the potential to treat other lysosomal diseases., (Copyright © 2020 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.)

Mucopolysaccharidosis type I (MPS I) is a genetic disease caused by a deficiency of the lysosomal hydrolase α-L-iduronidase (IDUA). IDUA degrades two types of glycosaminoglycans (GAGs): heparan and dermatan sulfates, important components of extracellular matrix, with signaling and structural functions. The accumulation of GAGs results in progressive physiological impairments in a variety of tissues, making MPS I a complex and multisystemic disease. Due the advent of therapeutic strategies which have increased patients' life expectancy, our group have been investigating the effect of IDUA deficiency on the reproductive system. In the present study, we aimed to characterize some of the accessory glands of the male reproductive tract in an MPS I mouse model. We used 6-month-old Idua+/+ and Idua-/- male mice to evaluate the histology of the seminal vesicles and prostate. Interstitial deposits of GAGs and collagen fibers were also observed. Seminal vesicles were smaller in the Idua-/- group, regardless of the normal staining pattern of the epithelial cells, marked with antiandrogen receptor. The prostate of Idua-/- mice presented necrotic acini and increased deposition of collagen fibers in the interstitium. All glands presented evident deposits of GAGs in the extracellular matrix, especially inside vacuolated interstitial cells. We concluded that, at this stage of the disease, the prostate is the most damaged accessory gland and may therefore, be the first to manifest functional impairments during disease progression.

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.

Mucopolysaccharidosis type I (MPS I) is a lysosomal storage disease caused by a deficiency of the lysosomal hydrolase, α-L-iduronidase (IDUA). IDUA degrades heparan and dermatan sulfates, two types of glycosaminoglycan (GAG), important signalling and structural molecules of the extracellular matrix. Because many cell types store GAGs, MPS I has been investigated in human and animal models. Enzyme replacement therapy is available for MPS I patients and has improved their life expectancy, allowing them to achieve reproductive age. The aim of this study was to evaluate epididymal and sperm morphology and function in a murine model of MPS I. We used C57BL Idua+/+ and Idua-/- adult male mice (6 months old) to investigate epididymal morphology, sperm ultrastructure, GAG characterisation and mating competence. Epithelial GAG storage, especially in the cauda epididymidis, was seen in Idua-/- mice. Regardless of the morphologic change and GAG storage found in the cauda epididymis, sperm morphology and motility were normal, similar to wild types. In the interstitium, vacuolated cells were found in addition to deposits of GAGs. Mating was not impaired in Idua-/- males and litter sizes were similar between groups. At the time point of the disease evaluated, the deficiency in IDUA affected the morphology of the epididymis in male Idua-/- mice, whereas sperm appearance and motility and the male's capacity to mate and impregnate females were preserved.

Allogeneic hematopoietic cell transplantation (HCT) benefits children with Hurler syndrome (MPS-IH). However, survivors remain burdened by substantial MPS-IH related residual disease. We studied the feasibility, safety and biochemical impact of augmentative recombinant intravenous enzyme replacement therapy (IV-ERT) post transplantation. Ten children with MPS-IH and ≥2 years from successful HCT underwent IV-ERT for 2 years' duration. Patients were monitored for anti-drug antibody (ADA) development, including inhibitory capacity and changes in urinary excretion of glycosaminoglycans (uGAG). Three patients demonstrated low-level ADA at baseline, though all children tolerated IV-ERT well. Eight patients developed ADA over the 2-year study, with 3 (38%) meeting criteria for an inhibitory ADA response. The aggregate cohort experienced a reduction in uGAG from baseline to study end, which was enhanced in children with low or no ADA response. Conversely, children with inhibitory ADA showed increase in uGAG over time. IV-ERT in previously transplanted children with MPS-IH appears safe and can reduce uGAG, although this is reversed by the presence of inhibitory ADA. These data show a biochemical change after initiation of post-HCT IV-ERT, but the occurrence of ADA and inhibitory antibodies are a concern and should be monitored in future efficacy trials. This trial was registered at www.clinicaltrials.gov , NCT01173016, 07/30/2010.

Mucopolysaccharidosis type IH (MPS IH, Hurler syndrome) is a progressive, multisystem autosomal recessive lysosomal storage disorder resulting in the consequent accumulation of glycosaminoglycans. It is well recognized that early hematopoietic stem cell transplantation (HSCT) prevents neurocognitive decline in MPS IH. We followed the divergent clinical course in two Japanese siblings with MPS IH. The elder sister (proband) received a diagnosis of MPS IH at 6 months old. At the time of this diagnosis enzyme replacement therapy (ERT) was not available in Japan. She developed severe and recurrent respiratory disease and died at 1 year 10 months of age. Her younger sister also received a diagnosis of MPS IH, but at 18 days of age, and started ERT at 34 days of age. ERT continued until 8 months of age and prevented the progression of somatic manifestations of MPS IH. She received HSCT at 9 months old. Five years after HSCT she had no symptoms of MPS IH except for mild signs of dysostosis multiplex and mild cardiac valvular disease. Her neurological function was generally preserved compared with her elder sister. The prognosis and quality of life differed significantly between the sisters. Therefore, early HSCT can preserve neurocognition by preventing the neurodegeneration from MPS IH. In addition, ERT initiated during the asymptomatic period prevented the patient from developing somatic manifestations and enabled successful HSCT in this case., (Copyright © 2019. Published by Elsevier B.V.)

Mucopolysaccharidosis type I (MPS I) is a severe disease due to deficiency of the lysosomal hydrolase α-L-iduronidase (IDUA) and the subsequent accumulation of the glycosaminoglycans (GAG), leading to progressive, systemic disease and a shortened lifespan. Current treatment options consist of hematopoietic stem cell transplantation, which carries significant mortality and morbidity risk, and enzyme replacement therapy, which requires lifelong infusions of replacement enzyme; neither provides adequate therapy, even in combination. A novel in vivo genome-editing approach is described in the murine model of Hurler syndrome. A corrective copy of the IDUA gene is inserted at the albumin locus in hepatocytes, leading to sustained enzyme expression, secretion from the liver into circulation, and subsequent uptake systemically at levels sufficient for correction of metabolic disease (GAG substrate accumulation) and prevention of neurobehavioral deficits in MPS I mice. This study serves as a proof-of-concept for this platform-based approach that should be broadly applicable to the treatment of a wide array of monogenic diseases., (Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.)

Mucopolysaccharidosis type I (MPS I) is a multisystemic disorder caused by the deficiency of alpha-L-iduronidase (IDUA) that leads to intracellular accumulation of glycosaminoglycans (GAG). In the present study we aimed to use cationic liposomes carrying the CRISPR/Cas9 plasmid and a donor vector for in vitro and in vivo MPS I gene editing, and compare to treatment with naked plasmids. The liposomal formulation was prepared by microfluidization. Complexes were obtained by the addition of DNA at +4/-1 charge ratio. The overall results showed complexes of about 110 nm, with positive zeta potential of +30 mV. The incubation of the complexes with fibroblasts from MPS I patients led to a significant increase in IDUA activity and reduction of lysosomal abnormalities. Hydrodynamic injection of the liposomal complex in newborn MPS I mice led to a significant increase in serum IDUA levels for up to six months. The biodistribution of complexes after hydrodynamic injection was markedly detected in the lungs and heart, corroborating the results of increased IDUA activity and decreased GAG storage especially in these tissues, while the group that received the naked plasmids presented increased enzyme activity especially in the liver. Furthermore, animals treated with the liposomal formulation presented improvement in cardiovascular parameters, one of the main causes of death observed in MPS I patients. We conclude that the IDUA production in multiple organs had a significant beneficial effect on the characteristics of MPS I disease, which may bring hope to gene therapy of Hurler patients., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Mucopolysaccharidosis type I (MPS I) is caused by deficiency of α-l-iduronidase (IDUA), a lysosomal enzyme involved in the breakdown and recycling of glycosaminoglycans (GAGs). Although enzyme replacement therapy is available, the efficacy of the treatment for neuropathic manifestations is limited. To facilitate drug discovery and model disease pathophysiology, we generated neural stem cells (NSCs) from MPS I patient-derived induced pluripotent stem cells (iPSCs). The NSCs exhibited characteristic disease phenotypes with deficiency of IDUA, accumulation of GAGs and enlargement of lysosomes, in agreement with the severity of clinical subgroups of MPS I. Transcriptome profiling of NSCs revealed 429 genes that demonstrated a more extensive change in expression in the most severe Hurler syndrome subgroup compared to the intermediate Hurler-Scheie or the least severe Scheie syndrome subgroups. Clustering and pathway analysis revealed high concordance of the severity of neurological defects with marked dysregulation of GAG biosynthesis, GAG degradation, lysosomal function and autophagy. Gene ontology (GO) analysis identified a dramatic upregulation of the autophagy pathway, especially in the Hurler syndrome subgroup. We conclude that GAG accumulation in the patient-derived cells disrupts lysosomal homeostasis, affecting multiple related cellular pathways in response to IDUA deficiency. These dysregulated processes likely lead to enhanced autophagy and progressively severe disease states. Our study provides potentially useful targets for clinical biomarker development, disease diagnosis and prognosis, and drug discovery.

Preparation of substituent-diverse, triazole-iduronic acid hybrid molecules by click reaction of an azido iduronic acid derivative with randomly chosen alkynes is described. Library members were screened for their ability to inhibit α-l-iduronidase, and hit molecules and analogues were then investigated for their ability to stabilize rh-α-IDUA in a thermal denaturation study. This work resulted in the discovery of the first small molecules that can be used to stabilize exogenous rh-α-IDUA protein in vitro.

Glycosaminoglycan (GAG) catabolism involves endo-hydrolysis of polysaccharides followed by the sequential removal of the non-reducing end residue from the resulting oligosaccharides by exo-enzymes. In the inherited metabolic disorder, mucopolysaccharidosis type I (MPS I), a deficiency in the exo-enzyme, α-l-iduronidase, prevents removal of α-l-iduronic acid residues from the non-reducing end of the GAGs, heparan sulphate (HS) and dermatan sulphate (DS). The excretion of partially degraded HS and DS in urine of MPS I patients has long been recognized, but the question of whether they do indeed reflect GAG load in a particular tissue has not been addressed - an important issue in the context of biomarkers for assessment of disease burden in MPS I. Therefore, we measured specific low molecular weight HS and DS oligosaccharides with terminal α-l-iduronic acid residues, in the brain, liver, kidney, serum and urine, and correlated these findings with total GAG in the MPS I mouse model. Six oligosaccharides were identified in the urine, ranging from di- to pentasaccharides. Of these, five were observed in the kidney, four in the liver and brain, with the three most abundant in urine also seen in serum. These oligosaccharides accounted for just 0.1-2% of total GAG, with a disaccharide showing the best correlation with total GAG. The oligosaccharides and total GAG were most abundant in the liver, with the least observed in the brain. The concentration of oligosaccharides as a percentage of total GAG in urine was similar to that observed in the kidney, and both revealed a similar ratio of HS:DS, suggesting that the oligosaccharide storage pattern in urine is a reflection of that in the kidney. Serum, liver and brain had a similar ratio of HS:DS, which was lower to that seen in the urine and kidney. The distribution of oligosaccharides when ranked from most to least abundant, was also the same between serum, liver and brain suggesting that serum more closely reflects the oligosaccharides of the brain and liver and may therefore be a more informative measurement of disease burden than urine. The accumulation of HS and DS oligosaccharides was observed in the brain as early as one month of age, with the disaccharide showing a continuous increase with age. This demonstrates the progressive nature of the disease and as such this disaccharide could prove to be a useful biomarker to measure disease burden in MPS I., (Copyright © 2017 Elsevier Inc. All rights reserved.)

This work aimed to investigate the potential effect of cyclodextrin encapsulation on intrinsic ability of daidzein (DAD) and genistein (GEN) to inhibit the glycosaminoglycan (GAG) synthesis in fibroblasts originating from patients with mucopolysaccharidosis (MPS), type II and III. DAD or GEN encapsulation with either 2-hydroxypropyl-β-cyclodextrin or sulphobuthylether-β-cyclodextrin were achieved by neat grinding and were characterised by thermal analysis, X-ray powder diffraction, scanning electron microscopy and solubility testing which confirmed the complexes formation with increased solubility with respect to starting compounds. Both isoflavones, as well as their co-ground cyclodextrin complexes reduced GAG levels in the fibroblasts of MPS II and MPS III patients from 54.8-77.5%, in a dose dependent manner, without any significant cytotoxic effect. Cyclodextrin encapsulation did not change the intrinsically high effect of both DAD and GEN on the GAG level reduction in the treated cells, thus could be considered as a part of combination therapies of MPS.

The mucopolysaccharidoses (MPS) are rare genetic disorders marked by severe somatic and neurological symptoms. Development of treatments for the neurological manifestations of MPS has been hindered by the lack of objective measures of central nervous system disease burden. Identification of biomarkers for central nervous system disease in MPS patients would facilitate the evaluation of new agents in clinical trials. High throughput metabolite screening of cerebrospinal fluid (CSF) samples from a canine model of MPS I revealed a marked elevation of the polyamine, spermine, in affected animals, and gene therapy studies demonstrated that reduction of CSF spermine reflects correction of brain lesions in these animals. In humans, CSF spermine was elevated in neuropathic subtypes of MPS (MPS I, II, IIIA, IIIB), but not in subtypes in which cognitive function is preserved (MPS IVA, VI). In MPS I patients, elevated CSF spermine was restricted to patients with genotypes associated with CNS disease and was reduced following hematopoietic stem cell transplantation, which is the only therapy currently capable of improving cognitive outcomes. Additional studies in cultured neurons from MPS I mice showed that elevated spermine was essential for the abnormal neurite overgrowth exhibited by MPS neurons. These findings offer new insights into the pathogenesis of CNS disease in MPS patients, and support the use of spermine as a new biomarker to facilitate the development of next generation therapeutics for MPS., (© The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Umbilical cord blood (UCB) is a promising source of stem cells to use in early haematopoietic stem cell transplantation (HSCT) approaches for several genetic diseases that can be diagnosed at birth. Mucopolysaccharidosis type I (MPS-I) is a progressive multi-system disorder caused by deficiency of lysosomal enzyme α-L-iduronidase, and patients treated with allogeneic HSCT at the onset have improved outcome, suggesting to administer such therapy as early as possible. Given that the best characterized MPS-I murine model is an immunocompetent mouse, we here developed a transplantation system based on murine UCB. With the final aim of testing the therapeutic efficacy of UCB in MPS-I mice transplanted at birth, we first defined the features of murine UCB cells and demonstrated that they are capable of multi-lineage haematopoietic repopulation of myeloablated adult mice similarly to bone marrow cells. We then assessed the effectiveness of murine UCB cells transplantation in busulfan-conditioned newborn MPS-I mice. Twenty weeks after treatment, iduronidase activity was increased in visceral organs of MPS-I animals, glycosaminoglycans storage was reduced, and skeletal phenotype was ameliorated. This study explores a potential therapy for MPS-I at a very early stage in life and represents a novel model to test UCB-based transplantation approaches for various diseases.

Background: Mucopolysaccharidosis type I (MPS I) is an autosomal recessive disease due to deficiency of α-L-iduronidase (IDUA), a lysosomal enzyme that degrades glycosaminoglycans (GAG) heparan and dermatan sulfate. To achieve optimal clinical outcomes, early and proper treatment is essential, which requires early diagnosis and phenotype severity prediction., Results: To establish a genotype/phenotype correlation of MPS I disease, a combination of bioinformatics tools including SIFT, PolyPhen, I-Mutant, PROVEAN, PANTHER, SNPs&GO and PHD-SNP are utilized. Through analyzing single nucleotide polymorphisms (SNPs) by these in silico approaches, 28 out of 285 missense SNPs were predicted to be damaging. By integrating outcomes from these in silico approaches, a prediction algorithm (sensitivity 94%, specificity 80%) was thereby developed. Three dimensional structural analysis of 5 candidate SNPs (P533R, P496R, L346R, D349G, T374P) were performed by SWISS PDB viewer, which revealed specific structural changes responsible for the functional impacts of these SNPs. Additionally, SNPs in the untranslated region were analyzed by UTRscan and PolymiRTS. Moreover, by investigating known pathogenic mutations and relevant patient phenotypes in previous publications, phenotype severity (severe, intermediate or mild) of each mutation was deduced., Conclusions: Collectively, these results identified potential candidate SNPs with functional significance for studying MPS I disease. This study also demonstrates the effectiveness, reliability and simplicity of these in silico approaches in addressing complexity of underlying genetic basis of MPS I disease. Further, a step-by-step guideline for phenotype prediction of MPS I disease is established, which can be broadly applied in other lysosomal diseases or genetic disorders.

Mucopolysaccharidosis type I (MPSI) is caused by mutations in the gene encoding the lysosomal enzyme, α-l-iduronidase (IDUA), and patients with MPSI are currently treated with IDUA enzyme replacement therapy (ERT). However, the majority of MPSI patients have severe CNS involvement, and conventional ERT does not treat the brain. The failure of ERT to treat the brain is believed to be due to the lack of IDUA transport through the blood-brain barrier (BBB). However, BBB transport of IDUA has not been directly measured, to date. BBB transport of IDUA may be enhanced by fusion of the enzyme to a monoclonal antibody (mAb) against the human insulin receptor (HIR). The HIRMAb binds the insulin receptor on the BBB to trigger transport into the brain and acts as a molecular Trojan horse to deliver IDUA to brain cells. Therefore, the purpose of the present investigation was to compare, side-by-side, the BBB transport of IDUA alone and the HIRMAb-IDUA fusion protein in the Rhesus monkey in vivo. Each protein was radio-iodinated by conjugation with the [ 125 I]-Bolton-Hunter reagent and injected intravenously (IV) in the primate. The uptake by brain, and peripheral organs, was measured by whole body autoradiography. The results show there is no transport of IDUA alone into the brain, but that the brain uptake of the HIRMAb-IDUA fusion protein is high, 1.2% injected dose/brain. There is comparable uptake of the IDUA and the HIRMAb-IDUA fusion protein by peripheral organs, where uptake is primarily controlled by the mannose 6-phosphate receptor. The work suggests that treatment of MPSI with the HIRMAb-IDUA fusion protein will be as effective as IDUA in peripheral organs, but offer the benefit of treatment of the central nervous system in MPSI.

Gangliosides are the most complex oligosaccharide-containing glycosphingolipids defined by the presence of sialic acid and although present in all tissues, predominate in the brain. Considering their importance in neural development, it is unsurprising that ganglioside metabolism is altered in neurodegenerative diseases. The severe form of mucopolysaccharidosis type I, Hurler syndrome (HS), is characterised by progressive loss of neuronal function through largely undefined mechanisms. Here, we sought to interrogate brain gangliosides in a murine model of HS and further, assessed whether dietary modulation of lipid metabolism effected correction of the metabolic abnormalities. The simple gangliosides, G M 2 , G M 3 , G D 2 and G D 3 were elevated in the five subregions examined - brain stem, cerebellum, cortex, hippocampus, subcortex - in HS mice as early as 2 months of age compared with their wild type counterparts. Their elevation persisted at 6 months of age, imparting protracted neurological development as these simple gangliosides have usually subsided by this stage of brain development. Their immediate synthetic precursor, lactosylceramide, was also elevated, suggesting that their increase arises at this metabolic intermediary, as dihydroceramide, ceramide and monohexosylceramide were unaffected. Dietary linoleic acid supplementation significantly reduced G M 2 and G M 3 , and furthermore, improved exploratory behaviour as assessed by the open field test, highlighting the possibility of further exploring dietary intervention as a therapeutic consideration., (© 2017 International Society for Neurochemistry.)

Mucopolysaccharidosis type I (MPS I) is an autosomal disease caused by alpha-l-iduronidase (IDUA) deficiency. This study used IDUA knockout mice as a model to evaluate whether parameters such as dose of plasmid and time of treatment could influence the transfection efficiency of complexes formed with PEGylated cationic nanoemulsions and plasmid (pIDUA), which contains the gene that encodes for IDUA. Formulations were composed of medium chain triglycerides, 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine, 1,2-distearoyl- sn -glycero-3-phosphoethanolamine- N -(amino[polyethylene glycol]-2000), 1,2-dioleoyl-sn-glycero-3-trimethylammonium propane (DOTAP), glycerol, and water and were prepared by the adsorption or encapsulation of preformed pIDUA-DOTAP complexes by high-pressure homogenization. A progressive increase in IDUA expression was observed with an increase in the dose and time of transfection for mice treated with both complexes (adsorbed and encapsulated), especially in the liver. Regardless of the complex administered, a significant increase in IDUA activity was detected in lungs and liver compared with nontreated MPS I when a dose of 60 μg was administered and IDUA activity was measured 7 days postadministration. Tissue sections of major organs showed no presence of cell necrosis, inflammatory infiltrate, or an increase in apoptosis. Furthermore, immunohistochemistry for CD68 showed no difference in the number of macrophage cells in treated and nontreated animals, indicating the absence of inflammatory reaction caused by the treatment. The data set obtained in this study allowed establishing that factors such as dose and time can influence transfection efficiency in different degrees and that these complexes did not lead to any lethal effect in the MPS I murine model used., Competing Interests: Disclosure The authors report no conflicts of interest in this work.

Mucopolysaccharidosis type I (MPS I) is a lysosomal disorder characterized by a deficiency of alpha-L-iduronidase and storage of undegraded glycosaminoglycans (GAGs). Clinical findings of the disease include heart failure, and patients often need valve replacement. It has been shown that, later in life, MPS I mice develop those abnormalities, but to date, there have not been studies on the progression and pathogenesis of the disease. Therefore, in the present study, we evaluated heart function in normal and MPS I male mice from 2 to 8 months of age. Echocardiographic analysis showed left ventricular enlargement with progressive reduction in ejection fraction, fractional area change, and left ventricular fractional shortening in the MPS I hearts at 6 and 8 months of age and a reduction in acceleration time/ejection time ratio of the pulmonary artery starting at 6 months of age, which suggests pulmonary vascular resistance. Histological and biochemical analysis confirmed progressive GAG storage from 2 months of age and onwards in the myocardium and heart valves, which had also increased in thickness. Additionally, macrophages were present in the MPS I heart tissue. Collagen content was reduced in the MPS I mouse valves. Cathepsin B, an enzyme that is known to be able to degrade collagen and is involved in heart dilatation, displayed a marked elevation in activity in the MPS I mice and could be responsible for the heart dilatation and valves alterations observed. Our results suggest that the MPS I mice have progressive heart failure and valve disease, which may be caused by cathepsin B overexpression., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Mucopolysaccharidosis type I (MPS I) is due to deficiency of α-l-iduronidase (IDUA) and subsequent storage of undegraded glycosaminoglycans (GAG). The severe form of the disease, known as Hurler syndrome, is characterized by mental retardation and neurodegeneration of unknown etiology. To identify potential biomarkers and unveil the neuropathology mechanism of MPS I disease, two-dimensional polyacrylamide gel electrophoresis (PAGE) and nanoliquid chromatography-tandem mass spectrometry (nanoLC-MS/MS) were applied to compare proteome profiling of brains from MPS I and control mice (5-month old). A total of 2055 spots were compared, and 25 spots (corresponding to 50 different proteins) with a fold change ≥3.5 and a p value <0.05 between MPS I and control mice were further analyzed by nanoLC-MS/MS. These altered proteins could be divided into three major groups based on Gene Ontology (GO) terms: proteins involved in metabolism, neurotransmission and cytoskeleton. Cytoskeletal proteins including ACTA1, ACTN4, TUBB4B and DNM1 were significantly downregulated. STXBP1, a regulator of synaptic vesicle fusion and docking was also downregulated, indicating impaired synaptic transmission. Additionally, proteins regulating Ca 2+ and H + homeostasis including ATP6V1B2 and RYR3 were downregulated, which may be related to disrupted autophagic and endocytotic pathways. Notably, there is no altered expression in proteins associated with cell death, ubiquitin or inflammation. These results for the first time highlight the important role of alterations in metabolism pathways, intracellular ionic homeostasis and the cytoskeleton in the neuropathology of MPS I disease. The proteins identified in this study would provide potential biomarkers for diagnostic and therapeutic studies of MPS I., (Copyright © 2016 Elsevier Inc. All rights reserved.)

The prevalence of aortic root dilatation (ARD) in mucopolysaccharidosis (MPS) is not well documented. We investigated aortic root measurements in 34 MPS patients at the Children's Hospital of Orange County (CHOC). The diagnosis, treatment status, age, gender, height, weight and aortic root parameters (aortic valve annulus (AVA), sinuses of Valsalva (SoV), and sinotubular junction (STJ)) were extracted by retrospective chart review and echocardiographic measurements. Descriptive statistics, ANOVA, and paired post-hoc t -tests were used to summarize the aortic dimensions. Exact binomial 95% confidence intervals (CIs) were constructed for ARD, defined as a z- score greater than 2 at the SoV. The patient age ranged from 3.4-25.9 years (mean 13.3 ± 6.1), the height from 0.87-1.62 meters (mean 1.24 ± 0.21), and the weight from 14.1-84.5 kg (mean 34.4 ± 18.0). The prevalence of dilation at the AVA was 41% (14/34; 95% CI: 25%-59%); at the SoV was 35% (12/34; 95% CI: 20%-54%); and at the STJ was 30% (9/30; 95% CI: 15%-49%). The highest prevalence of ARD was in MPS IVa (87.5%). There was no significant difference between mean z-scores of MPS patients who received treatment with hematopoietic stem cell transplantation (HSCT) or enzyme replacement therapy (ERT) vs. untreated MPS patients at the AVA ( z = 1.9 ± 2.5 vs. z = 1.5 ± 2.4; p = 0.62), SoV ( z = 1.2 ± 1.6 vs. z = 1.3 ± 2.2; p = 0.79), or STJ ( z = 1.0 ± 1.8 vs. z = 1.2 ± 1.6; p = 0.83). The prevalence of ARD was 35% in our cohort of MPS I-VII patients. Thus, we recommend screening for ARD on a routine basis in this patient population., Competing Interests: The authors declare no conflict of interest.

Current treatment options for MPS I have limited effects on some organs, including the skeletal system. In MPS animal models pentosan polysulphate (PPS) reduces the concentrations of glycosaminoglycans (GAGs) in tissues and body fluids and improves cartilaginous and osseous pathologies. The goals of this study were to investigate primarily the safety and secondary the clinical effects, concerning mobility and pain, of PPS treatment in MPS I patients. Four MPS I-Hurler-Scheie/-Scheie patients aged 35.6 ± 6.4 years with one male were included in the study. All patients were on enzyme replacement therapy since 9.45 ± 3.75 years. PPS was applied subcutaneously in two patients with 1 mg/kg and in two patients with 2 mg/kg, weekly for 12 weeks and then biweekly for 12 weeks. The 24-week treatment with PPS was well tolerated by all patients. Urinary GAG concentrations were reduced from 4.13 ± 1.17 at baseline to 2.69 ± 0.36 mg/mmol creatinine after 24-week treatment with 1 mg/kg PPS, and from 6.71 ± 0.62 to 2.65 ± 0.09 mg/mmol creatinine with 2 mg/kg PPS. An improvement in range of motion was noted in three out of four patients. The pain intensity score was reduced from 4.5 ± 1.77 at baseline to 1.8 ± 0.47 after 24-week treatment with 1 mg/kg PPS; patients with 2 mg/kg PPS already had minimal pain at the start of the study. In conclusion, PPS treatment in a small number of adult MPS I patients was well tolerated and resulted in a significant reduction of urinary GAG excretion and in an improvement of joint mobility and pain.

The mucopolysaccharidoses (MPSs) comprise a group of lysosomal storage disorders characterized by deficient degradation and subsequent accumulation of glycosaminoglycans (GAGs). Progressive bone and joint disease are a major cause of morbidity, and current therapeutic strategies have limited effect on these symptoms. By elucidating pathophysiological mechanisms underlying bone disease, new therapeutic targets may be identified. Longitudinal growth is regulated by interaction between GAGs and growth factors. Because GAGs accumulate in the MPSs, we hypothesized that altered interaction between growth factors and GAGs contribute to the pathogenesis of MPS bone disease. In this study, binding between GAGs from MPS I chondrocytes and fibroblast growth factor 2 (FGF2) was not significantly different from binding of FGF2 to GAGs from control chondrocytes. FGF2 signaling, however, was increased in MPS I chondrocytes after incubation with FGF2, as compared to control chondrocytes. Using bone cultures, we demonstrated decreased growth of WT mouse bones after incubation with FGF2, but no effect on MPS I bone growth. However, MPS I bones showed decreased growth in the presence of GAGs from MPS I chondrocytes. Finally, we demonstrate altered GAG distribution in MPS I chondrocytes, and altered GAG, FGF2 and Indian hedgehog distribution in growth plates from MPS I mice. In summary, our results suggest that altered interaction and distribution of growth factors and accumulated GAGs may contribute to the pathogenesis of MPS bone disease. In the future, targeting growth factor regulation or the interaction between in growth factors and GAGs might be a promising therapeutic strategy for MPS bone disease., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Lysosomal Storage Disorders (LSDs) are a group of metabolic syndromes, each one due to the deficit of one lysosomal enzyme. Many LSDs affect most of the organ systems and overall about 75% of the patients present neurological impairment. Enzyme Replacement Therapy, although determining some systemic clinical improvements, is ineffective on the CNS disease, due to enzymes' inability to cross the blood-brain barrier (BBB). With the aim to deliver the therapeutic enzymes across the BBB, we here assayed biodegradable and biocompatible PLGA-nanoparticles (NPs) in two murine models for LSDs, Mucopolysaccharidosis type I and II (MPS I and MPS II). PLGA-NPs were modified with a 7-aminoacid glycopeptide (g7), yet demonstrated to be able to deliver low molecular weight (MW) molecules across the BBB in rodents. We specifically investigated, for the first time, the g7-NPs ability to transfer a model drug (FITC-albumin) with a high MW, comparable to the enzymes to be delivered for LSDs brain therapy. In vivo experiments, conducted on wild-type mice and knockout mouse models for MPS I and II, also included a whole series of control injections to obtain a broad preliminary view of the procedure efficiency. Results clearly showed efficient BBB crossing of albumin in all injected mice, underlying the ability of NPs to deliver high MW molecules to the brain. These results encourage successful experiments with enzyme-loaded g7-NPs to deliver sufficient amounts of the drug to the brain district on LSDs, where exerting a corrective effect on the pathological phenotype.

Background: We previously demonstrated the therapeutic benefits of pentosan polysulfate (PPS) in a rat model of mucopolysaccharidosis (MPS) type VI. Reduction of inflammation, reduction of glycosaminoglycan (GAG) storage, and improvement in the skeletal phenotype were shown. Herein, we evaluate the long-term safety and therapeutic effects of PPS in a large animal model of a different MPS type, MPS I dogs. We focused on the arterial phenotype since this is one of the most consistent and clinically significant features of the model., Methodology/principal Findings: MPS I dogs were treated with daily oral or biweekly subcutaneous (subQ) PPS at a human equivalent dose of 1.6 mg/kg for 17 and 12 months, respectively. Safety parameters were assessed at 6 months and at the end of the study. Following treatment, cytokine and GAG levels were determined in fluids and tissues. Assessments of the aorta and carotid arteries also were performed. No drug-related increases in liver enzymes, coagulation factors, or other adverse effects were observed. Significantly reduced IL-8 and TNF-alpha were found in urine and cerebrospinal fluid (CSF). GAG reduction was observed in urine and tissues. Increases in the luminal openings and reduction of the intimal media thickening occurred in the carotids and aortas of PPS-treated animals, along with a reduction of storage vacuoles. These results were correlated with a reduction of GAG storage, reduction of clusterin 1 staining, and improved elastin integrity. No significant changes in the spines of the treated animals were observed., Conclusions: PPS treatment led to reductions of pro-inflammatory cytokines and GAG storage in urine and tissues of MPS I dogs, which were most evident after subQ administration. SubQ administration also led to significant cytokine reductions in the CSF. Both treatment groups exhibited markedly reduced carotid and aortic inflammation, increased vessel integrity, and improved histopathology. We conclude that PPS may be a safe and useful therapy for MPS I, either as an adjunct or as a stand-alone treatment that reduces inflammation and GAG storage.

Mucopolysaccharidosis type I (MPS I) is a rare autosomal recessive disease caused by alpha-L-iduronidase deficiency in which heparan and dermatan sulfate degradation is compromised. Besides primary lysosomal glycosaminoglycan accumulation, further changes in cellular functions have also been described in several murine MPS models. Herein, we evaluated alterations in hematopoiesis and its implications on the production of mature progeny in a MPS I murine model. Despite the significant increase in hematopoietic stem cells, a reduction in common myeloid progenitors and granulocyte-macrophage progenitor cells was observed in Idua -/- mice bone marrow. Furthermore, no alterations in number, viability nor activation of cell death mechanisms were observed in Idua -/- mice mature macrophages but they presented higher sensitivity to apoptotic induction after staurosporine treatment. In addition, changes in Ca(2+) signaling and a reduction in phagocytosis ability were also found. In summary, our results revealed significant intracellular changes in mature Idua -/- macrophages related to alterations in Idua -/- mice hematopoiesis, revealing a disruption in cell homeostasis. These results provide new insights into physiopathology of MPS I., (© 2015 Wiley Periodicals, Inc.)

Mucopolysaccharidosis type I (MPS I) is an autosomal disease caused by alpha-L-iduronidase deficiency. This study proposed the use of cationic nanoemulsions as non-viral vectors for a plasmid (pIDUA) containing the gene that codes for alpha-L-iduronidase. Nanoemulsions composed of medium chain triglycerides (MCT)/1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE)/1,2-dioleoyl-sn-glycero-3-trimethylammonium propane (DOTAP)/1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[amino(polyethylene glycol)-2000] (DSPE-PEG) were prepared by high pressure homogenization. Formulations were prepared by the adsorption or encapsulation of preformed pIDUA-DOTAP complexes into the oil core of nanoemulsions at different charge ratios. pIDUA complexed was protected from enzymatic degradation by DNase I. The physicochemical characteristics of complexes in protein-containing medium were mainly influenced by the presence of DSPE-PEG. Bragg reflections corresponding to a lamellar organization were identified for blank formulations by energy dispersive X-ray diffraction, which could not be detected after pIDUA complexation. The intravenous injection of these formulations in MPS I knockout mice led to a significant increase in IDUA activity (fluorescence assay) and expression (RT-qPCR) in different organs, especially the lungs and liver. These findings were more significant for formulations prepared at higher charge ratios (+4/-), suggesting a correlation between charge ratio and transfection efficiency. The present preclinical results demonstrated that these nanocomplexes represent a potential therapeutic option for the treatment of MPS I., (Copyright © 2015. Published by Elsevier B.V.)

Background: The lysosomal storage disorder, mucopolysaccharidosis I (MPS I), commonly manifests with upper airway obstruction and sleep disordered breathing (SDB). The success of current therapies, including haematopoietic stem cell transplantation (HSCT) and enzyme replacement therapy (ERT) may be influenced by a number of factors and monitored using biomarkers of metabolic correction. We describe the pattern of SDB seen in the largest MPS I cohort described to date and determine therapies and biomarkers influencing the severity of long-term airway disease., Methods: Therapeutic, clinical and biomarker data, including longitudinal outcome parameters from 150 sleep oximetry studies were collected in 61 MPS I (44 Hurler, 17 attenuated) patients between 6 months pre to 16 years post-treatment (median follow-up 22 months). The presence and functional nature of an immune response to ERT was determined using ELISA and a cellular uptake inhibition assay. Multivariate analysis was performed to determine significant correlators of airway disease., Results: The incidence of SDB in our cohort is 68%, while 16% require therapeutic intervention for airway obstruction. A greater rate of progression (73%) and requirement for intervention is seen amongst ERT patients in contrast to HSCT treated individuals (24%). Multivariate analysis identifies poorer metabolic clearance, as measured by a rise in the biomarker urinary dermatan sulphate: chondroitin sulphate (DS:CS) ratio, as a significant correlator of increased presence and severity of SDB in MPS I patients (p = 0.0017, 0.008). Amongst transplanted Hurler patients, delivered enzyme (leukocyte iduronidase) at one year is significantly raised in those without SDB (p = 0.004). Cellular uptake inhibitory antibodies in ERT treated patients correlate with reduced substrate clearance and occurrence of severe SDB (p = 0.001)., Conclusion: We have identified biochemical and therapeutic factors modifying airway disease across the phenotypic spectrum in MPS I. Interventions maximising substrate reduction correlate with improved long-term SDB, while inhibitory antibodies impact on biochemical and clinical outcomes. Monitoring and tolerisation strategies should be re-evaluated to improve detection and minimise the inhibitory antibody response to ERT in MPS I and other lysosomal storage diseases. Future studies should consider the use of sleep disordered breathing as an objective parameter of clinical and metabolic improvement.

Mucopolysaccharidosis type I (MPS I) is due to deficient alpha-L-iduronidase (IDUA) which leads to storage of undegraded glycosaminoglycans (GAG). The severe form of the disease is characterized by mental retardation of unknown etiology. Trying to unveil the mechanisms that lead to cognitive impairment in MPS I, we studied alterations in the proteome from MPS I mouse hippocampus. Eight-month old mice presented increased LAMP-1 expression, GAG storage in neurons and glial cells, and impaired aversive and non-aversive memory. Shotgun proteomics was performed and 297 proteins were identified. Of those, 32 were differentially expressed. We found elevation in proteins such as cathepsins B and D; however their increase did not lead to cell death in MPS I brains. Glial fibrillary acid protein (GFAP) was markedly elevated, and immunohistochemistry confirmed a neuroinflammatory process that could be responsible for neuronal dysfunction. We didn't observe any differences in ubiquitin expression, as well as in other proteins related to protein folding, suggesting that the ubiquitin system is working properly. Finally, we observed alterations in several proteins involved in synaptic plasticity, including overexpression of post synaptic density-95 (PSD95) and reduction of microtubule-associated proteins 1A and 1B. These results together suggest that the cognitive impairment in MPS I mice is not due to massive cell death, but rather to neuronal dysfunction caused by multiple processes, including neuroinflammation and alterations in synaptic plasticity., (Copyright © 2015. Published by Elsevier Inc.)

Progressive skeletal and connective tissue disease represents a significant clinical burden in all of the mucopolysaccharidoses. Despite the introduction of enzyme replacement strategies for many of the mucopolysaccharidoses, symptomatology related to bone and joint disease appears to be recalcitrant to current therapies. In order to address these unmet medical needs a clearer understanding of skeletal and connective tissue disease pathogenesis is required. Historically the pathogenesis of the mucopolysaccharidoses has been assumed to directly relate to progressive storage of glycosaminoglycans. It is now apparent for many lysosomal storage disorders that more complex pathogenic mechanisms underlie patients' clinical symptoms. We have used proteomic and genome wide expression studies in the murine mucopolysaccharidosis I model to identify early pathogenic events occurring in micro-dissected growth plate tissue. Studies were conducted using 3 and 5-week-old mice thus representing a time at which no obvious morphological changes of bone or joints have taken place. An unbiased iTRAQ differential proteomic approach was used to identify candidates followed by validation with multiple reaction monitoring mass spectrometry and immunohistochemistry. These studies reveal significant decreases in six key structural and signaling extracellular matrix proteins; biglycan, fibromodulin, PRELP, type I collagen, lactotransferrin, and SERPINF1. Genome-wide expression studies in embryonic day 13.5 limb cartilage and 5 week growth plate cartilage followed by specific gene candidate qPCR studies in the 5week growth plate identified fourteen significantly deregulated mRNAs (Adamts12, Aspn, Chad, Col2a1, Col9a1, Hapln4, Lum, Matn1, Mmp3, Ogn, Omd, P4ha2, Prelp, and Rab32). The involvement of biglycan, PRELP and fibromodulin; all members of the small leucine repeat proteoglycan family is intriguing, as this protein family is implicated in the pathogenesis of late onset osteoarthritis. Taken as a whole, our data indicates that alteration of the extracellular matrix represents a very early event in the pathogenesis of the mucopolysaccharidoses and implies that biomechanical failure of chondro-osseous tissue may underlie progressive bone and joint disease symptoms. These findings have important therapeutic implications., (Copyright © 2014 Elsevier Inc. All rights reserved.)

The authors describe the first mother-infant pair to complete an on-going, prospective, open-label, Phase 4 trial (ALIU) UU3, NCT00418821) determining the safety of laronidase enzyme replacement therapy (ERT) in pregnant women with mucopolysaccharidosis type I (MPS I) and their breastfed infants. The mother, a 32-year-old with attenuated MPS I (Scheie syndrome), received laronidase for three years and continued treatment throughout her second pregnancy and while lactating. A healthy 2.5 kg male was delivered by elective cesarean section at 37 weeks. He was breastfed for three months. No laronidase was detected in breast milk. The infant never developed anti-laronidase IgM antibodies, never had inhibitory antibody activity in a cellular uptake assay, and always had normal urinary glycosaminoglycan (GAG) levels. No drug-related adverse events were reported. At 2.5 years of age, the boy is healthy with normal growth and development. In this first prospectively monitored mother-infant pair, laronidase during pregnancy and breastfeeding was uneventful.

Background: Mucopolysaccharidose type I (MPSI) is a lysosomal monogenic disease caused by mutations in the gene for α- L-iduronidase (IDUA). MPSI patients need a constant supply of IDUA to alleviate progression of the disease. IDUA gene transfer using integrative vectors might provide a definitive solution and support advancement to clinical trials, although studies have not yet been satisfactory. To achieve a stable IDUA gene expression in vivo, phiC31 was tested in the present study., Methods: Several plasmid vectors were constructed and IDUA-/- mice were treated with cyclophosphamide and transfected with these vectors hydrodynamically via tail veins. IDUA expression was monitored over time. Treated and nontreated mice underwent an open-field test at age 8 months, and IDUA activity and glycosaminoglycan (GAG) content of tissues were evaluated., Results: High levels of IDUA activity were detected initially (>1000 U/ml), although these levels decayed over time. The reinjection of vectors produced a similar profile of IDUA decay. Three out of six treated mice had IDUA activity in the livers, and also showed lower GAG content, reduced lysosomes and better locomotion. To investigate unsustained IDUA production, wild-type mice were submitted to the same gene therapy procedure, which generated a similar profile of IDUA decay. Anti-IDUA antibody was detected in the sera of these animals. In addition, we also found three methylated sites in the cytomegalovirus promoter region., Conclusions: phiC31-mediated gene therapy resulted in an important improvement in IDUA-/- mice, including locomotion, although the obstacles that need to be overcome to enable long-term gene therapy for MPSI are also noted., (Copyright © 2015 John Wiley & Sons, Ltd.)