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Lysosomal storage disorders (LSDs) are predominantly enzyme deficiencies leading to substrate accumulation, causing progressive damage to multiple organs. To date, a crucial part of diagnosing LSDs is measuring enzymatic activity in leucocytes, plasma, or dried blood spots (DBS). Here, we present results from a proof-of-principle study, evaluating an innovative digital microfluidics (DMF) platform, referred to as SEEKER®, that can measure the activity of the following four lysosomal enzymes from DBS: α-L-iduronidase (IDUA) for mucopolysaccharidosis I (MPS I), acid α-glucosidase (GAA) for Pompe disease, ß-glucosidase (GBA) for Gaucher disease, and α-galactosidase A (GLA) for Fabry disease. Over 900 DBS were analysed from newborns, children, and adults. DMF successfully detected known patients with MPS I, Pompe disease, and Gaucher disease, and known males with Fabry disease. This is the first demonstration of this multiplexed DMF platform for identification of patients with LSDs in a specialised diagnostic enzyme laboratory environment. We conclude that this DMF platform is relatively simple, high-throughput, and could be readily accommodated into a specialised laboratory as a first-tier test for MPS I, Pompe disease, and Gaucher disease for all patients, and Fabry disease for male patients only.

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Hurler syndrome, a type of Mucopolysaccharidosis type I, is an inherited disorder caused by the accumulation of glycosaminoglycans (GAG) due to a deficiency in lysosomal α-L-iduronidase (IDUA), resulting in multiorgan dysfunction. In many patients with Hurler syndrome, IDUA proteins are not produced due to nonsense mutations in their genes; therefore, readthrough-inducing compounds, such as gentamycin, are expected to restore IDUA proteins by skipping the premature termination codon. In the present study, we synthesized a series of chromenopyridine derivatives to identify novel readthrough-inducing compounds. The readthrough-inducing activities of synthesized compounds were examined by measuring cellular IDUA activities and GAG concentrations in Hurler syndrome patient-derived cells. Compounds with a difluorophenyl group at the 2-position of chromenopyridine, a cyclobutyl group at the 3-position, and a basic side chain or basic fused ring exhibited excellent readthrough-inducing activities. KY-640, a chromenopyridine derivative with a tetrahydroisoquinoline sub-structure, increased the cellular IDUA activities of patient-derived cells by 3.2-fold at 0.3 µM and significantly reduced GAG concentrations, and also significantly increased enzyme activity in mouse models, suggesting its therapeutic potential in patients with Hurler syndrome.

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The readthrough mechanism, which skips the premature termination codon and restores the biosynthesis of the defective enzyme, is an emerging therapeutic tactic for nonsense mutation-related diseases, such as Hurler syndrome, a type of mucopolysaccharidosis. In the present study, novel triaryl derivatives were synthesized and their readthrough-inducing activities were evaluated by a luciferase reporter assay with a partial α-L-iduronidase (IDUA) DNA sequence containing the Q70X nonsense mutation found in Hurler syndrome and by measuring the enzyme activity of IDUA knockout cells transfected with the mutant IDUA gene. KY-516, a representative compound in which the meta position carboxyl group of the left ring of the clinically used ataluren was converted to the para position sulfamoylamino group, the central ring to triazole, and the right ring to cyanobenzene, exhibited the most potent readthrough-inducing activity in the Q70X/luciferase reporter assay. In Q70X mutant IDUA transgenic cells, KY-516 significantly increased enzyme activity at 0.1 µM. After the oral administration of KY-516 (10 mg/kg), the highest plasma concentration of KY-516 was above 5 µM in rats. These results indicate that KY-516, a novel triaryl derivative, exhibits potent readthrough-inducing activity and has potential as a therapeutic agent for Hurler syndrome.

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Mucopolysaccharidosis I (MPS I), a lysosomal storage disease caused by dysfunction of α-L-iduronidase (IDUA), is characterized by the deposition of dermatan sulfate (DS) and heparan sulfate (HS) throughout the body, which causes several somatic and central nervous symptoms. Although enzyme-replacement therapy (ERT) is currently available to treat MPS I, it does not alleviate central nervous disorders, as it cannot penetrate the blood-brain barrier. Here we evaluate the brain delivery, efficacy, and safety of JR-171, a fusion protein comprising humanized anti-human transferrin receptor antibody Fab and IDUA, using monkeys and MPS I mice. Intravenously administered JR-171 was distributed in major organs, including the brain, and reduced DS and HS concentrations in the central nervous system and peripheral tissues. JR-171 exerted similar effects on peripheral disorders similar to conventional ERT and further reversed brain pathology in MPS I mice. We found that JR-171 improved spatial learning ability, which was seen to deteriorate in the vehicle-treated mice. Further, no safety concerns were noted in repeat-dose toxicity studies in monkeys. This study provides nonclinical evidence that JR-171 might potentially prevent and even improve disease conditions in patients with neuronopathic MPS I without serious safety concerns.

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INTRODUCTION: Mucopolysaccharidosis type I (MPS I) is a lysosomal storage disease present in 1:100,000 newborns. Variants in the IDUA (alpha-L-iduronidase) gene decrease the enzyme activity for glycosaminoglycans metabolism. MPS I patients exhibit clinical manifestations that fall on the Hurler, Hurler-Scheie, and Scheie syndrome spectrum. CASE PRESENTATION: We present a male Mexican patient with respiratory exacerbations requiring recurrent hospitalizations. He showed macrocephaly, coarse facies, hepatomegaly, umbilical hernia, and dorsal kyphosis. The sequencing of the IDUA gene revealed the following genotype: c.46_57del12/c.1205G>A. He received combined therapy with hematopoietic stem cell transplantation and enzyme replacement. Mexican case reports were analyzed to estimate the prevalence of the associated genetic variants. CONCLUSION: Despite the challenges of managing this rare disease in Mexico, our patient benefited from the combined therapy. The discrete clinical manifestations and prompt evaluation by a geneticist were crucial in establishing a diagnosis, enabling an early intervention by a multidisciplinary team. The combination of ERT before and after HSCT provided health benefits to our patient.

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Abstract Mucopolysaccharidosis (MPS) is a group of metabolic disorders caused by the deficiency or complete absence of certain lysosomal enzymes responsible for the breakdown of mucopolysaccharides, causing an accumulation of glycosaminoglycans (GAGs) throughout the body. Mucopolysaccharidosis type I (MPS I), also called Hurler syndrome, is an autosomal recessive lysosomal storage disorder resulting from a deficiency of the enzyme α-L-iduronidase. This report aims to present the clinical findings and diagnosis of a 21-month-old female with no history of similar cases in their previous generations. The diagnosis was considered based on the clinical and radiological characteristics of Hurler syndrome (HS) and confirmed biochemically, becoming the first confirmed case in the Dominican Republic.

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Mucopolysaccharidosis type I (MPS I) is an autosomal-recessive metabolic disorder caused by an enzyme deficiency of lysosomal alpha-l-iduronidase (IDUA). Haematopoietic stem cell transplantation (HSCT) is the therapeutic option of choice in MPS I patients younger than 2.5 years, which has a positive impact on neurocognitive development. However, impaired growth remains a problem. In this monocentric study, 14 patients with MPS I (mean age 1.72 years, range 0.81-3.08) were monitored according to a standardised follow-up program after successful allogeneic HSCT. A detailed anthropometric program was carried out to identify growth patterns and to determine predictors of growth in these children. All patients are alive and in outpatient care (mean follow-up 8.1 years, range 0.1-16.0). Progressively lower standard deviation scores (SDS) were observed for body length (mean SDS -1.61; -4.58 - 3.29), weight (-0.56; -3.19 - 2.95), sitting height (-3.28; -7.37 - 0.26), leg length (-1.64; -3.88 - 1.49) and head circumference (0.91; -2.52 - 6.09). Already at the age of 24 months, significant disproportions were detected being associated with increasing deterioration in growth for age. Younger age at HSCT, lower counts for haemoglobin and platelets, lower potassium, higher donor-derived chimerism, higher counts for leukocytes and recruitment of a matched unrelated donor (MUD) positively correlated with body length (p ≤ 0.05). In conclusion, this study characterised predictors and aspects of growth patterns in children with MPS I after HSCT, underlining that early HSCT of MUD is essential for slowing body disproportion.

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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.

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Mucopolysaccharidosis type I (MPS I) is a rare monogenic disease in which glycosaminoglycans' abnormal metabolism leads to the storage of heparan sulfate and dermatan sulfate in various tissues. It causes its damage and impairment. Patients with the severe form of MPS I usually do not live up to the age of ten. Currently, the therapy is based on multidisciplinary care and enzyme replacement therapy or hematopoietic stem cell transplantation. Applying gene therapy might benefit the MPS I patients because it overcomes the typical limitations of standard treatments. Nanoparticles, including nanoemulsions, are used more and more in medicine to deliver a particular drug to the target cells. It allows for creating a specific, efficient therapy method in MPS I and other lysosomal storage disorders. This article briefly presents the basics of nanoemulsions and discusses the current state of knowledge about their usage in mucopolysaccharidosis type I.

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Mucopolysaccharidosis (MPS) I is a lysosomal storage disease characterized by deficient activity of the enzyme alpha-L-iduronidase, leading to abnormal accumulation of heparan and dermatan sulfate glycosaminoglycans in cells and tissues. Patients commonly exhibit progressive skeletal abnormalities, in part due to failures of endochondral ossification during postnatal growth. Previously, using the naturally-occurring canine model, we showed that bone and cartilage cells in MPS I exhibit elevated lysosomal storage from an early age and that animals subsequently exhibit significantly diminished vertebral trabecular bone formation. Wnts are critical regulators of endochondral ossification that depend on glycosaminoglycans for signaling. The objective of this study was to examine whether lithium, a glycogen synthase kinase-3 inhibitor and stimulator of Wnt/beta-catenin signaling, administered during postnatal growth could attenuate progression of vertebral trabecular bone disease in MPS I. MPS I dogs were treated orally with therapeutic levels of lithium carbonate from 14 days to 6 months-of-age. Untreated heterozygous and MPS I dogs served as controls. Serum was collected at 3 and 6 months for assessment of bone turnover markers. At the study end point, thoracic vertebrae were excised and assessed using microcomputed tomography and histology. Lithium-treated animals exhibited significantly improved trabecular spacing, number and connectivity density, and serum bone-specific alkaline phosphatase levels compared to untreated animals. Growth plates from lithium-treated animals exhibited increased numbers of hypertrophic chondrocytes relative to both untreated MPS I and heterozygous animals. These findings suggest that bone and cartilage cells in MPS I are still capable of responding to exogenous osteogenic signals even in the presence of significant lysosomal storage, and that targeted osteogenic therapies may represent a promising approach for attenuating bone disease progression in MPS I.

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Introducción: el síndrome de Hurler es la forma más grave de la mucopolisacaridosis I. El depósito de heparán y dermatán sulfato en las meninges favorece el desarrollo de hidrocefalia y, a su vez, de meningoencefaloceles. Caso clínico: se describe el caso de una paciente de 23 años con este síndrome y un encefalocele nasosinusal intervenido mediante cirugía endoscópica nasosinusal y posterior refuerzo mediante un colgajo pericraneal. Discusión: el abordaje endoscópico de los meningoencefaloceles nasales ha crecido notablemente en los últimos años debido a una tasa de éxito elevada tanto para la eliminación de dicha lesión, como para el adecuado control de la fístula de líquido cefalorraquídeo consecuente. Conclusiones: actualmente, la cirugía endoscópica nasosinusal es una herramienta muy útil para el tratamiento de patologías de base de cráneo y reduce la morbilidad causada por el abordaje transcraneal.

Introduction: Hurler syndrome is the most severe version of mucopolysaccharidosis I. The storage of dermatan and heparin sulfate in meninges allows the development of hydrocephalus and meningoencephaloceles. Case report: We report a 23-year-old female with this syndrome and a sinonasal encephalocele operated by endoscopic sinonasal surgery and subsequent pericranial flap as support. Discussion: Endoscopic sinonasal surgery has grown in last years in relation with treatment of sinonasal meningoencephaloceles due to a high rate of success removing the lesion and closing the subsequent cerebrospinal fluid fistula. Conclusion: Currently, the endoscopic sinonasal surgery has become a useful tool in the management of skull base pathologies, and reduces the morbidity due to a transcranial approach.

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The impact of hematopoietic stem cell transplantation (HSCT) on growth in patients diagnosed with mucopolysaccharidosis I Hurler (MPS-IH) has been historically regarded as unsatisfactory. Nevertheless, the growth patterns recorded in transplanted patients have always been compared to those of healthy children. The objective of this study was to verify the impact of HSCT on MPS-IH long term growth achievements. The auxological data of 15 patients were assessed longitudinally and compared both to the WHO growth centiles for healthy individuals and to recently published curves of untreated MPS-IH children. Despite a progressive decrease after HSCT when estimated with reference to the WHO growth charts, median height SDS showed a progressive and statistically significant increase when comparing the stature recorded at each timepoint in our population to the curves of untreated MPS-IH individuals (from -0.39 SDS at t0 to +1.35 SDS 5 years after HSCT, p value < 0.001 and to +3.67 SDS at the age of 9 years, p value < 0.0001). In conclusion, though not efficient enough to restore a normal growth pattern in MPS-IH patients, we hereby demonstrate that HSCT positively affects growth and provides transplanted patients with a remarkable height gain compared to untreated gender- and age- matched individuals.

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Mucopolysaccharidosis (MPS) type I is a rare lysosomal storage disorder caused by an accumulation of glycosaminoglycans (GAGs) resulting in multisystem disease. Neurological morbidity includes hydrocephalus, spinal cord compression, and cognitive decline. While many neurological symptoms have been described, stroke is not a widely-recognized manifestation of MPS I. Accordingly, patients with MPS I are not routinely evaluated for stroke, and there are no guidelines for managing stroke in patients with this disease. We report the case of a child diagnosed with MPS I who presented with overt stroke and repeated neurological symptoms with imaging findings for severe ventriculomegaly, infarction, and bilateral terminal carotid artery stenosis. Direct intracranial pressure evaluation proved negative for hydrocephalus. The patient was subsequently treated with cerebral revascularization and at a 3-year follow-up, the patient reported no further neurological events or new ischemia on cerebral imaging. Cerebral arteriopathy in patients with MPS I may be associated with GAG accumulation within the cerebrovascular system and may predispose patients to recurrent strokes. However, further studies are required to elucidate the etiology of cerebrovascular arteriopathy in the setting of MPS I. Although the natural history of steno-occlusive arteriopathy in patients with MPS I remains unclear, our findings suggest that cerebral revascularization is a safe treatment option that may mitigate the risk of future strokes and should be strongly considered within the overall management guidelines for patients with MPS I.

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Mutations in the IDUA gene cause deficiency of the lysosomal enzyme alpha-l-iduronidase (IDUA), which leads to a rare disease known as mucopolysaccharidosis type I. More than 300 pathogenic variants of the IDUA gene have been reported to date, but not much is known about the distribution of mutations in different populations and ethnic groups due to the low prevalence of the disease. This article presents the results of a molecular genetic study of 206 patients with mucopolysaccharidosis type I (MPS I) from the Russian Federation (RF) and other republics of the former Soviet Union. Among them, there were 173 Russian (Slavic) patients, 9 Tatars, and 24 patients of different nationalities from other republics of the former Soviet Union. Seventy-three different pathogenic variants in the IDUA gene were identified. The common variant NM_000203.5:c.208C>T was the most prevalent mutant allele among Russian and Tatar patients. The common variant NM_000203.5:c.1205G>A accounted for only 5.8% mutant alleles in Russian patients. Both mutations were very rare or absent in patients from other populations. The pathogenic variant NM_000203.5:c.187C>T was the major allele in patients of Turkic origin (Altaian, Uzbeks, and Kyrgyz). Specific own pathogenic alleles in the IDUA gene were identified in each of these ethnic groups. The identified features are important for understanding the molecular origin of the disease, predicting the risk of its development and creating optimal diagnostic and treatment tools for specific regions and ethnic groups.

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Mucopolysaccharidosis (MPS) syndrome is an inherited metabolic disorder. In more than half of the patients with MPS syndrome, heart valve involvement is reported; however, combined aortic and mitral valve stenosis in MPS syndrome type I-S is very rare. We describe a 39-year-old man with severe mitral and aortic valve stenosis due to MPS syndrome type I-S. Transthoracic and transesophageal echocardiography revealed severe thickening and calcification in the aortic and mitral valves with severe left ventricular hypertrophy. The coronary arteries were normal in angiography.

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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.

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ABSTRACT Objective: To report the stabilization of urinary glycosaminoglicans (GAG) excretion and clinical improvements in patients with mucopolysaccharidosis type I (MPS I) under an alternative dose regimen of laronidase of 1.2 mg/kg every other week. Methods: We participated in a dose-optimization trial for laronidase in MPS-I patients using four alternative regimens: 0.58 mg/kg every week, 1.2 mg/kg every two weeks, 1.2 mg/kg every week and 1.8 mg/kg every other week (EOW). After the trial ended, the patients resumed the recommended dose and regimen of 0.58 mg/kg every week. Under this regimen, some patients presented difficulties in venous access and were unable to commute weekly to the treatment center. Therefore, we used an alternative regimen that consisted of 1.2 mg/kg EOW in eight patients. A retrospective study of medical records of MPS-I patients who underwent both enzyme replacement therapy (ERT) regimens, of 0.58 mg/kg every week and 1.2 mg/kg EOW, was done. Results: Patients remained clinically stable under the alternative regimen, did not present elevation of urinary GAG nor any adverse event. Conclusions: The switch of dose regimen to 1.2 mg/kg EOW of laronidase was safe, and did not cause any clinical worsening in patients who had been previously under standard dose ERT.

RESUMO Objetivo: Descrever a manutenção dos níveis de glicosaminoglicano (GAG) excretados na urina e da estabilização clínica em pacientes com mucopolissacaridose do tipo I (MPS I) com o uso da laronidase num regime de dose alternativo de 1,2 mg/kg a cada duas semanas. Método: Alguns pacientes do nosso serviço participaram de um estudo de otimização de dose da laronidase para o tratamento da MPS I no qual foram comparados quatro esquemas terapêuticos: 0,58 mg/kg/semana, 1,2 mg/kg a cada duas semanas, 1,2 mg/kg/semana e 1,8 mg/kg a cada duas semanas. Após o término do estudo, todos os pacientes passaram a receber a terapia de reposição enzimática (TRE) na dose padrão de bula, que é de 0,58 mg/kg/semana, e nesse regime alguns pais se queixaram da dificuldade em comparecer ao centro todas as semanas, além da dificuldade de se obter acesso para punção venosa. Com base nessas queixas, oito pacientes passaram a receber a TRE no regime alternativo de 1,2 mg/kg a cada duas semanas. Foi feito o estudo retrospectivo de dados de prontuário de pacientes com MPS I que fizeram TRE com laronidase nas doses 0,58 mg/kg/semana e 1,2 mg/kg a cada duas semanas. Resultados: Os pacientes mantiveram-se clinicamente estáveis, não apresentaram aumento dos níveis de GAG urinários nem eventos adversos durante o regime alternativo de dose. Conclusões: A mudança para o esquema de 1,2 mg/kg de laronidase a cada duas semanas foi segura e não acarretou piora clínica nos pacientes que já estavam em TRE na dose padrão.

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Although disease progression in mucopolysaccharidosis type I (MPS-I) can be attenuated by hematopoietic cell transplantation (HCT), it is increasingly recognized that residual disease is substantial. Biomarkers that would allow us to evaluate the efficacy of HCT (and upcoming new therapies) in nonhematologic tissues are needed. Current biomarkers, including the iduronidase (IDUA) activity in leukocytes, are not suitable for this purpose because they are assessed in tissues of hematologic origin and may not reflect enzyme availability in nonhematologic tissues. Saliva is a nonhematologic body fluid that can be collected easily and noninvasively. We hypothesized that the extent of recovery of IDUA activity in saliva after HCT could provide a better understanding of the penetration of donor-derived enzyme into nonhematologic compartments. This study in 20 patients with MPS-I shows that the measurement of IDUA activity in saliva is possible and allows diagnosis of IDUA deficiency (P < .0001), with values a magnitude further deviating from the normal range than when assayed in corresponding dried blood spots (DBSs). Furthermore, it could possibly differentiate between phenotypes (P = .045). More importantly, patients exhibit strikingly low values of IDUA in saliva after HCT, far below the normal range of control subjects (P = .013), contrasting the normal IDUA levels in DBSs. We postulate that the limited recovery of donor-derived IDUA activity in saliva after treatment reflects the situation in poorly responding nonhematologic tissue compartments, unveiling enzyme delivery as a weak spot of the current therapy. Salivary IDUA activity could be used as a biomarker for the evaluation of the effect of new therapies in well-vascularized nonhematologic tissues.

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KEY MESSAGE: Arabidopsis N-glycan processing mutants provide the basis for tailoring recombinant enzymes for use as replacement therapeutics to treat lysosomal storage diseases, including N-glycan mannose phosphorylation to ensure lysosomal trafficking and efficacy. Functional recombinant human alpha-L-iduronidase (IDUA; EC 3.2.1.76) enzymes were generated in seeds of the Arabidopsis thaliana complex-glycan-deficient (cgl) C5 background, which is deficient in the activity of N-acetylglucosaminyl transferase I, and in seeds of the Arabidopsis gm1 mutant, which lacks Golgi α-mannosidase I (GM1) activity. Both strategies effectively prevented N-glycan maturation and the resultant N-glycan structures on the consensus sites for N-glycosylation of the human enzyme revealed high-mannose N-glycans of predominantly Man5 (cgl-IDUA) or Man6-8 (gm1-IDUA) structures. Both forms of IDUA were equivalent with respect to their kinetic parameters characterized by cleavage of the artificial substrate 4-methylumbelliferyl-iduronide. Because recombinant lysosomal enzymes produced in plants require the addition of mannose-6-phosphate (M6P) in order to be suitable for lysosomal delivery in human cells, we characterized the two IDUA proteins for their amenability to downstream in vitro mannose phosphorylation mediated by a soluble form of the human phosphotransferase (UDP-GlcNAc: lysosomal enzyme N-acetylglucosamine [GlcNAc]-1-phosphotransferase). Gm1-IDUA exhibited a slight advantage over the cgl-IDUA in the in vitro M6P-tagging process, with respect to having a better affinity (i.e. lower K m) for the soluble phosphotransferase. This may be due to the greater number of mannose residues comprising the high-mannose N-glycans of gm1-IDUA. Our elite cgl- line produces IDUA at > 5.7% TSP (total soluble protein); screening of the gm1 lines showed a maximum yield of 1.5% TSP. Overall our findings demonstrate the relative advantages and disadvantages associated with the two platforms to create enzyme replacement therapeutics for lysosomal storage diseases.

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