Hematopoietic Stem- and Progenitor-Cell Gene Therapy for Hurler Syndrome.

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.).

Mucopolysaccharidosis Type I: A Review of the Natural History and Molecular Pathology.

Mucopolysaccharidosis type I (MPS I) is a rare autosomal recessive inherited disease, caused by deficiency of the enzyme α-L-iduronidase, resulting in accumulation of the glycosaminoglycans (GAGs) dermatan and heparan sulfate in organs and tissues. If untreated, patients with the severe phenotype die within the first decade of life. Early diagnosis is crucial to prevent the development of fatal disease manifestations, prominently cardiac and respiratory disease, as well as cognitive impairment. However, the initial symptoms are nonspecific and impede early diagnosis. This review discusses common phenotypic manifestations in the order in which they develop. Similarities and differences in the three animal models for MPS I are highlighted. Earliest symptoms, which present during the first 6 months of life, include hernias, coarse facial features, recurrent rhinitis and/or upper airway obstructions in the absence of infection, and thoracolumbar kyphosis. During the next 6 months, loss of hearing, corneal clouding, and further musculoskeletal dysplasias develop. Finally, late manifestations including lower airway obstructions and cognitive decline emerge. Cardiac symptoms are common in MPS I and can develop in infancy. The underlying pathogenesis is in the intra- and extracellular accumulation of partially degraded GAGs and infiltration of cells with enlarged lysosomes causing tissue expansion and bone deformities. These interfere with the proper arrangement of collagen fibrils, disrupt nerve fibers, and cause devastating secondary pathophysiological cascades including inflammation, oxidative stress, and other disruptions to intracellular and extracellular homeostasis. A greater understanding of the natural history of MPS I will allow early diagnosis and timely management of the disease facilitating better treatment outcomes.

Morphologic description of male reproductive accessory glands in a mouse model of mucopolysaccharidosis type I (MPS I).

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.

Evidence that glycosaminoglycan storage and collagen deposition in the cauda epididymidis does not impair sperm viability in the Mucopolysaccharidosis type I mouse model.

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.

Elevated cerebral spinal fluid biomarkers in children with mucopolysaccharidosis I-H.

Mucopolysaccharidosis (MPS) type-IH is a lysosomal storage disease that results from mutations in the IDUA gene causing the accumulation of glycosaminoglycans (GAGs). Historically, children with the severe phenotype, MPS-IH (Hurler syndrome) develop progressive neurodegeneration with death in the first decade due to cardio-pulmonary complications. New data suggest that inflammation may play a role in MPS pathophysiology. To date there is almost no information on the pathophysiologic changes within the cerebral spinal fluid (CSF) of these patients. We evaluated the CSF of 25 consecutive patients with MPS-IH. While CSF glucose and total protein were within the normal range, we found a significantly mean elevated CSF opening pressure at 24 cm H2O (range 14-37 cm H2O). We observed a 3-fold elevation in CSF heparan sulfate and a 3-8 fold increase in MPS-IH specific non-reducing ends, I0S0 and I0S6. Cytokine analyses in CSF of children with MPS-IH showed significantly elevated inflammatory markers including: MCP-1 SDF-1a, IL-Ra, MIP-1b, IL-8, and VEGF in comparison to unaffected children. This is the largest report of CSF characteristics in children with MPS-IH. Identification of key biomarkers may provide further insight into the inflammatory-mediated mechanisms related to MPS diseases and perhaps lead to improved targeted therapies.

Neonatal tolerance induction enables accurate evaluation of gene therapy for MPS I in a canine model.

High fidelity animal models of human disease are essential for preclinical evaluation of novel gene and protein therapeutics. However, these studies can be complicated by exaggerated immune responses against the human transgene. Here we demonstrate that dogs with a genetic deficiency of the enzyme α-l-iduronidase (IDUA), a model of the lysosomal storage disease mucopolysaccharidosis type I (MPS I), can be rendered immunologically tolerant to human IDUA through neonatal exposure to the enzyme. Using MPS I dogs tolerized to human IDUA as neonates, we evaluated intrathecal delivery of an adeno-associated virus serotype 9 vector expressing human IDUA as a therapy for the central nervous system manifestations of MPS I. These studies established the efficacy of the human vector in the canine model, and allowed for estimation of the minimum effective dose, providing key information for the design of first-in-human trials. This approach can facilitate evaluation of human therapeutics in relevant animal models, and may also have clinical applications for the prevention of immune responses to gene and protein replacement therapies.

12 year follow up of enzyme-replacement therapy in two siblings with attenuated mucopolysaccharidosis I: the important role of early treatment.

BACKGROUND: Mucopolysaccharidosis type I is an autosomal recessive disorder caused by deficiency of α-L-iduronidase and characterized by a progressive course with multisystem involvement. Clinically, Mucopolysaccharidosis type I is classified into two forms: severe (Hurler syndrome), which presents in infancy and is characterized by rapid progressive neurological involvement and attenuated (Hurler/Scheie and Scheie syndromes), which presents with slower progression and absent to mild nervous system involvement. The specific treatment for attenuated Mucopolysaccharidosis type I consists of enzyme-replacement therapy with laronidase (human recombinant α-L-iduronidase, Aldurazyme). We present here the clinical and laboratory results in an 12-year-old patient affected by the attenuated form of Mucopolysaccharidosis type I treated by enzyme-replacement therapy from the age of 5 months, compared with his 17 year old affected sister, who started therapy at 5 years of age. CASE PRESENTATION: Clinical evaluation of these siblings shows that initiation of therapy prior of the onset of clinically detectable disease resulted in considerable improvement in outcome in the young sibling. After 12 years of enzyme-replacement therapy, facial appearance, linear growth rate, and liver and spleen volumes were normal; moreover, the degree of joint disease, vertebral, and cardiac valvular involvement were only minimal compared with those of his sister. CONCLUSION: This study demonstrates that early diagnosis and early initiation of enzyme-replacement therapy substantially modify the natural history of the attenuated form of Mucopolysaccharidosis type I.

Selective normalisation of regional brain bis(monoacylglycero)phosphate in the mucopolysaccharidosis 1 (Hurler) mouse.

Bis(monoacylglycero)phosphate (BMP) is a glycerophospholipid highly enriched in the lysosomal network and elevated in lysosomal diseases. To correct this elevation, BMP synthesis was manipulated by dietary fatty acid supplementation and the impact on subregional brain BMP and pathology assessed in the mouse model of mucopolysaccharidosis 1 (Hurler syndrome (HS)). There was widespread elevation of BMP in HS mice across all six sub-regions - brain stem, cortex, cerebellum, hippocampus, olfactory bulb and the sub-cortex - with 22:6/22:6 the most abundant species. Linoleic acid normalised total BMP in all regions except the cortex and cerebellum, although there were differences in fatty acid species; the major finding a decrease in 22:6- and a concomitant increase in 22:5-containing species. A battery of behaviour assessments showed that in the water cross maze both HS and wild type mice performed less well on the linoleic acid diet, and that both HS and wild type mice on the linoleic acid diet performed similarly and better in the exploratory open field test. This may be a consequence of differential subregional BMP composition in the brain. The effects of high fat and docosahexaenoic/eicosapentaenoic acid enriched diets were generally unremarkable. Although major pathologies were not completely abrogated, much of the neurobehavioural testing was confounded by skeletal pathology that did not resolve. This is the first detailed characterisation of subregional brain BMP species informing on the ability to manipulate this phospholipid in the brain, and as such, may hold promise as an adjunct therapy not only for HS but also for other lysosomal diseases.

Determination of frequencies of alleles, associated with the pseudodeficiency of lysosomal hydrolases, in population of Ukraine.

The pseudodeficiency of lysosomal hydrolases described as a significant reduction in enzyme activi­ty in vitro in clinically healthy individuals, can lead to diagnostic errors in the process of biochemical analysis of lysosomal storage disease in case of its combination with pathology of another origin. Pseudodeficiency is mostly caused by some non-pathogenic changes in the corresponding gene. These changes lead to the in vitro lability of the enzyme molecule, whereas in vivo the enzyme retains its functional activity. To assess the prevalence of the most common lysosomal hydrolases pseudodeficiency alleles in Ukraine, we have determined the frequency of alleles c.1055A>G and c.* 96A>G in the ARSA gene, substitutions c.739C>T (R247W) and c.745C>T (R249W) in the HEXA gene, c.1726G>A (G576S) and c.2065G>A (E689K) in the GAA gene, c.937G>T (D313Y) in the GLA1 gene and c.898G>A (A300T) in the IDUA gene in a group of 117 healthy individuals from different regions of the country and 14 heterozygous carriers of pathogenic mutations in the HEXA gene (parents of children with confirmed diagnosis of Tay-Sachs disease). The total frequency of haplotypes, associated with arylsulfatase A pseudodeficiency, in healthy people in Ukraine (c.1055G/c.*96G and c.1055G/c.*96A haplotypes) was 10.3%. The frequency of c.739C>T (R247W) allele, associated with hexo­saminidase A pseudodeficiency, among Tay-Sachs carriers from Ukraine was 7.1%. The total frequency of α-glucosidase pseudodeficiency haplotypes in healthy individuals in Ukraine (c.1726A/c.2065A and c.1726G/c.2065A haplotypes) was 2.6%. No person among examined individuals with the substitution c.937G>T (D313Y) in the GLA1 gene and c.898G>A (A300T) in the IDUA gene was found. The differential diagnostics of lysosomal storage diseases requires obligatory determination of the presence of the pseudodeficiency alleles, particularly the ones with high incidence in the total population. Ignoring phenomenon of pseudodeficiency may lead to serious diagnostic errors.

Impaired bone remodeling and its correction by combination therapy in a mouse model of mucopolysaccharidosis-I.

Mucopolysaccharidosis-I (MPS-I) is a lysosomal storage disease (LSD) caused by inactivating mutations of IDUA, encoding the glycosaminoglycan-degrading enzyme α-l-iduronidase. Although MPS-I is associated with skeletal abnormalities, the impact of IDUA deficiency on bone remodeling is poorly defined. Here we report that Idua-deficient mice progressively develop a high bone mass phenotype with pathological lysosomal storage in cells of the osteoblast lineage. Histomorphometric quantification identified shortening of bone-forming units and reduced osteoclast numbers per bone surface. This phenotype was not transferable into wild-type mice by bone marrow transplantation (BMT). In contrast, the high bone mass phenotype of Idua-deficient mice was prevented by BMT from wild-type donors. At the cellular level, BMT did not only normalize defects of Idua-deficient osteoblasts and osteocytes but additionally caused increased osteoclastogenesis. Based on clinical observations in an individual with MPS-I, previously subjected to BMT and enzyme replacement therapy (ERT), we treated Idua-deficient mice accordingly and found that combining both treatments normalized all histomorphometric parameters of bone remodeling. Our results demonstrate that BMT and ERT profoundly affect skeletal remodeling of Idua-deficient mice, thereby suggesting that individuals with MPS-I should be monitored for their bone remodeling status, before and after treatment, to avoid long-term skeletal complications.

Decreased performance in IDUA knockout mouse mimic limitations of joint function and locomotion in patients with Hurler syndrome.

BACKGROUND: Mucopolysaccharidosis type I (MPS I) is caused by the deficiency of alpha-L-iduronidase (IDUA), which is involved in the degradation of glycosaminoglycans (GAGs), such as heparan sulfate and dermatan sulfate in the lysosome. It has been reported that joint symptoms are almost universal in MPS I patients, and even in the case of attenuated disease, they are the first symptom that brings a child to medical attention. However, functional tests and biological markers have not been published for the evaluation of the limitations in joint and locomotion in animal model-mimicking MPS. METHODS: We generated IDUA knockout (KO) mice to observe whether they present impairment of joint function. KO mice were characterized phenotypically and tested dual-energy X-ray absorptiometry analysis (DEXA), open-field, rotarod, and grip strength. RESULTS: The IDUA KO mice, generated by disruption between exon 6 and exon 9, exhibited clinical and laboratory findings, such as high urinary GAGs excretion, GAGs accumulation in various tissues, and significantly increased bone mineral density (BMD) in both female and male mice in the DEXA of the femur and whole bone. Remarkably, we observed a decrease in grasp function, decreased performance in the rotarod test, and hypo-activity in the open-field test, which mimic the limitations of joint mobility and decreased motor performance in the 6-min walk test in patients with MPS I. CONCLUSIONS: We generated a new IDUA KO mouse, tested open field, rotarod and grip strength and demonstrated decrease in grip strength, decreased performance and hypo-activity, which may be useful for investigating therapeutic approaches, and studying the pathogenesis of joint and locomotion symptoms in MPS I.

Toward reducing immunogenicity of enzyme replacement therapy: altering the specificity of human ß-glucuronidase to compensate for α-iduronidase deficiency.

Enzyme replacement therapy (ERT) is an effective treatment for many patients with lysosomal storage disorders caused by deficiency in enzymes involved in cell metabolism. However, immune responses that develop against the administered enzyme in some patients can hinder therapeutic efficacy and cause serious side effects. Here we investigated the feasibility of a general approach to decrease ERT immunogenicity by altering the specificity of a normal endogenous enzyme to compensate for a defective enzyme. We sought to identify human ß-glucuronidase variants that display α-iduronidase activity, which is defective in mucopolysaccharidosis (MPS) type I patients. A human ß-glucuronidase library was screened by the Enzyme Cleavable Surface-Tethered All-purpose Screen sYstem to isolate variants that exhibited 100-290-fold greater activity against the α-iduronidase substrate 4-methylumbelliferyl α-l-iduronide and 7900-24 500-fold enzymatic specificity shift when compared with wild-type ß-glucuronidase. In vitro treatment of MPS I cells with the ß-glucuronidase variants significantly restored lysosome appearance similar to treatment with α-iduronidase. Our study suggests that ß-glucuronidase variants can be isolated to display α-iduronidase activity and produce significant phenotype improvement of MPS I cells. This strategy may represent a possible approach to produce enzymes that display therapeutic benefits with potentially less immunogenicity.

Neonatal Systemic AAV Induces Tolerance to CNS Gene Therapy in MPS I Dogs and Nonhuman Primates.

The potential host immune response to a nonself protein poses a fundamental challenge for gene therapies targeting recessive diseases. We demonstrate in both dogs and nonhuman primates that liver-directed gene transfer using an adeno-associated virus (AAV) vector in neonates induces a persistent state of immunological tolerance to the transgene product, substantially improving the efficacy of subsequent vector administration targeting the central nervous system (CNS). We applied this approach to a canine model of mucopolysaccharidosis type I (MPS I), a progressive neuropathic lysosomal storage disease caused by deficient activity of the enzyme α-l-iduronidase (IDUA). MPS I dogs treated systemically in the first week of life with a vector expressing canine IDUA did not develop antibodies against the enzyme and exhibited robust expression in the CNS upon intrathecal AAV delivery at 1 month of age, resulting in complete correction of brain storage lesions. Newborn rhesus monkeys treated systemically with AAV vector expressing human IDUA developed tolerance to the transgene, resulting in high cerebrospinal fluid (CSF) IDUA expression and no antibody induction after subsequent CNS gene therapy. These findings suggest that inducing tolerance to the transgene product during a critical period in immunological development can improve the efficacy and safety of gene therapy.

Diagnosing lysosomal storage disorders: mucopolysaccharidosis type I.

Mucopolysaccharidosis type I (MPS I) is a lysosomal storage disorder due to deficiency of alpha iduronidase (IDUA). Progressive storage of dermatan and heparan sulfate throughout the body lead to a multiorgan presentation including short stature, dysostosis multiplex, corneal clouding, hearing loss, coarse facies, hepatosplenomegaly, and intellectual disability. Diagnosis of MPS I is based on IDUA enzyme analysis in leukocytes or dried blood spots (DBS) followed by molecular confirmation of the IDUA gene mutations in individuals with low enzyme activity. The advent of mass spectrometry methods for enzyme analysis in DBS has enabled high-throughput screening for MPS I in symptomatic individuals and newborn infants. The following unit provides the detailed analytical protocol for measurement of IDUA activity in DBS using tandem mass spectrometry.

Shotgun proteomics reveals possible mechanisms for cognitive impairment in Mucopolysaccharidosis I mice.

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.

High-dose enzyme replacement therapy in murine Hurler syndrome.

Mucopolysaccharidosis type I (MPS I) is an autosomal recessive disease that is systemic, including progressive neurodegeneration, mental retardation and death before the age of 10 years. MPS I results from deficiency of α-L-iduronidase (IDUA) in lysosomes and subsequent accumulation of glycosaminoglycans (GAG). Clinical enzyme replacement therapy (ERT) with intravenous laronidase reverses some aspects of MPS I disease (e.g., hepatomegaly, splenomegaly, glycosaminoglycanuria) and ameliorates others (e.g., pulmonary function, cardiac disease, arthropathy, exercise tolerance). However, neurologic benefits are thought to be negligible because the blood-brain barrier (BBB) blocks enzyme from reaching the central nervous system (CNS). We considered the possibility that a very high dose of intravenous laronidase might be able to traverse the BBB in small quantities, and provide some metabolic correction in the brain. To address this question, high-dose laronidase was administered (11.6 mg/kg, once per week, 4 weeks) to adult MPS I mice. IDUA enzyme activity in the cortex of treated mice increased to 97% of that in wild type mice (p<0.01). GAG levels in cortex were reduced by 63% of that from untreated MPS I mice (p<0.05). Further, immunohistochemical analysis showed that treatment reduced secondary GM3-ganglioside accumulation in treated MPS I mice. Water T-maze tests showed that the learning abnormality in MPS I mice was reduced (p<0.0001). In summary, repeated, high-dose ERT facilitated laronidase transit across the BBB, reduced GAG accumulation within the CNS, and rescued cognitive impairment.

Effect of recombinant human growth hormone on changes in height, bone mineral density, and body composition over 1-2 years in children with Hurler or Hunter syndrome.

Patients with Hurler or Hunter syndrome typically have moderate to severe growth deficiencies despite therapy with allogeneic hematopoietic stem cell transplantation and/or enzyme replacement therapy. It is unknown whether treatment with recombinant human growth hormone (hGH) can improve growth in these children. The objectives of this study were to determine the effects of hGH on growth, bone mineral density (BMD), and body composition in children with Hurler or Hunter syndrome enrolled in a longitudinal observational study. The difference in annual change in outcomes between hGH treated and untreated subjects was estimated by longitudinal regression models that adjusted for age, Tanner stage, and sex where appropriate. We report on 23 participants who completed at least 2 annual study visits (10 [43%] treated with hGH): Hurler syndrome (n=13) average age of 9.8 ± 3.1 years (range 5.3-13.6 years; 54% female) and Hunter syndrome (n=10) average age of 12.0 ± 2.7 years (range 7.0-17.0 years; 0% female). As a group, children with Hurler or Hunter syndrome treated with hGH had no difference in annual change in height (growth velocity) compared to those untreated with hGH. Growth velocity in hGH treated individuals ranged from -0.4 to 8.1cm/year and from 0.3 to 6.6 cm/year in the untreated individuals. Among children with Hunter syndrome, 100% (N=4) of those treated but only 50% of those untreated with hGH had an annual increase in height standard deviation score (SDS). Of the individuals treated with hGH, those with GHD had a trend towards higher annualized growth velocity compared to those without GHD (6.5 ± 1.9 cm/year vs. 3.5 ± 2.1cm/year; p=.050). Children treated with hGH had greater annual gains in BMD and lean body mass. In conclusion, although as a group we found no significant difference in growth between individuals treated versus not treated with hGH, individual response was highly variable and we are unable to predict who will respond to treatment. Thus, a trial of hGH may be appropriate in children with Hurler or Hunter syndrome, severe short stature, and growth failure. However, efficacy of hGH therapy should be evaluated after 1 year and discontinued if there is no increase in growth velocity or height SDS. Finally, the long-term benefits of changes in body composition with hGH treatment in this population are unknown.

Mucopolysaccharidosis type I and craniosynostosis.

Mucopolysaccharidosis type-I is caused by a deficiency of the lysosomal enzyme α-L-iduronidase, resulting in gradual deposition of glycosaminoglycans in multiple body organs, affecting physical appearance and system functioning. We present the first reported case associating MPS-I (Hurler-Scheie subtype) with craniosynostosis. A 2.5-year-old girl presented initially with macrocrania. On clinical and radiological examinations we noted a scaphocephaly with dysmorphic facial features of MPS confirmed later on. Intracranial hypertension was documented at fundoscopy (papilloedema) and ICP monitoring, and then surgically treated. This association of scaphocephaly and MPS-I highlights the importance of a meticulous physical examination performed by craniofacial, metabolic and ophthalmologic teams.

An algorithm to predict phenotypic severity in mucopolysaccharidosis type I in the first month of life.

INTRODUCTION: Mucopolysaccharidosis type I (MPS I) is a progressive multisystem lysosomal storage disease caused by deficiency of the enzyme α-L-iduronidase (IDUA). Patients present with a continuous spectrum of disease severity, and the most severely affected patients (Hurler phenotype; MPS I-H) develop progressive cognitive impairment. The treatment of choice for MPS I-H patients is haematopoietic stem cell transplantation, while patients with the more attenuated phenotypes benefit from enzyme replacement therapy. METHODS: Thirty patients were included in this study. Genotypes were collected from all patients and all patients were phenotypically categorized at an age of > 18 months based on the clinical course of the disease. In 18 patients, IDUA activity in fibroblast cultures was measured using an optimized IDUA assay. Clinical characteristics from the first month of life were collected from 23 patients. RESULTS: Homozygosity or compound heterozygosity for specific mutations which are associated with MPS I-H, discriminated a subset of patients with MPS I-H from patients with more attenuated phenotypes (specificity 100%, sensitivity 82%). Next, we found that enzymatic analysis of IDUA activity in fibroblasts allowed identification of patients affected by MPS I-H. Therefore, residual IDUA activity in fibroblasts was introduced as second step in the algorithm. Patients with an IDUA activity of < 0.32 nmol x mg(-1) × hr(-1) invariably were MPS I-H patients, while an IDUA activity of > 0.66 nmol × mg(-1) × hr(-1) was only observed in more attenuated patients. Patients with an intermediate IDUA activity could be further classified by the presence of differentiating clinical characteristics, resulting in a model with 100% sensitivity and specificity for this cohort of patients. CONCLUSION: Using genetic, biochemical and clinical characteristics, all potentially available in the newborn period, an algorithm was developed to predict the MPS I phenotype, allowing timely initiation of the optimal treatment strategy after introduction of NBS.

Characterization of joint disease in mucopolysaccharidosis type I mice.

Mucopolysaccharidoses (MPS) are lysosomal storage disorders characterized by mutations in enzymes that degrade glycosaminoglycans (GAGs). Joint disease is present in most forms of MPS, including MPS I. This work aimed to describe the joint disease progression in the murine model of MPS I. Normal (wild-type) and MPS I mice were sacrificed at different time points (from 2 to 12 months). The knee joints were collected, and haematoxylin-eosin staining was used to evaluate the articular architecture. Safranin-O and Sirius Red staining was used to analyse the proteoglycan and collagen content. Additionally, we analysed the expression of the matrix-degrading metalloproteinases (MMPs), MMP-2 and MMP-9, using immunohistochemistry. We observed progressive joint alterations from 6 months, including the presence of synovial inflammatory infiltrate, the destruction and thickening of the cartilage extracellular matrix, as well as proteoglycan and collagen depletion. Furthermore, we observed an increase in the expression of MMP-2 and MMP-9, which could conceivably explain the degenerative changes. Our results suggest that the joint disease in MPS I mice may be caused by a degenerative process due to increase in proteases expression, leading to loss of collagen and proteoglycans. These results may guide the development of ancillary therapies for joint disease in MPS I.