Studies on cartilage and bone disease in Mucopolysaccharidoses and Mucolipidoses

Mucopolysaccharidosis (MPS) and Mucolipidosis (ML II and III) are lysosomal storage disorders with multisystem involvement. In MPS, deficiencies of glycosaminoglycans (GAGs) degrading enzymes lead to intralysosomal GAG storage. In the MLs, defective trafficking of lysosomal enzymes to the lysosome, leads to accumulation of a combination of GAGs and several other complex molecules. GAGs are degraded by enzymes, in part extracellularly and in part intracellularly in the lysosomes after uptake through endocytosis. Intralysosomal storage in MPS and ML patients gives rise to loss of cellular function by disturbed autophagy, polyubiquitination, mitochondrial dysfunction, inflammation, apoptosis, and loss of lysosomal membrane integrity, followed by tissue damage and organ dysfunction. These events eventually determine the clinical symptoms observed in the patients. Skeletal abnormalities are common in MPS and ML patients and originate from intralysosomal storage in cells of the cartilage, bones and ligaments. A major problem of these tissues is that they are difficult to treat as vascularization is poor and cell renewal (division) is slow. The existing therapies for these diseases are unable to fully correct or prevent the abnormalities occurring in bones and cartilage. To enable development of new therapies it is crucial to understand the processes involved in abnormal cartilage and bone development as observed in MPS and ML and relate them to normal skeletal development. The aim of this thesis is to create a better understanding of the etiology and pathophysiology of cartilage and bone development in patients with mucopolysaccharidosis and mucolipidosis, the clinical course and therapeutic challenges of skeletal disease in these disorders

Severe tracheal and bronchial collapse in adults with type II mucopolysaccharidosis

Background: Mucopolysaccharidosis type II (MPSII) patients frequently suffer from dyspnoea caused by restrictive airway disease due to skeletal abnormalities as well as glycosaminoglycans (GAG) accumulation at different levels of the airway, including the trachea. In this study we describe the extent of the tracheal and bronchial narrowing, the changes in airway diameter during respiration and the effects of these obstructions on respiratory function in adult MPSII patients. Methods: Five adult MPSII patients (mean age 40 years) were included. Pulmonary function tests and in- and expiratory chest CT scans were obtained. Cross-sectional areas of trachea and main bronchi were measured at end-inspiration and -expiration and percentage collapse was calculated. Results: There was diffuse narrowing of the entire intra-thoracic trachea and main bronchi and severe expiratory collapse of the trachea in all patients. At 1 cm above the aortic arch the median % collapse of the trachea was 68 (range 60 to 77 %), at the level of the aortic arch 64 (range 21-93 %), for the main bronchi this was 58 (range 26-66 %) on the left and 44 (range 9-76 %) on the right side. The pulmonary function tests showed that this airway collapse results in obstructive airway disease in all patients, which was severe (forced expiratory volume <50 % of predicted) in four out of five patients. Conclusion: In adult MPS II patients, central airways diameters are strikingly reduced and upon expiration there is extensive collapse of the trachea and main bronchi. This central airways obstruction explains the severe respiratory symptoms in MPSII patients

Clinical trial of laronidase in Hurler syndrome after hematopoietic cell transplantation.

BackgroundMucopolysaccharidosis I (MPS IH) is a lysosomal storage disease treated with hematopoietic cell transplantation (HCT) because it stabilizes cognitive deterioration, but is insufficient to alleviate all somatic manifestations. Intravenous laronidase improves somatic burden in attenuated MPS I. It is unknown whether laronidase can improve somatic disease following HCT in MPS IH. The objective of this study was to evaluate the effects of laronidase on somatic outcomes of patients with MPS IH previously treated with HCT.MethodsThis 2-year open-label pilot study of laronidase included ten patients (age 5-13 years) who were at least 2 years post-HCT and donor engrafted. Outcomes were assessed semi-annually and compared to historic controls.ResultsThe two youngest participants had a statistically significant improvement in growth compared to controls. Development of persistent high-titer anti-drug antibodies (ADA) was associated with poorer 6-min walk test (6MWT) performance; when patients with high ADA titers were excluded, there was a significant improvement in the 6MWT in the remaining seven patients.ConclusionsLaronidase seemed to improve growth in participants <8 years old, and 6MWT performance in participants without ADA. Given the small number of patients treated in this pilot study, additional study is needed before definitive conclusions can be made

Classical galactosemia: neuropsychological and psychosocial functioning beyond intellectual abilities

BACKGROUND: Despite early diagnosis and treatment, Classical Galactosemia (CG) patients frequently develop long-term complications, such as cognitive impairment. Available literature primarily reports on general intellectual abilities and shows a substantially lower Full Scale Intelligence Quotient (FSIQ) in CG patients than in the general population. Both problems in social functioning as well as internalizing problems are often reported in CG patients. The combination of intelligence, cognitive functioning, beh

Pathway to diagnosis and burden of illness in mucopolysaccharidosis type VII- A European caregiver survey

Background: Mucopolysaccharidosis type VII (Sly disease, MPS VII), is an ultra-rare, multi-symptom disease with variable clinical presentations which can present challenges with diagnosis, management and care. We believe this survey is the first to explore the patient experience through direct questioning of the caregivers of 13 individuals with MPS VII. Methods: This European survey, using a specifically designed questionnaire, was conducted in order to describe the pathway to diagnosis and the burden of illness of MPS VII. Information on early symptoms, clinicians seen, and current symptoms was collected. Questions on the caregivers' ability to work and the use and availability of health, social and educational support were included. Results: Caregivers of 13 patients from Germany, Spain, The Netherlands and Turkey responded to the survey. Five patients with non-immune hydrops fetalis (NIHF) were diagnosed with MPS VII at a mean age of 1.9 years (median 0.3 years, range 0.2 to 6 years). Those without NIHF (n = 7) were diagnosed at a mean age of 6.1 years (median 6.0 years, range 1.9 to 14 years). The symptoms most likely to raise a suspicion of MPS VII, excluding NIHF, did not appear until a median age of at least three years. Over one half of patients required assistance with daily living and mobility. Reduction of the working hours of caregivers was often necessary (46.2% reduced hours, 30.8% stopped working). Patients attended frequent medical appointments (12.7/year), over 80% had surgery and 30% had been hospitalised for respiratory issues. While support for learning and behavioural needs was generally available, support for mobility was not available to 50% of patients. Half of the respondents (6/12) said they were not offered genetic counselling. Conclusions: For children that do not present with NIHF, diagnosis can take several years as early symptoms can be non-specific and mistaken for other conditions. Increased awareness of the early signs of disease and more information for parents/caregivers at diagnosis are needed. MPS VII poses significant burden to patients, caregivers, healthcare, social and educational services. Access to information and support varies across Europe and the availability of genetic counselling is limited in some countries

The Galactose Index measured in fibroblasts of GALT deficient patients distinguishes variant patients detected by newborn screening from patients with classical phenotypes

Background: The high variability in clinical outcome of patients with Classical Galactosemia (CG) is poorly understood and underlines the importance of prognostic biomarkers, which are currently lacking. The aim of this study was to investigate if residual galactose metabolism capacity is associated with clinical and biochemical outcomes in CG patients with varying geno- and phenotypes. Methods: Galactose Metabolite Profiling (GMP) was used to determine residual galactose metabolism in fibroblasts of CG patients. The association between the galactose index (GI) defined as the ratio of the measured metabolites [U13C]Gal-1-P/ [13C6]UDP-galactose, and both intellectual and neurological outcome and galactose-1-phosphate (Gal-1-P) levels was investigated. Results: GMP was performed in fibroblasts of 28 patients and 3 control subjects. The GI of the classical phenotype patients (n = 22) was significantly higher than the GI of four variant patients detected by newborn screening (NBS) (p = .002), two homozygous p.Ser135Leu patients (p = .022) and three controls (p = .006). In the classical phenotype patients, 13/18 (72%) had a poor intellectual outcome (IQ < 85) and 6/12 (50%) had a movement disorder. All the NBS detected variant patients (n = 4) had a normal intellectual outcome (IQ ≥ 85) and none of them has a movement disorder. In the classical phenotype patients, there was no significant difference in GI between patients with a poor and normal clinical outcome. The NBS detected variant patients had significantly lower GI levels and thus higher residual galactose metabolism than patients with classical phenotypes. There was a clear correlation between Gal-1-P levels in erythrocytes and the GI (p = .001). Conclusions: The GI was able to distinguish CG patients with varying geno- and phenotypes and correlated with Gal-1-P. The data of the NBS detected variant patients demonstrated that a higher residual galactose metabolism may result in a more favourable clinical outcome. Further research is needed to enable individual prognostication and treatment in all CG patients

Hip Morphology in Mucolipidosis Type II

Mucolipidosis type II (MLII) is a rare lysosomal storage disorder caused by defective trafficking of lysosomal enzymes. Severe skeletal manifestations are a hallmark of the disease including hip dysplasia. This study aims to describe hip morphology and the natural course of hip pathologies in MLII by systematic evaluation of plain radiographs, ultrasounds and magnetic resonance imaging (MRI). An international two-centered study was performed by retrospective chart review. All MLII patients with at least one pelvic radiograph were included. A total of 16 patients were followed over a mean of 3.5 years (range 0.2–10.7 years). Typical age-dependent radiographic signs identified were femoral cloaking (7/16), rickets/hyperparathyroidism-like changes (6/16) and constrictions of the supra-acetabular part of the os ilium (16/16) and the femoral neck (7/16). The course of acetabular and migration indexes (AI, MI) significantly increased in female patients. However, in the overall group, there was no relevant progression of acetabular dysplasia with a mean AI of 23.0 (range 5◦–41◦ ) and 23.7◦ (range 5◦–40◦ ) at the first and last assessments, respectively. Better knowledge on hip morphology in MLII could lead to earlier diagnosis, improved clinical management and enables assessment of effects of upcoming therapies on the skeletal system

Analytical assay validation for acute myeloid leukemia measurable residual disease assessment by multiparametric flow cytometry

Background: Measurable residual disease (MRD) assessed by multiparametric flow cytometry (MFC) has gained importance in clinical decision-making for acute myeloid leukemia (AML) patients. However, complying with the recent In Vitro Diagnostic Regulations (IVDR) in Europe and Food and Drug Administration (FDA) guidance in the United States requires rigorous validation prior to their use in investigational clinical trials and diagnostics. Validating AML MRD-MFC assays poses challenges due to the unique underlying disease biology and paucity of patient specimens. In this study, we describe an experimental framework for validation that meets regulatory expectations. Methods: Our validation efforts focused on evaluating assay accuracy, analytical specificity, analytical and functional sensitivity (limit of blank (LoB), detection (LLoD) and quantitation (LLoQ)), precision, linearity, sample/reagent stability and establishing the assay background frequencies. Results: Correlation between different MFC methods was highly significant (r = 0.99 for %blasts and r = 0.93 for %LAIPs). The analysis of LAIP specificity accurately discriminated from negative control cells. The assay demonstrated a LoB of 0.03, LLoD of 0.04, and LLoQ of 0.1%. Precision experiments yielded highly reproducible results (Coefficient of Variation &lt;20%). Stability experiments demonstrated reliable measurement of samples up to 96 h from collection. Furthermore, the reference range of LAIP frequencies in non-AML patients was below 0.1%, ranging from 0.0% to 0.04%.Conclusion: In this manuscript, we present the validation of an AML MFC-MRD assay using BM/PB patient specimens, adhering to best practices. Our approach is expected to assist other laboratories in expediting their validation activities to fulfill recent health authority guidelines.</p

Measurement of χ c1 and χ c2 production with s√ = 7 TeV pp collisions at ATLAS

The prompt and non-prompt production cross-sections for the χ c1 and χ c2 charmonium states are measured in pp collisions at s√ = 7 TeV with the ATLAS detector at the LHC using 4.5 fb−1 of integrated luminosity. The χ c states are reconstructed through the radiative decay χ c → J/ψγ (with J/ψ → μ + μ −) where photons are reconstructed from γ → e + e − conversions. The production rate of the χ c2 state relative to the χ c1 state is measured for prompt and non-prompt χ c as a function of J/ψ transverse momentum. The prompt χ c cross-sections are combined with existing measurements of prompt J/ψ production to derive the fraction of prompt J/ψ produced in feed-down from χ c decays. The fractions of χ c1 and χ c2 produced in b-hadron decays are also measured