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The lack of a validated severity scoring system for individuals with Zellweger spectrum disorders (ZSD) hampers optimal patient care and reliable research. Here, we describe the development of such severity score and its validation in a large, well-characterized cohort of ZSD individuals. We developed a severity scoring system based on the 14 organs that typically can be affected in ZSD. A standardized and validated method was used to classify additional care needs in individuals with neurodevelopmental disabilities (Capacity Profile [CAP]). Thirty ZSD patients of varying ages were scored by the severity score and the CAP. The median score was 9 (range 6-19) with a median scoring age of 16.0 years (range 2-36 years). The ZSD severity score was significantly correlated with all 5 domains of the CAP, most significantly with the sensory domain (r = 0.8971, P =

Context 4H or POLR3-related leukodystrophy is an autosomal recessive disorder typically characterized by hypomyelination, hypodontia, and hypogonadotropic hypogonadism, caused by biallelic pathogenic variants in POLR3A, POLR3B, POLR1C, and POLR3K. The endocrine and growth abnormalities associated with this disorder have not been thoroughly investigated to date. Objective To systematically characterize endocrine abnormalities of patients with 4H leukodystrophy. Design An international cross-sectional study was performed on 150 patients with genetically confirmed 4H leukodystrophy between 2015 and 2016. Endocrine and growth abnormalities were evaluated, and neurological and other non-neurological features were reviewed. Potential genotype/phenotype associations were also investigated. Setting This was a multicenter retrospective study using information collected from 3 predominant centers. Patients A total of 150 patients with 4H leukodystrophy and pathogenic variants in POLR3A, POLR3B, or POLR1C were included. Main Outcome Measures Variables used to evaluate endocrine and growth abnormalities included pubertal history, hormone levels (estradiol, testosterone, stimulated LH and FSH, stimulated GH, IGF-I, prolactin, ACTH, cortisol, TSH, and T4), and height and head circumference charts. Results The most common endocrine abnormalities were delayed puberty (57/74; 77% overall, 64% in males, 89% in females) and short stature (57/93; 61%), when evaluated according to physician assessment. Abnormal thyroid function was reported in 22% (13/59) of patients. Conclusions Our results confirm pubertal abnormalities and short stature are the most common endocrine features seen in 4H leukodystrophy. However, we noted that endocrine abnormalities are typically underinvestigated in this patient population. A prospective study is required to formulate evidence-based recommendations for management of the endocrine manifestations of this disorder., Canadian Institutes of Health Research [201610PJT-377869, MOP-G2-341146-159133-BRIDG]; Fondation Les Amis d'Elliot; Leuco-Action; Fondation Lueur d'Espoir pour Ayden; Fondation le Tout pour Loo; Reseau de Medecine Genetique Appliquee of the Fonds de Recherche du Quebec-Sante; Compute Canada; Fonds de Recherche du Quebec en Sante (FRQS) Doctoral Scholarship; Fondation du Grand defi Pierre Lavoie Doctoral Scholarship; McGill Faculty of Medicine F. S.B. Miller Fellowship; Research Institute of the McGill University Health Centre Desjardins Studentship in Child Health Research; Directorate of Higher Education Overseas Scholarship-Dikti Scholarship, Ministry of National Education, Republic of Indonesia; CIHR [201603PJT-148695]; BC Children's Hospital Foundation through its intramural Investigator Grant Award Program (IGAP); National Institute for Neurological Disorders and Stroke [R01NS082094]; Jakob Kamens Chair in Translational Neurotherapeutics; Fonds de Recherche du Quebec-Sante (FRQS); Canadian Institutes of Health Research; European Reference Network for Rare Neurological Disorders (ERN-RND) [739510], This study was supported by grants from the Canadian Institutes of Health Research (201610PJT-377869, MOP-G2-341146-159133-BRIDG), Fondation Les Amis d'Elliot, Leuco-Action, Fondation Lueur d'Espoir pour Ayden, Fondation le Tout pour Loo, and Reseau de Medecine Genetique Appliquee of the Fonds de Recherche du Quebec-Sante to G. Bernard. This research was enabled in part by support provided by Compute Canada (www.computecanada.ca).S.Perrier is supported by the Fonds de Recherche du Quebec en Sante (FRQS) Doctoral Scholarship, the Fondation du Grand defi Pierre Lavoie Doctoral Scholarship, the McGill Faculty of Medicine F. S.B. Miller Fellowship, and the Research Institute of the McGill University Health Centre Desjardins Studentship in Child Health Research. F. K.C. is a recipient of the Directorate of Higher Education Overseas Scholarship-Dikti Scholarship, Ministry of National Education, Republic of Indonesia. W.T. G. received funding from the CIHR (201603PJT-148695) and is supported by the BC Children's Hospital Foundation through its intramural Investigator Grant Award Program (IGAP). B.L. F. was supported by the National Institute for Neurological Disorders and Stroke (R01NS082094). A.V. receives funding from the Jakob Kamens Chair in Translational Neurotherapeutics. G. Bernard has received a Research Scholar Junior 1 award from the Fonds de Recherche du Quebec-Sante (FRQS) (2012-2016) and the New Investigator Salary Award from the Canadian Institutes of Health Research (2017-2022). I.K.M., M. Synofzik, D. Tonduti, B.P.v.d.W., M.S. V.d.K., and N.I.W. are members of the European Reference Network for Rare Neurological Disorders (ERN-RND), project ID 739510.

A frozen shoulder is characterized by pain and stiffness caused by a thickened, contracted capsule of the glenohumeral joint, resulting in marked disabilities in daily life. The pathophysiology of frozen starts with an inflammatory process which triggers a cascade leading to capsular tissue fibrosis. The majority of frozen shoulder patients can be treated non-surgical. Intra-articular corticosteroids should be given early, because they can counteract the inflammatory cascade and decrease the differentiation of fibroblasts into myofibroblasts, but can not undo the capsular fibrosis which has already been formed. Non-surgical treatment should be the initial treatment of choice for FS. This should consist of the combination of an early intra-articular corticosteroid injection, followed by physiotherapy guided by tissue irritability. If conservative treatment is insufficient, manipulation under anesthesia can improve range of motion, pain and function within six weeks to three months. We suggest to use the following criteria before proceeding with MUA: - Unable to cope with the pain and stiffness of a FS - Clinical signs of a FS in stage 2 with external rotation being less than 50% compared to the contralateral shoulder - Decrease of pain in relation to stage 1, and pain mainly at the end range of motion - Failure to respond to an intra-articular injection - A minimal duration of six months of conservative treatment including an intra-articular corticosteroid injection and physiotherapy Future research should focus on prognostic factors predicting the natural course of FS, and advance medical therapies to interrupt the involved inflammatory signalling pathways.

X-linked adrenoleukodystrophy (ALD) is a truly puzzling inborn error of metabolism. Although all patients have ABCD1 mutations, the clinical course is variable and unpredictable in individual patients. In this thesis we continue the effort to unravel the natural history of ALD by delineating the clinical spectrum, improving diagnostics, working towards clinical trial readiness and searching for surrogate outcome measures. Key elements of the clinical spectrum of ALD include primary adrenal insufficiency, cerebral inflammatory demyelination and myelopathy. We retrospectively describe lifetime prevalence of adrenal insufficiency and propose age-dependent follow-up recommendations for the adrenal function. In addition, we evaluate cognitive function in the absence of cerebral disease, and, in a case series, we are the first to describe patients with reactivation of spontaneously arrested cerebral lesions. Timely diagnosis of key manifestations of ALD can prevent severe morbidity and mortality and therefore, implementation of newborn screening has been initiated. We compare C26:0-carnitine to C26:0-lysoPC for the diagnosis of ALD in newborns. Moreover, we evaluate if C26:0-lysoPC and C26:0-carnitine are superior in comparison to routine measurements for the diagnosis of ALD in women. Besides adrenal insufficiency and brain involvement, myelopathy is the key manifestation of ALD in adulthood. We evaluate the rate of progression of myelopathy in both women and men. Moreover, we evaluate the potential of diffusion MRI and optical coherence tomography as surrogate outcome measures for myelopathy. This thesis provides new quantitative data on natural history and will hopefully make life a little less unexpected for ALD patients.

Pontocerebellar Hypoplasia (PCH) is a rare heterogeneous group of neurodegenerative disorders, often with a prenatal onset. Patients have severe hypoplasia or atrophy of cerebellum and pons, with variable involvement of supratentorial structures. Although clinical and neuroradiological features are variable, most patients suffer from severe motor and cognitive impairments. PCH often leads to death during infancy. Based on distinct clinical features and genetic causes, current classification comprises eleven types of PCH. This thesis focuses on the identification of novel disease genes and the description of new phenotype-genotype correlations in PCH. In addition, the neuroradiological features of the most frequent subtypes of PCH (PCH1B/PCH2A) are studied in detail. The expression patterns of the PCH related genes in the developing and adult brain will be shortly discussed in an effort to explain the specific cerebellar involvement in PCH. Finally, the clinical applicability of the ever expanding PCH classification, which now includes a large variety of phenotypes, will be considered.

This thesis describes translational studies in patients with a Zellweger spectrum disorder (ZSD), aimed at providing a framework for the in vitro development of therapies for ZSDs and for testing potential therapies in future clinical trials. ZSDs are a heterogeneous group of disorders characterized by a defect in peroxisome formation and are caused by mutations in one of the PEX genes. Because of the defect in peroxisome formation, multiple metabolic pathways are impaired resulting in metabolic abnormalities. Currently, no curative therapy is available. We studied whether autophagy inhibitors are capable of restoring peroxisomal function in ZSD cells and found no improvement of peroxisomal function by inhibition of autophagy. Moreover, we studied the clinical and biochemical effects of cholic acid treatment in a cohort of ZSD patients. Cholic acid therapy led to a decrease in toxic bile acid intermediates in plasma of the majority of patients, but gave rise to a worsening of liver tests in patients with advanced liver disease. No improvement of clinically relevant parameters could be observed. Furthermore, we describe the validation of two potential new biomarkers for ZSDs: C26:0-lysophosphatidylcholine and C26:0-carnitine in dried blood spots from a large cohort of ZSD patients and healthy individuals. ZSDs comprise a spectrum of disorders in which patients can present with a broad range of symptoms and widely varying clinical severity. The lack of a validated severity scoring system for patients with a ZSD hampers clinical research in this disease. In this thesis, we present the first severity scoring system for ZSD and its validation in a large, well-characterized cohort of ZSD patients.

X-linked adrenoleukodystrophy (ALD) is the most common leukodystrophy with a birth incidence of 1:14,700. ALD is characterized by impaired beta-oxidation and enhanced elongation of very long-chain fatty acids (VLCFA) due to a defect in the peroxisomal transmembrane protein ABCD1. The first part of this theses is focused on improving the diagnosis of ALD. To this end, a method is described which can be used to measure beta-oxidation and de novo synthesis in living cells by using stable-isotope labelled substrates. Diagnosing women with ALD can be challenging since 20% have normal VLCFA plasma levels. This thesis shows that C26:0-lysoPC (the biomarker currently used to detect ALD in newborn screening programs) is a more accurate biomarker to diagnose women with ALD. Furthermore, C26:0-carnitine was identified as a good biomarker for ALD in an ALD mouse model and adult male ALD patients but failed as a biomarker for newborns and women with ALD. The exact mechanism behind VLCFA toxicity has not been resolved at the cellular level yet. This thesis shows that exposure to VLCFA activates the endoplasmic reticulum (ER) stress response and causes lipoapoptosis in fibroblasts from ALD patients. Interestingly, only saturated VLCFA but not mono-unsaturated VLCFA activate the ER stress response. Via activation of stearoyl-CoA desaturase 1, it is possible to redirect the saturated VLCFA synthesis towards the mono-unsaturated VLCFA synthesis. This results in a normalization of the saturated VLCFA levels in fibroblasts from ALD patients.

Pontocerebellar hypoplasia (PCH) is a recessive neurodegenerative disease with, in most cases, a prenatal onset. The patients suffer from severe intellectual and motor impairments. The majority of patients dies in childhood. This thesis describes novel genes and genotype-phenotype correlations in PCH. In chapter 2 we confirm the presence of PCH and disorders of sex development as an isolated disease (PCH subtype 7). Chapter 3 focuses on genotype-phenotype correlations in EXOSC3-related PCH. We describe a cohort of twelve families and show that specific mutations can predict disease progression. Chapter 4 describes CLP1 as a novel PCH-causing gene and investigates the function of CLP1 in brain development using a zebrafish model. Chapter 5 discusses (t)RNA processing deficiencies as possible underlying disease mechanism in PCH.

Zellweger spectrum disorders (ZSDs) are inherited metabolic diseases characterized by a (partial) deficiency of peroxisomal function, leading to accumulation of several toxic metabolites in organs and blood. Currently, there is no curative therapy for the diseases and intervention is supportive and based on symptoms. In this thesis we have studied the effect of a possible novel therapy, arginine, in skin fibroblasts of mildly affected ZSD patients. Supplementing arginine to the culture medium of these skin fibroblasts resulted in an increase in the amount of peroxisomes as well as peroxisomal functions. Moreover the effect of cholic acid was studied for the first time in a large cohort of patients. We have demonstrated that cholic acid lowers the concentration of specific toxic metabolites in plasma in the majority of the patients. We also generated a new mouse model for the disease, which resembles the relatively milder human phenotype. In the near future, this model can be used to study disease pathogenesis at the organ level and test future therapies. Research in this thesis emphasizes that ZSDs should no longer be considered solely as paediatric diseases, but rather as slowly progressive diseases with patients surviving into adulthood, presenting with age specific (neurological) symptoms as described in our cohort study.