Folate receptor α deficiency - Myelin-sensitive MRI as a reliable biomarker to monitor the efficacy and long-term outcome of a new therapeutic approach.

Cerebral folate transport deficiency, caused by a genetic defect in folate receptor α, is a devastating neurometabolic disorder that, if untreated, leads to epileptic encephalopathy, psychomotor decline and hypomyelination. Currently, there are limited data on effective dosage and duration of treatment, though early diagnosis and therapy with folinic acid appears critical. The aim of this long-term study was to identify new therapeutic approaches and novel biomarkers for assessing efficacy, focusing on myelin-sensitive MRI. Clinical, biochemical, structural and quantitative MRI parameters of seven patients with genetically confirmed folate receptor α deficiency were acquired over 13 years. Multimodal MRI approaches comprised MR-spectroscopy (MRS), magnetization transfer (MTI) and diffusion tensor imaging (DTI) sequences. Patients started oral treatment immediately following diagnosis or in an interval of up to 2.5 years. Escalation to intravenous and intrathecal administration was performed in the absence of effects. Five patients improved, one with a presymptomatic start of therapy remained symptom-free, and one with inconsistent treatment deteriorated. While CSF 5-methyltetrahydrofolate and MRS parameters normalized immediately after therapy initiation, myelin-sensitive MTI and DTI measures correlated with gradual clinical improvement and ongoing myelination under therapy. Early initiation of treatment at sufficient doses, considering early intrathecal applications, is critical for favorable outcome. The majority of patients showed clinical improvements that correlated best with MTI parameters, allowing individualized monitoring of myelination recovery. Presymptomatic therapy seems to ensure normal development and warrants newborn screening. Furthermore, the quantitative parameters of myelin-sensitive MRI for therapy assessments can now be used for hypomyelination disorders in general.

Cln5 represents a new type of cysteine-based S-depalmitoylase linked to neurodegeneration.

Genetic CLN5 variants are associated with childhood neurodegeneration and Alzheimer's disease; however, the molecular function of ceroid lipofuscinosis neuronal protein 5 (Cln5) is unknown. We solved the Cln5 crystal structure and identified a region homologous to the catalytic domain of members of the N1pC/P60 superfamily of papain-like enzymes. However, we observed no protease activity for Cln5; and instead, we discovered that Cln5 and structurally related PPPDE1 and PPPDE2 have efficient cysteine palmitoyl thioesterase (S-depalmitoylation) activity using fluorescent substrates. Mutational analysis revealed that the predicted catalytic residues histidine-166 and cysteine-280 are critical for Cln5 thioesterase activity, uncovering a new cysteine-based catalytic mechanism for S-depalmitoylation enzymes. Last, we found that Cln5-deficient neuronal progenitor cells showed reduced thioesterase activity, confirming live cell function of Cln5 in setting S-depalmitoylation levels. Our results provide new insight into the function of Cln5, emphasize the importance of S-depalmitoylation in neuronal homeostasis, and disclose a new, unexpected enzymatic function for the N1pC/P60 superfamily of proteins.

CD9 and folate receptor overexpression are not sufficient for VSV-G-independent lentiviral transduction.

Extracellular vesicles have become a research focus for their potential as therapeutic vehicles that carry cargo substances. Extracellular vesicles may origin from the endosomal compartment and share several characteristics with the envelope of lentiviruses. A previous study reported that constitutive expression of the tetraspanin CD9, an extracellular vesicle marker, not only increases vesicle secretion from cells, but has also a positive effect on lentiviral transduction efficiency. Moreover, it was shown that expression of CD9 on the viral envelope in absence of viral glycoproteins was sufficient for the transduction of mammalian cells. In this study, we investigate the effect of CD9 and folate receptor alpha, a GPI-anchored protein, on biosynthesis and transduction efficiency of vesicles carrying lentiviral vectors. We demonstrate that neither CD9 nor FRα nor the combination of both were able to mediate a significant transduction of therapeutic vesicles carrying lentiviral RNA. Further studies are required to identify endogenous mammalian proteins that can be used for pseudotyping of viral envelopes to improve viral targeting without inducing immune responses.

WARS2 mutations cause dopa-responsive early-onset parkinsonism and progressive myoclonus ataxia.

INTRODUCTION: Sixteen subjects with biallelic WARS2 variants encoding the tryptophanyl mitochondrial aminoacyl-tRNA synthetase, presenting with a neonatal- or infantile-onset mitochondrial disease, have been reported to date. Here we present six novel cases with WARS2-related diseases and expand the spectrum to later onset phenotypes including dopa-responsive early-onset parkinsonism and progressive myoclonus-ataxia. METHODS: Six individuals from four families underwent whole-exome sequencing within research and diagnostic settings. Following the identification of a genetic defect, in-depth phenotyping and protein expression studies were performed. RESULTS: A relatively common (gnomAD MAF = 0.0033) pathogenic p.(Trp13Gly) missense variant in WARS2 was detected in trans in all six affected individuals in combination with different pathogenic alleles (exon 2 deletion in family 1; p.(Leu100del) in family 2; p.(Gly50Asp) in family 3; and p.(Glu208*) in family 4). Two subjects presented with action tremor around age 10-12 years and developed tremor-dominant parkinsonism with prominent neuropsychiatric features later in their 20s. Two subjects presented with a progressive myoclonus-ataxia dominant phenotype. One subject presented with spasticity, choreo-dystonia, myoclonus, and speech problems. One subject presented with speech problems, ataxia, and tremor. Western blotting analyses in patient-derived fibroblasts showed a markedly decreased expression of the full-length WARS2 protein in both subjects carrying p.(Trp13Gly) and an exon-2 deletion in compound heterozygosity. CONCLUSIONS: This study expands the spectrum of the disease to later onset phenotypes of early-onset tremor-dominant parkinsonism and progressive myoclonus-ataxia phenotypes.

Variants in Mitochondrial ATP Synthase Cause Variable Neurologic Phenotypes.

OBJECTIVE: ATP synthase (ATPase) is responsible for the majority of ATP production. Nevertheless, disease phenotypes associated with mutations in ATPase subunits are extremely rare. We aimed at expanding the spectrum of ATPase-related diseases. METHODS: Whole-exome sequencing in cohorts with 2,962 patients diagnosed with mitochondrial disease and/or dystonia and international collaboration were used to identify deleterious variants in ATPase-encoding genes. Findings were complemented by transcriptional and proteomic profiling of patient fibroblasts. ATPase integrity and activity were assayed using cells and tissues from 5 patients. RESULTS: We present 10 total individuals with biallelic or de novo monoallelic variants in nuclear ATPase subunit genes. Three unrelated patients showed the same homozygous missense ATP5F1E mutation (including one published case). An intronic splice-disrupting alteration in compound heterozygosity with a nonsense variant in ATP5PO was found in one patient. Three patients had de novo heterozygous missense variants in ATP5F1A, whereas another 3 were heterozygous for ATP5MC3 de novo missense changes. Bioinformatics methods and populational data supported the variants' pathogenicity. Immunohistochemistry, proteomics, and/or immunoblotting revealed significantly reduced ATPase amounts in association to ATP5F1E and ATP5PO mutations. Diminished activity and/or defective assembly of ATPase was demonstrated by enzymatic assays and/or immunoblotting in patient samples bearing ATP5F1A-p.Arg207His, ATP5MC3-p.Gly79Val, and ATP5MC3-p.Asn106Lys. The associated clinical profiles were heterogeneous, ranging from hypotonia with spontaneous resolution (1/10) to epilepsy with early death (1/10) or variable persistent abnormalities, including movement disorders, developmental delay, intellectual disability, hyperlactatemia, and other neurologic and systemic features. Although potentially reflecting an ascertainment bias, dystonia was common (7/10). INTERPRETATION: Our results establish evidence for a previously unrecognized role of ATPase nuclear-gene defects in phenotypes characterized by neurodevelopmental and neurodegenerative features. ANN NEUROL 2022;91:225-237.

Human COQ4 deficiency: delineating the clinical, metabolic and neuroimaging phenotypes.

BACKGROUND: Human coenzyme Q4 (COQ4) is essential for coenzyme Q10 (CoQ10) biosynthesis. Pathogenic variants in COQ4 cause childhood-onset neurodegeneration. We aimed to delineate the clinical spectrum and the cellular consequences of COQ4 deficiency. METHODS: Clinical course and neuroradiological findings in a large cohort of paediatric patients with COQ4 deficiency were analysed. Functional studies in patient-derived cell lines were performed. RESULTS: We characterised 44 individuals from 36 families with COQ4 deficiency (16 newly described). A total of 23 different variants were identified, including four novel variants in COQ4. Correlation analyses of clinical and neuroimaging findings revealed three disease patterns: type 1: early-onset phenotype with neonatal brain anomalies and epileptic encephalopathy; type 2: intermediate phenotype with distinct stroke-like lesions; and type 3: moderate phenotype with non-specific brain pathology and a stable disease course. The functional relevance of COQ4 variants was supported by in vitro studies using patient-derived fibroblast lines. Experiments revealed significantly decreased COQ4 protein levels, reduced levels of cellular CoQ10 and elevated levels of the metabolic intermediate 6-demethoxyubiquinone. CONCLUSION: Our study describes the heterogeneous clinical presentation of COQ4 deficiency and identifies phenotypic subtypes. Cell-based studies support the pathogenic characteristics of COQ4 variants. Due to the insufficient clinical response to oral CoQ10 supplementation, alternative treatment strategies are warranted.

Preoperative clinical symptomatology and stroke burden in pediatric moyamoya angiopathy: Defining associated risk variables.

OBJECTIVE: To evaluate the preoperative clinical and magnetic resonance imaging and angiography (MRI-MRA) characteristics in pediatric moyamoya patients. METHODS: Analysis included 100 children with moyamoya angiopathy referred to our moyamoya center for preoperative evaluation. Clinical symptoms, neurological status using Pediatric Stroke Outcome Measurement (PSOM) and degree of disability on modified Rankin scale score (mRS) were evaluated. MRI-MRA evaluation included the assessment of ischemic lesions and involvement of posterior circulation. Data were analyzed for moyamoya disease (MMD), moyamoya syndrome (MMS) and age at disease onset. RESULTS: Stroke was a common presentation in both MMD and MMS patients. TIAs and headaches/migraine were more frequent in MMD. There was no evidence of a difference in stroke burden on MRI as well as in PCA involvement between the two subgroups. Children <2 years had higher odds of having a stroke (OR 15.5, 95% CI 3.8-62.4, p < 0.001), recurrent stroke (OR 11.8, 95%CI 2.9-46.7, p < 0.001) and unfavorable mRS (≥2) (OR 4.2, 95% CI 1.3-13.7, p = 0.01) when compared to those >5 years of age. There was some evidence of association of PCA involvement with recurrent strokes (OR 3.1, CI 1.0-9.6, p = 0.05), a poor PSOM (OR 3.0, 95% CI 1.1-8.2, p = 0.04) and mRS (OR 3.1, 95% CI 1.2-8.3, p = 0.02). CONCLUSION: Stroke seems to be a common presentation in both MMD and MMS patients. Early age at symptom onset and involvement of posterior circulation seem to be important risk factors for a high stroke burden and an unfavorable PSOM and mRS.

Early childhood neurodevelopmental outcome after open prenatal spina bifida aperta repair.

AIM: To investigate neurodevelopmental outcome of children with open prenatal spina bifida aperta (SBA) repair. METHOD: Prenatal SBA repair was performed in 130 fetuses at the Zurich Center between 2010 and 2019. Seventy-seven children underwent 1 year assessment with the Griffiths Mental Developmental Scales (Griffiths) and 65 with the Bayley Scales of Infant and Toddler Development, Third Edition (Bayley-III) at 2 years. Anatomical and functional level and ambulation status were assessed. Descriptive statistics and multiple linear regression analyses for risk factors were performed. RESULTS: The Bayley-III cognition composite score in children with prenatal SBA repair was within normal limits but lower compared to population norms (mean=95.15, SD=14.683 vs norm=100, SD=15, p=0.01). Fine motor development (mean=9.58, SD=2.744, p=0.227) was typical while gross motor development was lower than the norm (mean=3.02, SD=2.758 vs norm=10, SD=3, p<0.001). Griffiths developmental quotient subscales correlated significantly with corresponding Bayley-III scores (all p<0.001, r=0.519-0.594). At 2 years, 50.8% could walk. INTERPRETATION: Children with non-trial open prenatal SBA repair show favourable cognitive outcome in the low-average range at 1 and 2 years of age. While gross motor function remained delayed, fine motor function was age appropriate. The correlation between Griffiths and Bayley-III allows a prediction about neurodevelopmental outcome at the age of 1 year. What this paper adds Children with non-trial open prenatal spina bifida repair show favourable cognitive outcome. Gross motor function remains impaired, while fine motor function is age appropriate. At 2 years of age, 50.8% of children were walking. Neurodevelopmental testing correlated between 1 (Griffiths Mental Developmental Scales) and 2 (Bayley Scales of Infant and Toddler Development, Third Edition) years.

PIGG variant pathogenicity assessment reveals characteristic features within 19 families.

PURPOSE: Phosphatidylinositol Glycan Anchor Biosynthesis, class G (PIGG) is an ethanolamine phosphate transferase catalyzing the modification of glycosylphosphatidylinositol (GPI). GPI serves as an anchor on the cell membrane for surface proteins called GPI-anchored proteins (GPI-APs). Pathogenic variants in genes involved in the biosynthesis of GPI cause inherited GPI deficiency (IGD), which still needs to be further characterized. METHODS: We describe 22 individuals from 19 unrelated families with biallelic variants in PIGG. We analyzed GPI-AP surface levels on granulocytes and fibroblasts for three and two individuals, respectively. We demonstrated enzymatic activity defects for PIGG variants in vitro in a PIGG/PIGO double knockout system. RESULTS: Phenotypic analysis of reported individuals reveals shared PIGG deficiency-associated features. All tested GPI-APs were unchanged on granulocytes whereas CD73 level in fibroblasts was decreased. In addition to classic IGD symptoms such as hypotonia, intellectual disability/developmental delay (ID/DD), and seizures, individuals with PIGG variants of null or severely decreased activity showed cerebellar atrophy, various neurological manifestations, and mitochondrial dysfunction, a feature increasingly recognized in IGDs. Individuals with mildly decreased activity showed autism spectrum disorder. CONCLUSION: This in vitro system is a useful method to validate the pathogenicity of variants in PIGG and to study PIGG physiological functions.

High-grade extracellular vesicles preparation by combined size-exclusion and affinity chromatography.

Extracellular vesicles (EVs) have recently gained growing interest for their diagnostic and therapeutic potential. Despite this, few protocols have been reported for the isolation of EVs with preserved biological function. Most EV purification methods include a precipitation step that results in aggregation of vesicles and most available techniques do not efficiently separate the various types of EVs such as exosomes and ectosomes, which are involved in distinct biological processes. For this reason, we developed a new two-step fast performance liquid chromatography (FPLC) protocol for purification of large numbers of EVs. The method comprises size exclusion chromatography followed by immobilized metal affinity chromatography, which is enabled by expression of poly-histidine tagged folate receptor α in the parental cells. Characterisation and comparison of the EVs obtained by this method to EVs purified by differential centrifugation, currently the most common method to isolate EVs, demonstrated higher purity and more selective enrichment of exosomes in EV preparations using our FPLC method, as assessed by comparison of marker proteins and density distribution. Our studies reveal new possibilities for the isolation of defined subpopulations of EVs with preserved biological function that can easily be upscaled for production of larger amounts of EVs.

De novo variants in neurodevelopmental disorders-experiences from a tertiary care center.

Up to 40% of neurodevelopmental disorders (NDDs) such as intellectual disability, developmental delay, autism spectrum disorder, and developmental motor abnormalities have a documented underlying monogenic defect, primarily due to de novo variants. Still, the overall burden of de novo variants as well as novel disease genes in NDDs await discovery. We performed parent-offspring trio exome sequencing in 231 individuals with NDDs. Phenotypes were compiled using human phenotype ontology terms. The overall diagnostic yield was 49.8% (n = 115/231) with de novo variants contributing to more than 80% (n = 93/115) of all solved cases. De novo variants affected 72 different-mostly constrained-genes. In addition, we identified putative pathogenic variants in 16 genes not linked to NDDs to date. Reanalysis performed in 80 initially unsolved cases revealed a definitive diagnosis in two additional cases. Our study consolidates the contribution and genetic heterogeneity of de novo variants in NDDs highlighting trio exome sequencing as effective diagnostic tool for NDDs. Besides, we illustrate the potential of a trio-approach for candidate gene discovery and the power of systematic reanalysis of unsolved cases.

CIC de novo loss of function variants contribute to cerebral folate deficiency by downregulating FOLR1 expression.

Background Cerebral folate deficiency (CFD) syndrome is characterised by a low concentration of 5-methyltetrahydrofolate in cerebrospinal fluid, while folate levels in plasma and red blood cells are in the low normal range. Mutations in several folate pathway genes, including FOLR1 (folate receptor alpha, FRα), DHFR (dihydrofolate reductase) and PCFT (proton coupled folate transporter) have been previously identified in patients with CFD. Methods In an effort to identify causal mutations for CFD, we performed whole exome sequencing analysis on eight CFD trios and identified eight de novo mutations in seven trios. Results Notably, we found a de novo stop gain mutation in the capicua (CIC) gene. Using 48 sporadic CFD samples as a validation cohort, we identified three additional rare variants in CIC that are putatively deleterious mutations. Functional analysis indicates that CIC binds to an octameric sequence in the promoter regions of folate transport genes: FOLR1, PCFT and reduced folate carrier (Slc19A1; RFC1). The CIC nonsense variant (p.R353X) downregulated FOLR1 expression in HeLa cells as well as in the induced pluripotent stem cell (iPSCs) derived from the original CFD proband. Folate binding assay demonstrated that the p.R353X variant decreased cellular binding of folic acid in cells. Conclusion This study indicates that CIC loss of function variants can contribute to the genetic aetiology of CFD through regulating FOLR1 expression. Our study described the first mutations in a non-folate pathway gene that can contribute to the aetiology of CFD.

Impact of interventional and non-interventional variables on anthropometric long-term development in glutaric aciduria type 1: A national prospective multi-centre study.

Glutaric aciduria type 1 (GA1) is a rare neurometabolic disorder, caused by inherited deficiency of glutaryl-CoA dehydrogenase, mostly affecting the brain. Early identification by newborn screening (NBS) significantly improves neurologic outcome. It has remained unclear whether recommended therapy, particular low lysine diet, is safe or negatively affects anthropometric long-term outcome. This national prospective, observational, multi-centre study included 79 patients identified by NBS and investigated effects of interventional and non-interventional parameters on body weight, body length, body mass index (BMI) and head circumference as well as neurological parameters. Adherence to recommended maintenance and emergency treatment (ET) had a positive impact on neurologic outcome and allowed normal anthropometric development until adulthood. In contrast, non-adherence to ET, resulting in increased risk of dystonia, had a negative impact on body weight (mean SDS -1.07; P = .023) and body length (mean SDS -1.34; P = -.016). Consistently, longitudinal analysis showed a negative influence of severe dystonia on weight and length development over time (P < .001). Macrocephaly was more often found in female (mean SDS 0.56) than in male patients (mean SDS -0.20; P = .049), and also in individuals with high excreter phenotype (mean SDS 0.44) compared to low excreter patients (mean SDS -0.68; P = .016). In GA1, recommended long-term treatment is effective and allows for normal anthropometric long-term development up to adolescence, with gender- and excreter type-specific variations. Delayed ET and severe movement disorder result in poor anthropometric outcome.

Pontocerebellar Hypoplasia: a Pattern Recognition Approach.

Pontocerebellar hypoplasias (PCH) represent a heterogeneous group of very rare disorders with reduced volume of pons and cerebellum. The term is purely descriptive and does not imply a genetic progressive disease. Currently (as of Jan 01, 2020), 13 different types are listed in OMIM (Online Mendelian Inheritance in Man), associated with 19 different genes. However, a large group of similar imaging patterns is known, and it is unclear why some are labeled as PCH, while others are not. The latter include CASK- and VLDLR-associated disorders, some tubulinopathies, certain dystroglycanopathies, a few congenital disorders of glycosylation (CDG) syndromes, several forms associated with rare variants (e.g., DCK1, WDR81, ITPR1), and "cerebellar disruption of prematurity"-an acquired etiology. The objective of this paper is to elaborate a pattern recognition approach, mainly imaging-based, to facilitate a timely and accurate diagnosis, to narrow the differential diagnosis, and to enable targeted additional (genetic) investigations. We describe magnetic resonance imaging (MRI) findings and offer "checklists" for infratentorial findings (e.g., non-lobulated vermis, dragonfly pattern of the cerebellum, cerebellar cysts, brainstem kinking, longitudinal grooves along the brainstem, flat pons) as well as for supratentorial anomalies (e.g., agenesis of corpus callosum, optic atrophy, simplified gyral pattern, and hypomyelination). The clinical context and laboratory investigations need to be considered as well. We also provide a "checklist" for clinical features. A systematic analysis of imaging and clinical features can assist in narrowing the differential diagnosis and permitting more targeted genetic testing. Some imaging patterns are diagnostic.

CASPR2 autoimmunity in children expanding to mild encephalopathy with hypertension.

OBJECTIVE: To delineate autoimmune disease in association with contactin-associated protein 2 (CASPR2) antibodies in childhood, we reviewed the clinical phenotype of children with CASPR2 antibodies. METHODS: Retrospective assessment of patients recruited through laboratories specialized in autoimmune CNS disease. RESULTS: Ten children with serum CASPR2 antibodies were identified (age at manifestation 18 months to 17 years). Eight children with CASPR2 antibody titers from ≥1:160 to 1:5,120 had complex autoimmune diseases with an age-dependent clinical phenotype. Two children with structural epilepsy due to CNS malformations harbored nonspecific low-titer CASPR2 antibodies (serum titers 1:80). The clinical symptoms of the 8 children with high-titer CASPR2 antibodies were general weakness (8/8), sleep dysregulation (8/8), dysautonomia (8/8) encephalopathy (7/8), neuropathic pain (7/8), neuromyotonia (3/8), and flaccid paresis (3/8). Adolescents (3/8) showed pain, neuromyotonia, and encephalopathy, whereas younger children (5/8) displayed severe hypertension, encephalopathy, and hormonal dysfunction mimicking a systemic disease. No tumors were identified. Motor symptoms remitted with immunotherapy. Mild behavioral changes persisted in 1 child, and autism spectrum disorder was diagnosed during follow-up in a young boy. CONCLUSION: High-titer CASPR2 antibodies are associated with Morvan syndrome in children as young as 2 years. However, CASPR2 autoimmunity mimics systemic disease and hypertensive encephalopathy in children younger than 7 years. The outcome following immunotherapy was mostly favorable; long-term behavioral impairment may occur in younger children.

Folate Receptor α-Modified Nanoparticles for Targeting of the Central Nervous System.

Effective and timely delivery of therapeutic agents from the systemic circulation to the central nervous system (CNS) is often precluded by the blood-brain barrier (BBB) and the blood-cerebrospinal fluid barrier (BCSFB). A new pathway of folate uptake mediated by folate receptor alpha (FRα, molecular weight of 28.29 kg mol-1) occurring in various epithelial cells of the CNS (e.g., choroid plexus) was described. Aiming to investigate this mechanism for the delivery of nanomedicines to the CNS, in this work, we initially produced nanoparticles (NPs) made of a highly hydrophobic poly(ethylene glycol)-b-poly(ε-caprolactone) (PEG-b-PCL) block copolymer functionalized with an amine moiety in the edge of the PEG block by a simple nanoprecipitation method. Hydrophilic PEG blocks migrated to the NP surface during formation, exposing primary amine groups that were used to conjugate the targeting ligand, FRα. The size of the NPs was in the 58-98 nm range and standard deviation (S.D., a measure of the size population peak width) of 26-41 nm, as measured by dynamic light scattering (DLS). The FRα conjugation yield ranged between 50% and 75% (determined indirectly by the bicinchoninic acid protein assay). Pristine and FRα-modified NPs showed good compatibility with primary human choroid plexus epithelial cells (HCPEpiCs). The uptake of FRα-conjugated NPs by HCPEpiCs was qualitatively evaluated in vitro using inverted optical fluorescence and confocal microscopy. FRα-modified NPs were internalized by HCPEpiCs to a greater extent than the unmodified counterparts. Then, their permeability was characterized in standard and inverted HCPEpiC monolayers. In both cases, NPs surface modified with the FRα and complexed to folic acid (FA) showed significantly higher apparent permeability coefficient (Papp) values than the pristine ones. Finally, the biodistribution of unmodified and FRα-FA-modified NPs following intravenous (i.v.) administration was compared in ICR mice. Results indicated that conjugation of the FRα-FA complex to the NP surface promotes higher accumulation in the brain, highlighting the promise of FRα-FA-modified NPs to serve as a platform for the targeting of active molecules to the CNS from the systemic circulation.

Elucidation of the phenotypic spectrum and genetic landscape in primary and secondary microcephaly.

PURPOSE: Microcephaly is a sign of many genetic conditions but has been rarely systematically evaluated. We therefore comprehensively studied the clinical and genetic landscape of an unselected cohort of patients with microcephaly. METHODS: We performed clinical assessment, high-resolution chromosomal microarray analysis, exome sequencing, and functional studies in 62 patients (58% with primary microcephaly [PM], 27% with secondary microcephaly [SM], and 15% of unknown onset). RESULTS: We found severity of developmental delay/intellectual disability correlating with severity of microcephaly in PM, but not SM. We detected causative variants in 48.4% of patients and found divergent inheritance and variant pattern for PM (mainly recessive and likely gene-disrupting [LGD]) versus SM (all dominant de novo and evenly LGD or missense). While centrosome-related pathways were solely identified in PM, transcriptional regulation was the most frequently affected pathway in both SM and PM. Unexpectedly, we found causative variants in different mitochondria-related genes accounting for ~5% of patients, which emphasizes their role even in syndromic PM. Additionally, we delineated novel candidate genes involved in centrosome-related pathway (SPAG5, TEDC1), Wnt signaling (VPS26A, ZNRF3), and RNA trafficking (DDX1). CONCLUSION: Our findings enable improved evaluation and genetic counseling of PM and SM patients and further elucidate microcephaly pathways.

Mitochondrial biogenesis is transcriptionally repressed in lysosomal lipid storage diseases.

Perturbations in mitochondrial function and homeostasis are pervasive in lysosomal storage diseases, but the underlying mechanisms remain unknown. Here, we report a transcriptional program that represses mitochondrial biogenesis and function in lysosomal storage diseases Niemann-Pick type C (NPC) and acid sphingomyelinase deficiency (ASM), in patient cells and mouse tissues. This mechanism is mediated by the transcription factors KLF2 and ETV1, which are both induced in NPC and ASM patient cells. Mitochondrial biogenesis and function defects in these cells are rescued by the silencing of KLF2 or ETV1. Increased ETV1 expression is regulated by KLF2, while the increase of KLF2 protein levels in NPC and ASM stems from impaired signaling downstream sphingosine-1-phosphate receptor 1 (S1PR1), which normally represses KLF2. In patient cells, S1PR1 is barely detectable at the plasma membrane and thus unable to repress KLF2. This manuscript provides a mechanistic pathway for the prevalent mitochondrial defects in lysosomal storage diseases. Editorial note: This article has been through an editorial process in which the authors decide how to respond to the issues raised during peer review. The Reviewing Editor's assessment is that all the issues have been addressed (see decision letter).

Newborn screening: A disease-changing intervention for glutaric aciduria type 1.

OBJECTIVE: Untreated individuals with glutaric aciduria type 1 (GA1) commonly present with a complex, predominantly dystonic movement disorder (MD) following acute or insidious onset striatal damage. Implementation of GA1 into newborn screening (NBS) programs has improved the short-term outcome. It remains unclear, however, whether NBS changes the long-term outcome and which variables are predictive. METHODS: This prospective, observational, multicenter study includes 87 patients identified by NBS, 4 patients missed by NBS, and 3 women with GA1 identified by positive NBS results of their unaffected children. RESULTS: The study population comprises 98.3% of individuals with GA1 identified by NBS in Germany during 1999-2016. Overall, cumulative sensitivity of NBS is 95.6%, but it is lower (84%) for patients with low excreter phenotype. The neurologic outcome of patients missed by NBS is as poor as in the pre-NBS era, and the clinical phenotype of diagnosed patients depends on the quality of therapeutic interventions rather than noninterventional variables. Presymptomatic start of treatment according to current guideline recommendations clearly improves the neurologic outcome (MD: 7% of patients), whereas delayed emergency treatment results in acute onset MD (100%), and deviations from maintenance treatment increase the risk of insidious onset MD (50%). Independent of the neurologic phenotype, kidney function tends to decline with age, a nonneurologic manifestation not predicted by any variable included in this study. INTERPRETATION: NBS is a beneficial, disease-changing intervention for GA1. However, improved neurologic outcome critically depends on adherence to recommended therapy, whereas kidney dysfunction does not appear to be impacted by recommended therapy. Ann Neurol 2018;83:970-979.