Genome Sequencing for Cases Unsolved by Exome Sequencing: Identifying a Single-Exon Deletion in TBCK in a Case from 30 Years Ago.

In patients with neurodevelopmental disorders (NDDs), exome sequencing (ES), the diagnostic gold standard, reveals an underlying monogenic condition in only approximately 40% of cases. We report the case of a female patient with profound NDD who died 30 years ago at the age of 3 years and for whom genome sequencing (GS) now identified a single-exon deletion in TBCK previously missed by ExomeDepth, the copy number variation (CNV) detection algorithm in ES.Deoxyribonucleic acid (DNA) was extracted from frozen muscle tissue of the index patient and the parents' blood. Genome data were analyzed for structural variants and single nucleotide variants (SUVs)/indels as part of the Bavarian Genomes consortium project.Biallelic variants in TBCK, which are linked to the autosomal recessive disorder TBCK syndrome, were detected in the affected individual: a novel frameshift variant and a deletion of exon 23, previously established as common but underrecognized pathogenic variant in individuals with TBCK syndrome. While in the foregoing ES analysis, calling algorithms for (SNVs)/indels were able to identify the frameshift variant, ExomeDepth failed to call the intragenic deletion.Our case illustrates the added value of GS for the detection of single-exon deletions for which calling from ES data remains challenging and confirms that the deletion of exon 23 in TBCK may be underdiagnosed in patients with NDDs. Furthermore, it shows the importance of "molecular or genetic autopsy" allowing genetic risk counseling for family members as well as the end of a diagnostic odyssey of 30 years.

SNUPN deficiency causes a recessive muscular dystrophy due to RNA mis-splicing and ECM dysregulation.

SNURPORTIN-1, encoded by SNUPN, plays a central role in the nuclear import of spliceosomal small nuclear ribonucleoproteins. However, its physiological function remains unexplored. In this study, we investigate 18 children from 15 unrelated families who present with atypical muscular dystrophy and neurological defects. Nine hypomorphic SNUPN biallelic variants, predominantly clustered in the last coding exon, are ascertained to segregate with the disease. We demonstrate that mutant SPN1 failed to oligomerize leading to cytoplasmic aggregation in patients' primary fibroblasts and CRISPR/Cas9-mediated mutant cell lines. Additionally, mutant nuclei exhibit defective spliceosomal maturation and breakdown of Cajal bodies. Transcriptome analyses reveal splicing and mRNA expression dysregulation, particularly in sarcolemmal components, causing disruption of cytoskeletal organization in mutant cells and patient muscle tissues. Our findings establish SNUPN deficiency as the genetic etiology of a previously unrecognized subtype of muscular dystrophy and provide robust evidence of the role of SPN1 for muscle homeostasis.

A Homozygous PTRHD1 Missense Variant (p.Arg122Gln) in an Individual with Intellectual Disability, Generalized Epilepsy, and Juvenile Parkinsonism.

Biallelic variants in PTRHD1 have been associated with autosomal recessive intellectual disability, spasticity, and juvenile parkinsonism, with few reported cases. Here, we present the clinical and genetic findings of a female of Austrian origin exhibiting infantile neurodevelopmental abnormalities, intellectual disability, and childhood-onset parkinsonian features, consistent with the established phenotypic spectrum. Notably, she developed genetic generalized epilepsy at age 4, persisting into adulthood. Using diagnostic exome sequencing, we identified a homozygous missense variant (c.365G > A, p.(Arg122Gln)) in PTRHD1 (NM_001013663). In summary, our findings not only support the existing link between biallelic PTRHD1 variants and parkinsonism with neurodevelopmental abnormalities but also suggest a potential extension of the phenotypic spectrum to include generalized epilepsy.

De novo variants in RNF213 are associated with a clinical spectrum ranging from Leigh syndrome to early-onset stroke.

PURPOSE: RNF213, encoding a giant E3 ubiquitin ligase, has been recognized for its role as a key susceptibility gene for moyamoya disease. Case reports have also implicated specific variants in RNF213 with an early-onset form of moyamoya disease with full penetrance. We aimed to expand the phenotypic spectrum of monogenic RNF213-related disease and to evaluate genotype-phenotype correlations. METHODS: Patients were identified through reanalysis of exome sequencing data of an unselected cohort of unsolved pediatric cases and through GeneMatcher or ClinVar. Functional characterization was done by proteomics analysis and oxidative phosphorylation enzyme activities using patient-derived fibroblasts. RESULTS: We identified 14 individuals from 13 unrelated families with (de novo) missense variants in RNF213 clustering within or around the Really Interesting New Gene (RING) domain. Individuals presented either with early-onset stroke (n = 11) or with Leigh syndrome (n = 3). No genotype-phenotype correlation could be established. Proteomics using patient-derived fibroblasts revealed no significant differences between clinical subgroups. 3D modeling revealed a clustering of missense variants in the tertiary structure of RNF213 potentially affecting zinc-binding suggesting a gain-of-function or dominant negative effect. CONCLUSION: De novo missense variants in RNF213 clustering in the E3 RING or other regions affecting zinc-binding lead to an early-onset syndrome characterized by stroke or Leigh syndrome.

Next-generation sequencing and comprehensive data reassessment in 263 adult patients with neuromuscular disorders: insights into the gray zone of molecular diagnoses.

BACKGROUND: Neuromuscular disorders (NMDs) are heterogeneous conditions with a considerable fraction attributed to monogenic defects. Despite the advancements in genomic medicine, many patients remain without a diagnosis. Here, we investigate whether a comprehensive reassessment strategy improves the diagnostic outcomes. METHODS: We analyzed 263 patients with NMD phenotypes that underwent diagnostic exome or genome sequencing at our tertiary referral center between 2015 and 2023. We applied a comprehensive reassessment encompassing variant reclassification, re-phenotyping and NGS data reanalysis. Multivariable logistic regression was performed to identify predictive factors associated with a molecular diagnosis. RESULTS: Initially, a molecular diagnosis was identified in 53 cases (20%), while an additional 23 (9%) had findings of uncertain significance. Following comprehensive reassessment, the diagnostic yield increased to 23%, revealing 44 distinct monogenic etiologies. Reasons for newly obtained molecular diagnoses were variant reclassifications in 7 and NGS data reanalysis in 3 cases including one recently described disease-gene association (DNAJB4). Male sex reduced the odds of receiving a molecular diagnosis (OR 0.42; 95%CI 0.21-0.82), while a positive family history (OR 5.46; 95%CI 2.60-11.76) and a myopathy phenotype (OR 2.72; 95%CI 1.11-7.14) increased the likelihood. 7% were resolved through targeted genetic testing or classified as acquired etiologies. CONCLUSION: Our findings reinforce the use of NGS in NMDs of suspected monogenic origin. We show that a comprehensive reassessment enhances diagnostic accuracy. However, one needs to be aware that genetic diagnoses are often made with uncertainty and can even be downgraded based on new evidence.

Biallelic variants in SLC4A10 encoding a sodium-dependent bicarbonate transporter lead to a neurodevelopmental disorder.

PURPOSE: SLC4A10 encodes a plasma membrane-bound transporter, which mediates Na+-dependent HCO3- import, thus mediating net acid extrusion. Slc4a10 knockout mice show collapsed brain ventricles, an increased seizure threshold, mild behavioral abnormalities, impaired vision, and deafness. METHODS: Utilizing exome/genome sequencing in families with undiagnosed neurodevelopmental disorders and international data sharing, 11 patients from 6 independent families with biallelic variants in SLC4A10 were identified. Clinico-radiological and dysmorphology assessments were conducted. A minigene assay, localization studies, intracellular pH recordings, and protein modeling were performed to study the possible functional consequences of the variant alleles. RESULTS: The families harbor 8 segregating ultra-rare biallelic SLC4A10 variants (7 missense and 1 splicing). Phenotypically, patients present with global developmental delay/intellectual disability and central hypotonia, accompanied by variable speech delay, microcephaly, cerebellar ataxia, facial dysmorphism, and infrequently, epilepsy. Neuroimaging features range from some non-specific to distinct neuroradiological findings, including slit ventricles and a peculiar form of bilateral curvilinear nodular heterotopia. In silico analyses showed 6 of 7 missense variants affect evolutionarily conserved residues. Functional analyses supported the pathogenicity of 4 of 7 missense variants. CONCLUSION: We provide evidence that pathogenic biallelic SLC4A10 variants can lead to neurodevelopmental disorders characterized by variable abnormalities of the central nervous system, including altered brain ventricles, thus resembling several features observed in knockout mice.

Exome sequencing (ES) of a pediatric cohort with chronic endocrine diseases: a single-center study (within the framework of the TRANSLATE-NAMSE project).

BACKGROUND: Endocrine disorders are heterogeneous and include a significant number of rare monogenic diseases. METHODS: We performed exome sequencing (ES) in 106 children recruited from a single center within the TRANSLATE­NAMSE project. They were categorized into subgroups: proportionate short stature (PSS), disproportionate short stature (DSS), hypopituitarism (H), differences in sexual development (DSD), syndromic diseases (SD) and others. RESULTS: The overall diagnostic yield was 34.9% (n = 37/106), including 5 patients with variants in candidate genes, which have contributed to collaborations to identify gene-disease associations. The diagnostic yield varied significantly between subgroups: PSS: 16.6% (1/6); DSS: 18.8% (3/16); H: 17.1% (6/35); DSD: 37.5% (3/8); SD: 66.6% (22/33); others: 25% (2/8). Confirmed diagnoses included 75% ultrarare diseases. Three patients harbored more than one disease-causing variant, resulting in dual diagnoses. CONCLUSIONS: ES is an effective tool for genetic diagnosis in pediatric patients with complex endocrine diseases. An accurate phenotypic description, including comprehensive endocrine diagnostics, as well as the evaluation of variants in multidisciplinary case conferences involving geneticists, are necessary for personalized diagnostic care. Here, we illustrate the broad spectrum of genetic endocrinopathies that have led to the initiation of specific treatment, surveillance, and family counseling.

Genetic landscape of pediatric acute liver failure of indeterminate origin.

BACKGROUND AIMS: Pediatric acute liver failure (PALF) is a life-threatening condition. In Europe, main causes are viral infections (12-16%) and inherited metabolic diseases (14-28%). Yet, in up to 50% of cases the underlying etiology remains elusive, challenging clinical management, including liver transplantation. We systematically studied indeterminate PALF cases referred for genetic evaluation by whole-exome sequencing (WES), and analyzed phenotypic and biochemical markers, and the diagnostic yield of WES in this condition. METHODS: With this international, multicenter observational study, patients (0-18 y) with indeterminate PALF were analyzed by WES. Data on the clinical and biochemical phenotype were retrieved and systematically analyzed. RESULTS: In total, 260 indeterminate PALF patients from 19 countries were recruited between 2011 and 2022, of whom 59 had recurrent PALF (RALF). WES established a genetic diagnosis in 37% of cases (97/260). Diagnostic yield was highest in children with PALF in the first year of life (46%), and in children with RALF (64%). Thirty-six distinct disease genes were identified. Defects in NBAS (n=20), MPV17 (n=8) and DGUOK (n=7) were the most frequent findings. When categorizing, most frequent were mitochondrial diseases (45%), disorders of vesicular trafficking (28%) and cytosolic aminoacyl-tRNA synthetase deficiencies (10%). One-third of patients had a fatal outcome. Fifty-six patients received liver transplants. CONCLUSION: This study elucidates a large contribution of genetic causes in PALF of indeterminate origin with an increasing spectrum of disease entities. The high proportion of diagnosed cases and potential treatment implications argue for exome or in future rapid genome sequencing in PALF diagnostics.

Stretch-activated ion channel TMEM63B associates with developmental and epileptic encephalopathies and progressive neurodegeneration.

By converting physical forces into electrical signals or triggering intracellular cascades, stretch-activated ion channels allow the cell to respond to osmotic and mechanical stress. Knowledge of the pathophysiological mechanisms underlying associations of stretch-activated ion channels with human disease is limited. Here, we describe 17 unrelated individuals with severe early-onset developmental and epileptic encephalopathy (DEE), intellectual disability, and severe motor and cortical visual impairment associated with progressive neurodegenerative brain changes carrying ten distinct heterozygous variants of TMEM63B, encoding for a highly conserved stretch-activated ion channel. The variants occurred de novo in 16/17 individuals for whom parental DNA was available and either missense, including the recurrent p.Val44Met in 7/17 individuals, or in-frame, all affecting conserved residues located in transmembrane regions of the protein. In 12 individuals, hematological abnormalities co-occurred, such as macrocytosis and hemolysis, requiring blood transfusions in some. We modeled six variants (p.Val44Met, p.Arg433His, p.Thr481Asn, p.Gly580Ser, p.Arg660Thr, and p.Phe697Leu), each affecting a distinct transmembrane domain of the channel, in transfected Neuro2a cells and demonstrated inward leak cation currents across the mutated channel even in isotonic conditions, while the response to hypo-osmotic challenge was impaired, as were the Ca2+ transients generated under hypo-osmotic stimulation. Ectopic expression of the p.Val44Met and p.Gly580Cys variants in Drosophila resulted in early death. TMEM63B-associated DEE represents a recognizable clinicopathological entity in which altered cation conductivity results in a severe neurological phenotype with progressive brain damage and early-onset epilepsy associated with hematological abnormalities in most individuals.

Null and missense mutations of ERI1 cause a recessive phenotypic dichotomy in humans.

ERI1 is a 3'-to-5' exoribonuclease involved in RNA metabolic pathways including 5.8S rRNA processing and turnover of histone mRNAs. Its biological and medical significance remain unclear. Here, we uncover a phenotypic dichotomy associated with bi-allelic ERI1 variants by reporting eight affected individuals from seven unrelated families. A severe spondyloepimetaphyseal dysplasia (SEMD) was identified in five affected individuals with missense variants but not in those with bi-allelic null variants, who showed mild intellectual disability and digital anomalies. The ERI1 missense variants cause a loss of the exoribonuclease activity, leading to defective trimming of the 5.8S rRNA 3' end and a decreased degradation of replication-dependent histone mRNAs. Affected-individual-derived induced pluripotent stem cells (iPSCs) showed impaired in vitro chondrogenesis with downregulation of genes regulating skeletal patterning. Our study establishes an entity previously unreported in OMIM and provides a model showing a more severe effect of missense alleles than null alleles within recessive genotypes, suggesting a key role of ERI1-mediated RNA metabolism in human skeletal patterning and chondrogenesis.

Episignature analysis of moderate effects and mosaics.

DNA methylation classifiers ("episignatures") help to determine the pathogenicity of variants of uncertain significance (VUS). However, their sensitivity is limited due to their training on unambiguous cases with strong-effect variants so that the classification of variants with reduced effect size or in mosaic state may fail. Moreover, episignature evaluation of mosaics as a function of their degree of mosaicism has not been developed so far. We improved episignatures with respect to three categories. Applying (i) minimum-redundancy-maximum-relevance feature selection we reduced their length by up to one order of magnitude without loss of accuracy. Performing (ii) repeated re-training of a support vector machine classifier by step-wise inclusion of cases in the training set that reached probability scores larger than 0.5, we increased the sensitivity of the episignature-classifiers by 30%. In the newly diagnosed patients we confirmed the association between DNA methylation aberration and age at onset of KMT2B-deficient dystonia. Moreover, we found evidence for allelic series, including KMT2B-variants with moderate effects and comparatively mild phenotypes such as late-onset focal dystonia. Retrained classifiers also can detect mosaics that previously remained below the 0.5-threshold, as we showed for KMT2D-associated Kabuki syndrome. Conversely, episignature-classifiers are able to revoke erroneous exome calls of mosaicism, as we demonstrated by (iii) comparing presumed mosaic cases with a distribution of artificial in silico-mosaics that represented all the possible variation in degree of mosaicism, variant read sampling and methylation analysis.

PSMC3 proteasome subunit variants are associated with neurodevelopmental delay and type I interferon production.

A critical step in preserving protein homeostasis is the recognition, binding, unfolding, and translocation of protein substrates by six AAA-ATPase proteasome subunits (ATPase-associated with various cellular activities) termed PSMC1-6, which are required for degradation of proteins by 26S proteasomes. Here, we identified 15 de novo missense variants in the PSMC3 gene encoding the AAA-ATPase proteasome subunit PSMC3/Rpt5 in 23 unrelated heterozygous patients with an autosomal dominant form of neurodevelopmental delay and intellectual disability. Expression of PSMC3 variants in mouse neuronal cultures led to altered dendrite development, and deletion of the PSMC3 fly ortholog Rpt5 impaired reversal learning capabilities in fruit flies. Structural modeling as well as proteomic and transcriptomic analyses of T cells derived from patients with PSMC3 variants implicated the PSMC3 variants in proteasome dysfunction through disruption of substrate translocation, induction of proteotoxic stress, and alterations in proteins controlling developmental and innate immune programs. The proteostatic perturbations in T cells from patients with PSMC3 variants correlated with a dysregulation in type I interferon (IFN) signaling in these T cells, which could be blocked by inhibition of the intracellular stress sensor protein kinase R (PKR). These results suggest that proteotoxic stress activated PKR in patient-derived T cells, resulting in a type I IFN response. The potential relationship among proteosome dysfunction, type I IFN production, and neurodevelopment suggests new directions in our understanding of pathogenesis in some neurodevelopmental disorders.

Biallelic variants in CRIPT cause a Rothmund-Thomson-like syndrome with increased cellular senescence.

PURPOSE: Rothmund-Thomson syndrome (RTS) is characterized by poikiloderma, sparse hair, small stature, skeletal defects, cancer, and cataracts, resembling features of premature aging. RECQL4 and ANAPC1 are the 2 known disease genes associated with RTS in >70% of cases. We describe RTS-like features in 5 individuals with biallelic variants in CRIPT (OMIM 615789). METHODS: Two newly identified and 4 published individuals with CRIPT variants were systematically compared with those with RTS using clinical data, computational analysis of photographs, histologic analysis of skin, and cellular studies on fibroblasts. RESULTS: All CRIPT individuals fulfilled the diagnostic criteria for RTS and additionally had neurodevelopmental delay and seizures. Using computational gestalt analysis, CRIPT individuals showed greatest facial similarity with individuals with RTS. Skin biopsies revealed a high expression of senescence markers (p53/p16/p21) and the senescence-associated ß-galactosidase activity was elevated in CRIPT-deficient fibroblasts. RECQL4- and CRIPT-deficient fibroblasts showed an unremarkable mitotic progression and unremarkable number of mitotic errors and no or only mild sensitivity to genotoxic stress by ionizing radiation, mitomycin C, hydroxyurea, etoposide, and potassium bromate. CONCLUSION: CRIPT causes an RTS-like syndrome associated with neurodevelopmental delay and epilepsy. At the cellular level, RECQL4- and CRIPT-deficient cells display increased senescence, suggesting shared molecular mechanisms leading to the clinical phenotypes.

De novo missense variants in RRAGC lead to a fatal mTORopathy of early childhood.

PURPOSE: Mechanistic target of rapamycin (mTOR) complex 1 (mTORC1) regulates cell growth in response to nutritional status. Central to the mTORC1 function is the Rag-GTPase heterodimer. One component of the Rag heterodimer is RagC (Ras-related GTP-binding protein C), which is encoded by the RRAGC gene. METHODS: Genetic testing via trio exome sequencing was applied to identify the underlying disease cause in 3 infants with dilated cardiomyopathy, hepatopathy, and brain abnormalities, including pachygyria, polymicrogyria, and septo-optic dysplasia. Studies in patient-derived skin fibroblasts and in a HEK293 cell model were performed to investigate the cellular consequences. RESULTS: We identified 3 de novo missense variants in RRAGC (NM_022157.4: c.269C>A, p.(Thr90Asn), c.353C>T, p.(Pro118Leu), and c.343T>C, p.(Trp115Arg)), which were previously reported as occurring somatically in follicular lymphoma. Studies of patient-derived fibroblasts carrying the p.(Thr90Asn) variant revealed increased cell size, as well as dysregulation of mTOR-related p70S6K (ribosomal protein S6 kinase 1) and transcription factor EB signaling. Moreover, subcellular localization of mTOR was decoupled from metabolic state. We confirmed the key findings for all RRAGC variants described in this study in a HEK293 cell model. CONCLUSION: The above results are in line with a constitutive overactivation of the mTORC1 pathway. Our study establishes de novo missense variants in RRAGC as cause of an early-onset mTORopathy with unfavorable prognosis.

The constitutional gain-of-function variant p.Glu1099Lys in NSD2 is associated with a novel syndrome.

NSD2 dimethylates histone H3 at lysine 36 (H3K36me2) and is located in the Wolf-Hirschhorn syndrome (WHS) critical region. Recent descriptions have delineated loss-of-function (LoF) variants in NSD2 with a distinct disorder. The oncogenic missense variant p.Glu1099Lys occurs somatically in leukemia and has a gain-of-function (GoF) effect. We describe two individuals carrying p.Glu1099Lys as heterozygous de novo germline variant identified by exome sequencing (ES) of blood DNA and subsequently confirmed in two ectodermal tissues. Clinically, these individuals are characterized by intellectual disability, coarse/ square facial gestalt, abnormalities of the hands, and organomegaly. Public cell lines with NSD2 GoF variants had increased K36me2, DNA promoter methylation, and dysregulated RNA expression. NSD2 GoF caused by p.Glu1099Lys is associated with a novel phenotype different from WHS and Rauch-Steindl syndrome (RAUST).

Loss-of-function variants in MYCBP2 cause neurobehavioural phenotypes and corpus callosum defects.

The corpus callosum is a bundle of axon fibres that connects the two hemispheres of the brain. Neurodevelopmental disorders that feature dysgenesis of the corpus callosum as a core phenotype offer a valuable window into pathology derived from abnormal axon development. Here, we describe a cohort of eight patients with a neurodevelopmental disorder characterized by a range of deficits including corpus callosum abnormalities, developmental delay, intellectual disability, epilepsy and autistic features. Each patient harboured a distinct de novo variant in MYCBP2, a gene encoding an atypical really interesting new gene (RING) ubiquitin ligase and signalling hub with evolutionarily conserved functions in axon development. We used CRISPR/Cas9 gene editing to introduce disease-associated variants into conserved residues in the Caenorhabditis elegans MYCBP2 orthologue, RPM-1, and evaluated functional outcomes in vivo. Consistent with variable phenotypes in patients with MYCBP2 variants, C. elegans carrying the corresponding human mutations in rpm-1 displayed axonal and behavioural abnormalities including altered habituation. Furthermore, abnormal axonal accumulation of the autophagy marker LGG-1/LC3 occurred in variants that affect RPM-1 ubiquitin ligase activity. Functional genetic outcomes from anatomical, cell biological and behavioural readouts indicate that MYCBP2 variants are likely to result in loss of function. Collectively, our results from multiple human patients and CRISPR gene editing with an in vivo animal model support a direct link between MYCBP2 and a human neurodevelopmental spectrum disorder that we term, MYCBP2-related developmental delay with corpus callosum defects (MDCD).

Case Report: Stüve-Wiedemann syndrome-a rare cause of persistent pulmonary hypertension of the newborn.

Introduction: Persistent pulmonary hypertension of the newborn (PPHN) is a life-threatening condition characterized by hypoxemia due to elevated pulmonary vascular resistance. PPHN commonly arises secondary to various underlying conditions, including infection, meconium aspiration, and respiratory distress syndrome. Management includes pulmonary vasodilators, mechanical ventilation, oxygen supplementation, vasopressors, and volume replacement. Stüve-Wiedemann syndrome (SWS), a rare genetic disorder characterized by bone dysplasia, respiratory distress, hyperthermia, and swallowing difficulties, may present with pulmonary hypertension, indicating a poor prognosis. Case description: A term female neonate presented with secondary respiratory failure and severe PPHN of unknown etiology on the second day of life, necessitating intubation. Clinical findings included facial dysmorphia, camptodactyly, skeletal anomalies, and generalized muscular hypotonia. High-frequency oscillation ventilation and surfactant administration yielded marginal improvement. On the third day of life, a severe pulmonary hypertensive crisis necessitated inhaled and systemic pulmonary vasodilators along with volume and catecholamine therapy. Whole exome sequencing revealed a homozygous mutation in the leukemia inhibitory factor receptor (LIFR) gene, consistent with Stüve-Wiedemann syndrome. Discussion/conclusion: The case underscores the importance of considering and prompting evaluation of rare genetic causes in the differential diagnosis of PPHN, especially when other abnormalities are present and conventional therapies prove inadequate. Therapeutic strategies must account for the different pathophysiology of primary PPHN including vascular remodeling, as seen in SWS, which may not respond to pulmonary vasodilators typically employed in secondary PPHN due to vasoconstriction. In this case, the patient responded well to treatment for primary PPHN, but the use of high-frequency oscillation ventilation and surfactant was not helpful.

Mitochondrial diseases mimicking autoimmune diseases of the CNS and good response to steroids initially.

INTRODUCTION: Neuroimmunological diseases such as autoimmune encephalitis (AE) or acquired demyelinating syndromes (ADS), can present with neurological symptoms and imaging features that are indistinguishable from mitochondrial diseases (MD) in particular at initial presentation. METHODS: Retrospective analysis of the clinical, laboratory and neuroimaging features of five patients who presented with signs of a neuroimmunological disease but all had pathological pathogenic variants in genes related to mitochondrial energy metabolism. RESULTS: Four patients presented with an acute neurological episode reminiscent of a possible AE and one patient with a suspected ADS at initial presentation. MRI findings were compatible with neuroimmunological diseases in all patients. In two children cerebrospinal fluid (CSF) studies revealed a mildly elevated cell count, two had elevated CSF lactate, none had oligoclonal bands (OCBs). All patients improved rapidly with intravenous steroids or immunoglobulins. Four patients had one or more relapses. Three patients showed worsening of their neurological symptoms with subsequent episodes and one patient died. Relapses in conjunction with new and progressive neurological symptoms, led to additional work-up which finally resulted in different genetic diagnosis of MD in all patients (MT-TL1, MT-ND5, APOA1-BP, HPDL, POLG). DISCUSSION: We would like to draw attention to a subset of patients with MD initially presenting with signs and symptoms mimicking neuroimmunological. Absence of CSF pleocytosis, elevated CSF lactate and progressive, relapsing course should trigger further (genetic) investigations in search of a MD even in patients with good response initially to immunomodulating therapies.

Heterozygous variants in MYH10 associated with neurodevelopmental disorders and congenital anomalies with evidence for primary cilia-dependent defects in Hedgehog signaling.

PURPOSE: Nonmuscle myosin II complexes are master regulators of actin dynamics that play essential roles during embryogenesis with vertebrates possessing 3 nonmuscle myosin II heavy chain genes, MYH9, MYH10, and MYH14. As opposed to MYH9 and MYH14, no recognizable disorder has been associated with MYH10. We sought to define the clinical characteristics and molecular mechanism of a novel autosomal dominant disorder related to MYH10. METHODS: An international collaboration identified the patient cohort. CAS9-mediated knockout cell models were used to explore the mechanism of disease pathogenesis. RESULTS: We identified a cohort of 16 individuals with heterozygous MYH10 variants presenting with a broad spectrum of neurodevelopmental disorders and variable congenital anomalies that affect most organ systems and were recapitulated in animal models of altered MYH10 activity. Variants were typically de novo missense changes with clustering observed in the motor domain. MYH10 knockout cells showed defects in primary ciliogenesis and reduced ciliary length with impaired Hedgehog signaling. MYH10 variant overexpression produced a dominant-negative effect on ciliary length. CONCLUSION: These data presented a novel genetic cause of isolated and syndromic neurodevelopmental disorders related to heterozygous variants in the MYH10 gene with implications for disrupted primary cilia length control and altered Hedgehog signaling in disease pathogenesis.

A reverse genetics and genomics approach to gene paralog function and disease: Myokymia and the juxtaparanode.

The leucine-rich glioma-inactivated (LGI) family consists of four highly conserved paralogous genes, LGI1-4, that are highly expressed in mammalian central and/or peripheral nervous systems. LGI1 antibodies are detected in subjects with autoimmune limbic encephalitis and peripheral nerve hyperexcitability syndromes (PNHSs) such as Isaacs and Morvan syndromes. Pathogenic variations of LGI1 and LGI4 are associated with neurological disorders as disease traits including familial temporal lobe epilepsy and neurogenic arthrogryposis multiplex congenita 1 with myelin defects, respectively. No human disease has been reported associated with either LGI2 or LGI3. We implemented exome sequencing and family-based genomics to identify individuals with deleterious variants in LGI3 and utilized GeneMatcher to connect practitioners and researchers worldwide to investigate the clinical and electrophysiological phenotype in affected subjects. We also generated Lgi3-null mice and performed peripheral nerve dissection and immunohistochemistry to examine the juxtaparanode LGI3 microarchitecture. As a result, we identified 16 individuals from eight unrelated families with loss-of-function (LoF) bi-allelic variants in LGI3. Deep phenotypic characterization showed LGI3 LoF causes a potentially clinically recognizable PNHS trait characterized by global developmental delay, intellectual disability, distal deformities with diminished reflexes, visible facial myokymia, and distinctive electromyographic features suggestive of motor nerve instability. Lgi3-null mice showed reduced and mis-localized Kv1 channel complexes in myelinated peripheral axons. Our data demonstrate bi-allelic LoF variants in LGI3 cause a clinically distinguishable disease trait of PNHS, most likely caused by disturbed Kv1 channel distribution in the absence of LGI3.