Mechanism of KMT5B haploinsufficiency in neurodevelopment in humans and mice.

Pathogenic variants in KMT5B, a lysine methyltransferase, are associated with global developmental delay, macrocephaly, autism, and congenital anomalies (OMIM# 617788). Given the relatively recent discovery of this disorder, it has not been fully characterized. Deep phenotyping of the largest (n = 43) patient cohort to date identified that hypotonia and congenital heart defects are prominent features that were previously not associated with this syndrome. Both missense variants and putative loss-of-function variants resulted in slow growth in patient-derived cell lines. KMT5B homozygous knockout mice were smaller in size than their wild-type littermates but did not have significantly smaller brains, suggesting relative macrocephaly, also noted as a prominent clinical feature. RNA sequencing of patient lymphoblasts and Kmt5b haploinsufficient mouse brains identified differentially expressed pathways associated with nervous system development and function including axon guidance signaling. Overall, we identified additional pathogenic variants and clinical features in KMT5B-related neurodevelopmental disorder and provide insights into the molecular mechanisms of the disorder using multiple model systems.

Novel pathogenic variants and quantitative phenotypic analyses of Robinow syndrome: WNT signaling perturbation and phenotypic variability.

Robinow syndrome (RS) is a genetically heterogeneous disorder with six genes that converge on the WNT/planar cell polarity (PCP) signaling pathway implicated (DVL1, DVL3, FZD2, NXN, ROR2, and WNT5A). RS is characterized by skeletal dysplasia and distinctive facial and physical characteristics. To further explore the genetic heterogeneity, paralog contribution, and phenotypic variability of RS, we investigated a cohort of 22 individuals clinically diagnosed with RS from 18 unrelated families. Pathogenic or likely pathogenic variants in genes associated with RS or RS phenocopies were identified in all 22 individuals, including the first variant to be reported in DVL2. We retrospectively collected medical records of 16 individuals from this cohort and extracted clinical descriptions from 52 previously published cases. We performed Human Phenotype Ontology (HPO) based quantitative phenotypic analyses to dissect allele-specific phenotypic differences. Individuals with FZD2 variants clustered into two groups with demonstrable phenotypic differences between those with missense and truncating alleles. Probands with biallelic NXN variants clustered together with the majority of probands carrying DVL1, DVL2, and DVL3 variants, demonstrating no phenotypic distinction between the NXN-autosomal recessive and dominant forms of RS. While phenotypically similar diseases on the RS differential matched through HPO analysis, clustering using phenotype similarity score placed RS-associated phenotypes in a unique cluster containing WNT5A, FZD2, and ROR2 apart from non-RS-associated paralogs. Through human phenotype analyses of this RS cohort and OMIM clinical synopses of Mendelian disease, this study begins to tease apart specific biologic roles for non-canonical WNT-pathway proteins.

Expanding the phenotype of ASXL3-related syndrome: A comprehensive description of 45 unpublished individuals with inherited and de novo pathogenic variants in ASXL3.

The study aimed at widening the clinical and genetic spectrum of ASXL3-related syndrome, a neurodevelopmental disorder, caused by truncating variants in the ASXL3 gene. In this international collaborative study, we have undertaken a detailed clinical and molecular analysis of 45 previously unpublished individuals with ASXL3-related syndrome, as well as a review of all previously published individuals. We have reviewed the rather limited functional characterization of pathogenic variants in ASXL3 and discuss current understanding of the consequences of the different ASXL3 variants. In this comprehensive analysis of ASXL3-related syndrome, we define its natural history and clinical evolution occurring with age. We report familial ASXL3 pathogenic variants, characterize the phenotype in mildly affected individuals and discuss nonpenetrance. We also discuss the role of missense variants in ASXL3. We delineate a variable but consistent phenotype. The most characteristic features are neurodevelopmental delay with consistently limited speech, significant neuro-behavioral issues, hypotonia, and feeding difficulties. Distinctive features include downslanting palpebral fissures, hypertelorism, tubular nose with a prominent nasal bridge, and low-hanging columella. The presented data will inform clinical management of individuals with ASXL3-related syndrome and improve interpretation of new ASXL3 sequence variants.

Expanding the genotypic and phenotypic spectrum in a diverse cohort of 104 individuals with Wiedemann-Steiner syndrome.

Wiedemann-Steiner syndrome (WSS) is an autosomal dominant disorder caused by monoallelic variants in KMT2A and characterized by intellectual disability and hypertrichosis. We performed a retrospective, multicenter, observational study of 104 individuals with WSS from five continents to characterize the clinical and molecular spectrum of WSS in diverse populations, to identify physical features that may be more prevalent in White versus Black Indigenous People of Color individuals, to delineate genotype-phenotype correlations, to define developmental milestones, to describe the syndrome through adulthood, and to examine clinicians' differential diagnoses. Sixty-nine of the 82 variants (84%) observed in the study were not previously reported in the literature. Common clinical features identified in the cohort included: developmental delay or intellectual disability (97%), constipation (63.8%), failure to thrive (67.7%), feeding difficulties (66.3%), hypertrichosis cubiti (57%), short stature (57.8%), and vertebral anomalies (46.9%). The median ages at walking and first words were 20 months and 18 months, respectively. Hypotonia was associated with loss of function (LoF) variants, and seizures were associated with non-LoF variants. This study identifies genotype-phenotype correlations as well as race-facial feature associations in an ethnically diverse cohort, and accurately defines developmental trajectories, medical comorbidities, and long-term outcomes in individuals with WSS.

Predominant and novel de novo variants in 29 individuals with ALG13 deficiency: Clinical description, biomarker status, biochemical analysis, and treatment suggestions.

Asparagine-linked glycosylation 13 homolog (ALG13) encodes a nonredundant, highly conserved, X-linked uridine diphosphate (UDP)-N-acetylglucosaminyltransferase required for the synthesis of lipid linked oligosaccharide precursor and proper N-linked glycosylation. De novo variants in ALG13 underlie a form of early infantile epileptic encephalopathy known as EIEE36, but given its essential role in glycosylation, it is also considered a congenital disorder of glycosylation (CDG), ALG13-CDG. Twenty-four previously reported ALG13-CDG cases had de novo variants, but surprisingly, unlike most forms of CDG, ALG13-CDG did not show the anticipated glycosylation defects, typically detected by altered transferrin glycosylation. Structural homology modeling of two recurrent de novo variants, p.A81T and p.N107S, suggests both are likely to impact the function of ALG13. Using a corresponding ALG13-deficient yeast strain, we show that expressing yeast ALG13 with either of the highly conserved hotspot variants rescues the observed growth defect, but not its glycosylation abnormality. We present molecular and clinical data on 29 previously unreported individuals with de novo variants in ALG13. This more than doubles the number of known cases. A key finding is that a vast majority of the individuals presents with West syndrome, a feature shared with other CDG types. Among these, the initial epileptic spasms best responded to adrenocorticotropic hormone or prednisolone, while clobazam and felbamate showed promise for continued epilepsy treatment. A ketogenic diet seems to play an important role in the treatment of these individuals.

SCN3A-Related Neurodevelopmental Disorder: A Spectrum of Epilepsy and Brain Malformation.

OBJECTIVE: Pathogenic variants in SCN3A, encoding the voltage-gated sodium channel subunit Nav1.3, cause severe childhood onset epilepsy and malformation of cortical development. Here, we define the spectrum of clinical, genetic, and neuroimaging features of SCN3A-related neurodevelopmental disorder. METHODS: Patients were ascertained via an international collaborative network. We compared sodium channels containing wild-type versus variant Nav1.3 subunits coexpressed with ß1 and ß2 subunits using whole-cell voltage clamp electrophysiological recordings in a heterologous mammalian system (HEK-293T cells). RESULTS: Of 22 patients with pathogenic SCN3A variants, most had treatment-resistant epilepsy beginning in the first year of life (16/21, 76%; median onset, 2 weeks), with severe or profound developmental delay (15/20, 75%). Many, but not all (15/19, 79%), exhibited malformations of cortical development. Pathogenic variants clustered in transmembrane segments 4 to 6 of domains II to IV. Most pathogenic missense variants tested (10/11, 91%) displayed gain of channel function, with increased persistent current and/or a leftward shift in the voltage dependence of activation, and all variants associated with malformation of cortical development exhibited gain of channel function. One variant (p.Ile1468Arg) exhibited mixed effects, with gain and partial loss of function. Two variants demonstrated loss of channel function. INTERPRETATION: Our study defines SCN3A-related neurodevelopmental disorder along a spectrum of severity, but typically including epilepsy and severe or profound developmental delay/intellectual disability. Malformations of cortical development are a characteristic feature of this unusual channelopathy syndrome, present in >75% of affected individuals. Gain of function at the channel level in developing neurons is likely an important mechanism of disease pathogenesis. ANN NEUROL 2020;88:348-362.

Further delineation of the clinical spectrum of KAT6B disorders and allelic series of pathogenic variants.

PURPOSE: Genitopatellar syndrome and Say-Barber-Biesecker-Young-Simpson syndrome are caused by variants in the KAT6B gene and are part of a broad clinical spectrum called KAT6B disorders, whose variable expressivity is increasingly being recognized. METHODS: We herein present the phenotypes of 32 previously unreported individuals with a molecularly confirmed diagnosis of a KAT6B disorder, report 24 new pathogenic KAT6B variants, and review phenotypic information available on all published individuals with this condition. We also suggest a classification of clinical subtypes within the KAT6B disorder spectrum. RESULTS: We demonstrate that cerebral anomalies, optic nerve hypoplasia, neurobehavioral difficulties, and distal limb anomalies other than long thumbs and great toes, such as polydactyly, are more frequently observed than initially reported. Intestinal malrotation and its serious consequences can be present in affected individuals. Additionally, we identified four children with Pierre Robin sequence, four individuals who had increased nuchal translucency/cystic hygroma prenatally, and two fetuses with severe renal anomalies leading to renal failure. We also report an individual in which a pathogenic variant was inherited from a mildly affected parent. CONCLUSION: Our work provides a comprehensive review and expansion of the genotypic and phenotypic spectrum of KAT6B disorders that will assist clinicians in the assessment, counseling, and management of affected individuals.

Mosaicism in ASXL3-related syndrome: Description of five patients from three families.

De novo pathogenic variants in the additional sex combs-like 3 (ASXL3) gene cause a rare multi-systemic neurodevelopmental disorder. There is growing evidence that germline and somatic mosaicism are more common and play a greater role in genetic disorders than previously acknowledged. There is one previous report of ASXL3-related syndrome caused by de novo pathogenic variants in two siblings suggesting gonadal mosaicism. In this report, we present five patients with ASXL3-related syndrome, describing two families comprising two non-twin siblings harbouring apparent de novo pathogenic variants in ASXL3. Parents were clinically unaffected and there was no evidence of mosaicism from genomic DNA on exome-trio data, suggesting germline mosaicism in one of the parents. We also describe clinical details of a patient with typical features of ASXL3-related syndrome and mosaic de novo pathogenic variant in ASXL3 in 30-35% of both blood and saliva sample on trio-exome sequencing. We expand the known genetic basis of ASXL3-related syndromes and discuss mosaicism as a disease mechanism in five patients from three unrelated families. The findings of this report highlight the importance of taking gonadal mosaicism into consideration when counselling families regarding recurrence risk. We also discuss postzygotic mosaicism as a cause of fully penetrant ASXL3-related syndrome.

The natural history of infantile mitochondrial DNA depletion syndrome due to RRM2B deficiency.

PURPOSE: Mitochondrial DNA (mtDNA) depletion syndrome (MDDS) encompasses a group of genetic disorders of mtDNA maintenance. Mutation of RRM2B is an uncommon cause of infantile-onset encephalomyopathic MDDS. Here we describe the natural history of this disease. METHODS: Multinational series of new genetically confirmed cases from six pediatric centers. RESULTS: Nine new cases of infantile-onset RRM2B deficiency, and 22 previously published cases comprised a total cohort of 31 patients. Infants presented at a mean of 1.95 months with truncal hypotonia, generalized weakness, and faltering growth. Seizures evolved in 39% at a mean of 3.1 months. Non-neurological manifestations included respiratory distress/failure (58%), renal tubulopathy (55%), sensorineural hearing loss (36%), gastrointestinal disturbance (32%), eye abnormalities (13%), and anemia (13%). Laboratory features included elevated lactate (blood, cerebrospinal fluid (CSF), urine, magnetic resonance (MR), spectroscopy), ragged-red and cytochrome c oxidase-deficient fibers, lipid myopathy, and multiple oxidative phosphorylation enzyme deficiencies in skeletal muscle. Eight new RRM2B variants were identified. Patients with biallelic truncating variants had the worst survival. Overall survival was 29% at 6 months and 16% at 1 year. CONCLUSIONS: Infantile-onset MDDS due to RRM2B deficiency is a severe disorder with characteristic clinical features and extremely poor prognosis. Presently management is supportive as there is no effective treatment. Novel treatments are urgently needed.

EPG5 Variants with Modest Functional Impact Result in an Ameliorated and Primarily Neurological Phenotype in a 3.5-Year-Old Patient with Vici Syndrome.

Congenital disorders of autophagy are multisystem disorders with significant neurological involvement. Ectopic p-granules protein 5 (EPG5)-associated Vici syndrome is a prototypical congenital disorder of autophagy and presents with the cardinal features of agenesis of the corpus callosum, cataracts, cardiomyopathy, immunodeficiency, and oculocutaneous hypopigmentation. The majority of EPG5 variants leading to Vici syndrome are null alleles with only a few missense variants published to date. Here we report a 3.5-year-old male with compound heterozygous EPG5 variants [NM_020964.2: c.772G > T/c.5943-9_5943-5del]. His clinical presentation deviates notably from classic Vici syndrome with a lack of hypopigmentation, cataracts, immunodeficiency, cardiomyopathy, or failure to thrive. Neurological manifestations within the known disease spectrum include early-onset global developmental delay, hypotonia, and postnatal microcephaly. Seizures, hearing loss, or optic nerve atrophy are absent, however. Magnetic resonance imaging demonstrates a thin but fully formed corpus callosum. Based on the ameliorated and primarily neurological phenotype, we hypothesized that the functional impact of the EPG5 variants present would be milder with a higher amount of residual EPG5 expression. Analyses of EPG5 messenger ribonucleic acid (mRNA) in the patient and his parents were performed to examine expression level and splicing; mRNA from a healthy control and a patient with classic Vici syndrome was also included. Aberrant splicing due to the intronic mutation was detected, but no loss of expression. In contrast, we observed a 50% reduction in mRNA expression in classic Vici syndrome patient fibroblasts. These results support a model of disease severity, which correlates to the dosage of EPG5 expression.

Genotype-first analysis of a generally healthy population cohort supports genetic testing for diagnosis of hereditary angioedema of unknown cause.

BACKGROUND: Hereditary angioedema (HAE) is a potentially life-threatening group of conditions that is often underdiagnosed or misdiagnosed. As HAE is typically diagnosed by detecting C1 inhibitor deficiency, there is a critical need for methods that can identify affected individuals with normal C1 inhibitor. The recent discovery of associations between PLG K330E and ANGPT1 A119S and HAE of unknown genetic cause (HAE-U), has raised the possibility that genetic evaluation could be used to diagnose HAE-U in patients with unexplained angioedema or non-confirmatory laboratory testing. CASE PRESENTATION: We analyzed genome sequences from a generally healthy population cohort of 2820 adults and identified PLG K330E in one individual. Subsequent review of this participant's medical history revealed symptoms clinically attributed to allergy of unknown etiology but that are consistent with published descriptions of HAE patients carrying the PLG K330E variant. The participant, a 31 year old female, reported lip and tongue angioedema, without wheals, which did not respond to treatment with steroids or antihistamines. CONCLUSIONS: The genotype-first approach demonstrated that detection of PLG K330E in undiagnosed or misdiagnosed individuals can identify patients actually affected with HAE-U. The genetic diagnosis will facilitate selection of appropriate treatment, discontinuation of therapies ineffective for this condition, and timely diagnosis of affected family members. The results support a role of PLG K330E in the pathogenesis of HAE and suggest that genetic testing be considered as an approach to diagnose patients with unexplained angioedema.

Pilot of urgent care center evaluation for acute coronary syndrome.

OBJECTIVES: Patients with chest pain and concern for potential coronary ischemia are frequently referred to the emergency department (ED), resulting in substantial resource utilization and cost. The objective of this study was to implement a protocol for urgent care center (UCC) evaluation of potential acute coronary syndrome (ACS) and describe its performance. STUDY DESIGN: This is a descriptive, retrospective review of consecutive cases included in a protocol for UCC evaluation of ACS. METHODS: Consecutive patient encounters from 4 urgent care facilities of our regional integrated health system were reviewed from a period spanning 4.5 months of the 2017 calendar year. The primary outcome was avoidance of an ED visit within 30 days of the index visit, and the primary safety outcome was serious adverse events (AEs) occurring in the UCC setting. RESULTS: There were 802 patients evaluated, with a median age of 55 years, and 58% were female. Seventy-three (9.1%) patients were referred to the ED or hospitalized for any reason at the index visit, 10 (1.2%) of whom were ultimately diagnosed with ACS. Within 30 days, 56 (7.7%) of the remaining 729 patients had ED visits or hospitalization for any reason, 2 (0.2%) of whom received a diagnosis of ACS. Overall, 673 (83.9%) patients were managed without any ED visit. No serious AEs were recorded. CONCLUSIONS: Our initial pilot data demonstrate the feasibility of an outpatient UCC evaluation for ACS without refuting the underlying premise of safety.

Endosomal trafficking defects in patient cells with KIAA1109 biallelic variants.

The uncharacterized gene KIAA1 109 has recently been associated with a congenital neurological malformation disorder that variably presents with arthrogryposis, craniofacial and/or cardiac abnormalities. We have identified two additional patients with compound heterozygous KIAA1109 variants presenting with the same neurological malformations. The mechanism whereby KIAA1109 loss of function causes this spectrum of disorders was the primary focus of our studies. We hypothesized that KIAA1109 function could be conserved relative to the fly gene tweek and examined endocytosis and endosome recycling in patient fibroblasts. Furthermore, we examined the structure of the cytoskeleton and cilia based on functional overlap with endocytosis and several known etiologies for neuronal migration disorders. Utilizing primary dermal fibroblasts from one patient and a healthy donor, we performed immunofluorescence and endocytosis assays to examine the endosomal, cytoskeletal, and ciliary cellular phenotypes. We found notable abnormalities in endosomal trafficking and endosome recycling pathways. We also observed changes in the actin cytoskeleton and cilia structural dynamics. We conclude that the function of KIAA1109 in humans may indeed overlap with the function of the Drosophila ortholog, resulting in perturbations to endosomal trafficking and the actin cytoskeleton. These alterations have ripple effects, altering many pathways that are critical for proper neuronal migration and embryonic development.

Rett syndrome (MECP2) and succinic semialdehyde dehydrogenase (ALDH5A1) deficiency in a developmentally delayed female.

BACKGROUND: We present a patient with Rett syndrome (RTT; MECP2) and autosomal-recessive succinic semialdehyde dehydrogenase deficiency (SSADHD; ALDH5A1 (aldehyde dehydrogenase 5a1 = SSADH), in whom the current phenotype exhibits features of SSADHD (hypotonia, global developmental delay) and RTT (hand stereotypies, gait anomalies). METHODS: γ-Hydroxybutyric acid (GHB) was quantified by UPLC-tandem mass spectrometry, while mutation analysis followed standard methodology of whole-exome sequencing. RESULTS: The biochemical hallmark of SSADHD, GHB was increased in the proband's dried bloodspot (DBS; 673 µM; previous SSADHD DBSs (n = 7), range 124-4851 µM); control range (n = 2,831), 0-78 µM. The proband was compound heterozygous for pathogenic ALDH5A1 mutations (p.(Asn418IlefsTer39); maternal; p.(Gly409Asp); paternal) and a de novo RTT nonsense mutation in MECP2 (p.Arg255*). CONCLUSION: The major inhibitory neurotransmitter, γ-aminobutyric acid (GABA), is increased in SSADHD but normal in RTT, although there are likely regional changes in GABA receptor distribution. GABAergic anomalies occur in both disorders, each featuring an autism spectrum phenotype. What effect the SSADHD biochemical anomalies (elevated GABA, GHB) might play in the neurodevelopmental/epileptic phenotype of our patient is currently unknown.

A case study of atypical Larsen syndrome with absent hallmark joint dislocations.

BACKGROUND: A family with skeletal and craniofacial anomalies is presented. Whole-exome sequencing (WES) analysis indicated a diagnosis of Larsen syndrome, although their clinical presentation does not include the hallmark joint dislocations typically observed in Larsen syndrome. METHODS: Patient consent for the sharing of de-identified clinical and genetic information, along with use of photographs for publication, was obtained. WES and variant segregation analysis by WES were performed by commercial laboratory, GeneDx (Gaithersburg, MD), on peripheral blood samples from the proband, her brother, and her parents using methods detailed on their website for test XomeDx Whole Exome Sequencing Trio (https://www.genedx.com/test-catalog/available-tests/xomedx-whole-exome-sequencing-trio/). WES uses next-generation sequencing (NGS) technology to assess for variants within the coding regions, or exons, of approximately 23,000 genes. For the FLNB gene (NM_001457.3), 100% of the coding region was covered at a minimum of 10x. GeneDx uses Sanger sequencing to confirm NGS variants. RESULTS: WES revealed a heterozygous pathogenic variant, p.Glu227Lys (c.679G>A), in the FLNB gene in three out of the four family members tested. This variant is associated with Larsen syndrome, a skeletal dysplasia condition with a wide range of phenotypic variability that usually includes congenital joint dislocations. CONCLUSION: This is a highly unusual presentation of Larsen syndrome in which the identifying hallmark trait is absent in the patients' phenotypes.

Rapidly growing, multifocal, benign choroid plexus tumor in an infant: case report.

Choroid plexus papillomas (CPPs) are rare, benign tumors that can arise in young children. Most pediatric patients present with signs of hydrocephalus and require immediate treatment. The natural history of choroid plexus tumors in children without hydrocephalus is poorly defined. In this report, the authors present the very rare case of a child without hydrocephalus but with two intraventricular choroid plexus tumors discovered shortly after birth. Initial imaging had been performed for seizures and showed agenesis of the corpus callosum and enhancing tumors in the third and left lateral ventricles. Sequential imaging demonstrated rapid growth of both tumors. The lateral tumor was removed when the child was 3 months of age. A histological examination of the specimen showed benign features with an elevated mitotic rate. Given the patient's age of under 3 years, the diagnosis was WHO grade I CPP. The third ventricle tumor grew rapidly. A second surgery was performed and this tumor was resected. Again, the pathological diagnosis was WHO grade I CPP. The authors present this rare case and discuss the current relevant literature.

Biochemical signatures mimicking multiple carboxylase deficiency in children with mutations in MT-ATP6.

Elevations of specific acylcarnitines in blood reflect carboxylase deficiencies, and have utility in newborn screening for life-threatening organic acidemias and other inherited metabolic diseases. In this report, we describe a newly-identified association of biochemical features of multiple carboxylase deficiency in individuals harboring mitochondrial DNA (mtDNA) mutations in MT-ATP6 and in whom organic acidemias and multiple carboxylase deficiencies were excluded. Using retrospective chart review, we identified eleven individuals with abnormally elevated propionylcarnitine (C3) or hydroxyisovalerylcarnitine (C5OH) with mutations in MT-ATP6, most commonly m.8993T>G in high heteroplasmy or homoplasmy. Most patients were ascertained on newborn screening; most had normal enzymatic or molecular genetic testing to exclude biotinidase and holocarboxylase synthetase deficiencies. MT-ATP6 is associated with some cases of Leigh disease; clinical outcomes in our cohort ranged from death from neurodegenerative disease in early childhood to clinically and developmentally normal after several years of follow-up. These cases expand the biochemical phenotype associated with MT-ATP6 mutations, especially m.8993T>G, to include acylcarnitine abnormalities mimicking carboxylase deficiency states. Clinicians should be aware of this association and its implications for newborn screening, and consider mtDNA sequencing in patients exhibiting similar acylcarnitine abnormalities that are biotin-unresponsive and in whom other enzymatic deficiencies have been excluded.