Autosomal Recessive Infantile Hyaline Fibromatosis Identified Using Artificial Intelligence-Assisted Rapid Whole Genome Sequencing: A Rare, Multisystemic, Hereditary Disorder.

Infantile hyaline fibromatosis syndrome (HFS) is an ultra-rare genetic condition characterized by the deposition of hyaline material in the skin, muscle, and viscera. Potential complications include debilitating joint contractures, coarse facial features, recurrent infections, failure to thrive, and death. Here, we present the case of a six-month-old infant with a history of painful extremity contractures, global developmental delay, neck hemangioma, and feeding intolerance presenting to our institution with abdominal distension. The multi-systemic, rapidly progressing, severe nature of her symptoms prompted consultation with inpatient pediatric genetics. Per their recommendation, rapid whole-genome sequencing (rWGS) was done with Fabric GEM®-assisted artificial intelligence (Fabric Genomics, Oakland, California, United States) at Rady Children's Hospital Institute for Genomic Medicine (San Diego, California, United States), revealing homozygous pathogenic variant c.652T>C; P.Cys218Arg in the ANTXR2 gene consistent with HFS. This case was significant not only for its rarity, but also its early manifestation of symptoms, wide range of affected body systems, and severity of symptoms, which together present a fascinating diagnostic dilemma for future clinicians that should be taken into consideration. It also highlights the increasing utility of AI-assisted rWGS as a diagnostic tool for medically complex patients with unknown multisystemic hereditary conditions.

DOCK3-Associated Neurodevelopmental Disorder-Clinical Features and Molecular Basis.

The protein product of DOCK3 is highly expressed in neurons and has a role in cell adhesion and neuronal outgrowth through its interaction with the actin cytoskeleton and key cell signaling molecules. The DOCK3 protein is essential for normal cell growth and migration. Biallelic variants in DOCK3 associated with complete or partial loss of function of the gene were recently reported in six patients with intellectual disability and muscle hypotonia. Only one of the reported patients had congenital malformations outside of the CNS. Further studies are necessary to better determine the prevalence of DOCK3-associated neurodevelopmental disorders and the frequency of non-CNS clinical manifestations in these patients. Since deficiency of the DOCK3 protein product is now an established pathway of this neurodevelopmental condition, supplementing the deficient gene product using a gene therapy approach may be an efficient treatment strategy.

Efficient Delivery of FMR1 across the Blood Brain Barrier Using AAVphp Construct in Adult FMR1 KO Mice Suggests the Feasibility of Gene Therapy for Fragile X Syndrome.

Background Fragile X syndrome (FXS) is the most common inherited cause of intellectual disability and autism. Gene therapy may offer an efficient method to ameliorate the symptoms of this disorder. Methods An AAVphp.eb-hSyn-mFMR1IOS7 vector and an empty control were injected into the tail vein of adult Fmr1 knockout (KO) mice and wildtype (WT) controls. The KO mice were injected with 2 × 1013 vg/kg of the construct. The control KO and WT mice were injected with an empty vector. Four weeks following treatment, the animals underwent a battery of tests: open field, marble burying, rotarod, and fear conditioning. The mouse brains were studied for levels of the Fmr1 product FMRP. Results: No significant levels of FMRP were found outside the CNS in the treated animals. The gene delivery was highly efficient, and it exceeded the control FMRP levels in all tested brain regions. There was also improved performance in the rotarod test and partial improvements in the other tests in the treated KO animals. Conclusion: These experiments demonstrate efficient, brain-specific delivery of Fmr1 via peripheral administration in adult mice. The gene delivery led to partial alleviation of the Fmr1 KO phenotypical behaviors. FMRP oversupply may explain why not all behaviors were significantly affected. Since AAV.php vectors are less efficient in humans than in the mice used in the current experiment, studies to determine the optimal dose using human-suitable vectors will be necessary to further demonstrate feasibility.

The association between mosaicism type and cognitive and behavioral functioning among males with fragile X syndrome.

Mosaicism in fragile X syndrome (FXS) refers to two different FMR1 allele variations: size mosaicism represents different numbers of CGG repeats between the two alleles, such that in addition to a full mutation allele there is an allele in the normal or premutation range of CGG repeats, while methylation mosaicism indicates whether a full-mutation allele is fully or partially methylated. The present study explored the association between mosaicism type and cognitive and behavioral functioning in a large sample of males 3 years and older (n = 487) with FXS, participating in the Fragile X Online Registry with Accessible Research Database. Participants with methylation mosaicism were less severely cognitively affected as indicated by a less severe intellectual disability rating, higher intelligence quotient and adaptive behavior score, and lower social impairment score. In contrast, the presence of size mosaicism was not significantly associated with better cognitive and behavioral outcomes than full mutation. Our findings suggest that methylation mosaicism is associated with better cognitive functioning and adaptive behavior and less social impairment. Further research could assess to what extent these cognitive and behavioral differences depend on molecular diagnostic methods and the impact of mosaicism on prognosis of individuals with FXS.

Height and BMI in fragile X syndrome: A longitudinal assessment.

OBJECTIVE: Previously reported data regarding growth parameters in individuals with fragile X syndrome (FXS) are inconsistent. A longitudinal analysis of height and BMI in a large number of individuals with FXS was conducted. METHODS: Age- and sex-specific z scores for height and BMI of 1,223 individuals with FXS were calculated based on published normative data. Mixed-effect linear regression models were fit separately for males and females, and z scores for height and weight were regressed against age and adjusted for intellectual disability (ID) and psychotropic medication use. RESULTS: Mean height z score for both sexes decreased with age and was lower than normative data. Mean BMI z score was greater than normative data in both sexes, and this disparity increased with age. BMI z score in females was greater for those with moderate or severe ID than those with no or mild ID. Individuals taking antipsychotics had higher BMI z scores than those taking no or other medications; those taking anticonvulsants or stimulants had lower BMI z scores. CONCLUSIONS: Individuals with FXS are at elevated risk for overweight and obesity. The risk is higher in individuals taking antipsychotics and among females with severe ID. These findings warrant increased attention to obesity prevention for all individuals with FXS.

Autosomal recessive SLC30A9 variants in a proband with a cerebrorenal syndrome and no parental consanguinity.

An SLC30A9-associated cerebrorenal syndrome was first reported in consanguineous Bedouin kindred by Perez et al. in 2017. Although the function of the gene has not yet been fully elucidated, it may be implicated in Wnt signaling and nuclear regulation, as well as in cell and mitochondrial zinc regulation. In this research report, we present a female proband with two distinct, inherited autosomal recessive loss-of-function SLC30A9 variants from unrelated parents. To our knowledge, this is the first reported case of a possible SLC30A9-associated cerebrorenal syndrome in a nonconsanguineous family. Furthermore, a limited statistical analysis was conducted to identify possible allele frequency differences between populations. Our findings provide further support for an SLC30A9-associated cerebrorenal syndrome and may help clarify the gene's function through its possible disease association.

FMR1 and Autism, an Intriguing Connection Revisited.

Autism Spectrum Disorder (ASD) represents a distinct phenotype of behavioral dysfunction that includes deficiencies in communication and stereotypic behaviors. ASD affects about 2% of the US population. It is a highly heritable spectrum of conditions with substantial genetic heterogeneity. To date, mutations in over 100 genes have been reported in association with ASD phenotypes. Fragile X syndrome (FXS) is the most common single-gene disorder associated with ASD. The gene associated with FXS, FMR1 is located on chromosome X. Accordingly, the condition has more severe manifestations in males. FXS results from the loss of function of FMR1 due to the expansion of an unstable CGG repeat located in the 5'' untranslated region of the gene. About 50% of the FXS males and 20% of the FXS females meet the Diagnostic Statistical Manual 5 (DSM-5) criteria for ASD. Among the individuals with ASD, about 3% test positive for FXS. FMRP, the protein product of FMR1, is a major gene regulator in the central nervous system. Multiple pathways regulated by FMRP are found to be dysfunctional in ASD patients who do not have FXS. Thus, FXS presents the opportunity to study cellular phenomena that may have wider applications in the management of ASD and to develop new strategies for ASD therapy.

Pharmacological inhibition of the primary endocannabinoid producing enzyme, DGL-α, induces autism spectrum disorder-like and co-morbid phenotypes in adult C57BL/J mice.

Accumulating evidence links dysfunction in the endocannabinoid system (ECS) with the pathology of neurodevelopmental disorders, particularly autism spectrum disorder (ASD). Variants in ECS genes CNR1 and DAGLA are associated with neurological phenotypes in humans. The endocannabinoids (eCBs), 2-AG and AEA, which act at the primary cannabinoid receptor (CB1), mediate behaviors relevant to neurodevelopmental disorders. The overlap between these eCBs is poorly understood. Most ECS studies have focused on stress responses, anxiety, and epilepsy, however, its role in social behavior and communication has only recently come under investigation. This represents a critical gap in our understanding of the ECS and its relationship to ASD. Furthermore, the increasing prevalence of ASD and a lack of therapeutics emphasize a crucial need for novel therapeutic targets. To this aim, we used an inhibitor of the eCB producing enzyme DGL-α, DO34, and the CB1 inverse agonist, rimonabant, to evaluate the role of the primary eCB, 2-AG, in ASD. Adult male C57BL/6J mice were used in a series of behavioral paradigms which assessed social behavior, social communication, repetitive behaviors, anxiety and locomotor activity. DO34 and rimonabant increased anxiety-like behavior, while only DO34 induced hyperactivity, social deficits, and repetitive self-grooming behavior. These data indicate that reduced 2-AG bioavailability, or CB1 inhibition, each induce unique respective behavioral phenotypes relevant to neurodevelopmental disorders, particularly ASD. This suggests fundamental differences in CB1 signaling via 2-AG and the CB1 receptor itself, particularly for social behaviors, and that 2-AG signaling may represent a target for the development of novel therapeutics. LAY SUMMARY: Endocannabinoids play a critical role in the developing nervous system. Alterations in the endocannabinoid system are linked to neurodevelopmental disorders. Studies suggest these variants may play a critical role in the core symptoms of autism spectrum disorder. In this study, pharmacological inhibition of the primary endocannabinoid producing enzyme, DGL-α, induced a constellation of deficits in behavioral domains associated with autism.

Pharmacological inhibition of BKCa channels induces a specific social deficit in adult C57BL6/J mice.

Genetic variants in large conductance voltage and calcium sensitive potassium (BKCa) channels have associations with neurodevelopmental disorders such as autism spectrum disorder, fragile X syndrome, and intellectual disability. In the case of fragile X syndrome, early preclinical studies suggest that BKCa channels may be a promising treatment target for neurodevelopmental disorders. While BKCa channel dysfunction has been investigated within the context of fragile X syndrome, it is unknown whether interference with BKCa channel function is inductive for deficits in behavioral domains relevant to neurodevelopmental disorders. This represents a critical gap in our knowledge regarding the relationship between BKCa dysfunction and neurodevelopmental disorders. To explore this concept, we used the BKCa channel antagonist paxilline to evaluate the role of BKCa channel function in phenotypes of neurodevelopmental disorders. Here we used adult male C57BL/6J mice and a series of behavioral paradigms which assessed anxiety-like behavior, locomotor activity, social behavior, and repetitive self-grooming. We found that acute inhibition with paxilline induced a specific social deficit, but not anxiety-like behavior, or hyperactivity. These findings demonstrate proof-of-concept regarding a relationship between BKCa channel impairment and social behavior. Although this is a limited characterization of the BKCa channel in autistic-like behaviors, it provides evidence for this link. Future studies which examine the effective dose range of paxilline and exhaustive assays of behavior relevant to neurodevelopmental disorders will be needed to delineate the parametric space of the paxilline effect, particularly during critical periods of development, and its potential for therapeutic use. (PsycInfo Database Record (c) 2021 APA, all rights reserved).

Autosomal dominant neuronal ceroid lipofuscinosis: Clinical features and molecular basis.

The neuronal ceroid lipofuscinoses (NCLs) are at least 13 distinct progressive neurodegenerative disorders unified by the accumulation of lysosomal auto-fluorescent material called lipofuscin. The only form that occurs via autosomal-dominant inheritance exhibits adult onset and is sometimes referred to as Parry type NCL. The manifestations may include behavioral symptoms followed by seizures, ataxia, dementia, and early death. Mutations in the gene DNAJC5 that codes for the presynaptic co-chaperone cysteine string protein-α (CSPα) were recently reported in sporadic adult-onset cases and in families with dominant inheritance. The mutant CSPα protein may lead to disease progression by both loss and gain of function mechanisms. Iron chelation therapy may be considered as a possible pharmaceutical intervention based on our recent mechanism-based proposal of CSPα oligomerization via ectopic Fe-S cluster-binding, summarized in this review.

Soy-Based Infant Formula is Associated with an Increased Prevalence of Comorbidities in Fragile X Syndrome.

A large number of adults and children consume soy in various forms, but little information is available regarding potential neurological side effects. Prior work indicates an association between the consumption of soy-based diets and seizure prevalence in mouse models of neurological disease and in children with autism. Herein, we sought to evaluate potential associations between the consumption of soy-based formula during infancy and disease comorbidities in persons with fragile X syndrome (FXS), while controlling for potentially confounding issues, through a retrospective case-control survey study of participants with FXS enrolled in the Fragile X Online Registry with Accessible Research Database (FORWARD). There was a 25% usage rate of soy-based infant formula in the study population. We found significant associations between the consumption of soy-based infant formula and the comorbidity of autism, gastrointestinal problems (GI) and allergies. Specifically, there was a 1.5-fold higher prevalence of autism, 1.9-fold GI problems and 1.7-fold allergies in participants reporting the use of soy-based infant formula. The major reason for starting soy-based infant formula was GI problems. The average age of seizure and allergy onset occurred long after the use of soy-based infant formula. We conclude that early-life feeding with soy-based infant formula is associated with the development of several disease comorbidities in FXS.

Diagnosis of late-infantile neuronal ceroid lipofuscinosis using dried blood spot-based assay for TPPI enzyme activity: TPPI diagnostic assay from DBS.

BACKGROUND: The neuronal ceroid lipofuscinosis 2 (NCL2) or classic late-infantile neuronal ceroid lipofuscinosis (LINCL) is a neurogenetic disorder caused by mutations in the TPPI gene, which codes for the lysosomal tripeptidyl peptidase 1 (TPPI) EC 3.4.14.9. Loss of functional TPPI activity results in progressive visual and neurological symptoms starting at around 1-2 years of age causing early death. METHODS: We report a DBS-based TPPI assay that cleaves a synthetic tetrapeptide substrate generating a product that is detected by HPLC. Probands and carriers were identified with 100% accuracy (7 probands, 30 carriers, 13 controls). RESULTS: The assay detected a single TPPI activity at a lower pH towards the substrate tested. TPPI activity measurable when extracted at lower pH while inactive at neutral pH showed steady increase for at least 8 h incubation. No loss in TPPI activity was observed when DBS were stored for at least 2 weeks either in freezer, refrigerator, room temperature or 42 °C. CONCLUSION: A sequence variant causing Arg339Gln substitution in a proband had 12% TPPI. TPPI activity can be reliably measured in DBS, giving an opportunity to diagnose NCL2 at birth and refer patients for enzyme replacement or other therapies for earliest intervention, or alternatively offers a second-tier confirmatory test.

Aggregation of mutant cysteine string protein-α via Fe-S cluster binding is mitigated by iron chelators.

Point mutations in cysteine string protein-α (CSPα) cause dominantly inherited adult-onset neuronal ceroid lipofuscinosis (ANCL), a rapidly progressing and lethal neurodegenerative disease with no treatment. ANCL mutations are proposed to trigger CSPα aggregation/oligomerization, but the mechanism of oligomer formation remains unclear. Here we use purified proteins, mouse primary neurons and patient-derived induced neurons to show that the normally palmitoylated cysteine string region of CSPα loses palmitoylation in ANCL mutants. This allows oligomerization of mutant CSPα via ectopic binding of iron-sulfur (Fe-S) clusters. The resulting oligomerization of mutant CSPα causes its mislocalization and consequent loss of its synaptic SNARE-chaperoning function. We then find that pharmacological iron chelation mitigates the oligomerization of mutant CSPα, accompanied by partial rescue of the downstream SNARE defects and the pathological hallmark of lipofuscin accumulation. Thus, the iron chelators deferiprone (L1) and deferoxamine (Dfx), which are already used to treat iron overload in humans, offer a new approach for treating ANCL.

De novo and inherited variants in ZNF292 underlie a neurodevelopmental disorder with features of autism spectrum disorder.

PURPOSE: Intellectual disability (ID) and autism spectrum disorder (ASD) are genetically heterogeneous neurodevelopmental disorders. We sought to delineate the clinical, molecular, and neuroimaging spectrum of a novel neurodevelopmental disorder caused by variants in the zinc finger protein 292 gene (ZNF292). METHODS: We ascertained a cohort of 28 families with ID due to putatively pathogenic ZNF292 variants that were identified via targeted and exome sequencing. Available data were analyzed to characterize the canonical phenotype and examine genotype-phenotype relationships. RESULTS: Probands presented with ID as well as a spectrum of neurodevelopmental features including ASD, among others. All ZNF292 variants were de novo, except in one family with dominant inheritance. ZNF292 encodes a highly conserved zinc finger protein that acts as a transcription factor and is highly expressed in the developing human brain supporting its critical role in neurodevelopment. CONCLUSION: De novo and dominantly inherited variants in ZNF292 are associated with a range of neurodevelopmental features including ID and ASD. The clinical spectrum is broad, and most individuals present with mild to moderate ID with or without other syndromic features. Our results suggest that variants in ZNF292 are likely a recurrent cause of a neurodevelopmental disorder manifesting as ID with or without ASD.

VAC14 syndrome in two siblings with retinitis pigmentosa and neurodegeneration with brain iron accumulation.

Whole-exome sequencing was used to identify the genetic etiology of a rapidly progressing neurological disease present in two of six siblings with early childhood onset of severe progressive spastic paraparesis and learning disabilities. A homozygous mutation (c.2005G>T, p, V669L) was found in VAC14, and the clinical phenotype is consistent with the recently described VAC14-related striatonigral degeneration, childhood-onset syndrome (SNDC) (MIM#617054). However, the phenotype includes a distinct clinical presentation of retinitis pigmentosa (RP), which has not previously been reported in association with VAC14 mutations. Brain magnetic resonance imaging (MRI) revealed abnormal magnetic susceptibility in the globus pallidus, which can be seen in neurodegeneration with brain iron accumulation (NBIA). RP is a group of inherited retinal diseases with phenotypic/genetic heterogeneity, and the pathophysiologic basis of RP is not completely understood but is thought to be due to a primary retinal photoreceptor cell degenerative process. Most cases of RP are seen in isolation (nonsyndromic); this is a report of RP in two siblings with VAC14-associated syndrome, and it is suggested that a connection between RP and VAC14-associated syndrome should be explored in future studies.

De Novo Missense Variants in WDR37 Cause a Severe Multisystemic Syndrome.

While genetic causes are known for many syndromes involving developmental anomalies, a large number of individuals with overlapping phenotypes remain undiagnosed. Using exome-sequencing analysis and review of matchmaker databases, we have discovered four de novo missense variants predicted to affect the N-terminal region of WDR37-p.Ser119Phe, p.Thr125Ile, p.Ser129Cys, and p.Thr130Ile-in unrelated individuals with a previously unrecognized syndrome. Features of WDR37 syndrome include the following: ocular anomalies such as corneal opacity/Peters anomaly, coloboma, and microcornea; dysmorphic facial features; significant neurological impairment with structural brain defects and seizures; poor feeding; poor post-natal growth; variable skeletal, cardiac, and genitourinary defects; and death in infancy in one individual. WDR37 encodes a protein of unknown function with seven predicted WD40 domains and no previously reported human pathogenic variants. Immunocytochemistry and western blot studies showed that wild-type WDR37 is localized predominantly in the cytoplasm and mutant proteins demonstrate similar protein levels and localization. CRISPR-Cas9-mediated genome editing generated zebrafish mutants with novel missense and frameshift alleles: p.Ser129Phe, p.Ser129Cys (which replicates one of the human variants), p.Ser129Tyr, p.Lys127Cysfs, and p.Gln95Argfs. Zebrafish carrying heterozygous missense variants demonstrated poor growth and larval lethality, while heterozygotes with frameshift alleles survived to adulthood, suggesting a potential dominant-negative mechanism for the missense variants. RNA-seq analysis of zebrafish embryos carrying a missense variant detected significant upregulation of cholesterol biosynthesis pathways. This study identifies variants in WDR37 associated with human disease and provides insight into its essential role in vertebrate development and possible molecular functions.

Genomic Copy Number Variations in the Autism Clinic-Work in Progress.

The development of advanced technology for microarray-based chromosomal studies helped discover increased prevalence of genomic copy number variants (CNVs) in individuals with autism spectrum disorder (ASD). Chromosomal microarray analysis (CMA) is now an important tool for clinical investigations in patients with ASD. While this technology helps identify high proportion of CNV positive individuals among patients with autism, the clinical interpretation of such genomic rearrangements is often challenged by inconsistent genotype-phenotype correlations. Possible explanations of such inconsistencies may involve complex interactions of potentially pathogenic CNV with additional (secondary) CNVs or single nucleotide variants (SNVs). Other involved factors may include gender-specific effects or environmental contributions. Development of risk models for interpreting such complex interactions may be necessary in order to provide better informed genetic counseling to the affected families.

Folate receptor autoantibodies are prevalent in children diagnosed with autism spectrum disorder, their normal siblings and parents.

Folate deficiency can affect fetal and neonatal brain development Considering the reported association of Folate receptor alpha (FRα) autoantibodies (Abs) with autism and developmental disorders, we sought to confirm this in families of 82 children with ASD, 53 unaffected siblings, 65 fathers, and 70 mothers, along with 52 unrelated normal controls. Overall, 76% of the affected children, 75% of the unaffected siblings, 69% of fathers and 59% of mothers were positive for either blocking or binding Ab, whereas the prevalence of this Ab in the normal controls was 29%. The Ab was highly prevalent in affected families including unaffected siblings. The appearance of these antibodies may have a familial origin but the risk of developing ASD is likely influenced by other mitigating factors since some siblings who had the antibodies were not affected. The antibody response appears heritable with the blocking autoantibody in the parents and affected child increasing the risk of ASD. Autism Res 2018, 11: 707-712. © 2018 International Society for Autism Research, Wiley Periodicals, Inc. LAY SUMMARY: Folate is an essential nutrient during fetal and infant development. Autoantibodies against the folate receptor alpha can block folate transport from the mother to the fetus and to the brain in infants. Children diagnosed with autism and their immediate family members were evaluated for the prevalence of folate receptor autoantibodies. The autoantibody was highly prevalent in affected families with similar distribution in parents, normal siblings and affected children. The presence of these antibodies appears to have a familial origin and may contribute to developmental deficits when combined with other factors.

Longitudinal telomere shortening and early Alzheimer's disease progression in adults with down syndrome.

Telomere shortening was shown to parallel Alzheimer's disease (AD) associated dementia. By using a dual PNA Probe system we have developed a practical method for comparing telomere length in T-lymphocyte interphases from individuals with Down syndrome (DS) with and without "mild cognitive impairment" (MCI-DS) and demonstrated that telomere length can serve as a valid biomarker for the onset of MCI-DS in this high-risk population. To verify progressive cognitive decline we have now examined sequential changes in telomere length in 10 adults with DS (N = 4 Female, N = 6 Male) developing MCI-DS. Cases were selected blind to telomere length from a sample of adults with DS previously enrolled in a prospective longitudinal study at 18-month intervals with clinical and telomere assessments: (1) MCI-DS group data were collected approximately three years prior to development of MCI-DS; (2) 18 months later; (3) when MCI-DS was first observed. These telomere measures were compared to those from another 10 adults with DS matched by sex and approximate age but without indications of MCI-DS (Controls). PNA (peptide nucleic acid) probes for telomeres together with a chromosome two centromere probe were used. Findings indicated telomere shortening over time for both Cases and Controls. Group differences emerged by 18-months prior to recognition of MCI-DS onset and completely non-overlapping distributions of telomere measures were observed by the time of MCI-DS onset. This study adds to accumulating evidence of the value of telomere length, as an early biomarker of AD progression in adults with Down syndrome.