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Autism spectrum disorder (ASD) exhibits an ∼4:1 male-to-female sex bias and is characterized by early-onset impairment of social/communication skills, restricted interests, and stereotyped behaviors. Disruption of the Xp22.11 locus has been associated with ASD in males. This locus includes the three-exon PTCHD1, an adjacent multi-isoform long noncoding RNA (lncRNA) named PTCHD1-AS (spanning ∼1 Mb), and a poorly characterized single-exon RNA helicase named DDX53 that is intronic to PTCHD1-AS. While the relationship between PTCHD1/PTCHD1-AS and ASD is being studied, the role of DDX53 has not been comprehensively examined, in part because there is no apparent functional murine ortholog. Through clinical testing, here, we identified 8 males and 2 females with ASD from 8 unrelated families carrying rare, predicted damaging or loss-of-function variants in DDX53. Additionally, we identified a family consisting of a male proband and his affected mother with high-functioning autism, both harboring a gene deletion involving DDX53 and exons of the noncoding RNA PTCHD1-AS. Then, we examined databases, including the Autism Speaks MSSNG and Simons Foundation Autism Research Initiative, as well as population controls. We identified 26 additional individuals with ASD harboring 19 mostly maternally inherited, rare, damaging DDX53 variations, including two variants detected in families from the original clinical analysis. Our findings in humans support a direct link between DDX53 and ASD, which will be important in clinical genetic testing. These same autism-related findings, coupled with the observation that a functional orthologous gene is not found in mice, may also influence the design and interpretation of murine modeling of ASD., Competing Interests: Declaration of interests At the time of this study and its publication, S.W.S. served on the scientific advisory committee of Population Bio. Intellectual property from aspects of his research held at The Hospital for Sick Children are licensed to Athena Diagnostics and Population Bio. These relationships did not influence data interpretation or presentation during this study but are disclosed for potential future considerations. S.V.M. is an employee of GeneDx, LLC. H.F.P. is on the research advisory boards and speaker bureau for Takeda Pharmaceutical, AvroBio, Amicus Therapeutics, Sanofi, Alexion Therapeutics, Denali Therapeutics, and Acer Therapeutics., (Copyright © 2024 American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)

Autism Spectrum Disorder (ASD) exhibits an ~4:1 male-to-female sex bias and is characterized by early-onset impairment of social/communication skills, restricted interests, and stereotyped behaviors. Disruption of the Xp22.11 locus has been associated with ASD in males. This locus includes the three-exon PTCHD1 gene, an adjacent multi-isoform long noncoding RNA (lncRNA) named PTCHD1-AS (spanning ~1Mb), and a poorly characterized single-exon RNA helicase named DDX53 that is intronic to PTCHD1-AS . While the relationship between PTCHD1/PTCHD1-AS and ASD is being studied, the role of DDX53 has not been examined, in part because there is no apparent functional murine orthologue. Through clinical testing, here, we identified 6 males and 1 female with ASD from 6 unrelated families carrying rare, predicted-damaging or loss-of-function variants in DDX53 . Then, we examined databases, including the Autism Speaks MSSNG and Simons Foundation Autism Research Initiative, as well as population controls. We identified 24 additional individuals with ASD harboring rare, damaging DDX53 variations, including the same variants detected in two families from the original clinical analysis. In this extended cohort of 31 participants with ASD (28 male, 3 female), we identified 25 mostly maternally-inherited variations in DDX53 , including 18 missense changes, 2 truncating variants, 2 in-frame variants, 2 deletions in the 3' UTR and 1 copy number deletion. Our findings in humans support a direct link between DDX53 and ASD, which will be important in clinical genetic testing. These same autism-related findings, coupled with the observation that a functional orthologous gene is not found in mouse, may also influence the design and interpretation of murine-modelling of ASD., Competing Interests: Competing interests At the time of this study and its publication, S.W.S. served on the Scientific Advisory Committee of Population Bio. Intellectual property from aspects of his research held at The Hospital for Sick Children are licensed to Athena Diagnostics and Population Bio. These relationships did not influence data interpretation or presentation during this study but are disclosed for potential future considerations. SVM is an employee of GeneDx, LLC. HFP is on the research advisory boards and speaker bureau for Takeda Pharmaceutical, AvroBio, Amicus Therapeutics, Sanofi, Alexion Therapeutics, Denali Therapeutics and Acer Therapeutics. All other authors declare no conflict of interest.

Objective: We designed a quality improvement (QI) project to improve rates of documented folic acid supplementation counseling for adolescent females with epilepsy, consistent with a quality measure from the American Academy of Neurology and American Epilepsy Society. Our SMART aim was to increase the percentage of visits at which folic acid counseling was addressed from our baseline rate of 23% to 50% by July 1, 2020., Methods: This initiative was conducted in female patients ≥12 years old with epilepsy who were prescribed daily antiseizure medication and were seen by the 13 providers in our Neurology QI Program. Using provider interviews, we undertook a root cause analysis of low counseling rates and identified the following main factors: insufficient time during clinic visit to counsel, lack of provider knowledge, and forgetting to counsel. Countermeasures were designed to address these main root causes and were implemented through iterative plan-do-study-act (PDSA) cycles. Interventions included provider education and features within the electronic health record, which were introduced sequentially, culminating in the creation of a best practice advisory (BPA). We performed biweekly chart reviews of visits for applicable patients to establish baseline performance rate and track progress over time. We used a statistical process control p-chart to analyze the outcome measure of documented counseling. As a balancing measure, clinicians were surveyed using the Technology Adoption Model survey to assess acceptance of the BPA., Results: From September 2019 to August 2022, the QI team improved rates of documented folic acid counseling from 23% to 73% through several PDSA cycles. This level of performance has been sustained over time. The most successful and sustainable intervention was the BPA. Provider acceptance of the BPA was overall positive., Significance: We successfully used QI methodology to improve and sustain our rates of documented folic acid supplementation counseling for adolescent females with epilepsy., (© 2023 International League Against Epilepsy.)

Introduction: Phenotypic spectrum of SLC6A1 -related neurodevelopmental disorders ( SLC6A1 -NDD) includes intellectual disability (ID), autistic spectrum disorders (ASD), epilepsy, developmental delay, beginning from early infancy or after seizure onset, and other neurological features such as hypotonia and movement disorders. Data on familial phenotypic heterogeneity have been rarely reported, thus in our study we aimed to investigate intrafamilial phenotypic variability in families with SLC6A1 variants., Methods: We collected clinical, laboratory and genetic data on 39 individuals, including 17 probands, belonging to 13 families harboring inherited variants of SLC6A1 . Data were collected through an international network of Epilepsy and Genetic Centers., Results: Main clinical findings in the whole cohort of 39 subjects were: (a) epilepsy, mainly presenting with generalized seizures, reported in 71% of probands and 36% of siblings or first/second-degree relatives. Within a family, the same epilepsy type (generalized or focal) was observed; (b) ID reported in 100% and in 13% of probands and siblings or first/second-degree relatives, respectively; (c) learning disabilities detected in 28% of the SLC6A1 carriers, all of them were relatives of a proband; (d) around 51% of the whole cohort presented with psychiatric symptoms or behavioral disorders, including 82% of the probands. Out of the 19 patients with psychiatric symptoms, ASD were diagnosed in 40% of them; (e) neurological findings (primarily tremor and speech difficulties) were observed 38.5% of the whole cohort, including 10 probands. Our families harbored 12 different SLC6A1 variants, one was a frameshift, two stop-gain, while the remaining were missense. No genotype-phenotype associations were identified., Discussion: Our study showed that first-or second-degree relatives presented with a less severe phenotype, featuring mainly mild intellectual and/or learning disabilities, at variance with the probands who suffered from moderate to severe ID, generalized, sometimes intractable, epileptic seizures, behavioral and psychiatric disorders. These findings may suggest that a proportion of individuals with mild SLC6A1 -NDD might be missed, in particular those with an older age where genetic testing is not performed. Further studies on intrafamilial phenotypic variability are needed to confirm our results and possibly to expand the phenotypic spectrum of these disorders and benefit genetic counseling., Competing Interests: KG has consulted for Taysha Gene Therapy, Jaguar Gene Therapy, Astellas Gene Therapy, and all Stripes for unrelated work. She also serves as co-chair of the Clinical Advisory Board (unpaid) for the non-profit COMBINEDBrain, which involves collaborations on SLC6A1-NDD. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Kassabian, Fenger, Willems, Aledo-Serrano, Linnankivi, McDonnell, Lusk, Jepsen, Bayat, Kattentidt, Vidal, Valero-Lopez, Alarcon-Martinez, Goodspeed, van Slegtenhorst, Barakat, Møller, Johannesen and Rubboli.)

PPFIA3 encodes the Protein-Tyrosine Phosphatase, Receptor-Type, F Polypeptide-Interacting Protein Alpha-3 (PPFIA3), which is a member of the LAR protein-tyrosine phosphatase-interacting protein (liprin) family involved in synaptic vesicle transport and presynaptic active zone assembly. The protein structure and function are well conserved in both invertebrates and vertebrates, but human diseases related to PPFIA3 dysfunction are not yet known. Here, we report 14 individuals with rare mono-allelic PPFIA3 variants presenting with features including developmental delay, intellectual disability, hypotonia, autism, and epilepsy. To determine the pathogenicity of PPFIA3 variants in vivo , we generated transgenic fruit flies expressing either human PPFIA3 wildtype (WT) or variant protein using GAL4-UAS targeted gene expression systems. Ubiquitous expression with Actin-GAL4 showed that the PPFIA3 variants had variable penetrance of pupal lethality, eclosion defects, and anatomical leg defects. Neuronal expression with elav-GAL4 showed that the PPFIA3 variants had seizure-like behaviors, motor defects, and bouton loss at the 3 rd instar larval neuromuscular junction (NMJ). Altogether, in the fly overexpression assays, we found that the PPFIA3 variants in the N-terminal coiled coil domain exhibited stronger phenotypes compared to those in the C-terminal region. In the loss-of-function fly assay, we show that the homozygous loss of fly Liprin- α leads to embryonic lethality. This lethality is partially rescued by the expression of human PPFIA3 WT, suggesting human PPFIA3 protein function is partially conserved in the fly. However, the PPFIA3 variants failed to rescue lethality. Altogether, the human and fruit fly data reveal that the rare PPFIA3 variants are dominant negative loss-of-function alleles that perturb multiple developmental processes and synapse formation.

Heterozygous pathogenic variants in DNM1 cause developmental and epileptic encephalopathy (DEE) as a result of a dominant-negative mechanism impeding vesicular fission. Thus far, pathogenic variants in DNM1 have been studied with a canonical transcript that includes the alternatively spliced exon 10b. However, after performing RNA sequencing in 39 pediatric brain samples, we find the primary transcript expressed in the brain includes the downstream exon 10a instead. Using this information, we evaluated genotype-phenotype correlations of variants affecting exon 10a and identified a cohort of eleven previously unreported individuals. Eight individuals harbor a recurrent de novo splice site variant, c.1197-8G>A (GenBank: NM_001288739.1), which affects exon 10a and leads to DEE consistent with the classical DNM1 phenotype. We find this splice site variant leads to disease through an unexpected dominant-negative mechanism. Functional testing reveals an in-frame upstream splice acceptor causing insertion of two amino acids predicted to impair oligomerization-dependent activity. This is supported by neuropathological samples showing accumulation of enlarged synaptic vesicles adherent to the plasma membrane consistent with impaired vesicular fission. Two additional individuals with missense variants affecting exon 10a, p.Arg399Trp and p.Gly401Asp, had a similar DEE phenotype. In contrast, one individual with a missense variant affecting exon 10b, p.Pro405Leu, which is less expressed in the brain, had a correspondingly less severe presentation. Thus, we implicate variants affecting exon 10a as causing the severe DEE typically associated with DNM1-related disorders. We highlight the importance of considering relevant isoforms for disease-causing variants as well as the possibility of splice site variants acting through a dominant-negative mechanism., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2022 American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)