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Craniosynostosis (CS) is a major birth defect resulting from premature fusion of cranial sutures. Nonsyndromic CS occurs more frequently than syndromic CS, with sagittal nonsyndromic craniosynostosis (sNCS) presenting as the most common CS phenotype. Previous genome-wide association and targeted sequencing analyses of sNCS have identified multiple associated loci, with the strongest association on chromosome 20. Herein, we report the first whole-genome sequencing study of sNCS using 63 proband-parent trios. Sequencing data for these trios were analyzed using the transmission disequilibrium test (TDT) and rare variant TDT (rvTDT) to identify high-risk rare gene variants. Sequencing data were also examined for copy number variants (CNVs) and de novo variants. TDT analysis identified a highly significant locus at 20p12.3, localized to the intergenic region between BMP2 and the noncoding RNA gene LINC01428. Three variants (rs6054763, rs6054764, rs932517) were identified as potential causal variants due to their probability of being transcription factor binding sites, deleterious combined annotation dependent depletion scores, and high minor allele enrichment in probands. Morphometric analysis of cranial vault shape in an unaffected cohort validated the effect of these three single nucleotide variants (SNVs) on dolichocephaly. No genome-wide significant rare variants, de novo loci, or CNVs were identified. Future efforts to identify risk variants for sNCS should include sequencing of larger and more diverse population samples and increased omics analyses, such as RNA-seq and ATAC-seq.

BACKGROUND: Genomic sequencing technology allows for identification of reproductive couples with an increased chance, as compared with that in the general population, of having a child with an autosomal recessive or X-linked genetic condition. METHODS: We investigated the feasibility, acceptability, and outcomes of a nationwide, couple-based genetic carrier screening program in Australia as part of the Mackenzie's Mission project. Health care providers offered screening to persons before pregnancy or early in pregnancy. The results obtained from testing at least 1281 genes were provided to the reproductive couples. We aimed to ascertain the psychosocial effects on participants, the acceptability of screening to all participants, and the reproductive choices of persons identified as having an increased chance of having a child with a condition for which we screened. RESULTS: Among 10,038 reproductive couples enrolled in the study, 9107 (90.7%) completed screening, and 175 (1.9%) were newly identified as having an increased chance of having a child with a genetic condition for which we screened. These conditions involved pathogenic variants in 90 different genes; 74.3% of the conditions were autosomal recessive. Three months after receiving the results, 76.6% of the couples with a newly identified increased chance had used or planned to use reproductive interventions to avoid having an affected child. Those newly identified as having an increased chance had greater anxiety than those with a low chance. The median level of decisional regret was low in all result groups, and 98.9% of participants perceived screening to be acceptable. CONCLUSIONS: Couple-based reproductive genetic carrier screening was largely acceptable to participants and was used to inform reproductive decision making. The delivery of screening to a diverse and geographically dispersed population was feasible. (Funded by the Medical Research Future Fund of the Australian government; ClinicalTrials.gov

Sequence-based genetic testing identifies causative variants in ~ 50% of individuals with developmental and epileptic encephalopathies (DEEs). Aberrant changes in DNA methylation are implicated in various neurodevelopmental disorders but remain unstudied in DEEs. We interrogate the diagnostic utility of genome-wide DNA methylation array analysis on peripheral blood samples from 582 individuals with genetically unsolved DEEs. We identify rare differentially methylated regions (DMRs) and explanatory episignatures to uncover causative and candidate genetic etiologies in 12 individuals. Using long-read sequencing, we identify DNA variants underlying rare DMRs, including one balanced translocation, three CG-rich repeat expansions, and four copy number variants. We also identify pathogenic variants associated with episignatures. Finally, we refine the CHD2 episignature using an 850 K methylation array and bisulfite sequencing to investigate potential insights into CHD2 pathophysiology. Our study demonstrates the diagnostic yield of genome-wide DNA methylation analysis to identify causal and candidate variants as 2% (12/582) for unsolved DEE cases.

Telomere maintenance 2 (TELO2), Tel2 interacting protein 2 (TTI2), and Tel2 interacting protein 1 (TTI1) are the three components of the conserved Triple T (TTT) complex that modulates activity of phosphatidylinositol 3-kinase-related protein kinases (PIKKs), including mTOR, ATM, and ATR, by regulating the assembly of mTOR complex 1 (mTORC1). The TTT complex is essential for the expression, maturation, and stability of ATM and ATR in response to DNA damage. TELO2- and TTI2-related bi-allelic autosomal-recessive (AR) encephalopathies have been described in individuals with moderate to severe intellectual disability (ID), short stature, postnatal microcephaly, and a movement disorder (in the case of variants within TELO2). We present clinical, genomic, and functional data from 11 individuals in 9 unrelated families with bi-allelic variants in TTI1. All present with ID, and most with microcephaly, short stature, and a movement disorder. Functional studies performed in HEK293T cell lines and fibroblasts and lymphoblastoid cells derived from 4 unrelated individuals showed impairment of the TTT complex and of mTOR pathway activity which is improved by treatment with Rapamycin. Our data delineate a TTI1-related neurodevelopmental disorder and expand the group of disorders related to the TTT complex. © 2023 American Society of Human Genetics

Sequence-based genetic testing currently identifies causative genetic variants in ∼50% of individuals with developmental and epileptic encephalopathies (DEEs). Aberrant changes in DNA methylation are implicated in various neurodevelopmental disorders but remain unstudied in DEEs. Rare epigenetic variations ("epivariants") can drive disease by modulating gene expression at single loci, whereas genome-wide DNA methylation changes can result in distinct "episignature" biomarkers for monogenic disorders in a growing number of rare diseases. Here, we interrogate the diagnostic utility of genome-wide DNA methylation array analysis on peripheral blood samples from 516 individuals with genetically unsolved DEEs who had previously undergone extensive genetic testing. We identified rare differentially methylated regions (DMRs) and explanatory episignatures to discover causative and candidate genetic etiologies in 10 individuals. We then used long-read sequencing to identify DNA variants underlying rare DMRs, including one balanced translocation, three CG-rich repeat expansions, and two copy number variants. We also identify pathogenic sequence variants associated with episignatures; some had been missed by previous exome sequencing. Although most DEE genes lack known episignatures, the increase in diagnostic yield for DNA methylation analysis in DEEs is comparable to the added yield of genome sequencing. Finally, we refine an episignature for CHD2 using an 850K methylation array which was further refined at higher CpG resolution using bisulfite sequencing to investigate potential insights into CHD2 pathophysiology. Our study demonstrates the diagnostic yield of genome-wide DNA methylation analysis to identify causal and candidate genetic causes as ∼2% (10/516) for unsolved DEE cases.

Item does not contain fulltext, De novo variants are a leading cause of neurodevelopmental disorders (NDDs), but because every monogenic NDD is different and usually extremely rare, it remains a major challenge to understand the complete phenotype and genotype spectrum of any morbid gene. According to OMIM, heterozygous variants in KDM6B cause "neurodevelopmental disorder with coarse facies and mild distal skeletal abnormalities." Here, by examining the molecular and clinical spectrum of 85 reported individuals with mostly de novo (likely) pathogenic KDM6B variants, we demonstrate that this description is inaccurate and potentially misleading. Cognitive deficits are seen consistently in all individuals, but the overall phenotype is highly variable. Notably, coarse facies and distal skeletal anomalies, as defined by OMIM, are rare in this expanded cohort while other features are unexpectedly common (e.g., hypotonia, psychosis, etc.). Using 3D protein structure analysis and an innovative dual Drosophila gain-of-function assay, we demonstrated a disruptive effect of 11 missense/in-frame indels located in or near the enzymatic JmJC or Zn-containing domain of KDM6B. Consistent with the role of KDM6B in human cognition, we demonstrated a role for the Drosophila KDM6B ortholog in memory and behavior. Taken together, we accurately define the broad clinical spectrum of the KDM6B-related NDD, introduce an innovative functional testing paradigm for the assessment of KDM6B variants, and demonstrate a conserved role for KDM6B in cognition and behavior. Our study demonstrates the critical importance of international collaboration, sharing of clinical data, and rigorous functional analysis of genetic variants to ensure correct disease diagnosis for rare disorders.

Item does not contain fulltext, AMOTL1 encodes angiomotin-like protein 1, an actin-binding protein that regulates cell polarity, adhesion, and migration. The role of AMOTL1 in human disease is equivocal. We report a large cohort of individuals harboring heterozygous AMOTL1 variants and define a core phenotype of orofacial clefting, congenital heart disease, tall stature, auricular anomalies, and gastrointestinal manifestations in individuals with variants in AMOTL1 affecting amino acids 157-161, a functionally undefined but highly conserved region. Three individuals with AMOTL1 variants outside this region are also described who had variable presentations with orofacial clefting and multi-organ disease. Our case cohort suggests that heterozygous missense variants in AMOTL1, most commonly affecting amino acid residues 157-161, define a new orofacial clefting syndrome, and indicates an important functional role for this undefined region.

Overlapping clinical phenotypes and an expanding breadth and complexity of genomic associations are a growing challenge in the diagnosis and clinical management of Mendelian disorders. The functional consequences and clinical impacts of genomic variation may involve unique, disorder-specific, genomic DNA methylation episignatures. In this study, we describe 19 novel episignature disorders and compare the findings alongside 38 previously established episignatures for a total of 57 episignatures associated with 65 genetic syndromes. We demonstrate increasing resolution and specificity ranging from protein complex, gene, sub-gene, protein domain, and even single nucleotide-level Mendelian episignatures. We show the power of multiclass modeling to develop highly accurate and disease-specific diagnostic classifiers. This study significantly expands the number and spectrum of disorders with detectable DNA methylation episignatures, improves the clinical diagnostic capabilities through the resolution of unsolved cases and the reclassification of variants of unknown clinical significance, and provides further insight into the molecular etiology of Mendelian conditions.

Item does not contain fulltext, NOTCH1 belongs to the NOTCH family of proteins that regulate cell fate and inflammatory responses. Somatic and germline NOTCH1 variants have been implicated in cancer, Adams-Oliver syndrome, and cardiovascular defects. We describe 7 unrelated patients grouped by the presence of leukoencephalopathy with calcifications and heterozygous de novo gain-of-function variants in NOTCH1. Immunologic profiling showed upregulated CSF IP-10, a cytokine secreted downstream of NOTCH1 signaling. Autopsy revealed extensive leukoencephalopathy and microangiopathy with vascular calcifications. This evidence implicates that heterozygous gain-of-function variants in NOTCH1 lead to a chronic central nervous system (CNS) inflammatory response resulting in a calcifying microangiopathy with leukoencephalopathy. ANN NEUROL 2022;92:895-901.

Neurodevelopmental disorders are highly heterogenous conditions resulting from abnormalities of brain architecture and/or function. FBXW7 (F-box and WD-repeat-domain-containing 7), a recognized developmental regulator and tumor suppressor, has been shown to regulate cell-cycle progression and cell growth and survival by targeting substrates including CYCLIN E1/2 and NOTCH for degradation via the ubiquitin proteasome system. We used a genotype-first approach and global data-sharing platforms to identify 35 individuals harboring de novo and inherited FBXW7 germline monoallelic chromosomal deletions and nonsense, frameshift, splice-site, and missense variants associated with a neurodevelopmental syndrome. The FBXW7 neurodevelopmental syndrome is distinguished by global developmental delay, borderline to severe intellectual disability, hypotonia, and gastrointestinal issues. Brain imaging detailed variable underlying structural abnormalities affecting the cerebellum, corpus collosum, and white matter. A crystal-structure model of FBXW7 predicted that missense variants were clustered at the substrate-binding surface of the WD40 domain and that these might reduce FBXW7 substrate binding affinity. Expression of recombinant FBXW7 missense variants in cultured cells demonstrated impaired CYCLIN E1 and CYCLIN E2 turnover. Pan-neuronal knockdown of the Drosophila ortholog, archipelago, impaired learning and neuronal function. Collectively, the data presented herein provide compelling evidence of an F-Box protein-related, phenotypically variable neurodevelopmental disorder associated with monoallelic variants in FBXW7.

BACKGROUND: Children's interstitial and diffuse lung disease (chILD) is a complex heterogeneous group of lung disorders. Gene panel approaches have a reported diagnostic yield of ~ 12%. No data currently exist using trio exome sequencing as the standard diagnostic modality. We assessed the diagnostic utility of using trio exome sequencing in chILD. We prospectively enrolled children meeting specified clinical criteria between 2016 and 2020 from 16 Australian hospitals. Exome sequencing was performed with analysis of an initial gene panel followed by trio exome analysis. A subset of critically ill infants underwent ultra-rapid trio exome sequencing as first-line test. RESULTS: 36 patients [median (range) age 0.34 years (0.02-11.46); 11F] were recruited from multiple States and Territories. Five patients had clinically significant likely pathogenic/pathogenic variants (RARB, RPL15, CTCF, RFXANK, TBX4) and one patient had a variant of uncertain significance (VIP) suspected to contribute to their clinical phenotype, with VIP being a novel gene candidate. CONCLUSIONS: Trio exomes (6/36; 16.7%) had a better diagnostic rate than gene panel (1/36; 2.8%), due to the ability to consider a broader range of underlying conditions. However, the aetiology of chILD in most cases remained undetermined, likely reflecting the interplay between low penetrant genetic and environmental factors.

Whole genome sequencing (WGS) has the potential to outperform clinical microarrays for the detection of structural variants (SV) including copy number variants (CNVs), but has been challenged by high false positive rates. Here we present ClinSV, a WGS based SV integration, annotation, prioritization, and visualization framework, which identified 99.8% of simulated pathogenic ClinVar CNVs > 10 kb and 11/11 pathogenic variants from matched microarrays. The false positive rate was low (1.5–4.5%) and reproducibility high (95–99%). In clinical practice, ClinSV identified reportable variants in 22 of 485 patients (4.7%) of which 35–63% were not detectable by current clinical microarray designs. ClinSV is available at https://github.com/KCCG/ClinSV.

Objective: To assess the benefits and limitations of whole genome sequencing (WGS) compared to exome sequencing (ES) or multigene panel (MGP) in the molecular diagnosis of developmental and epileptic encephalopathies (DEE). Methods: We performed WGS of 30 comprehensively phenotyped DEE patient trios that were undiagnosed after first-tier testing, including chromosomal microarray and either research ES (n = 15) or diagnostic MGP (n = 15). Results: Eight diagnoses were made in the 15 individuals who received prior ES (53%): 3 individuals had complex structural variants; 5 had ES-detectable variants, which now had additional evidence for pathogenicity. Eleven diagnoses were made in the 15 MGP-negative individuals (68%); the majority (n = 10) involved genes not included in the panel, particularly in individuals with postneonatal onset of seizures and those with more complex presentations including movement disorders, dysmorphic features, or multiorgan involvement. A total of 42% of diagnoses were autosomal recessive or X-chromosome linked. Conclusion: WGS was able to improve diagnostic yield over ES primarily through the detection of complex structural variants (n = 3). The higher diagnostic yield was otherwise better attributed to the power of re-analysis rather than inherent advantages of the WGS platform. Additional research is required to assist in the assessment of pathogenicity of novel noncoding and complex structural variants and further improve diagnostic yield for patients with DEE and other neurogenetic disorders.

Contains fulltext : 229431.pdf (publisher's version ) (Open Access), ARGONAUTE-2 and associated miRNAs form the RNA-induced silencing complex (RISC), which targets mRNAs for translational silencing and degradation as part of the RNA interference pathway. Despite the essential nature of this process for cellular function, there is little information on the role of RISC components in human development and organ function. We identify 13 heterozygous mutations in AGO2 in 21 patients affected by disturbances in neurological development. Each of the identified single amino acid mutations result in impaired shRNA-mediated silencing. We observe either impaired RISC formation or increased binding of AGO2 to mRNA targets as mutation specific functional consequences. The latter is supported by decreased phosphorylation of a C-terminal serine cluster involved in mRNA target release, increased formation of dendritic P-bodies in neurons and global transcriptome alterations in patient-derived primary fibroblasts. Our data emphasize the importance of gene expression regulation through the dynamic AGO2-RNA association for human neuronal development.

We recently described a new neurodevelopmental syndrome (TAF1/MRXS33 intellectual disability [ID] syndrome) (MIM# 300966) caused by pathogenic variants involving the X-linked gene TATA-box binding protein associated factor 1 (TAF1), which participates in RNA polymerase II transcription. The initial study reported 11 families, and the syndrome was defined as presenting early in life with hypotonia, facial dysmorphia, and developmental delay that evolved into ID and/or autism spectrum disorder. We have now identified an additional 27 families through a genotype-first approach. Familial segregation analysis, clinical phenotyping, and bioinformatics were capitalized on to assess potential variant pathogenicity, and molecular modeling was performed for those variants falling within structurally characterized domains of TAF1. A novel phenotypic clustering approach was also applied, in which the phenotypes of affected individuals were classified using 51 standardized Human Phenotype Ontology terms. Phenotypes associated with TAF1 variants show considerable pleiotropy and clinical variability, but prominent among previously unreported effects were brain morphological abnormalities, seizures, hearing loss, and heart malformations. Our allelic series broadens the phenotypic spectrum of the TAF1/MRXS33 ID syndrome and the range of TAF1 molecular defects in humans. It also illustrates the challenges for determining the pathogenicity of inherited missense variants, particularly for a gene mapping to chromosome X.

Introduction: Implementation of rapid genomic testing in neonatal and paediatric intensive care units (NICUs/PICUs) is gathering momentum, and requires the development of systems capable of consistent delivery across multi-site networks. Method(s): We developed a rapid genomic diagnosis program involving 10 Australian hospitals and two laboratories with the aim of providing test results in <5 days for acutely unwell paediatric patients with suspected monogenic disorders. Rapid exome sequencing (rES) was performed as trios when possible, and analysis utilised multidisciplinary expertise. Experience was shared between clinical sites, laboratories, and professional groups to enable collective learning. Result(s): The program considered 123 patients for rES over 10 months, and approved 114 (93%). Five families declined testing (4.4%), and nine (7.9%) were withdrawn due to change in clinical circumstances. Of 100 patients tested, 51 received a diagnosis. Eleven of the diagnoses (21%) were made using approaches augmenting standard ES analysis: mitochondrial genome sequencing, ES-based copy number analysis, matchmaking of emerging genes, reverse phenotyping and RNA analysis. Median time from hospital admission to consent was 6 days (range 0-64 days); median time from sample receipt to clinical ES report was 3 days (range 2-7 days). The total cost of testing was AU$1,123,000/701,638 (AU$11,230/7,016 per case). Changes in management following a result occurred in 77% of diagnosed patients and 10% of undiagnosed patients. Conclusion(s): We demonstrate the feasibility of a national, highly integrated clinical-laboratory approach to rapid genomic diagnosis, which delivers results within a timeframe relevant to acute paediatrics, while optimising clinical utility and resource allocation.

Multiple studies have shown that genomic testing has a high diagnostic yield and an impact on clinical management for patients with suspected genetic conditions. Therefore, there has been a push worldwide to apply rapid genomic sequencing in critically ill neonatal and pediatric patients. The goal of this study was to investigate the practicality of applying ultrarapid genomic testing for critically ill neonatal and pediatric patients with suspected monogenic conditions in Australia. The study recruited a total of 108 patients prospectively from March 2018 to February 2019, and data were collected until May 2019. Of the 12 hospitals that acted as collaborating sites, 5 were women's hospitals, 3 women's and children's hospitals, and 4 children's hospitals. Eligible patients included those who were admitted to a neonatal or pediatric intensive care unit (NICU or PICU) and were referred to clinical genetics for a possible monogenic condition. Chromosomal microarray was a requirement before enrollment if there was a suspected chromosomal condition. Chromosomal microarrays were performed concurrently with ultrarapid exome sequencing when the probability of a positive result on microarray was low. Additionally, rapid mitochondrial genome sequencing was performed alongside ultrarapid exome sequencing if a mitochondrial condition was suspected. When possible, ultrarapid exome sequencing was performed in both parents as well as the child (trio). The primary outcome of this study measured the time from the last sample received to the ultrarapid exome sequencing report finalized. Other outcomes measured were the diagnostic yield, change in clinical management after the report was finalized, number of reports returned before hospital discharge, and time from admission to the report being finalized. Of the 108 patients enrolled, the median age was 28 days, with a range of 0 days to 17 years. Overall, 34% were female, 57% were NICU admissions, 33% were PICU admissions, and 9% wer

Pathogenic variants in CDH1, encoding epithelial cadherin (E-cadherin), have been implicated in hereditary diffuse gastric cancer (HDGC), lobular breast cancer, and both syndromic and non-syndromic cleft lip/palate (CL/P). Despite the large number of CDH1 mutations described, the nature of the phenotypic consequence of such mutations is currently not able to be predicted, creating significant challenges for genetic counselling. This study collates the phenotype and molecular data for available CDH1 variants that have been classified, using the American College of Medical Genetics and Genomics criteria, as at least ‘likely pathogenic’, and correlates their molecular and structural characteristics to phenotype. We demonstrate that CDH1 variant type and location differ between HDGC and CL/P, and that there is clustering of CL/P variants within linker regions between the extracellular domains of the cadherin protein. While these differences do not provide for exact prediction of the phenotype for a given mutation, they may contribute to more accurate assessments of risk for HDGC or CL/P for individuals with specific CDH1 variants.

Importance: Widespread adoption of rapid genomic testing in pediatric critical care requires robust clinical and laboratory pathways that provide equitable and consistent service across health care systems. Objective(s): To prospectively evaluate the performance of a multicenter network for ultra-rapid genomic diagnosis in a public health care system. Design, Setting, and Participant(s): Descriptive feasibility study of critically ill pediatric patients with suspected monogenic conditions treated at 12 Australian hospitals between March 2018 and February 2019, with data collected to May 2019. A formal implementation strategy emphasizing communication and feedback, standardized processes, coordination, distributed leadership, and collective learning was used to facilitate adoption. Exposures: Ultra-rapid exome sequencing. Main Outcomes and Measures: The primary outcome was time from sample receipt to ultra-rapid exome sequencing report. The secondary outcomes were the molecular diagnostic yield, the change in clinical management after the ultra-rapid exome sequencing report, the time from hospital admission to the laboratory report, and the proportion of laboratory reports returned prior to death or hospital discharge. Result(s): The study population included 108 patients with a median age of 28 days (range, 0 days to 17 years); 34% were female; and 57% were from neonatal intensive care units, 33% were from pediatric intensive care units, and 9% were from other hospital wards. The mean time from sample receipt to ultra-rapid exome sequencing report was 3.3 days (95% CI, 3.2-3.5 days) and the median time was 3 days (range, 2-7 days). The mean time from hospital admission to ultra-rapid exome sequencing report was 17.5 days (95% CI, 14.6-21.1 days) and 93 reports (86%) were issued prior to death or hospital discharge. A molecular diagnosis was established in 55 patients (51%). Eleven diagnoses (20%) resulted from using the following approaches to augment standard exome

LARS2 variants are associated with Perrault syndrome, characterized by premature ovarian failure and hearing loss, and with an infantile lethal multisystem disorder: Hydrops, lactic acidosis, sideroblastic anemia (HLASA) in one individual. Recently we reported LARS2 deafness with (ovario) leukodystrophy. Here we describe five patients with a range of phenotypes, in whom we identified biallelic LARS2 variants: three patients with a HLASA‐like phenotype, an individual with Perrault syndrome whose affected siblings also had leukodystrophy, and an individual with a reversible mitochondrial myopathy, lactic acidosis, and developmental delay. Three HLASA cases from two unrelated families were identified. All were males with genital anomalies. Two survived multisystem disease in the neonatal period; both have developmental delay and hearing loss. A 55‐year old male with deafness has not displayed neurological symptoms while his female siblings with Perrault syndrome developed leukodystrophy and died in their 30s. Analysis of muscle from a child with a reversible myopathy showed reduced LARS2 and mitochondrial complex I levels, and an unusual form of degeneration. Analysis of recombinant LARS2 variant proteins showed they had reduced aminoacylation efficiency, with HLASA‐associated variants having the most severe effect. A broad phenotypic spectrum should be considered in association with LARS2 variants

Defects in the motor domain of kinesin family member 1A (KIF1A), a neuron-specific ATP-dependent anterograde axonal transporter of synaptic cargo, are well-recognized to cause a spectrum of neurological conditions, commonly known as KIF1A-associated neurological disorders (KAND). Here, we report one mutation-negative female with classic Rett syndrome (RTT) harboring a de novo heterozygous novel variant [NP_001230937.1:p.(Asp248Glu)] in the highly conserved motor domain of KIF1A. In addition, three individuals with severe neurodevelopmental disorder along with clinical features overlapping with KAND are also reported carrying de novo heterozygous novel [NP_001230937.1:p.(Cys92Arg) and p.(Pro305Leu)] or previously reported [NP_001230937.1:p.(Thr99Met)] variants in KIF1A. In silico tools predicted these variants to be likely pathogenic, and 3D molecular modeling predicted defective ATP hydrolysis and/or microtubule binding. Using the neurite tip accumulation assay, we demonstrated that all novel KIF1A variants significantly reduced the ability of the motor domain of KIF1A to accumulate along the neurite lengths of differentiated SH-SY5Y cells. In vitro microtubule gliding assays showed significantly reduced velocities for the variant p.(Asp248Glu) and reduced microtubule binding for the p.(Cys92Arg) and p.(Pro305Leu) variants, suggesting a decreased ability of KIF1A to move along microtubules. Thus, this study further expanded the phenotypic characteristics of KAND individuals with pathogenic variants in the KIF1A motor domain to include clinical features commonly seen in RTT individuals.

Our previous genome-wide association study (GWAS) for sagittal nonsyndromic craniosynostosis (sNCS) provided important insights into the genetics of midline CS. In this study, we performed a GWAS for a second midline NCS, metopic NCS (mNCS), using 215 non-Hispanic white case-parent triads. We identified six variants with genome-wide significance (P ≤ 5 × 10–8): rs781716 (P = 4.71 × 10–9; odds ratio [OR] = 2.44) intronic to SPRY3; rs6127972 (P = 4.41 × 10–8; OR = 2.17) intronic to BMP7; rs62590971 (P = 6.22 × 10–9; OR = 0.34), located ~ 155 kb upstream from TGIF2LX; and rs2522623, rs2573826, and rs2754857, all intronic to PCDH11X (P = 1.76 × 10–8, OR = 0.45; P = 3.31 × 10–8, OR = 0.45; P = 1.09 × 10–8, OR = 0.44, respectively). We performed a replication study of these variants using an independent non-Hispanic white sample of 194 unrelated mNCS cases and 333 unaffected controls; only the association for rs6127972 (P = 0.004, OR = 1.45; meta-analysis P = 1.27 × 10–8, OR = 1.74) was replicated. Our meta-analysis examining single nucleotide polymorphisms common to both our mNCS and sNCS studies showed the strongest association for rs6127972 (P = 1.16 × 10–6). Our imputation analysis identified a linkage disequilibrium block encompassing rs6127972, which contained an enhancer overlapping a CTCF transcription factor binding site (chr20:55,798,821–55,798,917) that was significantly hypomethylated in mesenchymal stem cells derived from fused metopic compared to open sutures from the same probands. This study provides additional insights into genetic factors in midline CS.

Contains fulltext : 208676.pdf (publisher's version ) (Closed access), Cleft lip with or without cleft palate (CL/P) is generally viewed as a complex trait with multiple genetic and environmental contributions. In 70% of cases, CL/P presents as an isolated feature and/or deemed nonsyndromic. In the remaining 30%, CL/P is associated with multisystem phenotypes or clinically recognizable syndromes, many with a monogenic basis. Here we report the identification, via exome sequencing, of likely pathogenic variants in two genes that encode interacting proteins previously only linked to orofacial clefting in mouse models. A variant in GDF11 (encoding growth differentiation factor 11), predicting a p.(Arg298Gln) substitution at the Furin protease cleavage site, was identified in one family that segregated with CL/P and both rib and vertebral hypersegmentation, mirroring that seen in Gdf11 knockout mice. In the second family in which CL/P was the only phenotype, a mutation in FST (encoding the GDF11 antagonist, Follistatin) was identified that is predicted to result in a p.(Cys56Tyr) substitution in the region that binds GDF11. Functional assays demonstrated a significant impact of the specific mutated amino acids on FST and GDF11 function and, together with embryonic expression data, provide strong evidence for the importance of GDF11 and Follistatin in the regulation of human orofacial development.

Contains fulltext : 202800.pdf (publisher's version ) (Open Access), SMARCC2 (BAF170) is one of the invariable core subunits of the ATP-dependent chromatin remodeling BAF (BRG1-associated factor) complex and plays a crucial role in embryogenesis and corticogenesis. Pathogenic variants in genes encoding other components of the BAF complex have been associated with intellectual disability syndromes. Despite its significant biological role, variants in SMARCC2 have not been directly associated with human disease previously. Using whole-exome sequencing and a web-based gene-matching program, we identified 15 individuals with variable degrees of neurodevelopmental delay and growth retardation harboring one of 13 heterozygous variants in SMARCC2, most of them novel and proven de novo. The clinical presentation overlaps with intellectual disability syndromes associated with other BAF subunits, such as Coffin-Siris and Nicolaides-Baraitser syndromes and includes prominent speech impairment, hypotonia, feeding difficulties, behavioral abnormalities, and dysmorphic features such as hypertrichosis, thick eyebrows, thin upper lip vermilion, and upturned nose. Nine out of the fifteen individuals harbor variants in the highly conserved SMARCC2 DNA-interacting domains (SANT and SWIRM) and present with a more severe phenotype. Two of these individuals present cardiac abnormalities. Transcriptomic analysis of fibroblasts from affected individuals highlights a group of differentially expressed genes with possible roles in regulation of neuronal development and function, namely H19, SCRG1, RELN, and CACNB4. Our findings suggest a novel SMARCC2-related syndrome that overlaps with neurodevelopmental disorders associated with variants in BAF-complex subunits.

(The American Journal of Human Genetics 104, 542–552; March 7, 2019) In the original version of this article published on March 7, 2019, Łukasz Jaremko's name was unfortunately misspelled as Łukas Jaremko. It appears correctly here and online. The Journal and the authors apologize for this error.

The study aimed to explore with consanguineous couples in Australia the acceptability and perceived utility of whole exome reproductive carrier screening for autosomal recessive and X-linked recessive conditions. Semi-structured interviews with 21 consanguineous couples were conducted prior to the offer of screening. Interviews were coded, and thematic analysis was informed by an inductive approach. Three major themes were identified: experiences and attitudes of Australian consanguineous couples, childhood genetic conditions and beliefs, and the perceived utility of genomic screening. All but one couple had previously sought genetic advice, and a large majority of couples were aware of childhood conditions within their family or community. Thirteen couples perceived consanguinity as increasing the risk of having affected children. Nine spoke of premarital screening programs routinely conducted in their countries of origin. All supported the concept and availability of genomic reproductive carrier screening. Hypothetically, if found to be carriers of a severe childhood disorder, 13 couples reported they would test a pregnancy, and 12 of whom would consider termination of pregnancy or pre-implantation genetic diagnosis. Four couples would not test a pregnancy and two were unsure. A majority of couples would communicate potential at-risk status to family members, although there were some caveats. Fourteen couples chose to have exome screening and reported that they would utilize the results with the goal of preventing childhood conditions. Of these couples, nine (64%) had an affected child but were aware that testing may reveal they were at risk for a child with a different condition and five (71%) without an affected child. While from diverse ethnic and backgrounds, all couples practiced a religion and all but one couple were recruited from the same clinical genetics unit, with a likely higher genetic literacy and bias towards accepting genetic testing. However, the

Background/Aim: Implementation of rapid genomic testing in neonatal and pediatric intensive care units (NICUs/PICUs) is gathering momentum, and requires the development of systems capable of consistent delivery across multiple sites. Method(s): We developed a rapid genomic diagnosis program involving 10 Australian hospitals and two laboratories with the aim of providing test results in <5 days for acutely unwell pediatric patients with suspected monogenic disorders. Rapid exome sequencing (rES) was performed as trios when possible, and analysis utilized multidisciplinary expertise. Experience was shared between clinical sites, laboratories, and professional groups to enable collective learning. Result(s): The program considered 123 patients for rES over 10 months, and approved 114 (93%). Five families declined testing (4.4%), and nine (7.9%) were withdrawn due to change in clinical circumstances. Of 100 patients tested, 53 received a diagnosis. Twelve of the diagnoses (23%) were made using approaches augmenting standard ES analysis: mitochondrial genome sequencing, ES-based copy number analysis, matchmaking of emerging genes, reverse phenotyping and RNA analysis. Median time from hospital admission to consent was 6 days (range 0-64 days); median time from sample receipt to clinical ES report was 3 days (range 2-7 days). The total cost of testing was AU $1,123,000 (AU$11,230 per case). Changesin management following a result occurred in 77% of diagnosed patients and 10% of undiag-nosed patients. Conclusion(s): We demonstrate the feasibility of a national, highly integrated clinical-laboratory approach to rapid genomic diagnosis, which delivers results within a timeframe relevant to acute pediatrics, while optimizing clinical utility and resource allocation.

Background: Despite increasing scientific evidence regarding the utility of polygenic risk score (PRS) for families at high-risk of breast cancer, research findings are yet to be integrated into clinical practice. Before integrating polygenic information into clinical practice, it is important to understand the psychological implications. Aim(s): This prospective study aims to assess uptake of breast cancer PRS and ascertain the psychosocial and behavioral implications of receiving this information. Method(s): Eligible women are invited to participate and receive their breast cancer PRS. Eligibility: affected and unaffected women currently enrolled in the Variants in Practice Study, who have a high or low PRS, and a personal and/or family history of breast cancer where genetic testing for BRCA1/2 is negative. Participants complete three self-administered questionnaires: T1 prior to result, T2 two weeks and T3 one year post receipt of PRS. Result(s): As of April 2018, 147/173 (85%) participants reported interest in receiving their PRS with 25/173 (14%) declining. Logistic regression showed that uptake of PRS was associated with having daughters (OR = 4.43, 95% CI [1.36, 14.43], p = .013), higher uncertainty avoidance (OR = 0.6.30, 95% CI [2.65, 14.91], p <= .000) and higher response cost (OR = .85 (95% CI [0.72, 0.99], p = .043). Additional preliminary analysis showed a significant reduction in perceived risk (mean difference -0.20, p = .013), and general anxiety and depression (mean difference -1.56, p = .002) between T1 and T2. Conclusion(s): There is strong interest in receiving PRS among women at high-risk of breast cancer. Recruitment is ongoing, with additional data regarding short-term and longterm psychological and behavioral impact of receiving PRS to be collected.

A 17-year-old boy with a rapidly progressive movement disorder of unknown aetiology underwent diagnostic whole exome sequencing. A previously undescribed heterozygous nonsense mutation was identified in DNAJC5 p.(Gln160Ter) which is associated with neuronal lipofucinosis (CLN) type 4B. This variant is located 22bp from the 3' end of the penultimate coding exon of the gene and is therefore predicted to escape nonsense mediated mRNA decay and result in protein truncation/instability. Electron microscopy of a skin biopsy confirmed ultrastructural morphological changes consistent with the diagnosis of CLN. CLN4 is a rare form of CLN and is the only form of the disorder that is consistently inherited in an autosomal dominant manner. It is postulated that the mutant protein product CSPalpha causes a dominant-negative effect by dimerising with and interrupting the function of wild type CSPalpha. Segregation studies confirmed the presence of the pathogenic variant in the proband's unaffected mother consistent with current non-penetrance.Copyright © 2018

Purpose: Whole exome sequencing (WES) is rapidly becoming the standard of care for genetic services. We present the results of 500 clinical exomes performed at the Randwick Genetic Laboratory. Method(s): WES was performed in 204 probands with suspected Mendelian disorders, together with family members. Ampliseq RDY exome, libraries were analyzed on a Life Technologies Proton instrument. Data were analyzed using an in house pipeline with variant reporting following ACMG guidelines. Result(s): WES resulted in greater than 43% definitive findings with a small number of variants of uncertain significance. The diagnostic rate has increased over time, likely reflecting refinements in clinician referral practice as well as improved functionality of bioinformatics pipelines. Two families (1%)with Cantu and MoyaMoya syndromes had results which could lead to pharmacologic interventions. 33 families (16%) had De novo variants and seven (3.5%) were X-linked with significant implications for recurrence risk. Two prenatal diagnoses have been performed based on WES results. A diagnosis was made in 6/8 (75%) rapid turnaround studies, including two during pregnancy. Almost 10% of the diagnoses were unanticipated by the clinical teams and involved significant changes in diagnostic category. Four likely novel genes involved in known biological pathways were identified, enabling research participation. Conclusion(s): The minimum number of management changing outcomes was at least 20% within the 12 month period of this study. The number of secondary findings was small at about 1%. This demonstrates that WES is characterized by high levels of patient safety and clinical utility with likely cost effectiveness.

Rapid advances in genomic technologies have facilitated the identification pathogenic variants causing human disease. We report siblings with developmental and epileptic encephalopathy due to a novel, shared heterozygous pathogenic 13 bp duplication in SYNGAP1 (c.435_447dup, p.(L150Vfs*6)) that was identified by whole genome sequencing (WGS). The pathogenic variant had escaped earlier detection via two methodologies: whole exome sequencing and high-depth targeted sequencing. Both technologies had produced reads carrying the variant, however, they were either not aligned due to the size of the insertion or aligned to multiple major histocompatibility complex (MHC) regions in the hg19 reference genome, making the critical reads unavailable for variant calling. The WGS pipeline followed different protocols, including alignment of reads to the GRCh37 reference genome, which lacks the additional MHC contigs. Our findings highlight the benefit of using orthogonal clinical bioinformatic pipelines and all relevant inheritance patterns to re-analyze genomic data in undiagnosed patients.

We recently described a new neurodevelopmental syndrome (TAF1/MRXS33 intellectual disability [ID] syndrome) (MIM# 300966) caused by pathogenic variants involving the X‐linked gene TATA‐box binding protein associated factor 1 (TAF1), which participates in RNA polymerase II transcription. The initial study reported 11 families, and the syndrome was defined as presenting early in life with hypotonia, facial dysmorphia, and developmental delay that evolved into ID and/or autism spectrum disorder. We have now identified an additional 27 families through a genotype‐first approach. Familial segregation analysis, clinical phenotyping, and bioinformatics were capitalized on to assess potential variant pathogenicity, and molecular modeling was performed for those variants falling within structurally characterized domains of TAF1. A novel phenotypic clustering approach was also applied, in which the phenotypes of affected individuals were classified using 51 standardized Human Phenotype Ontology terms. Phenotypes associated with TAF1 variants show considerable pleiotropy and clinical variability, but prominent among previously unreported effects were brain morphological abnormalities, seizures, hearing loss, and heart malformations. Our allelic series broadens the phenotypic spectrum of the TAF1/MRXS33 ID syndrome and the range of TAF1 molecular defects in humans. It also illustrates the challenges for determining the pathogenicity of inherited missense variants, particularly for a gene mapping to chromosome X.