Mutations in ACTL6B Cause Neurodevelopmental Deficits and Epilepsy and Lead to Loss of Dendrites in Human Neurons.

We identified individuals with variations in ACTL6B, a component of the chromatin remodeling machinery including the BAF complex. Ten individuals harbored bi-allelic mutations and presented with global developmental delay, epileptic encephalopathy, and spasticity, and ten individuals with de novo heterozygous mutations displayed intellectual disability, ambulation deficits, severe language impairment, hypotonia, Rett-like stereotypies, and minor facial dysmorphisms (wide mouth, diastema, bulbous nose). Nine of these ten unrelated individuals had the identical de novo c.1027G>A (p.Gly343Arg) mutation. Human-derived neurons were generated that recaptured ACTL6B expression patterns in development from progenitor cell to post-mitotic neuron, validating the use of this model. Engineered knock-out of ACTL6B in wild-type human neurons resulted in profound deficits in dendrite development, a result recapitulated in two individuals with different bi-allelic mutations, and reversed on clonal genetic repair or exogenous expression of ACTL6B. Whole-transcriptome analyses and whole-genomic profiling of the BAF complex in wild-type and bi-allelic mutant ACTL6B neural progenitor cells and neurons revealed increased genomic binding of the BAF complex in ACTL6B mutants, with corresponding transcriptional changes in several genes including TPPP and FSCN1, suggesting that altered regulation of some cytoskeletal genes contribute to altered dendrite development. Assessment of bi-alleic and heterozygous ACTL6B mutations on an ACTL6B knock-out human background demonstrated that bi-allelic mutations mimic engineered deletion deficits while heterozygous mutations do not, suggesting that the former are loss of function and the latter are gain of function. These results reveal a role for ACTL6B in neurodevelopment and implicate another component of chromatin remodeling machinery in brain disease.

Disruption of RFX family transcription factors causes autism, attention-deficit/hyperactivity disorder, intellectual disability, and dysregulated behavior.

PURPOSE: We describe a novel neurobehavioral phenotype of autism spectrum disorder (ASD), intellectual disability, and/or attention-deficit/hyperactivity disorder (ADHD) associated with de novo or inherited deleterious variants in members of the RFX family of genes. RFX genes are evolutionarily conserved transcription factors that act as master regulators of central nervous system development and ciliogenesis. METHODS: We assembled a cohort of 38 individuals (from 33 unrelated families) with de novo variants in RFX3, RFX4, and RFX7. We describe their common clinical phenotypes and present bioinformatic analyses of expression patterns and downstream targets of these genes as they relate to other neurodevelopmental risk genes. RESULTS: These individuals share neurobehavioral features including ASD, intellectual disability, and/or ADHD; other frequent features include hypersensitivity to sensory stimuli and sleep problems. RFX3, RFX4, and RFX7 are strongly expressed in developing and adult human brain, and X-box binding motifs as well as RFX ChIP-seq peaks are enriched in the cis-regulatory regions of known ASD risk genes. CONCLUSION: These results establish a likely role of deleterious variation in RFX3, RFX4, and RFX7 in cases of monogenic intellectual disability, ADHD and ASD, and position these genes as potentially critical transcriptional regulators of neurobiological pathways associated with neurodevelopmental disease pathogenesis.

Breast cancer polygenic risk scores in the clinical cancer genetic counseling setting: Current practices and impact on patient management.

Multivariate risk models are commonly used in clinical practice to estimate a woman's lifetime risk for breast cancer and assist in implementation of appropriate screening and risk reduction strategies. More recently, breast cancer polygenic risk scores (PRS) have been derived and integrated into these models to further improve risk estimation. While breast cancer PRS have been offered by two clinical diagnostic laboratories since 2017, little is known about the extent to which genetic counselors are ordering breast cancer PRS or incorporating the results into patient management. This study surveyed U.S. cancer genetic counselors from October 2019 to January 2020 to identify and understand their current practices with breast cancer PRS, to determine the impact of breast cancer PRS on patient management, and to anticipate future genetic counselor practices with breast cancer PRS. Fewer than half of respondents (43%, 51/120) had ordered breast cancer PRS and approximately one-third (35%, 16/46) reported that the PRS had changed their medical management recommendations. The majority of cancer genetic counselors had not ordered PRS, most commonly due to (a) lack of clinical guidelines (90%, 60/67), (b) insufficient evidence of clinical utility (88%, 59/67), and (c) lack of availability for patients of non-European ancestry (70%, 47/67). Of genetic counselors who had not ordered breast cancer PRS, only 10% (7/68) did not believe they would order PRS in the future. This is the first study to characterize genetic counselors' experiences with breast cancer PRS. Results from this study indicate that although breast cancer PRS have been clinically available for patients for several years, most cancer genetic counselors are not yet convinced they are ready to be incorporated into patient care.

High Rate of Recurrent De Novo Mutations in Developmental and Epileptic Encephalopathies.

Developmental and epileptic encephalopathy (DEE) is a group of conditions characterized by the co-occurrence of epilepsy and intellectual disability (ID), typically with developmental plateauing or regression associated with frequent epileptiform activity. The cause of DEE remains unknown in the majority of cases. We performed whole-genome sequencing (WGS) in 197 individuals with unexplained DEE and pharmaco-resistant seizures and in their unaffected parents. We focused our attention on de novo mutations (DNMs) and identified candidate genes containing such variants. We sought to identify additional subjects with DNMs in these genes by performing targeted sequencing in another series of individuals with DEE and by mining various sequencing datasets. We also performed meta-analyses to document enrichment of DNMs in candidate genes by leveraging our WGS dataset with those of several DEE and ID series. By combining these strategies, we were able to provide a causal link between DEE and the following genes: NTRK2, GABRB2, CLTC, DHDDS, NUS1, RAB11A, GABBR2, and SNAP25. Overall, we established a molecular diagnosis in 63/197 (32%) individuals in our WGS series. The main cause of DEE in these individuals was de novo point mutations (53/63 solved cases), followed by inherited mutations (6/63 solved cases) and de novo CNVs (4/63 solved cases). De novo missense variants explained a larger proportion of individuals in our series than in other series that were primarily ascertained because of ID. Moreover, these DNMs were more frequently recurrent than those identified in ID series. These observations indicate that the genetic landscape of DEE might be different from that of ID without epilepsy.

Widening of the genetic and clinical spectrum of Lamb-Shaffer syndrome, a neurodevelopmental disorder due to SOX5 haploinsufficiency.

PURPOSE: Lamb-Shaffer syndrome (LAMSHF) is a neurodevelopmental disorder described in just over two dozen patients with heterozygous genetic alterations involving SOX5, a gene encoding a transcription factor regulating cell fate and differentiation in neurogenesis and other discrete developmental processes. The genetic alterations described so far are mainly microdeletions. The present study was aimed at increasing our understanding of LAMSHF, its clinical and genetic spectrum, and the pathophysiological mechanisms involved. METHODS: Clinical and genetic data were collected through GeneMatcher and clinical or genetic networks for 41 novel patients harboring various types ofSOX5 alterations. Functional consequences of selected substitutions were investigated. RESULTS: Microdeletions and truncating variants occurred throughout SOX5. In contrast, most missense variants clustered in the pivotal SOX-specific high-mobility-group domain. The latter variants prevented SOX5 from binding DNA and promoting transactivation in vitro, whereas missense variants located outside the high-mobility-group domain did not. Clinical manifestations and severity varied among patients. No clear genotype-phenotype correlations were found, except that missense variants outside the high-mobility-group domain were generally better tolerated. CONCLUSIONS: This study extends the clinical and genetic spectrum associated with LAMSHF and consolidates evidence that SOX5 haploinsufficiency leads to variable degrees of intellectual disability, language delay, and other clinical features.

Missense variants in TAF1 and developmental phenotypes: challenges of determining pathogenicity.

We recently described a new neurodevelopmental syndrome (TAF1/MRXS33 intellectual disability syndrome) (MIM# 300966) caused by pathogenic variants involving the X-linked gene TAF1, which participates in RNA polymerase II transcription. The initial study reported eleven families, and the syndrome was defined as presenting early in life with hypotonia, facial dysmorphia, and developmental delay that evolved into intellectual disability (ID) and/or autism spectrum disorder (ASD). 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 modelling 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 (HPO) 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 TAF1/MRXS33 intellectual disability 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 genes mapping to chromosome X. This article is protected by copyright. All rights reserved.

De novo mutation screening in childhood-onset cerebellar atrophy identifies gain-of-function mutations in the CACNA1G calcium channel gene.

Cerebellar atrophy is a key neuroradiological finding usually associated with cerebellar ataxia and cognitive development defect in children. Unlike the adult forms, early onset cerebellar atrophies are classically described as mostly autosomal recessive conditions and the exact contribution of de novo mutations to this phenotype has not been assessed. In contrast, recent studies pinpoint the high prevalence of pathogenic de novo mutations in other developmental disorders such as intellectual disability, autism spectrum disorders and epilepsy. Here, we investigated a cohort of 47 patients with early onset cerebellar atrophy and/or hypoplasia using a custom gene panel as well as whole exome sequencing. De novo mutations were identified in 35% of patients while 27% had mutations inherited in an autosomal recessive manner. Understanding if these de novo events act through a loss or a gain of function effect is critical for treatment considerations. To gain a better insight into the disease mechanisms causing these cerebellar defects, we focused on CACNA1G, a gene not yet associated with the early-onset form. This gene encodes the Cav3.1 subunit of T-type calcium channels highly expressed in Purkinje neurons and deep cerebellar nuclei. We identified four patients with de novo CACNA1G mutations. They all display severe motor and cognitive impairment, cerebellar atrophy as well as variable features such as facial dysmorphisms, digital anomalies, microcephaly and epilepsy. Three subjects share a recurrent c.2881G>A/p.Ala961Thr variant while the fourth patient has the c.4591A>G/p.Met1531Val variant. Both mutations drastically impaired channel inactivation properties with significantly slower kinetics (∼5 times) and negatively shifted potential for half-inactivation (>10 mV). In addition, these two mutations increase neuronal firing in a cerebellar nuclear neuron model and promote a larger window current fully inhibited by TTA-P2, a selective T-type channel blocker. This study highlights the prevalence of de novo mutations in early-onset cerebellar atrophy and demonstrates that A961T and M1531V are gain of function mutations. Moreover, it reveals that aberrant activity of Cav3.1 channels can markedly alter brain development and suggests that this condition could be amenable to treatment.

Genetic counseling in CHARGE syndrome: Diagnostic evaluation through follow up.

CHARGE syndrome (CS) is a complex genetic disorder causing multiple birth defects and sensory deficits (hearing, vision, balance, smell). Genetic counseling in CS must include not only the provision of factual information about CS, its cause, and inheritance, but also information about the developmental implications of CS features, referral to appropriate resources, and assistance with psychosocial adaptation to this information. CS should be considered in patients with any of the major diagnostic features: coloboma, choanal atresia, semicircular canal anomalies, or cranial nerve anomalies. The prime candidates in the differential are 22q11.2 deletion and Kabuki syndromes. Evaluation of features of CS, dysmorphology examination, and genetic testing can usually distinguish between the three conditions. Genetic counseling is important from early on, to help the family understand the process of genetic diagnosis, to interpret information coming from other specialists and to provide support and resources. Parents can easily be overwhelmed with the complexity of issues facing their child at diagnosis and in the future. CS is a substantial burden on a child, with high early mortality, multiple illnesses, hospitalizations and surgeries, and apparent medical fragility throughout life. The medical complexity of CS disrupts family life and contributes to delayed development. Multiple sensory deficits (impaired vision, hearing, and balance) further contribute to delayed motor and language development despite many individuals with CS having normal intelligence. Early referral to specialists in deafblindness and sensory deficits is essential. Resources are available to assist genetic counselors in diagnosis, follow-up, and management of patients with CS.

The Exome Clinic and the role of medical genetics expertise in the interpretation of exome sequencing results.

PURPOSE: Evaluation of the clinician's role in the optimal interpretation of clinical exome sequencing (ES) results. METHODS: Retrospective chart review of the first 155 patients who underwent clinical ES in our Exome Clinic and direct interaction with the ordering geneticist to evaluate the process of interpretation of results. RESULTS: The most common primary indication was neurodevelopmental problems (~66%), followed by multiple congenital anomalies (~10%). Based on sequencing data, the overall diagnostic yield was 36%. After assessment by the medical geneticist, incorporation of detailed phenotypic and molecular data, and utilization of additional diagnostic modalities, the final diagnostic yield increased to 43%. Seven patients in our cohort were included in initial case series that described novel genetic syndromes, and 23% of patients were involved in subsequent research studies directly related to their results or involved in efforts to move beyond clinical ES for diagnosis. Clinical management was directly altered due to the ES findings in 12% of definitively diagnosed cases. CONCLUSIONS: Our results emphasize the usefulness of ES, demonstrate the significant role of the medical geneticist in the diagnostic process of patients undergoing ES, and illustrate the benefits of postanalytical diagnostic work-up in solving the "diagnostic odyssey." Genet Med advance online publication 02 March 2017.

Mutations in the PH Domain of DNM1 are associated with a nonepileptic phenotype characterized by developmental delay and neurobehavioral abnormalities.

BACKGROUND: Dynamin 1 is a protein involved in the synaptic vesicle cycle, which facilitates the exocytosis of neurotransmitters necessary for normal signaling and development in the central nervous system. Pathogenic variants in DNM1 have been implicated in global developmental delay (DD), severe intellectual disability (ID), and notably, epileptic encephalopathy. All previously reported DNM1 pathogenic variants causing this severe phenotype occur in the GTPase and Middle domains of the dynamin 1 protein. METHODS: We used whole-exome sequencing to characterize the molecular basis of DD and autistic symptoms in two identical siblings. RESULTS: The twin siblings exhibit mild to moderate ID and autistic symptoms but no epileptic encephalopathy. Exome sequencing revealed a genetic variant, c.1603A>G (p.Lys535Glu), in the PH domain of dynamin 1. Previous in vitro studies showed that mutations at Lys535 inhibit endocytosis and impair PH loop binding to PIP2. CONCLUSIONS: Our data suggest a previously undescribed milder phenotype associated with a missense genetic variant in the PH domain of dynamin 1.