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The variability of SMARCA4‐related Coffin–Siris syndrome: Do nonsense candidate variants add to milder phenotypes?
Abstract
SMARCA4 encodes a central ATPase subunit in the BRG1‐/BRM‐associated factors (BAF) or polybromo‐associated BAF (PBAF) complex in humans, which is responsible in part for chromatin remodeling and transcriptional regulation. Variants in this and other genes encoding BAF/PBAF complexes have been implicated in Coffin–Siris Syndrome, a multiple congenital anomaly syndrome classically characterized by learning and developmental differences, coarse facial features, hypertrichosis, and underdevelopment of the fifth digits/nails of the hands and feet. Individuals with SMARCA4 variants have been previously reported and appear to display a variable phenotype. We describe here a cohort of 15 unrelated individuals with SMARCA4 variants from the Coffin–Siris syndrome/BAF pathway disorders registry who further display variability in severity and degrees of learning impairment and health issues. Within this cohort, we also report two individuals with novel nonsense variants who appear to have a phenotype of milder learning/behavioral differences and no organ‐system involvement.
... Синдром Коффіна-Сіріса є не тільки генетично гетерогенним розладом, але й характеризується широким спектром клінічних ознак [5]. Основними ознаками є затримка розвитку або когнітивних функцій (у всіх пацієнтів) від легкого до тяжкого ступеня, гіпоплазія або аплазія нігтя/дистальної фаланги п'ятого пальця (майже у всіх пацієнтів при народженні) та грубі риси обличчя, які посилюються з часом [1,8]. ...
... У першому випадку -згинальна деформація мізинця та широкі кисті. Недавні дослідження свідчать, що гіпоплазія або аплазія нігтя/дистальної фаланги п'ятого пальця не є обов'язковою знахідкою, а згинальну деформацію мізинця описано і в інших пацієнтів із варіантом у гені SMARCA4 [5]. ...
... Науковці зазначають, що ступінь затримки розвитку також залежить від залученого гена [5]. Так, особи зі змінами в генах, які кодують групу білків ARID, зазвичай мають більше мовленнєвих/комунікативних труднощів, ніж особи зі змінами в генах, пов'язаних із SMARC, які можуть мати більше аномалій внутрішніх органів [1,9]. ...
Coffin-Siris syndrome encompasses a group of genetic disorders with a wide spectrum of phenotypic features, characterized by varying degrees of intellectual disability, developmental delay, and other clinical signs. Among the most common features are the aplasia or hypoplasia of the distal phalanx or nail of the fifth finger and coarse facial features. The aim of the work is to increase awareness among physicians about the diagnosis of Coffin-Siris syndrome and expand knowledge of the spectrum of clinical features of the disease based on the presentation of two clinical cases. We presented two clinical cases of Coffin-Siris syndrome caused by variants in the SMARCA4 and DPF2 genes. In both cases, there is a delay in speech development, cognitive impairments of varying severity, behavioral changes, characteristic dysmorphic features, and changes in the hands and/or feet, especially in the little finger. Distal phalanx nail hypoplasia of the fifth finger was present in the case associated with a variant in the DPF2 gene. In the patient with a variant in the SMARCA4 gene, flexion deformity of the little finger and broad palms were observed. Pachydermodactyly and a congenital kidney defect were also diagnosed in this patient. Thus, genetic diagnosis allows for timely identification, preventing complications and avoiding unnecessary investigations. Accumulating clinical data on rare diseases and their systematization contribute to research development and, subsequently, to determining optimal therapeutic approaches. The identified genetic factor in the proband ensures genetic counseling for the family and determining risk indicators for pathologies in offspring. The research was carried out in accordance with the principles of the Helsinki Declaration. The informed consent of the patient was obtained for conducting the studies. No conflict of interests was declared by the authors.
... Las variantes en la línea germinal de SMARCA4 generan el CSS [6]. Adicionalmente, las alteraciones en SMARCA4 son una de las principales etiologías de neoplasias malignas (5-7%) por mutaciones inactivadoras [7], además se une a BRCA1 y regula la expresión de la proteína tumorogénica CD44. ...
... En este estudio el gen identificado es el SMARCA4, el cual es una subunidad ATPasa del complejo BAF/BAF [7], siendo todas sus variantes sin sentido en dominios funcionales altamente conservados de la proteína. Su fenotipo parece estar relacionado con un deterioro menos severo del crecimiento, siendo leve antes del nacimiento y de leve a moderado después de este, rasgos faciales menos toscos, frecuentemente dificultad para la alimentación [11] y se ha asociado fuertemente con varios tumores debido a su papel fundamental en la reparación del DNA, lo cual al producirse una mutación en la línea germinal predispone a mayor riesgo de neoplasias y a una segunda alteración somática; entre los tumores se encuentra el neuroblastoma, tumores teratoides, rabdoides, carcinoma de ovario de células pequeñas de tipo hipercalcémico, sarcoma uterino, carcinoma de próstata [7]. ...
... En este estudio el gen identificado es el SMARCA4, el cual es una subunidad ATPasa del complejo BAF/BAF [7], siendo todas sus variantes sin sentido en dominios funcionales altamente conservados de la proteína. Su fenotipo parece estar relacionado con un deterioro menos severo del crecimiento, siendo leve antes del nacimiento y de leve a moderado después de este, rasgos faciales menos toscos, frecuentemente dificultad para la alimentación [11] y se ha asociado fuertemente con varios tumores debido a su papel fundamental en la reparación del DNA, lo cual al producirse una mutación en la línea germinal predispone a mayor riesgo de neoplasias y a una segunda alteración somática; entre los tumores se encuentra el neuroblastoma, tumores teratoides, rabdoides, carcinoma de ovario de células pequeñas de tipo hipercalcémico, sarcoma uterino, carcinoma de próstata [7]. Nuestra variante no se ha asociado con bases de datos sobre neoplasias, pero si se evidencio que es una variante recurrente. ...
El síndrome de Coffin-Siris (CSS) es de herencia autosómica dominante y la mayoría de los casos reportados son de novo. En la literatura global existen reportados menos de 300 casos, dado la escasez en el número de casos su prevalencia e incidencia exactas son desconocidas. Sus manifestaciones clínicas se caracterizan por la triada de discapacidad intelectual, rasgos faciales toscos e hipoplasia del quinto dedo. En este estudio describiremos el abordaje diagnóstico de un caso pediátrico de síndrome de Coffin-Siris con variante probablemente patogénica en el gen SMARCA4 con manifestaciones clásicas de esta patología y otras no tan frecuentes como el paladar hendido.
... Up till now, 48 variants of SMARCA4, including 44 single nucleotide variants (SNVs) and four deletion, have been identified in patients with neurodevelopmental-related disorders, and these variants are clustered in the catalytic core region (amino acids 766-1246) (Aref-Eshghi et al., 2018;Bogershausen & Wollnik, 2018;Bramswig et al., 2015;Coe et al., 2019;Deciphering Developmental Disorders 2017;Dsouza et al., 2019;Kosaki et al., 2020;Kosho et al., 2014;Kosho et al., 2013;Li et al., 2020;Nair et al., 2018;Retterer et al., 2016;Santen et al., 2013;Sekiguchi et al., 2019;Tsurusaki et al., 2012;Tzeng et al., 2014;Wright et al., 2019;Zhao et al., 2018). Most of these patients showed ID, speech and motor delay, and more than half of the patients showed behavioral problems. ...
... Our patients showed ID and speech delay, similar to reported patients. In contrast, autism spectrum disorders were the dominant clinical symptom in our patients (87.5%, 7/8), while the percentages of patients with behavioral problems ranged from 27% to 64% in previous reports (Kosho et al., 2014;Li et al., 2020). ...
... A frameshift variant of D485Lfs*16 was detected in a patient with abnormalities of the nervous system (Retterer et al., 2016). Two nonsense variants (Q1104* and Y1530*) seem to cause a relatively mild phenotype reported in the Li et al. group with ADHD, but both of them had craniofacial and finger/toe anomalies (Li et al., 2020). However, the Fig. 3 The clinical phenotypes of the patients with detected SMARCA4 variants. ...
Autism spectrum disorders (ASDs) are a group of neurodevelopmental-related disorders with a high genetic risk. Recently, chromatin remodeling factors have been found to be related to ASDs. SMARCA4 is such a catalytic subunit of the chromatin-remodeling complex. In this report, we identified seven novel missense variants in the SMARCA4 gene from eight pediatric patients. All eight patients had moderate to severe intellectual disability, and seven showed autistic/likely autistic features. Compared with the patients reported in the literature, our patients were less likely to show craniofacial or finger/toe anomalies. Our findings further supported that SMARCA4 is associated with ASDs. We suggest that individuals with the abovementioned phenotypes should consider genetic testing.
... Additionally, whether BRG1 is critical for OL remyelination after demyelinating injury is not known. Gaining insights into the functions of BRG1 during development and repair is critical for better understanding its role in disorders, including Coffin-Siris syndrome (CSS), which is characterized by white matter defects (Li et al., 2020;Tsurusaki et al., 2012), and related conditions collectively referred to as Coffin-Siris and BRG1-or HRBM-associated factor related (CSS-BAF) disorders (Vasko et al., 2021;Zarate et al., 2016). ...
... This insight opens avenues for developing future therapies aimed at treating demyelinating diseases. Additionally, BRG1 mutations have been linked to Coffin-Siris syndrome (CSS) (Li et al., 2020;Tsurusaki et al., 2012), a rare congenital abnormality with clinical characteristics such as intellectual disability, progressive coarsening of the face, hypertrichosis, and frequent infections. Notably, CSS patients with BRG1 mutations frequently exhibit white matter defects and partial agenesis of the corpus callosum (Vasko et al., 2021), suggesting a critical role of BRG1 for white matter development in the CNS. ...
Chromatin-remodeling protein BRG1/SMARCA4 is pivotal for establishing oligodendrocyte (OL) lineage identity. However, its functions for oligodendrocyte-precursor cell (OPC) differentiation within the postnatal brain and during remyelination remain elusive. Here, we demonstrate that Brg1 loss profoundly impairs OPC differentiation in the brain with a comparatively lesser effect in the spinal cord. Moreover, BRG1 is critical for OPC remyelination after injury. Integrative transcriptomic/genomic profiling reveals that BRG1 exhibits a dual role by promoting OPC differentiation networks while repressing OL-inhibitory cues and proneuronal programs. Furthermore, we find that BRG1 interacts with EED/PRC2 polycomb-repressive-complexes to enhance H3K27me3-mediated repression at gene loci associated with OL-differentiation inhibition and neurogenesis. Notably, BRG1 depletion decreases H3K27me3 deposition, leading to the upregulation of BMP/WNT signaling and proneurogenic genes, which suppresses OL programs. Thus, our findings reveal a hitherto unexplored spatiotemporal-specific role of BRG1 for OPC differentiation in the developing CNS and underscore a new insight into BRG1/PRC2-mediated epigenetic regulation that promotes and safeguards OL lineage commitment and differentiation.
... BRG1 mutations are associated with many cancer types and are related to developmental disorders (11,12). Recent human genome analyses have revealed that BRG1 mutations are highly associated with neurodevelopmental diseases such as Coffin-Siris syndrome (CSS, MIM 135900) and autism spectrum disorders (13)(14)(15)(16)(17). CSS is a rare congenital disorder characterized by growth deficiency, intellectual disability, coarse facial appearance, feeding difficulties in infancy and hypoplasia of the fifth distal phalanges, fingernails and toenails (18). ...
... Other than BAF complex genes, PHD finger protein 6 (PHF6), SRY-box transcription factor 11 (SOX11) and SRY-box transcription factor 4 (SOX4) are additional causative genes of CSS (21)(22)(23). CSS patients with BRG1 mutations usually exhibit abnormal brain structures (16,17). Collectively, these data raise an interesting question about the role of BRG1 in cerebral development. ...
Mutations in genes encoding subunits of the BAF (BRG1/BRM-associated factor) complex cause various neurodevelopmental diseases. However, the underlying pathophysiology remains largely unknown. Here, we analyzed the function of Brg1, a core ATPase of BAF complexes, in the developing cerebral cortex. Loss of Brg1 causes several morphological defects resembling human malformations of cortical development (MCDs), including microcephaly, cortical dysplasia, cobblestone lissencephaly, and periventricular heterotopia. We demonstrated that neural progenitor cell (NPC) renewal, neuronal differentiation, neuronal migration, apoptotic cell death, pial basement membrane, and apical junctional complexes, which are associated with MCD formation, were impaired after Brg1 deletion. Furthermore, transcriptome profiling indicated that a large number of genes were deregulated. The deregulated genes were closely related to MCD formation, and most of these genes were bound by Brg1. Cumulatively, our study indicates an essential role of Brg1 in cortical development and provides a new possible pathogenesis underlying Brg1-based BAF complex-related neurodevelopmental disorders.
... variant is in the helicase ATP-binding domain, which lies between the mutational hotspot HAS domain and the helicase C-terminal domain. 58 One other sample with a variant in the helicase domain did not match this subsignature, indicating that this is a variant-specific and not domain-specific episignature. ...
... Gabriella Maria Squeo, 35 Miya St John, 23 Christel Thauvin-Robinet, 12,13,36,37 Giovanna Traficante, 3 Pleuntje J. van der Sluijs, 33 Samantha A. Vergano, 38,39 Niels Vos, 40 Kellie K. Walden, 10 Dimitar Azmanov, 41 Tugce Balci, 42,43 Siddharth Banka, 44,45 Jozef Gecz, 46,47 Peter Henneman, 28 Jennifer A. Lee, 10 Marcel M.A.M. Mannens, 28 Tony Roscioli, 48,49,50,51 Victoria Siu,42,43 David J. Amor, 23 Gareth Baynam, 29,52,53 Eric G. Bend,54 Kym Boycott, 55,56 Nicola Brunetti-Pierri, 6,7 Philippe M. Campeau, 32 John Christodoulou, 19 David Dyment, 57 Natacha Esber, 58 Jill A. Fahrner, 59 Mark D. Fleming, 60 David Genevieve, 15 Kristin D. Kerrnohan, 55,61 Alisdair McNeill, 62 Leonie A. Menke, 34 Giuseppe Merla, 35,63 Paolo Prontera, 64 Cheryl Rockman-Greenberg, 65 Charles Schwartz, 10 Steven A. Skinner, 10 Roger E. Stevenson, 10 Antonio Vitobello, 12,36 Marco Tartaglia, 8 Marielle Alders, 28 Matthew L. Tedder, 10 ...
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.
... Brain MRI was performed in 34 probands, and 14 (41%) had abnormal imaging findings, including delayed myelination, ventricle asymmetry, atrophy of the cerebral hemisphere, absent olfactory bulbs and tracts, dysmorphic inferior Introduction Neurodevelopmental disorders (NDDs) are a heterogeneous group of neurological and related neuropsychiatric conditions including intellectual disability (ID), autism spectrum disorder (ASD), and developmental and epileptic encephalopathies that manifest during childhood (1)(2)(3). Over the past 2 decades, there has been an impressive expansion with respect to gene discoveries for NDDs, revealing more than 1,500 genes in different signaling pathways (3,4), including many transcriptional regulators such as DNA/histone modifiers (5-7) and chromatin-regulatory protein complexes (8)(9)(10)(11). De novo variants are increasingly appreciated as playing a substantial causal role in the development of these disorders, and clinical laboratories now routinely report de novo variants in candidate genes not definitively associated with NDD (2,12), which further facilitates the gene discovery effort. Over the past decade, a genetic diagnosis was obtained, on average, in one-third of individuals with NDD, which is a fundamental step forward in offering biological insights into underlying molecular mechanisms and providing individuals who have an undiagnosed disorder with a prognosis, counseling for recurrence risk, and precision medicine considerations. ...
Pre-mRNA splicing is a highly coordinated process. While its dysregulation has been linked to neurological deficits, our understanding of the underlying molecular and cellular mechanisms remains limited. We implicated pathogenic variants in U2AF2 and PRPF19, encoding spliceosome subunits in neurodevelopmental disorders (NDDs), by identifying 46 unrelated individuals with 23 de novo U2AF2 missense variants (including seven recurrent variants in 30 individuals) and six individuals with de novo PRPF19 variants. Eight U2AF2 variants dysregulated splicing of a model substrate. Neuritogenesis was reduced in human neurons differentiated from human pluripotent stem cells carrying two U2AF2 hyper-recurrent variants. Neural loss of function of the Drosophila orthologs, U2af50 and Prp19, led to lethality, abnormal mushroom body (MB) patterning, and social deficits, differentially rescued by wild-type and mutant U2AF2 or PRPF19. Transcriptome profiling revealed splicing substrates or effectors (including Rbfox1, a third splicing factor), which rescued MB defects in U2af50 deficient flies. Upon re-analysis of negative clinical exomes followed by data sharing, we further identified six NDD patients carrying RBFOX1 missense variants which, by in vitro testing, showed loss of function. Our study implicates three splicing factors as NDD causative genes and establishes a genetic network with hierarchy underlying human brain development and function.
... 29 Interestingly, 34% of patients with CSS carrying a mutation in SMARCA4 also had hearing loss. 30 It is noteworthy that all previously reported mutations in SMARCA4 causing CSS were located in more proximal regions of the protein, particularly in and around the two helicase domains of BRG1, 31 whereas in our patients the mutation occurred near the BROMO domain, close to the c-terminus of the protein ( figure 2D). This may explain the absence of any of the severe clinical features of CSS in our patients who presented with isolated otosclerosis. ...
Background
Otosclerosis is a common cause of adult-onset progressive hearing loss, affecting 0.3%–0.4% of the population. It results from dysregulation of bone homeostasis in the otic capsule, most commonly leading to fixation of the stapes bone, impairing sound conduction through the middle ear. Otosclerosis has a well-known genetic predisposition including familial cases with apparent autosomal dominant mode of inheritance. While linkage analysis and genome-wide association studies suggested an association with several genomic loci and with genes encoding structural proteins involved in bone formation or metabolism, the molecular genetic pathophysiology of human otosclerosis is yet mostly unknown.
Methods
Whole-exome sequencing, linkage analysis, generation of CRISPR mutant mice, hearing tests and micro-CT.
Results
Through genetic studies of kindred with seven individuals affected by apparent autosomal dominant otosclerosis, we identified a disease-causing variant in SMARCA4 , encoding a key component of the PBAF chromatin remodelling complex. We generated CRISPR-Cas9 transgenic mice carrying the human mutation in the mouse SMARCA4 orthologue. Mutant Smarca4 +/E1548K mice exhibited marked hearing impairment demonstrated through acoustic startle response and auditory brainstem response tests. Isolated ossicles of the auditory bullae of mutant mice exhibited a highly irregular structure of the incus bone, and their in situ micro-CT studies demonstrated the anomalous structure of the incus bone, causing disruption in the ossicular chain.
Conclusion
We demonstrate that otosclerosis can be caused by a variant in SMARCA4 , with a similar phenotype of hearing impairment and abnormal bone formation in the auditory bullae in transgenic mice carrying the human mutation in the mouse SMARCA4 orthologue.
... Within a cohort of 15 unrelated individuals on the CSS pathway disorders registry, who were carriers of a SMARCA4 variant, they showed differences in the severity and number of learning disabilities and health problems. Two of them with novel nonsense variants appeared to acquire a non-organ/system affected phenotype with minor learning/behavioral differences [80]. A recent report described a Chinese woman presenting with on the CSS pathway disorders registry, who were carriers of a SMARCA4 variant, they showed differences in the severity and number of learning disabilities and health problems. ...
Different fates of neural stem/progenitor cells (NSPCs) and their progeny are determined by the gene regulatory network, where a chromatin-remodeling complex affects synergy with other regulators. Here, we review recent research progress indicating that the BRG1/BRM-associated factor (BAF) complex plays an important role in NSPCs during neural development and neural developmental disorders. Several studies based on animal models have shown that mutations in the BAF complex may cause abnormal neural differentiation, which can also lead to various diseases in humans. We discussed BAF complex subunits and their main characteristics in NSPCs. With advances in studies of human pluripotent stem cells and the feasibility of driving their differentiation into NSPCs, we can now investigate the role of the BAF complex in regulating the balance between self-renewal and differentiation of NSPCs. Considering recent progress in these research areas, we suggest that three approaches should be used in investigations in the near future. Sequencing of whole human exome and genome-wide association studies suggest that mutations in the subunits of the BAF complex are related to neurodevelopmental disorders. More insight into the mechanism of BAF complex regulation in NSPCs during neural cell fate decisions and neurodevelopment may help in exploiting new methods for clinical applications.
... [1][2][3][4][5][6] In the current literature, most patients present with no prenatal ultrasound (US) anomalies. Congenital diaphragmatic hernia, 7 hypoplastic left heart syndrome (HLHS), 8 and intrauterine growth restriction (IUGR) 9 have been occasionally described, whereas agenesis of the corpus callosum (ACC) 10 is observed more frequently. ...
... The majority of SMARCA4 variants associated with Coffin-Siris syndrome 4 are missense or small in-frame deletions and have been hypothesized to exert dominant-negative or gain-offunction effects (Kosho et al., 2014). However, recent reports have indicated that inactivating variants may cause a milder version of this syndrome (Errichiello et al., 2017;Li et al., 2020;Mitrakos et al., 2020). Subject 10 is a male with a right-sided CDH who died shortly after birth. ...
Congenital diaphragmatic hernia (CDH) can occur in isolation or in conjunction with other birth defects (CDH+). A molecular etiology can only be identified in a subset of CDH cases. This is due, in part, to an incomplete understanding of the genes that contribute to diaphragm development. Here, we used clinical and molecular data from 36 individuals with CDH+ who are cataloged in the DECIPHER database to identify genes that may play a role in diaphragm development and to discover new phenotypic expansions. Among this group, we identified individuals who carried putatively deleterious sequence or copy number variants affecting CREBBP, SMARCA4, UBA2, and USP9X. The role of these genes in diaphragm development was supported by their expression in the developing mouse diaphragm, their similarity to known CDH genes using data from a previously published and validated machine learning algorithm, and/or the presence of CDH in other individuals with their associated genetic disorders. Our results demonstrate how data from DECIPHER, and other public databases, can be used to identify new phenotypic expansions and suggest that CREBBP, SMARCA4, UBA2, and USP9X play a role in diaphragm development.
... [1][2][3][4][5][6] In the current literature, most patients present with no prenatal ultrasound (US) anomalies. Congenital diaphragmatic hernia, 7 hypoplastic left heart syndrome (HLHS), 8 and intrauterine growth restriction (IUGR) 9 have been occasionally described, whereas agenesis of the corpus callosum (ACC) 10 is observed more frequently. ...
Purpose
Genome-wide sequencing is increasingly being performed during pregnancy to identify the genetic cause of congenital anomalies. The interpretation of prenatally identified variants can be challenging and is hampered by our often limited knowledge of prenatal phenotypes. To better delineate the prenatal phenotype of Coffin-Siris syndrome (CSS), we collected clinical data from patients with a prenatal phenotype and a pathogenic variant in one of the CSS-associated genes.
Methods
Clinical data was collected through an extensive web-based survey.
Results
We included 44 patients with a variant in a CSS-associated gene and a prenatal phenotype; 9 of these patients have been reported before. Prenatal anomalies that were frequently observed in our cohort include hydrocephalus, agenesis of the corpus callosum, hypoplastic left heart syndrome, persistent left vena cava, diaphragmatic hernia, renal agenesis, and intrauterine growth restriction. Anal anomalies were frequently identified after birth in patients with ARID1A variants (6/14, 43%). Interestingly, pathogenic ARID1A variants were much more frequently identified in the current prenatal cohort (16/44, 36%) than in postnatal CSS cohorts (5%-9%).
Conclusion
Our data shed new light on the prenatal phenotype of patients with pathogenic variants in CSS genes.
... Mutations in genes that encode proteins of the SWI/SNF complex, called the BAF complex in mammals, have been found to be responsible for CSS by whole exome sequencing and pathway-based genetic testing [136]. Some studies have reported that germline variants in SMARCB1 and SMARCA4 were involved in the CSS phenotype [137,138]. Germline PVs of SMARCB1 (n = 14, 7%) and SMARCA4 (n = 32, 15%) were also reported to be identified among 208 CSS patients [139]. ...
Hereditary cancer syndromes, which are characterized by onset at an early age and an increased risk of developing certain tumors, are caused by germline pathogenic variants in tumor suppressor genes and are mostly inherited in an autosomal dominant manner. Therefore, hereditary cancer syndromes have been used as powerful models to identify and characterize susceptibility genes associated with cancer. Furthermore, clarification of the association between genotypes and phenotypes in one disease has provided insights into the etiology of other seemingly different diseases. Molecular genetic discoveries from the study of hereditary cancer syndrome have not only changed the methods of diagnosis and management, but have also shed light on the molecular regulatory pathways that are important in the development and treatment of sporadic tumors. The main cancer susceptibility syndromes that involve gynecologic cancers include hereditary breast and ovarian cancer syndrome as well as Lynch syndrome. However, in addition to these two hereditary cancer syndromes, there are several other hereditary syndromes associated with gynecologic cancers. In the present review, we provide an overview of the clinical features, and discuss the molecular genetics, of four rare hereditary gynecological cancer syndromes; Cowden syndrome, Peutz-Jeghers syndrome, DICER1 syndrome and rhabdoid tumor predisposition syndrome 2.
... Brahma-related gene 1 (Brg1, also known as Smarca4 and Baf190a) is one of the two mutually exclusive ATPase subunits of the SWI/SNF complex (Chiba et al., 1994;Wang et al., 1996). In humans, BRG1 mutations have been found in individuals suffering from intellectual disability disorders such as Coffin-Siris syndrome (Tsurusaki et al., 2012;Holsten et al., 2018;Sekiguchi et al., 2019;Li et al., 2020). Additionally, BRG1 has been proposed to play an important role in the pathogenesis of autism spectrum disorders (De Rubeis et al., 2014;Lim et al., 2017). ...
During development, gene expression is tightly controlled to facilitate the generation of the diverse cell types that form the central nervous system. Brahma-related gene 1 (Brg1, also known as Smarca4) is the catalytic subunit of the SWItch/sucrose nonfermentable (SWI/SNF) chromatin remodeling complex that regulates transcription. We investigated the role of Brg1 between embryonic day 6.5 (E6.5) and E14.5 in Sox2-positive neural stem cells (NSCs). Being without major consequences at E6.5 and E14.5, loss of Brg1 between E7.5 and E12.5 resulted in the formation of rosette-like structures in the subventricular zone, as well as morphological alterations and enlargement of neural retina (NR). Additionally, Brg1-deficient cells showed decreased survival in vitro and in vivo. Furthermore, we uncovered distinct changes in gene expression upon Brg1 loss, pointing towards impaired neuron functions, especially those involving synaptic communication and altered composition of the extracellular matrix. Comparison with mice deficient for integrase interactor 1 (Ini1, also known as Smarcb1) revealed that the enlarged NR was Brg1 specific and was not caused by a general dysfunction of the SWI/SNF complex. These results suggest a crucial role for Brg1 in NSCs during brain and eye development.
... A few additional cases of CSS have been linked to SMARCA4 PV. In a series of 15 patients with CSS and SMARCA4 PV, Li et al. reported two individuals with nonsense variants, associated with a milder phenotype; in the two male patients no tumor was reported by ages 9 and 15 years respectively [39]. By contrast, one patient with a mild CSS phenotype related to a truncating variant c.2935C > T;p.Arg979* developed a SCCOHT [40], again suggesting that detection of truncating SMARCA4 variants in females justifies surveillance for SCCOHT [40], suggesting that detection of this particular genotype in females justifies surveillance for SCCOHT. ...
The rhabdoid tumor (RT) predisposition syndromes 1 and 2 (RTPS1 and 2) are rare genetic conditions rendering young children vulnerable to an increased risk of RT, malignant neoplasms affecting the kidney, miscellaneous soft-part tissues, the liver and the central nervous system (Atypical Teratoid Rhabdoid Tumors, ATRT). Both, RTPS1&2 are due to pathogenic variants (PV) in genes encoding constituents of the BAF chromatin remodeling complex, i.e. SMARCB1 (RTPS1) and SMARCA4 (RTPS2). In contrast to other genetic disorders related to PVs in SMARCB1 and SMARCA4 such as Coffin-Siris Syndrome, RTPS1&2 are characterized by a predominance of truncating PVs, terminating transcription thus explaining a specific cancer risk. The penetrance of RTPS1 early in life is high and associated with a poor survival. However, few unaffected carriers may be encountered. Beyond RT, the tumor spectrum may be larger than initially suspected, and cancer surveillance offered to unaffected carriers (siblings or parents) and long-term survivors of RT is still a matter of discussion. RTPS2 exposes female carriers to an ill-defined risk of small cell carcinoma of the ovaries, hypercalcemic type (SCCOHT), which may appear in prepubertal females. RT surveillance protocols for these rare families have not been established. To address unresolved issues in the care of individuals with RTPS and to propose appropriate surveillance guidelines in childhood, the SIOPe Host Genome working group invited pediatric oncologists and geneticists to contribute to an expert meeting. The current manuscript summarizes conclusions of the panel discussion, including consented statements as well as non-evidence-based proposals for validation in the future.
Objective
Our objective was to examine the clinical and genetic features of Coffin-Siris syndrome resulting from a pathogenic variant in the SMARCA4 gene.
Methods
The clinical data and molecular genetic test results of a newbron with Coffin-Siris syndrome involving a pathogenic variant in the SMARCA4 gene were retrospectively analyzed, and the related literatures were reviewed.
Results
A newborn exhibited inspiratory dyspnea following birth and developmental anomalies (coarse appearance, thick hair, long eyelashes, broad nasal tip, flat nasal bridge, thin upper lip, thick lower lip, digital anomalies, cleft palate, supraglottic laryngeal chondromalacia, stenosis of the left upper bronchus and hypotonia). Whole exome sequencing revealed a heterozygous missense variant in SMARCA4 gene (NM_003072.5 c.3127C > T, p.Arg1043Trp). Parents did not find the above pathogenic variant, which was a new pathogenic variant. In addition to our case, we also retrieved 22 cases of Coffin-Siris Syndrome in SMARCA4 gene variation, which is a congenital multi-system dysfunction syndrome characterized by abnormal appearance and developmental retardation. The common otolaryngologic features of 23 patients with CSS in SMARCA4 gene variant included palate abnormalities, feeding difficulties, ear abnormalities and hearing loss.
Conclusion
Coffin-Siris syndrome is a rare genetic disease inherited in an autosomal-dominated manner. It is often associated with malformations in the otorhinolaryngologic system. This case has many common features with previously reported CSS cases with pathogenic variant in the SMARCA4 gene, which further characterizes the performance of the pathogenic variant, suggesting that palatal abnormalities may be a significant feature of the genotype. For patients with developmental abnormalities, whole-genome sequencing or whole-exome sequencing is particularly important to assist diagnosis. Currently, there is no known treatment for CSS, and individuals with CSS experience various complications affecting multiple systems.
Analysis of genomic DNA methylation by generating epigenetic signature profiles (episignatures) is increasingly being implemented in genetic diagnosis. Here we report our experience using episignature analysis to resolve both uncomplicated and complex cases of neurodevelopmental disorders (NDDs). We analyzed 97 NDDs divided into (1) a validation cohort of 59 patients with likely pathogenic/pathogenic variants characterized by a known episignature and (2) a test cohort of 38 patients harboring variants of unknown significance or unidentified variants. The expected episignature was obtained in most cases with likely pathogenic/pathogenic variants (53/59 [90%]), a revealing exception being the overlapping profile of two SMARCB1 pathogenic variants with ARID1A/B:c.6200, confirmed by the overlapping clinical features. In the test cohort, five cases showed the expected episignature, including (1) novel pathogenic variants in ARID1B and BRWD3; (2) a deletion in ATRX causing MRXFH1 X-linked mental retardation; and (3) confirmed the clinical diagnosis of Cornelia de Lange (CdL) syndrome in mutation-negative CdL patients. Episignatures analysis of the in BAF complex components revealed novel functional protein interactions and common episignatures affecting homologous residues in highly conserved paralogous proteins (SMARCA2 M856V and SMARCA4 M866V). Finally, we also found sex-dependent episignatures in X-linked disorders. Implementation of episignature profiling is still in its early days, but with increasing utilization comes increasing awareness of the capacity of this methodology to help resolve the complex challenges of genetic diagnoses.
Speech and language development are complex neurodevelopmental processes that are incompletely understood, yet current evidence suggests that speech and language disorders are prominent in those with disorders of chromatin regulation. This review aimed to unravel what is known about speech and language outcomes for individuals with chromatin-related neurodevelopmental disorders. A systematic literature search following PRISMA guidelines was conducted on 70 chromatin genes, to identify reports of speech/language outcomes across studies, including clinical reports, formal subjective measures, and standardised/objective measures. 3932 studies were identified and screened and 112 were systematically reviewed. Communication impairment was core across chromatin disorders, and specifically, chromatin writers and readers appear to play an important role in motor speech development. Identification of these relationships is important because chromatin disorders show promise as therapeutic targets due to the capacity for epigenetic modification. Further research is required using standardised and formal assessments to understand the nuanced speech/language profiles associated with variants in each gene, and the influence of chromatin dysregulation on the neurobiology of speech and language development.
Background
Whole-Exome Sequencing (WES) is a valuable tool for the molecular diagnosis of patients with a suspected genetic condition. In complex and heterogeneous diseases, the interpretation of WES variants is more challenging given the absence of diagnostic handles and other reported cases with overlapping clinical presentations.Objective
To describe candidate variants emerging from trio-WES and possibly associated with the clinical phenotype in clinically heterogeneous conditions.Methods
We performed WES in ten patients from eight families, selected because of the lack of a clear clinical diagnosis or suspicion, the presence of multiple clinical signs, and the negative results of traditional genetic tests.ResultsAlthough we identified ten candidate variants, reaching the diagnosis of these cases is challenging, given the complexity and the rarity of these syndromes and because affected genes are already associated with known genetic diseases only partially recapitulating patients’ phenotypes. However, the identification of these variants could shed light into the definition of new genotype–phenotype correlations. Here, we describe the clinical and molecular data of these cases with the aim of favoring the match with other similar cases and, hopefully, confirm our diagnostic hypotheses.Conclusion
This study emphasizes the major limitations associated with WES data interpretation, but also highlights its clinical utility in unraveling novel genotype–phenotype correlations in complex and heterogeneous undefined clinical conditions with a suspected genetic etiology.
Pathogenic variants in SMARCA4 cause Coffin-Siris syndrome (CSS) while those in SMAD6 lead to aortic valve disease and other dysmorphisms. We identified a 6-year-old Thai boy with features of CSS alongside unusual manifestations including very severe coarctation of the aorta (CoA) requiring coarctectomy in the neonatal period and bilateral radioulnar synostoses. Trio exome sequencing revealed that the patient harbored two de novo variants, a missense c.2475G > T, p.(Trp825Cys) in SMARCA4 and a nonsense c.652C > T, p.(Gln218Ter) in SMAD6. Both of which have never been previously reported. The clinical presentations in our patient are a result of the combinational features of each genetic variant: the SMARCA4 p.(Trp825Cys) variant leads to facial features of Coffin Siris syndrome and Dandy-Walker malformation, while the SMAD6 p.(Gln218Ter) variant underlies radioulnar synostosis. Interestingly, the severity of CoA in the proband is beyond the phenotypic spectra of each genetic variant and may be a result of the synergistic effects of both variants. Here, we reported a child with variants in SMARCA4 or SMAD6 with combined features of each plus a severe CoA, possibly due to an additive effect of each variant.
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 multi-class modelling 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 reclassification of variants of unknown clinical significance, and provides further insight into the molecular etiology of Mendelian conditions.
Coffin-Siris syndrome (CSS) is a developmental disability, caused by genomic variants in the gene SMARCA4, in addition to other known genes, but the full spectrum of SMARCA4 variants that can cause CSS is unknown with 40% of cases not having molecular confirmation. In this report, we identify a patient with CSS, a severe cardiac phenotype, and a novel SMARCA4 variant. There is no experimental structure of human SMARCA4, so we use molecular modeling techniques to generate a structural model of human SMARCA4. We then map known SMARCA4 variants causative of CSS and our novel variant to the model. We use the resulting information to support the interpretation that the novel variant is causative of disease in our patient. Modeling demonstrates that the variant found in our patient is in a region of SMARCA4 associated with DNA binding, as are the other known pathogenic SMARCA4 variants mapped. Because of this structural information, we discuss how these variants may be disease-causing through a dominant negative effect of disrupting DNA binding.
In neuroblastoma (NB), genetic alterations in chromatin remodeling (CRGs) and epigenetic modifier genes (EMGs) have been described. We sought to determine their frequency and clinical impact. Whole exome (WES)/whole genome sequencing (WGS) data and targeted sequencing (TSCA®) of exonic regions of 33 CRGs/EMGs were analyzed in tumor samples from 283 NB patients, with constitutional material available for 55 patients. The frequency of CRG/EMG variations in NB cases was then compared to the Genome Aggregation Database (gnomAD). The sequencing revealed SNVs/small InDels or focal CNAs of CRGs/EMGs in 20% (56/283) of all cases, occurring at a somatic level in 4 (7.2%), at a germline level in 12 (22%) cases, whereas for the remaining cases, only tumor material could be analyzed. The most frequently altered genes were ATRX (5%), SMARCA4 (2.5%), MLL3 (2.5%) and ARID1B (2.5%). Double events (SNVs/small InDels/CNAs associated with LOH) were observed in SMARCA4 (n = 3), ATRX (n = 1) and PBRM1 (n = 1). Among the 60 variations, 24 (8.4%) targeted domains of functional importance for chromatin remodeling or highly conserved domains but of unknown function. Variations in SMARCA4 and ATRX occurred more frequently in the NB as compared to the gnomAD control cohort (OR = 4.49, 95%CI: 1.63–9.97, p = 0.038; OR 3.44, 95%CI: 1.46–6.91, p = 0.043, respectively). Cases with CRG/EMG variations showed a poorer overall survival compared to cases without variations. Genetic variations of CRGs/EMGs with likely functional impact were observed in 8.4% (24/283) of NB. Our case–control approach suggests a role of SMARCA4 as a player of NB oncogenesis.
Background
Coffin–Siris syndrome (CSS) is characterized by intellectual disability, dysmorphic facial features, growth deficiency, microcephaly, and abnormalities of the fifth fingers/toes. CSS is caused by mutations in several genes of the BRG1‐associated factor pathway including SMARCA4.
Methods
Whole‐exome sequencing was performed on a 14‐year‐old female individual who presented with mild intellectual disability and dysmorphic features, tooth abnormalities, and short stature. She had brachydactyly but no aplasia or hypoplasia of the distal phalanx or nail of the fifth digit. She was also found to have retinal dystrophy that has not been previously reported in CSS.
Results
The individual presented herein was found to harbor a previously unreported de novo variant in SMARCA4.
Conclusion
This case expands the phenotypic spectrum of CSS manifestations.
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.
Coffin–Siris and Nicolaides–Baraitser syndromes (CSS and NCBRS) are Mendelian disorders caused by mutations in subunits of the BAF chromatin remodeling complex. We report overlapping peripheral blood DNA methylation epi-signatures in individuals with various subtypes of CSS (ARID1B, SMARCB1, and SMARCA4) and NCBRS (SMARCA2). We demonstrate that the degree of similarity in the epi-signatures of some CSS subtypes and NCBRS can be greater than that within CSS, indicating a link in the functional basis of the two syndromes. We show that chromosome 6q25 microdeletion syndrome, harboring ARID1B deletions, exhibits a similar CSS/NCBRS methylation profile. Specificity of this epi-signature was confirmed across a wide range of neurodevelopmental conditions including other chromatin remodeling and epigenetic machinery disorders. We demonstrate that a machine-learning model trained on this DNA methylation profile can resolve ambiguous clinical cases, reclassify those with variants of unknown significance, and identify previously undiagnosed subjects through targeted population screening.
Purpose:
Pathogenic variants in ARID1B are one of the most frequent causes of intellectual disability (ID) as determined by large-scale exome sequencing studies. Most studies published thus far describe clinically diagnosed Coffin-Siris patients (ARID1B-CSS) and it is unclear whether these data are representative for patients identified through sequencing of unbiased ID cohorts (ARID1B-ID). We therefore sought to determine genotypic and phenotypic differences between ARID1B-ID and ARID1B-CSS. In parallel, we investigated the effect of different methods of phenotype reporting.
Methods:
Clinicians entered clinical data in an extensive web-based survey.
Results:
79 ARID1B-CSS and 64 ARID1B-ID patients were included. CSS-associated dysmorphic features, such as thick eyebrows, long eyelashes, thick alae nasi, long and/or broad philtrum, small nails and small or absent fifth distal phalanx and hypertrichosis, were observed significantly more often (p < 0.001) in ARID1B-CSS patients. No other significant differences were identified.
Conclusion:
There are only minor differences between ARID1B-ID and ARID1B-CSS patients. ARID1B-related disorders seem to consist of a spectrum, and patients should be managed similarly. We demonstrated that data collection methods without an explicit option to report the absence of a feature (such as most Human Phenotype Ontology-based methods) tended to underestimate gene-related features.
Mutations in genes that encode proteins of the SWI/SNF complex, called BAF complex in mammals, cause a spectrum of disorders that ranges from syndromic intellectual disability to Coffin-Siris syndrome (CSS) to Nicolaides-Baraitser syndrome (NCBRS). While NCBRS is known to be a recognizable and restricted phenotype, caused by missense mutations in SMARCA2, the term CSS has been used lately for a more heterogeneous group of phenotypes that are caused by mutations in either of the genes ARID1B, ARID1A, ARID2, SMARCA4, SMARCB1, SMARCE1, SOX11, or DPF2. In this review, we summarize the current knowledge on the phenotypic traits and molecular causes of the above named conditions, consider the question whether a clinical distinction of the phenotypes is still adequate, and suggest the term “SWI/SNF-related intellectual disability disorders” (SSRIDDs). We will also outline important features to identify the ARID1B-related phenotype in the absence of classic CSS features, and discuss distinctive and overlapping features of the SSRIDD subtypes. Moreover, we will briefly review the function of the SWI/SNF complex in development and describe the mutational landscapes of the genes involved in SSRIDD.
SMARCA4 chromatin remodeling factor is mutated in 11% of Coffin-Siris syndrome (CSS) patients and in almost all small cell carcinoma of the ovary hypercalcemic type (SCCOHT) tumors. Missense mutations with gain-of-function or dominant-negative effects are associated with CSS, whereas inactivating mutations, leading to loss of SMARCA4 expression, have been exclusively found in SCCOHT. We applied whole exome sequencing to study a 15-year-old patient with mild CSS who concomitantly developed SCCOHT at age 13 years. Interestingly, our patient also showed congenital microphthalmia, never reported in CSS patients. We detected a de novo germline heterozygous nonsense mutation in the exon 19 of SMARCA4 (c.2935C>T;p.Arg979*), while a somatic frameshift mutation was identified in the exon 6 of the gene (c.1236_1236delC;p.Gln413Argfs*88), causing complete loss of SMARCA4 immunostaining in the tumor. The immunohistochemical findings are supported by the observation that the c.2935C>T mutant transcript was detected by RT-PCR at a much lower level than the wild-type allele in whole blood and lymphoblastoid cell line of the proband, confirming nonsense-mediated mRNA decay. Accordingly, immunoblotting demonstrated roughly half of SMARCA4 protein in the proband's cells as compared to controls. This study suggests that SMARCA4 constitutional mutations associated with CSS are not necessarily non-truncating, and that haploinsufficiency may explain milder CSS phenotypes, as previously reported for haploinsufficient ARID1B. In addition, our case supports the dual role of chromatin remodelers in developmental disorders and cancer, as well as the involvement of SMARCA4 in microphthalmia, confirming previous findings in mouse models and DECIPHER database. Finally, we speculate that mild CSS might be under-recognized in a proportion of SCCOHT patients harboring SMARCA4 mutations.
Small cell carcinoma of the ovary, hypercalcemic type, (SCCOHT) is the most common undifferentiated ovarian cancer in women aged under 40 years. SCCOHT is a monogenic disease, characterized by germline and somatic SMARCA4 mutations. Recent studies have stressed its morphological and clinical similarity to malignant rhabdoid tumours, which are usually caused by mutations in the related gene, SMARCB1. While familial tumours are rare, the incidence of germline mutations is relatively high, with up to 43% of SCCOHTs and 35% of rhabdoid tumours caused by germline mutations in SMARCA4 and SMARCB1, respectively. We report two new familial cases of SCCOHT. Affected members in both families and the associated tumours were found to carry SMARCA4 germline and somatic mutations, respectively, leading to loss of SMARCA4 protein expression in the tumours. Despite the rarity of familial SCCOHT, the high incidence of germline mutations is important to note, as without a family history of the disease, the hereditary nature of SCCOHT may be missed, especially if the mutation was inherited from the father or acquired de novo. The similarity between SCCOHT and rhabdoid tumours should be recognized, as infant carriers of SMARCA4 mutations may be at risk for these tumours in addition to SCCOHT.
During the last decade, a host of epigenetic mechanisms were found to contribute to cancer and other human diseases. Several genomic studies have revealed that ∼20% of malignancies have alterations of the subunits of polymorphic BRG-/BRM-associated factor (BAF) and Polybromo-associated BAF (PBAF) complexes, making them among the most frequently mutated complexes in cancer. Recurrent mutations arise in genes encoding several BAF/PBAF subunits, including ARID1A, ARID2, PBRM1, SMARCA4, and SMARCB1. These subunits share some degree of conservation with subunits from related ATP-dependent chromatin remodeling complexes in model organisms, in which a large body of work provides insight into their roles in cancer. Here, we review the roles of BAF- and PBAF-like complexes in these organisms, and relate these findings to recent discoveries in cancer epigenomics. We review several roles of BAF and PBAF complexes in cancer, including transcriptional regulation, DNA repair, and regulation of chromatin architecture and topology. More recent results highlight the need for new techniques to study these complexes.
Importance
Tumor genetic sequencing identifies potentially targetable genetic alterations with therapeutic implications. Analysis has concentrated on detecting tumor-specific variants, but recognition of germline variants may prove valuable as well.Objective
To estimate the burden of germline variants identified through routine clinical tumor sequencing.Design, Setting, and Participants
Patients with advanced cancer diagnoses eligible for studies of targeted agents at Memorial Sloan Kettering Cancer Center are offered tumor-normal sequencing with MSK-IMPACT, a 341-gene panel. We surveyed the germline variants seen in 187 overlapping genes with Mendelian disease associations in 1566 patients who had undergone tumor profiling between March and October 2014.Main Outcomes and Measures
The number of presumed pathogenic germline variants (PPGVs) and variants of uncertain significance per person in 187 genes associated with single-gene disorders and the proportions of individuals with PPGVs in clinically relevant gene subsets, in genes consistent with known tumor phenotypes, and in genes with evidence of second somatic hits in their tumors.Results
The mean age of the 1566 patients was 58 years, and 54% were women. Presumed pathogenic germline variants in known Mendelian disease-associated genes were identified in 246 of 1566 patients (15.7%; 95% CI, 14.0%-17.6%), including 198 individuals with mutations in genes associated with cancer susceptibility. Germline findings in cancer susceptibility genes were concordant with the individual’s cancer type in only 81 of 198 cases (40.9%; 95% CI, 34.3%-47.9%). In individuals with PPGVs retained in the tumor, somatic alteration of the other allele was seen in 39 of 182 cases (21.4%; 95% CI, 16.1%-28.0%), of which 13 cases did not show a known correlation of the germline mutation and a known syndrome. Mutations in non–cancer-related Mendelian disease genes were seen in 55 of 1566 cases (3.5%; 95% CI, 27.1%-45.4%). Almost every individual had more than 1 variant of uncertain significance (1565 of 1566 patients; 99.9%; 95% CI, 99.6%-99.9%).Conclusions and Relevance
Germline variants are common in individuals undergoing tumor-normal sequencing and may reveal otherwise unsuspected syndromic associations.
SWI/SNF complexes utilize BRG1 (also known as SMARCA4) or BRM (also known as SMARCA2) as alternative catalytic subunits with ATPase activity to remodel chromatin. These chromatin-remodeling complexes are required for mammalian development and are mutated in ~20% of all human primary tumors. Yet our knowledge of their tumor-suppressor mechanism is limited. To investigate the role of SWI/SNF complexes in the DNA-damage response (DDR), we used shRNAs to deplete BRG1 and BRM and then exposed these cells to a panel of 6 genotoxic agents. Compared to controls, the shRNA knockdown cells were hypersensitive to certain genotoxic agents that cause double-strand breaks (DSBs) associated with stalled/collapsed replication forks but not to ionizing radiation-induced DSBs that arise independently of DNA replication. These findings were supported by our analysis of DDR kinases, which demonstrated a more prominent role for SWI/SNF in the activation of the ATR-Chk1 pathway than the ATM-Chk2 pathway. Surprisingly, γH2AX induction was attenuated in shRNA knockdown cells exposed to a topoisomerase II inhibitor (etoposide) but not to other genotoxic agents including IR. However, this finding is compatible with recent studies linking SWI/SNF with TOP2A and TOP2BP1. Depletion of BRG1 and BRM did not result in genomic instability in a tumor-derived cell line but did result in nucleoplasmic bridges in normal human fibroblasts. Taken together, these results suggest that SWI/SNF tumor-suppressor activity involves a role in the DDR to attenuate replicative stress and genomic instability. These results may also help to inform the selection of chemotherapeutics for tumors deficient for SWI/SNF function.
To facilitate precision medicine and whole genome annotation, we developed a machine learning technique that scores how strongly genetic variants affect RNA splicing, whose alteration contributes to many diseases. Analysis of over 650,000 intronic and exonic variants reveals widespread patterns of mutation-driven aberrant splicing. Intronic disease mutations alter splicing nine times more often than common variants, and missense exonic disease mutations that least impact protein function are five times more likely to alter splicing than others. Tens of thousands of disease-causing mutations are detected, including those involved in cancers and spinal muscular atrophy. Examination of intronic and exonic variants found using whole genome sequencing of individuals with autism reveals mis-spliced genes with neurodevelopmental phenotypes. Our approach provides evidence for causal variants and should enable new discoveries in precision medicine.
Copyright © 2014, American Association for the Advancement of Science.
Small cell carcinoma of the ovary, hypercalcemic type (SCCOHT) is a rare, highly aggressive form of ovarian cancer primarily diagnosed in young women. We identified inactivating biallelic SMARCA4 mutations in 100% of the 12 SCCOHT tumors examined. Protein studies confirmed loss of SMARCA4 expression, suggesting a key role for the SWI/SNF chromatin-remodeling complex in SCCOHT.
Small cell carcinoma of the ovary, hypercalcemic type (SCCOHT) is the most common undifferentiated ovarian malignancy in women under 40 years of age. We sequenced the exomes of six individuals from three families with SCCOHT. After discovering segregating deleterious germline mutations in SMARCA4 in all three families, we tested DNA from a fourth affected family, which also carried a segregating SMARCA4 germline mutation. All the familial tumors sequenced harbored either a somatic mutation or loss of the wild-type allele. Immunohistochemical analysis of these cases and additional familial and non-familial cases showed loss of SMARCA4 (BRG1) protein in 38 of 40 tumors overall. Sequencing of cases with available DNA identified at least one germline or somatic deleterious SMARCA4 mutation in 30 of 32 cases. Additionally, the SCCOHT cell line BIN-67 had biallelic deleterious mutations in SMARCA4. Our findings identify alterations in SMARCA4 as the major cause of SCCOHT, which could lead to improvements in genetic counseling and new treatment approaches.
Introduction: Small cell carcinoma of the ovary of hypercalcemic type (SCCOHT) is arguably the most aggressive ovarian cancer. Most patients are diagnosed at an advanced stage, do not respond to chemotherapy, and die of disease within 1-2 years. It affects children and young women and is reported to occur in families. The cause of the disease is poorly understood. Therefore, we used next generation sequencing technology to identify the genetic basis of the disease.
Method: We performed whole genome, whole exome sequencing or targeted sequencing on tumours and germline samples from 17 SCCOHT patients and on the SCCOHT cell line BIN-67 and SCCOHT-1. Immunohistochemistry (IHC) was performed on formalin-fixed, paraffin-embedded tumors from 23 patients and on a tissue microarray of 485 primary ovarian tumours of other subtypes. BIN-67 cells harbouring biallelic inactivation of SMARCA4 were transduced with a lentivirus expressing wild type SMARCA4.
Result: We identified inactivating germline and somatic mutations in the SWI/SNF chromatin-remodeling gene SMARCA4 in 79% (11/14) of SCCOHT patients, 2 of whom bore germline mutations, and in both cell lines. SMARCA4 protein was lost in 87% (20/23) of SCCOHT tumours but in only 0.4% (2/485) of other primary ovarian tumours. Reintroduction of wild-type SMARCA4 into BIN-67 cells resulted in altered cell morphology and growth arrest.
Conclusions: The mutation spectrum, IHC profile and cell culture phenotype implicate SMARCA4 as a critical tumour suppressor in SCCOHT pathogenesis.
Citation Format: Pilar Ramos, Anthony N. Karnezis, David W. Craig, Aleksandar Sekulic, Megan L. Russell, William P.D. Hendricks, Jason J. Corneveaux, Michael T. Barrett, Karey Shumansky, Yidong Yang, Sohrab P. Shah, Leah M. Prentice, Marco A. Marra, Jeffrey Kiefer, Victoria L. Zismann, Troy A. McEachron, Bodour Salhia, Jaime Prat, Blaise A. Clarke, Joseph G. Pressey, John H. Farley, Stephen P. Anthony, Richard B.S. Roden, Heather E. Cunliffe, David G. Huntsman, Jeffrey M. Trent. Small cell carcinoma of the ovary, hypercalcemic type displays frequent inactivating germline and somatic mutations in SMARCA4 [abstract]. In: Proceedings of the 10th Biennial Ovarian Cancer Research Symposium; Sep 8-9, 2014; Seattle, WA. Philadelphia (PA): AACR; Clin Cancer Res 2015;21(16 Suppl):Abstract nr POSTER-BIOL-1327.
Ovarian tumors from two patients, compatible by histological and immunohistochemical criteria with small cell carcinoma of hypercalcemic type (SCCHT) (WT1+, EMA dispersed+, synaptophysin+ or dispersed+), were extensively sampled in order to find clues to their histogenesis. Subsequently, small foci of immature teratoma were found in both of them (in 1/122 and in 3/80 tumor sections). In one case, microfoci of yolk sac tumor were also present within the teratoma area as well as in the background of the small cell tumor population - in the primary tumor and in omental metastasis. We found a resemblance of the microscopic patterns of SCCHT and atypical teratoid/rhabdoid tumor (AT/RT) of the central nervous system, and this prompted us to evaluate INI-1 and SMARCA4 immunohistochemical expression, because their alternative loss is regarded as a molecular hallmark of AT/RT. INI-1 expression was retained, while that of SMARCA4 was lost. We therefore analyzed tumor DNA by PCR amplification and sequencing for mutations in the SMARCA4 gene (NG_011556.1), which were identified in both tumors (c.2184_2206del; nonsense c.3277C>T - both in one tumor; nonsense c.3760G>T in another tumor). These data suggest that SCCHT is most likely an embryonal tumor originating from immature teratoma and related to malignant rhabdoid tumor. Further analyses are necessary to determine whether the tumors diagnosed as SCCHT constitute a homogeneous group or represent more than one entity.
Genome sequencing projects are discovering millions of genetic variants in humans, and interpretation of their functional effects is essential for understanding the genetic basis of variation in human traits. Here we report sequencing and deep analysis of messenger RNA and microRNA from lymphoblastoid cell lines of 462 individuals from the 1000 Genomes Project-the first uniformly processed high-throughput RNA-sequencing data from multiple human populations with high-quality genome sequences. We discover extremely widespread genetic variation affecting the regulation of most genes, with transcript structure and expression level variation being equally common but genetically largely independent. Our characterization of causal regulatory variation sheds light on the cellular mechanisms of regulatory and loss-of-function variation, and allows us to infer putative causal variants for dozens of disease-associated loci. Altogether, this study provides a deep understanding of the cellular mechanisms of transcriptome variation and of the landscape of functional variants in the human genome.
Motivation:
Accurate alignment of high-throughput RNA-seq data is a challenging and yet unsolved problem because of the non-contiguous transcript structure, relatively short read lengths and constantly increasing throughput of the sequencing technologies. Currently available RNA-seq aligners suffer from high mapping error rates, low mapping speed, read length limitation and mapping biases.
Results:
To align our large (>80 billon reads) ENCODE Transcriptome RNA-seq dataset, we developed the Spliced Transcripts Alignment to a Reference (STAR) software based on a previously undescribed RNA-seq alignment algorithm that uses sequential maximum mappable seed search in uncompressed suffix arrays followed by seed clustering and stitching procedure. STAR outperforms other aligners by a factor of >50 in mapping speed, aligning to the human genome 550 million 2 × 76 bp paired-end reads per hour on a modest 12-core server, while at the same time improving alignment sensitivity and precision. In addition to unbiased de novo detection of canonical junctions, STAR can discover non-canonical splices and chimeric (fusion) transcripts, and is also capable of mapping full-length RNA sequences. Using Roche 454 sequencing of reverse transcription polymerase chain reaction amplicons, we experimentally validated 1960 novel intergenic splice junctions with an 80-90% success rate, corroborating the high precision of the STAR mapping strategy.
Availability and implementation:
STAR is implemented as a standalone C++ code. STAR is free open source software distributed under GPLv3 license and can be downloaded from http://code.google.com/p/rna-star/.
We describe a new computer program, SnpEff, for rapidly categorizing the effects of variants in genome sequences. Once a genome is sequenced, SnpEff annotates variants based on their genomic locations and predicts coding effects. Annotated genomic locations include intronic, untranslated region, upstream, downstream, splice site, or intergenic regions. Coding effects such as synonymous or non-synonymous amino acid replacement, start codon gains or losses, stop codon gains or losses, or frame shifts can be predicted. Here the use of SnpEff is illustrated by annotating ~356,660 candidate SNPs in ~117 Mb unique sequences, representing a substitution rate of ~1/305 nucleotides, between the Drosophila melanogaster w(1118); iso-2; iso-3 strain and the reference y(1); cn(1) bw(1) sp(1) strain. We show that ~15,842 SNPs are synonymous and ~4,467 SNPs are non-synonymous (N/S ~0.28). The remaining SNPs are in other categories, such as stop codon gains (38 SNPs), stop codon losses (8 SNPs), and start codon gains (297 SNPs) in the 5'UTR. We found, as expected, that the SNP frequency is proportional to the recombination frequency (i.e., highest in the middle of chromosome arms). We also found that start-gain or stop-lost SNPs in Drosophila melanogaster often result in additions of N-terminal or C-terminal amino acids that are conserved in other Drosophila species. It appears that the 5' and 3' UTRs are reservoirs for genetic variations that changes the termini of proteins during evolution of the Drosophila genus. As genome sequencing is becoming inexpensive and routine, SnpEff enables rapid analyses of whole-genome sequencing data to be performed by an individual laboratory.
Cellular memory is provided by two counteracting groups of chromatin proteins termed Trithorax group (TrxG) and Polycomb group (PcG) proteins. TrxG proteins activate transcription and are perhaps best known because of the involvement of the TrxG protein MLL in leukaemia. However, in terms of molecular analysis, they have lived in the shadow of their more famous counterparts, the PcG proteins. Recent advances have improved our understanding of TrxG protein function and demonstrated that the heterogeneous group of TrxG proteins is of critical importance in the epigenetic regulation of the cell cycle, senescence, DNA damage and stem cell biology.
High-throughput sequencing platforms are generating massive amounts of genetic variation data for diverse genomes, but it
remains a challenge to pinpoint a small subset of functionally important variants. To fill these unmet needs, we developed
the ANNOVAR tool to annotate single nucleotide variants (SNVs) and insertions/deletions, such as examining their functional
consequence on genes, inferring cytogenetic bands, reporting functional importance scores, finding variants in conserved regions,
or identifying variants reported in the 1000 Genomes Project and dbSNP. ANNOVAR can utilize annotation databases from the
UCSC Genome Browser or any annotation data set conforming to Generic Feature Format version 3 (GFF3). We also illustrate a
‘variants reduction’ protocol on 4.7 million SNVs and indels from a human genome, including two causal mutations for Miller
syndrome, a rare recessive disease. Through a stepwise procedure, we excluded variants that are unlikely to be causal, and
identified 20 candidate genes including the causal gene. Using a desktop computer, ANNOVAR requires ∼4 min to perform gene-based
annotation and ∼15 min to perform variants reduction on 4.7 million variants, making it practical to handle hundreds of human
genomes in a day. ANNOVAR is freely available at http://www.openbioinformatics.org/annovar/.
A rare subset of aggressive SMARCA4-deficient uterine sarcomas has been recently proposed, with only a limited number of cases having been previously described. Here, we identify 16 additional cases of SMARCA4-deficient uterine sarcoma from the database of a large, CLIA-certified and CAP-accredited, reference molecular laboratory, and we expand on their clinicopathological and genomic features. Median patient's age was 49 years (range 32–70). Most tumors were aggressive with distant metastasis. SMARCA4-deficient uterine sarcoma demonstrated predominantly rhabdoid or large epithelioid cells with abundant cytoplasm, but also had varying degrees of small cell and spindle cell morphology. Tumors were microsatellite stable and exhibited no other or only few co-occurring genomic alterations by comprehensive genomic profiling. We discovered one patient, who developed SMARCA4-deficient uterine sarcoma at the age of 55, had a germline SMARCA4 mutation, whose daughter had previously died of small cell carcinoma of the ovary, hypercalcemic type, at the age of 32. Our data support the notion that SMARCA4 inactivation is the driver oncogenic event of a morphologically and molecularly distinct form of uterine sarcoma. Identification of SMARCA4-deficient uterine sarcomas may be clinically important due to their aggressive behavior, germline association, and emerging targeted therapies.
Coffin–Siris syndrome (CSS; MIM 135900) is a multisystem congenital anomaly syndrome caused by mutations in the genes in the Brg‐1 associated factors (BAF) complex. Classically, individuals with CSS have been described with hypo‐ or aplasia of the fifth digit nails or phalanges (hence the term “fifth digit syndrome”). Other physical features seen include growth restriction, coarse facial features, hypertrichosis or hirsutism, sparse scalp hair, dental anomalies, and other organ‐system abnormalities. Varying degrees of developmental and intellectual delay are universal. To date, approximately 200 individuals have been described in the literature. With the advent of large‐scale genetic testing such as whole‐exome sequencing is becoming more available, more individuals are being found to have mutations in this pathway, and the phenotypic spectrum appears to be broadening. We report here a large cohort of 81 individuals with the diagnosis of CSS from the first parent‐reported CSS/BAF complex registry in an effort to describe this variation among individuals, the natural history of the syndrome, and draw some gene–phenotype correlations. We propose that changes in the BAF complex may represent a spectrum of disorders, including both ARID1B‐related nonsyndromic intellectual disability (ARID1B‐ID) and CSS with classic physical features. In addition, we offer surveillance and management recommendations based on the medical issues encountered in this cohort to help guide physicians and patients' families.
Coffin–Siris syndrome (CSS) is a rare congenital malformation syndrome, recently found to be caused by mutations in several genes encoding components of the BAF complex. To date, 109 patients have been reported with their mutations: SMARCB1 (12%), SMARCA4 (11%), SMARCE1 (2%), ARID1A (7%), ARID1B (65%), and PHF6 (2%). We review genotype-phenotype correlation of all previously reported patients with mutations in SMARCB1, SMARCA4, SMARCE1, and ARID1A through reassessment of their clinical and molecular findings. Cardinal features of CSS included variable degrees of intellectual disability (ID) predominantly affecting speech, sucking/feeding difficulty, and craniofacial (thick eyebrows, long eyelashes), digital (hypoplastic 5th fingers or toes, hypoplastic 5th fingernails or toenails), and other characteristics (hypertrichosis). In addition, patients with SMARCB1 mutations had severe neurodevelopmental deficits including severe ID, seizures, CNS structural abnormalities, and no expressive words as well as scoliosis. Especially, those with a recurrent mutation “p.Lys364del” represented strikingly similar phenotypes including characteristic facial coarseness. Patients with SMARCA4 mutations had less coarse craniofacial appearances and behavioral abnormalities. Patients with SMARCE1 mutations had a wide spectrum of manifestations from severe to moderate ID. Patients with ARID1A also had a wide spectrum of manifestations from severe ID and serous internal complications that could result in early death to mild ID. Mutations in SMARCB1, SMARCA4, and SMARCE1 are expected to exert dominant-negative or gain-of-function effects, whereas those in ARID1A are expected to exert loss-of-function effects. © 2014 Wiley Periodicals, Inc.
Here we provide compelling evidence that Next Generation Sequencing will revolutionise diagnostics. We reappraised a case from 1991, published in 1993, describing the unique occurrence of an ovarian immature teratoma arising in a young woman and a clonally distinct intracerebral immature teratoma developing in her daughter. We conducted whole exome sequencing on constitutional DNA from the mother and her daughter and identified a previously unreported nonsense mutation (c.3533G>A; p.Trp1178*) in the chromatin remodelling gene, SMARCA4, that was present in both individuals and was subject to nonsense-mediated decay. Tumour analysis by Sanger sequencing revealed a somatic SMARCA4 mutation, both in the mother (c.2438+1G>T), and in her daughter (c.3229C>T; p.Arg1077*), which are predicted to be truncating. As immature teratomas are classified as germ cell tumours, we performed a comprehensive mutation survey of 106 apparently sporadic germ cell tumours but did not find any other clearly deleterious SMARCA4 mutations. Recently, inactivating mutations in SMARCA4 have been found in two cases of rhabdoid tumour predisposition syndrome type 2. In the light of these findings, renewed efforts to locate previously unobtainable tumour samples were successfully undertaken. Histopathological and immunohistochemical re-analysis of the daughter's tumour revealed that it was indeed a rhabdoid tumour (atypical teratoid/rhabdoid tumour). In this context, the original pathology report of the mother's ovarian tumour was re-interpreted as describing a malignant rhabdoid tumour of the ovary. This report raises the question as to whether molecular genetic analysis should be included in tumour classification, alongside more traditional microscopy-based methods. The use of new sequencing technologies, particularly when applied to archived samples, will lead to many more "molecular re-diagnoses". This is the earliest known case of rhabdoid tumour predisposition syndrome type 2 and the only described case with an autosomal dominant pattern of inheritance, only discovered through an exome sequencing project.
The cBioPortal for Cancer Genomics (http://cbioportal.org) provides a Web resource for exploring, visualizing, and analyzing multidimensional cancer genomics data. The portal reduces molecular profiling data from cancer tissues and cell lines into readily understandable genetic, epigenetic, gene expression, and proteomic events. The query interface combined with customized data storage enables researchers to interactively explore genetic alterations across samples, genes, and pathways and, when available in the underlying data, to link these to clinical outcomes. The portal provides graphical summaries of gene-level data from multiple platforms, network visualization and analysis, survival analysis, patient-centric queries, and software programmatic access. The intuitive Web interface of the portal makes complex cancer genomics profiles accessible to researchers and clinicians without requiring bioinformatics expertise, thus facilitating biological discoveries. Here, we provide a practical guide to the analysis and visualization features of the cBioPortal for Cancer Genomics.
Rhabdoid tumors of early infancy are highly aggressive with consequent poor prognosis. Most cases show inactivation of the SMARCB1 (also known as INI1 and hSNF5) tumor suppressor, a core member of the ATP-dependent SWI/SNF chromatin-remodeling complex. Familial cases, described as rhabdoid tumor predisposition syndrome (RTPS), have been linked to heterozygous SMARCB1 germline mutations. We identified inactivation of another member of the SWI/SNF chromatin-remodeling complex, its ATPase subunit SMARCA4 (also known as BRG1), due to a SMARCA4/BRG1 germline mutation and loss of heterozygosity by uniparental disomy in the tumor cells of two sisters with rhabdoid tumors lacking SMARCB1 mutations. SMARCA4 is thus a second member of the SWI/SNF complex involved in cancer predisposition. Its general involvement in other tumor entities remains to be established.
& Coffin-Siris Syndrome International
- T Kosho
- N Okamoto
Kosho, T., Okamoto, N., & Coffin-Siris Syndrome International, C. (2014).