Publications by authors named "Charles E Schwartz"

186 Publications

Clinical epigenomics: genome-wide DNA methylation analysis for the diagnosis of Mendelian disorders.

Genet Med 2021 Feb 5. Epub 2021 Feb 5.

Amsterdam University Medical Center, University of Amsterdam, Department of Clinical Genetics, Amsterdam Reproduction and Development Research Institute, Amsterdam, The Netherlands.

Purpose: We describe the clinical implementation of genome-wide DNA methylation analysis in rare disorders across the EpiSign diagnostic laboratory network and the assessment of results and clinical impact in the first subjects tested.

Methods: We outline the logistics and data flow between an integrated network of clinical diagnostics laboratories in Europe, the United States, and Canada. We describe the clinical validation of EpiSign using 211 specimens and assess the test performance and diagnostic yield in the first 207 subjects tested involving two patient subgroups: the targeted cohort (subjects with previous ambiguous/inconclusive genetic findings including genetic variants of unknown clinical significance) and the screening cohort (subjects with clinical findings consistent with hereditary neurodevelopmental syndromes and no previous conclusive genetic findings).

Results: Among the 207 subjects tested, 57 (27.6%) were positive for a diagnostic episignature including 48/136 (35.3%) in the targeted cohort and 8/71 (11.3%) in the screening cohort, with 4/207 (1.9%) remaining inconclusive after EpiSign analysis.

Conclusion: This study describes the implementation of diagnostic clinical genomic DNA methylation testing in patients with rare disorders. It provides strong evidence of clinical utility of EpiSign analysis, including the ability to provide conclusive findings in the majority of subjects tested.
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http://dx.doi.org/10.1038/s41436-020-01096-4DOI Listing
February 2021

New Strategies for Clinical Trials in Autism Spectrum Disorder.

Rev Recent Clin Trials 2020 Nov 19. Epub 2020 Nov 19.

Greenwood Genetic Center, Greenwood, SC. United States.

Background: Autism spectrum disorder (ASD) is a complex neurodevelopmental condition that poses several challenges in terms of clinical diagnosis and investigation of molecular etiology. The lack of knowledge on the pathogenic mechanisms underlying ASD has hampered the clinical trials that so far have tried to target ASD behavioral symptoms. In order to improve our understanding of the molecular abnormalities associated with ASD, a deeper and more extensive genetic profiling of targeted individuals with ASD was needed.

Methods: The recent availability of new and more powerful sequencing technologies (third-generation sequencing) has allowed to develop novel strategies for characterization of comprehensive genetic profiles of individuals with ASD. In particular, this review will describe integrated approaches based on the combination of various omics technologies that will lead to a better stratification of targeted cohorts for the design of clinical trials in ASD.

Results: In order to analyze the big data collected by assays such as whole genome, epigenome, transcriptome, and proteome, it is critical to develop an efficient computational infrastructure. Machine learning models are instrumental to identify non-linear relationships between the omics technologies and therefore establish a functional informative network among the different data sources.

Conclusion: The potential advantage provided by these new integrated omics-based strategies is to better characterize the genetic background of ASD cohorts, identify novel molecular targets for drug development, and ultimately offer a more personalized approach in the design of clinical trials for ASD.
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http://dx.doi.org/10.2174/1574887115666201120093634DOI Listing
November 2020

Mutations in FAM50A suggest that Armfield XLID syndrome is a spliceosomopathy.

Nat Commun 2020 07 23;11(1):3698. Epub 2020 Jul 23.

Greenwood Genetic Center, Greenwood, SC, USA.

Intellectual disability (ID) is a heterogeneous clinical entity and includes an excess of males who harbor variants on the X-chromosome (XLID). We report rare FAM50A missense variants in the original Armfield XLID syndrome family localized in Xq28 and four additional unrelated males with overlapping features. Our fam50a knockout (KO) zebrafish model exhibits abnormal neurogenesis and craniofacial patterning, and in vivo complementation assays indicate that the patient-derived variants are hypomorphic. RNA sequencing analysis from fam50a KO zebrafish show dysregulation of the transcriptome, with augmented spliceosome mRNAs and depletion of transcripts involved in neurodevelopment. Zebrafish RNA-seq datasets show a preponderance of 3' alternative splicing events in fam50a KO, suggesting a role in the spliceosome C complex. These data are supported with transcriptomic signatures from cell lines derived from affected individuals and FAM50A protein-protein interaction data. In sum, Armfield XLID syndrome is a spliceosomopathy associated with aberrant mRNA processing during development.
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http://dx.doi.org/10.1038/s41467-020-17452-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7378245PMC
July 2020

Schimke XLID syndrome results from a deletion in BCAP31.

Am J Med Genet A 2020 09 18;182(9):2168-2174. Epub 2020 Jul 18.

Greenwood Genetic Center, Greenwood, South Carolina, USA.

A family with three affected males and a second family with a single affected male with intellectual disability, microcephaly, ophthalmoplegia, deafness, and Involuntary limb movements were reported by Schimke and Associates in 1984. The affected males with Schimke X-linked intellectual disability (XLID) syndrome (OMIM# 312840) had a similar facial appearance with deep-set eyes, downslanting palpebral fissures, hypotelorism, narrow nose and alae nasi, cupped ears and spacing of the teeth. Two mothers had mild hearing loss but no other manifestations of the disorder. The authors considered the disorder to be distinctive and likely X-linked. Whole genome sequencing in the single affected male available and the three carrier females from one of the families with Schimke XLID syndrome identified a 2 bp deletion in the BCAP31 gene. During the past decade, pathogenic alterations of the BCAP31 gene have been associated with deafness, dystonia, and central hypomyelination, an XLID condition given the eponym DDCH syndrome. A comparison of clinical findings in Schimke XLID syndrome and DDCH syndrome shows them to be the same clinical entity. The BCAP31 protein functions in endoplasmic reticulum-associated degradation to promote ubiquitination and destruction of misfolded proteins.
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http://dx.doi.org/10.1002/ajmg.a.61755DOI Listing
September 2020

Spermine synthase and MYC cooperate to maintain colorectal cancer cell survival by repressing Bim expression.

Nat Commun 2020 06 26;11(1):3243. Epub 2020 Jun 26.

Markey Cancer Center, University of Kentucky College of Medicine, Lexington, KY, 40506, USA.

Dysregulation of polyamine metabolism has been linked to the development of colorectal cancer (CRC), but the underlying mechanism is incompletely characterized. Here, we report that spermine synthase (SMS), a polyamine biosynthetic enzyme, is overexpressed in CRC. Targeted disruption of SMS in CRC cells results in spermidine accumulation, which inhibits FOXO3a acetylation and allows subsequent translocation to the nucleus to transcriptionally induce expression of the proapoptotic protein Bim. However, this induction is blunted by MYC-driven expression of miR-19a and miR-19b that repress Bim production. Pharmacological or genetic inhibition of MYC activity in SMS-depleted CRC cells dramatically induces Bim expression and apoptosis and causes tumor regression, but these effects are profoundly attenuated by silencing Bim. These findings uncover a key survival signal in CRC through convergent repression of Bim expression by distinct SMS- and MYC-mediated signaling pathways. Thus, combined inhibition of SMS and MYC signaling may be an effective therapy for CRC.
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http://dx.doi.org/10.1038/s41467-020-17067-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7320137PMC
June 2020

The Future of Clinical Diagnosis: Moving Functional Genomics Approaches to the Bedside.

Clin Lab Med 2020 06;40(2):221-230

Greenwood Genetic Center, JC Self Research Institute, 113 Gregor Mendel Circle, Greenwood, SC 29646, USA.

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http://dx.doi.org/10.1016/j.cll.2020.02.006DOI Listing
June 2020

Rare missense variant p.Ala505Ser in the ZAK protein observed in a patient with split-hand/foot malformation from a non-consanguineous pedigree.

J Int Med Res 2020 Apr;48(4):300060519879293

Department of Biomedical Sciences, University of South Carolina School of Medicine Greenville, Greenville, SC, United States.

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http://dx.doi.org/10.1177/0300060519879293DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7144677PMC
April 2020

Evaluation of DNA Methylation Episignatures for Diagnosis and Phenotype Correlations in 42 Mendelian Neurodevelopmental Disorders.

Am J Hum Genet 2020 03 27;106(3):356-370. Epub 2020 Feb 27.

Molecular Genetics Laboratory, Molecular Diagnostics Division, London Health Sciences Centre, London, ON N6A5W9, Canada; Department of Pathology and Laboratory Medicine, Western University, London, ON N6A3K7, Canada. Electronic address:

Genetic syndromes frequently present with overlapping clinical features and inconclusive or ambiguous genetic findings which can confound accurate diagnosis and clinical management. An expanding number of genetic syndromes have been shown to have unique genomic DNA methylation patterns (called "episignatures"). Peripheral blood episignatures can be used for diagnostic testing as well as for the interpretation of ambiguous genetic test results. We present here an approach to episignature mapping in 42 genetic syndromes, which has allowed the identification of 34 robust disease-specific episignatures. We examine emerging patterns of overlap, as well as similarities and hierarchical relationships across these episignatures, to highlight their key features as they are related to genetic heterogeneity, dosage effect, unaffected carrier status, and incomplete penetrance. We demonstrate the necessity of multiclass modeling for accurate genetic variant classification and show how disease classification using a single episignature at a time can sometimes lead to classification errors in closely related episignatures. We demonstrate the utility of this tool in resolving ambiguous clinical cases and identification of previously undiagnosed cases through mass screening of a large cohort of subjects with developmental delays and congenital anomalies. This study more than doubles the number of published syndromes with DNA methylation episignatures and, most significantly, opens new avenues for accurate diagnosis and clinical assessment in individuals affected by these disorders.
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http://dx.doi.org/10.1016/j.ajhg.2020.01.019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7058829PMC
March 2020

An intellectual disability syndrome with single-nucleotide variants in O-GlcNAc transferase.

Eur J Hum Genet 2020 06 20;28(6):706-714. Epub 2020 Feb 20.

Division of Gene Regulation and Expression and School of Life Sciences, University of Dundee, Dundee, UK.

Intellectual disability (ID) is a neurodevelopmental condition that affects ~1% of the world population. In total 5-10% of ID cases are due to variants in genes located on the X chromosome. Recently, variants in OGT have been shown to co-segregate with X-linked intellectual disability (XLID) in multiple families. OGT encodes O-GlcNAc transferase (OGT), an essential enzyme that catalyses O-linked glycosylation with β-N-acetylglucosamine (O-GlcNAc) on serine/threonine residues of thousands of nuclear and cytosolic proteins. In this review, we compile the work from the last few years that clearly delineates a new syndromic form of ID, which we propose to classify as a novel Congenital Disorder of Glycosylation (OGT-CDG). We discuss potential hypotheses for the underpinning molecular mechanism(s) that provide impetus for future research studies geared towards informed interventions.
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http://dx.doi.org/10.1038/s41431-020-0589-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7253464PMC
June 2020

(,)-1,12-Dimethylspermine can mitigate abnormal spermidine accumulation in Snyder-Robinson syndrome.

J Biol Chem 2020 03 29;295(10):3247-3256. Epub 2020 Jan 29.

Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins School of Medicine, Baltimore, Maryland 21287

Snyder-Robinson syndrome (SRS) is an X-linked intellectual disability syndrome caused by a loss-of-function mutation in the spermine synthase () gene. Primarily affecting males, the main manifestations of SRS include osteoporosis, hypotonic stature, seizures, cognitive impairment, and developmental delay. Because there is no cure for SRS, treatment plans focus on alleviating symptoms rather than targeting the underlying causes. Biochemically, the cells of individuals with SRS accumulate excess spermidine, whereas spermine levels are reduced. We recently demonstrated that SRS patient-derived lymphoblastoid cells are capable of transporting exogenous spermine and its analogs into the cell and, in response, decreasing excess spermidine pools to normal levels. However, dietary supplementation of spermine does not appear to benefit SRS patients or mouse models. Here, we investigated the potential use of a metabolically stable spermine mimetic, (,)-1,12-dimethylspermine (MeSPM), to reduce the intracellular spermidine pools of SRS patient-derived cells. MeSPM can functionally substitute for the native polyamines in supporting cell growth while stimulating polyamine homeostatic control mechanisms. We found that both lymphoblasts and fibroblasts from SRS patients can accumulate MeSPM, resulting in significantly decreased spermidine levels with no adverse effects on growth. MeSPM administration to mice revealed that MeSPM significantly decreases spermidine levels in multiple tissues. Importantly, MeSPM was detectable in brain tissue, the organ most affected in SRS, and was associated with changes in polyamine metabolic enzymes. These findings indicate that the (,)-diastereomer of 1,12-MeSPM represents a promising lead compound in developing a treatment aimed at targeting the molecular mechanisms underlying SRS pathology.
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http://dx.doi.org/10.1074/jbc.RA119.011572DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7062180PMC
March 2020

Phenotypic and molecular description of an individual with a disruptive variant in the SULF2 gene.

Clin Dysmorphol 2020 Jul;29(3):144-147

Department of Pediatrics, Section of Genetics and Metabolism, University of Arkansas for Medical Sciences, Little Rock, Arkansas, USA.

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http://dx.doi.org/10.1097/MCD.0000000000000309DOI Listing
July 2020

Clark-Baraitser syndrome is associated with a nonsense alteration in the autosomal gene TRIP12.

Am J Med Genet A 2020 03 8;182(3):595-596. Epub 2019 Dec 8.

J.C. Self Research Institute of Human Genetics, Greenwood Genetic Center, Greenwood, South Carolina.

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http://dx.doi.org/10.1002/ajmg.a.61443DOI Listing
March 2020

X-linked intellectual disability: Phenotypic expression in carrier females.

Clin Genet 2020 03 24;97(3):418-425. Epub 2019 Nov 24.

J.C. Self Research Institute of Human Genetics, Greenwood Genetic Center, Greenwood, South Carolina.

To better understand the landscape of female phenotypic expression in X-linked intellectual disability (XLID), we surveyed the literature for female carriers of XLID gene alterations (n = 1098) and combined this with experience evaluating XLID kindreds at the Greenwood Genetic Center (n = 341) and at the University of Adelaide (n = 157). One-hundred forty-four XLID genes were grouped into nine categories based on the level of female phenotypic expression, ranging from no expression to female only expression. For each gene, the clinical presentation, gene expression in blood, X-inactivation (XI) pattern, biological pathway involved, and whether the gene escapes XI were noted. Among the XLID conditions, 88 (61.1%) exhibited female cognitive phenotypic expression only, while 56 (38.9%) had no female phenotypic expression (n = 45), phenotype expression with normal cognition in females (n = 8), or unknown status for female phenotypic expression (n = 3). In twenty-four (16.6%) XLID genes, XI was consistently skewed in female carriers, in 54 (37.5%) XI showed variable skewing, and in 33 (22.9%) XI was consistently random. The XI pattern was unknown in 33 (22.9%) XLID conditions. Therefore, there is evidence of a female carrier phenotype in the majority of XLID conditions although how exactly XI patterns influence the female phenotype in XLID conditions remains unclear.
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http://dx.doi.org/10.1111/cge.13667DOI Listing
March 2020

Abnormalities in the genes that encode Large Amino Acid Transporters increase the risk of Autism Spectrum Disorder.

Mol Genet Genomic Med 2020 01 7;8(1):e1036. Epub 2019 Nov 7.

JC Self research Institute, Greenwood Genetic Center, Greenwood, SC, USA.

Background: Autism spectrum disorder (ASD) is a common neurodevelopmental disorder whose molecular mechanisms are largely unknown. Several studies have shown an association between ASD and abnormalities in the metabolism of amino acids, specifically tryptophan and branched-chain amino acids (BCAAs).

Methods: Ninety-seven patients with ASD were screened by Sanger sequencing the genes encoding the heavy (SLC3A2) and light subunits (SLC7A5 and SLC7A8) of the large amino acid transporters (LAT) 1 and 2. LAT1 and 2 are responsible for the transportation of tryptophan and BCAA across the blood-brain barrier and are expressed both in blood and brain. Functional studies were performed employing the Biolog Phenotype Microarray Mammalian (PM-M) technology to investigate the metabolic profiling in lymphoblastoid cell lines from 43 patients with ASD and 50 controls with particular focus on the amino acid substrates of LATs.

Results: We detected nine likely pathogenic variants in 11 of 97 patients (11.3%): three in SLC3A2, three in SLC7A5, and three in SLC7A8. Six variants of unknown significance were detected in eight patients, two of which also carrying a likely pathogenic variant. The functional studies showed a consistently reduced utilization of tryptophan, accompanied by evidence of reduced utilization of other large aromatic amino acids (LAAs), either alone or as part of a dipeptide.

Conclusion: Coding variants in the LAT genes were detected in 17 of 97 patients with ASD (17.5%). Metabolic assays indicate that such abnormalities affect the utilization of certain amino acids, particularly tryptophan and other LAAs, with potential consequences on their transport across the blood barrier and their availability during brain development. Therefore, abnormalities in the LAT1 and two transporters are likely associated with an increased risk of developing ASD.
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http://dx.doi.org/10.1002/mgg3.1036DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6978257PMC
January 2020

Histone demethylase KDM5C is a SAHA-sensitive central hub at the crossroads of transcriptional axes involved in multiple neurodevelopmental disorders.

Hum Mol Genet 2019 12;28(24):4089-4102

Institute of Genetics and Biophysics "Adriano Buzzati-Traverso", National Research Council (CNR), Naples, Italy.

A disproportional large number of neurodevelopmental disorders (NDDs) is caused by variants in genes encoding transcription factors and chromatin modifiers. However, the functional interactions between the corresponding proteins are only partly known. Here, we show that KDM5C, encoding a H3K4 demethylase, is at the intersection of transcriptional axes under the control of three regulatory proteins ARX, ZNF711 and PHF8. Interestingly, mutations in all four genes (KDM5C, ARX, ZNF711 and PHF8) are associated with X-linked NDDs comprising intellectual disability as a core feature. in vitro analysis of the KDM5C promoter revealed that ARX and ZNF711 function as antagonist transcription factors that activate KDM5C expression and compete for the recruitment of PHF8. Functional analysis of mutations in these genes showed a correlation between phenotype severity and the reduction in KDM5C transcriptional activity. The KDM5C decrease was associated with a lack of repression of downstream target genes Scn2a, Syn1 and Bdnf in the embryonic brain of Arx-null mice. Aiming to correct the faulty expression of KDM5C, we studied the effect of the FDA-approved histone deacetylase inhibitor suberanilohydroxamic acid (SAHA). In Arx-KO murine ES-derived neurons, SAHA was able to rescue KDM5C depletion, recover H3K4me3 signalling and improve neuronal differentiation. Indeed, in ARX/alr-1-deficient Caenorhabditis elegans animals, SAHA was shown to counteract the defective KDM5C/rbr-2-H3K4me3 signalling, recover abnormal behavioural phenotype and ameliorate neuronal maturation. Overall, our studies indicate that KDM5C is a conserved and druggable effector molecule across a number of NDDs for whom the use of SAHA may be considered a potential therapeutic strategy.
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http://dx.doi.org/10.1093/hmg/ddz254DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7002875PMC
December 2019

Disruptive mutations in TANC2 define a neurodevelopmental syndrome associated with psychiatric disorders.

Nat Commun 2019 10 15;10(1):4679. Epub 2019 Oct 15.

Department of Genome Sciences, University of Washington School of Medicine, Seattle, WA, 98195, USA.

Postsynaptic density (PSD) proteins have been implicated in the pathophysiology of neurodevelopmental and psychiatric disorders. Here, we present detailed clinical and genetic data for 20 patients with likely gene-disrupting mutations in TANC2-whose protein product interacts with multiple PSD proteins. Pediatric patients with disruptive mutations present with autism, intellectual disability, and delayed language and motor development. In addition to a variable degree of epilepsy and facial dysmorphism, we observe a pattern of more complex psychiatric dysfunction or behavioral problems in adult probands or carrier parents. Although this observation requires replication to establish statistical significance, it also suggests that mutations in this gene are associated with a variety of neuropsychiatric disorders consistent with its postsynaptic function. We find that TANC2 is expressed broadly in the human developing brain, especially in excitatory neurons and glial cells, but shows a more restricted pattern in Drosophila glial cells where its disruption affects behavioral outcomes.
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http://dx.doi.org/10.1038/s41467-019-12435-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6794285PMC
October 2019

The complete loss of function of the SMS gene results in a severe form of Snyder-Robinson syndrome.

Eur J Med Genet 2020 Apr 30;63(4):103777. Epub 2019 Sep 30.

APHP, UF de Génétique clinique, Centre de Référence Maladies Rares « Anomalies du développement et syndromes malformatifs », Hôpital Armand Trousseau, Paris, France.

Snyder-Robinson syndrome (SRS) is an X-linked syndromic intellectual disability condition caused by variants in the spermine synthase gene (SMS). The syndrome is characterized by facial dysmorphism, thin body build, kyphoscoliosis, osteoporosis, hypotonia, developmental delay and associated neurological features (seizures, unsteady gait, abnormal speech). Until now, only missense variants with a functionally characterized partial loss of function (LoF) have been described. Here we describe the first complete LoF variant, Met303Lysfs*, in a male patient with a severe form of Snyder-Robinson syndrome. He presented with multiple malformations and severly delayed development, and died at 4 months of age. Functional in vitro assays showed a complete absence of functional SMS protein. Taken together, our findings and those of previously reported patients confirm that pathogenic variants of SMS are indeed LoF and that there might exist a genotype-phenotype correlation between the type of variant and the severity of the syndrome.
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http://dx.doi.org/10.1016/j.ejmg.2019.103777DOI Listing
April 2020

An unusual cause for Coffin-Lowry syndrome: Three brothers with a novel microduplication in RPS6KA3.

Am J Med Genet A 2019 12 12;179(12):2357-2364. Epub 2019 Sep 12.

Department of Medical and Molecular Genetics, Indiana University School of Medicine, Indianapolis, Indiana.

Coffin-Lowry syndrome (CLS) is a rare X-linked disorder characterized by moderate to severe intellectual disability, hypotonia, craniofacial features, tapering digits, short stature, and skeletal deformities. Using whole exome sequencing and high-resolution targeted comparative genomic hybridization array analysis, we identified a novel microduplication encompassing exons five through nine of RPS6KA3 in three full brothers. Each brother presented with intellectual disability and clinical and radiographic features consistent with CLS. qRT-PCR analyses performed on mRNA from the peripheral blood of the three siblings revealed a marked reduction of RPS6KA3 levels suggesting a loss-of-function mechanism. PCR analysis of the patients' cDNA detected a band greater than expected for an exon 4-10 amplicon, suggesting this was likely a direct duplication that lies between exons 4 through 10, which was later confirmed by Sanger sequencing. This microduplication is only the third intragenic duplication of RPS6KA3, and the second and smallest reported to date thought to cause CLS. Our study further supports the clinical utility of methods such as next-generation sequencing and high-resolution genomic arrays to detect small intragenic duplications. These methods, coupled with expression studies and cDNA structural analysis have the capacity to confirm the diagnosis of CLS in these rare cases.
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http://dx.doi.org/10.1002/ajmg.a.61353DOI Listing
December 2019

Redefining the Etiologic Landscape of Cerebellar Malformations.

Am J Hum Genet 2019 09 29;105(3):606-615. Epub 2019 Aug 29.

Center for Integrative Brain Research, Seattle Children's Research Institute, Seattle, WA 98101, USA; Department of Pediatrics, University of Washington, Seattle, WA 98105, USA; Department of Neurology, University of Washington, Seattle, WA 98105, USA. Electronic address:

Cerebellar malformations are diverse congenital anomalies frequently associated with developmental disability. Although genetic and prenatal non-genetic causes have been described, no systematic analysis has been performed. Here, we present a large-exome sequencing study of Dandy-Walker malformation (DWM) and cerebellar hypoplasia (CBLH). We performed exome sequencing in 282 individuals from 100 families with DWM or CBLH, and we established a molecular diagnosis in 36 of 100 families, with a significantly higher yield for CBLH (51%) than for DWM (16%). The 41 variants impact 27 neurodevelopmental-disorder-associated genes, thus demonstrating that CBLH and DWM are often features of monogenic neurodevelopmental disorders. Though only seven monogenic causes (19%) were identified in more than one individual, neuroimaging review of 131 additional individuals confirmed cerebellar abnormalities in 23 of 27 genetic disorders (85%). Prenatal risk factors were frequently found among individuals without a genetic diagnosis (30 of 64 individuals [47%]). Single-cell RNA sequencing of prenatal human cerebellar tissue revealed gene enrichment in neuronal and vascular cell types; this suggests that defective vasculogenesis may disrupt cerebellar development. Further, de novo gain-of-function variants in PDGFRB, a tyrosine kinase receptor essential for vascular progenitor signaling, were associated with CBLH, and this discovery links genetic and non-genetic etiologies. Our results suggest that genetic defects impact specific cerebellar cell types and implicate abnormal vascular development as a mechanism for cerebellar malformations. We also confirmed a major contribution for non-genetic prenatal factors in individuals with cerebellar abnormalities, substantially influencing diagnostic evaluation and counseling regarding recurrence risk and prognosis.
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http://dx.doi.org/10.1016/j.ajhg.2019.07.019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6731369PMC
September 2019

Gene domain-specific DNA methylation episignatures highlight distinct molecular entities of ADNP syndrome.

Clin Epigenetics 2019 04 27;11(1):64. Epub 2019 Apr 27.

Department of Pathology and Laboratory Medicine, Western University, 800 Commissioner's Road E, London, ON, N6A 5W9, Canada.

Background: ADNP syndrome is a rare Mendelian disorder characterized by global developmental delay, intellectual disability, and autism. It is caused by truncating mutations in ADNP, which is involved in chromatin regulation. We hypothesized that the disruption of chromatin regulation might result in specific DNA methylation patterns that could be used in the molecular diagnosis of ADNP syndrome.

Results: We identified two distinct and partially opposing genomic DNA methylation episignatures in the peripheral blood samples from 22 patients with ADNP syndrome. The "epi-ADNP-1" episignature included ~ 6000 mostly hypomethylated CpGs, and the "epi-ADNP-2" episignature included ~ 1000 predominantly hypermethylated CpGs. The two signatures correlated with the locations of the ADNP mutations. Epi-ADNP-1 mutations occupy the N- and C-terminus, and epi-ADNP-2 mutations are centered on the nuclear localization signal. The episignatures were enriched for genes involved in neuronal system development and function. A classifier trained on these profiles yielded full sensitivity and specificity in detecting patients with either of the two episignatures. Applying this model to seven patients with uncertain clinical diagnosis enabled reclassification of genetic variants of uncertain significance and assigned new diagnosis when the primary clinical suspicion was not correct. When applied to a large cohort of unresolved patients with developmental delay (N = 1150), the model predicted three additional previously undiagnosed patients to have ADNP syndrome. DNA sequencing of these subjects, wherever available, identified pathogenic mutations within the gene domains predicted by the model.

Conclusions: We describe the first Mendelian condition with two distinct episignatures caused by mutations in a single gene. These highly sensitive and specific DNA methylation episignatures enable diagnosis, screening, and genetic variant classifications in ADNP syndrome.
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http://dx.doi.org/10.1186/s13148-019-0658-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6487024PMC
April 2019

ATP6AP2 variant impairs CNS development and neuronal survival to cause fulminant neurodegeneration.

J Clin Invest 2019 04 15;129(5):2145-2162. Epub 2019 Apr 15.

INSERM, UMR-S 1270, Paris, France.

Vacuolar H+-ATPase-dependent (V-ATPase-dependent) functions are critical for neural proteostasis and are involved in neurodegeneration and brain tumorigenesis. We identified a patient with fulminant neurodegeneration of the developing brain carrying a de novo splice site variant in ATP6AP2 encoding an accessory protein of the V-ATPase. Functional studies of induced pluripotent stem cell-derived (iPSC-derived) neurons from this patient revealed reduced spontaneous activity and severe deficiency in lysosomal acidification and protein degradation leading to neuronal cell death. These deficiencies could be rescued by expression of full-length ATP6AP2. Conditional deletion of Atp6ap2 in developing mouse brain impaired V-ATPase-dependent functions, causing impaired neural stem cell self-renewal, premature neuronal differentiation, and apoptosis resulting in degeneration of nearly the entire cortex. In vitro studies revealed that ATP6AP2 deficiency decreases V-ATPase membrane assembly and increases endosomal-lysosomal fusion. We conclude that ATP6AP2 is a key mediator of V-ATPase-dependent signaling and protein degradation in the developing human central nervous system.
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http://dx.doi.org/10.1172/JCI79990DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6486358PMC
April 2019

Diagnostic Utility of Genome-wide DNA Methylation Testing in Genetically Unsolved Individuals with Suspected Hereditary Conditions.

Am J Hum Genet 2019 04 28;104(4):685-700. Epub 2019 Mar 28.

Department of Pathology and Laboratory Medicine, Western University, London, ON N6A 3K7, Canada; Molecular Genetics Laboratory, Molecular Diagnostics Division, London Health Sciences Centre, London, ON N6A 5W9, Canada. Electronic address:

Conventional genetic testing of individuals with neurodevelopmental presentations and congenital anomalies (ND/CAs), i.e., the analysis of sequence and copy number variants, leaves a substantial proportion of them unexplained. Some of these cases have been shown to result from DNA methylation defects at a single locus (epi-variants), while others can exhibit syndrome-specific DNA methylation changes across multiple loci (epi-signatures). Here, we investigate the clinical diagnostic utility of genome-wide DNA methylation analysis of peripheral blood in unresolved ND/CAs. We generate a computational model enabling concurrent detection of 14 syndromes using DNA methylation data with full accuracy. We demonstrate the ability of this model in resolving 67 individuals with uncertain clinical diagnoses, some of whom had variants of unknown clinical significance (VUS) in the related genes. We show that the provisional diagnoses can be ruled out in many of the case subjects, some of whom are shown by our model to have other diseases initially not considered. By applying this model to a cohort of 965 ND/CA-affected subjects without a previous diagnostic assumption and a separate assessment of rare epi-variants in this cohort, we identify 15 case subjects with syndromic Mendelian disorders, 12 case subjects with imprinting and trinucleotide repeat expansion disorders, as well as 106 case subjects with rare epi-variants, a portion of which involved genes clinically or functionally linked to the subjects' phenotypes. This study demonstrates that genomic DNA methylation analysis can facilitate the molecular diagnosis of unresolved clinical cases and highlights the potential value of epigenomic testing in the routine clinical assessment of ND/CAs.
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http://dx.doi.org/10.1016/j.ajhg.2019.03.008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6451739PMC
April 2019

Dysregulations of sonic hedgehog signaling in MED12-related X-linked intellectual disability disorders.

Mol Genet Genomic Med 2019 04 6;7(4):e00569. Epub 2019 Feb 6.

Institute of Genetic Medicine and Department of Pediatrics, Johns Hopkins University, Baltimore, Maryland.

Background: Mutations in mediator of RNA polymerase II transcription subunit 12 homolog (MED12, OMIM 300188) cause X-linked intellectual disability (XLID) disorders including FG, Lujan, and Ohdo syndromes. The Gli3-dependent Sonic Hedgehog (SHH) signaling pathway has been implicated in the original FG syndrome and Lujan syndrome. How are SHH-signaling defects related to the complex clinical phenotype of MED12-associated XLID syndromes are not fully understood.

Methods: Quantitative RT-PCR was used to study expression levels of three SHH-signaling genes in lymophoblast cell lines carrying four MED12 mutations from four unrelated XLID families. Genotype and phenotype correlation studies were performed on these mutations.

Results: Three newly identified and one novel MED12 mutations in six affected males from four unrelated XLID families were studied. Three mutations (c.2692A>G; p.N898D, c.3640C>T; p.R1214C, and c.3884G>A; p.R1295H) are located in the LS domain and one (c.617G>A; p.R206Q) is in the L domain of MED12. These mutations involve highly conserved amino acid residues and segregate with ID and related congenital malformations in respective probands families. Patients with the LS-domain mutations share many features of FG syndrome and some features of Lujan syndrome. The patient with the L-domain mutation presented with ID and predominant neuropsychiatric features but little dysmorphic features of either FG or Lujan syndrome. Transcript levels of three Gli3-dependent SHH-signaling genes, CREB5, BMP4, and NEUROG2, were determined by quantitative RT-PCR and found to be significantly elevated in lymphoblasts from patients with three mutations in the MED12-LS domain.

Conclusions: These results support a critical role of MED12 in regulating Gli3-dependent SHH signaling and in developing ID and related congenital malformations in XLID syndromes. Differences in the expression profile of SHH-signaling genes potentially contribute to variability in clinical phenotypes in patients with MED12-related XLID disorders.
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http://dx.doi.org/10.1002/mgg3.569DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6465656PMC
April 2019

Polyamine Homeostasis in Snyder-Robinson Syndrome.

Med Sci (Basel) 2018 Dec 7;6(4). Epub 2018 Dec 7.

Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University, Baltimore, MD 21287, USA.

Loss-of-function mutations of the spermine synthase gene () result in Snyder-Robinson Syndrome (SRS), a recessive X-linked syndrome characterized by intellectual disability, osteoporosis, hypotonia, speech abnormalities, kyphoscoliosis, and seizures. As SMS catalyzes the biosynthesis of the polyamine spermine from its precursor spermidine, SMS deficiency causes a lack of spermine with an accumulation of spermidine. As polyamines, spermine, and spermidine play essential cellular roles that require tight homeostatic control to ensure normal cell growth, differentiation, and survival. Using patient-derived lymphoblast cell lines, we sought to comprehensively investigate the effects of SMS deficiency on polyamine homeostatic mechanisms including polyamine biosynthetic and catabolic enzymes, derivatives of the natural polyamines, and polyamine transport activity. In addition to decreased spermine and increased spermidine in SRS cells, ornithine decarboxylase activity and its product putrescine were significantly decreased. Treatment of SRS cells with exogenous spermine revealed that polyamine transport was active, as the cells accumulated spermine, decreased their spermidine level, and established a spermidine-to-spermine ratio within the range of wildtype cells. SRS cells also demonstrated elevated levels of tissue transglutaminase, a change associated with certain neurodegenerative diseases. These studies form a basis for further investigations into the leading biochemical changes and properties of -mutant cells that potentially represent therapeutic targets for the treatment of Snyder-Robinson Syndrome.
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http://dx.doi.org/10.3390/medsci6040112DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6318755PMC
December 2018

BAFopathies' DNA methylation epi-signatures demonstrate diagnostic utility and functional continuum of Coffin-Siris and Nicolaides-Baraitser syndromes.

Nat Commun 2018 11 20;9(1):4885. Epub 2018 Nov 20.

Department of Pathology and Laboratory Medicine, Western University, London, N6A 5W9, ON, Canada.

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.
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http://dx.doi.org/10.1038/s41467-018-07193-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6244416PMC
November 2018

UBE2A-related X-linked intellectual disability.

Clin Dysmorphol 2019 Jan;28(1):1-6

Greenwood Genetic Center, Greenwood, South Carolina, USA.

UBE2A-related X-linked intellectual disability is characterized by a distinctive facial phenotype (dense eyebrows and eyelashes, synophrys, hypertelorism, upslanted palpebral fissures, wide mouth, and thin lips), generalized hirsutism, hypoplastic genitalia, short stature, hypotonia, seizures, and severe intellectual disability. Five affected males in two families are described here and compared with the previously reported 17 males in eight families. The new cases are notable for the absence of nail dystrophy, previously considered a defining manifestation, and for the presence of hypogammaglobulinemia and adult-onset ataxia.
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http://dx.doi.org/10.1097/MCD.0000000000000242DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6279472PMC
January 2019

-GlcNAc transferase missense mutations linked to X-linked intellectual disability deregulate genes involved in cell fate determination and signaling.

J Biol Chem 2018 07 16;293(27):10810-10824. Epub 2018 May 16.

From the Department of Biochemistry and Molecular Biology, Complex Carbohydrate Research Center, University of Georgia, Athens, Georgia 30602,

It is estimated that ∼1% of the world's population has intellectual disability, with males affected more often than females. is an X-linked gene encoding for the enzyme GlcNAc transferase (OGT), which carries out the reversible addition of -acetylglucosamine (GlcNAc) to Ser/Thr residues of its intracellular substrates. Three missense mutations in the tetratricopeptide (TPR) repeats of OGT have recently been reported to cause X-linked intellectual disability (XLID). Here, we report the discovery of two additional novel missense mutations (c.775 G>A, p.A259T, and c.1016 A>G, p.E339G) in the TPR domain of OGT that segregate with XLID in affected families. Characterization of all five of these XLID missense variants of OGT demonstrates modest declines in thermodynamic stability and/or activities of the variants. We engineered each of the mutations into a male human embryonic stem cell line using CRISPR/Cas9. Investigation of the global GlcNAc profile as well as OGT and GlcNAc hydrolase levels by Western blotting showed no gross changes in steady-state levels in the engineered lines. However, analyses of the differential transcriptomes of the OGT variant-expressing stem cells revealed shared deregulation of genes involved in cell fate determination and liver X receptor/retinoid X receptor signaling, which has been implicated in neuronal development. Thus, here we reveal two additional mutations encoding residues in the TPR regions of OGT that appear causal for XLID and provide evidence that the relatively stable and active TPR variants may share a common, unelucidated mechanism of altering gene expression profiles in human embryonic stem cells.
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http://dx.doi.org/10.1074/jbc.RA118.002583DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6036218PMC
July 2018

X-linked intellectual disability update 2017.

Am J Med Genet A 2018 06 25;176(6):1375-1388. Epub 2018 Apr 25.

J.C. Self Research Institute of Human Genetics, Greenwood Genetic Center, Greenwood, South Carolina.

The X-chromosome comprises only about 5% of the human genome but accounts for about 15% of the genes currently known to be associated with intellectual disability. The early progress in identifying the X-linked intellectual disability (XLID)-associated genes through linkage analysis and candidate gene sequencing has been accelerated with the use of high-throughput technologies. In the 10 years since the last update, the number of genes associated with XLID has increased by 96% from 72 to 141 and duplications of all 141 XLID genes have been described, primarily through the application of high-resolution microarrays and next generation sequencing. The progress in identifying genetic and genomic alterations associated with XLID has not been matched with insights that improve the clinician's ability to form differential diagnoses, that bring into view the possibility of curative therapies for patients, or that inform scientists of the impact of the genetic alterations on cell organization and function.
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http://dx.doi.org/10.1002/ajmg.a.38710DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6049830PMC
June 2018