Publications by authors named "Miroslava Hančárová"

37 Publications

Variants in the degron of AFF3 are associated with intellectual disability, mesomelic dysplasia, horseshoe kidney, and epileptic encephalopathy.

Am J Hum Genet 2021 May;108(5):857-873

GeneDx, Gaithersburg, MD 20877, USA.

The ALF transcription factor paralogs, AFF1, AFF2, AFF3, and AFF4, are components of the transcriptional super elongation complex that regulates expression of genes involved in neurogenesis and development. We describe an autosomal dominant disorder associated with de novo missense variants in the degron of AFF3, a nine amino acid sequence important for its binding to ubiquitin ligase, or with de novo deletions of this region. The sixteen affected individuals we identified, along with two previously reported individuals, present with a recognizable pattern of anomalies, which we named KINSSHIP syndrome (KI for horseshoe kidney, NS for Nievergelt/Savarirayan type of mesomelic dysplasia, S for seizures, H for hypertrichosis, I for intellectual disability, and P for pulmonary involvement), partially overlapping the AFF4-associated CHOPS syndrome. Whereas homozygous Aff3 knockout mice display skeletal anomalies, kidney defects, brain malformations, and neurological anomalies, knockin animals modeling one of the microdeletions and the most common of the missense variants identified in affected individuals presented with lower mesomelic limb deformities like KINSSHIP-affected individuals and early lethality, respectively. Overexpression of AFF3 in zebrafish resulted in body axis anomalies, providing some support for the pathological effect of increased amount of AFF3. The only partial phenotypic overlap of AFF3- and AFF4-associated syndromes and the previously published transcriptome analyses of ALF transcription factors suggest that these factors are not redundant and each contributes uniquely to proper development.
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http://dx.doi.org/10.1016/j.ajhg.2021.04.001DOI Listing
May 2021

Missense variants in DPYSL5 cause a neurodevelopmental disorder with corpus callosum agenesis and cerebellar abnormalities.

Am J Hum Genet 2021 May 23;108(5):951-961. Epub 2021 Apr 23.

Service de Génétique, Centre Hospitalier Universitaire, 86021 Poitiers, France; Equipe d'Accueil 3808, Université de Poitiers, 86034 Poitiers, France.

The collapsin response mediator protein (CRMP) family proteins are intracellular mediators of neurotrophic factors regulating neurite structure/spine formation and are essential for dendrite patterning and directional axonal pathfinding during brain developmental processes. Among this family, CRMP5/DPYSL5 plays a significant role in neuronal migration, axonal guidance, dendrite outgrowth, and synapse formation by interacting with microtubules. Here, we report the identification of missense mutations in DPYSL5 in nine individuals with brain malformations, including corpus callosum agenesis and/or posterior fossa abnormalities, associated with variable degrees of intellectual disability. A recurrent de novo p.Glu41Lys variant was found in eight unrelated patients, and a p.Gly47Arg variant was identified in one individual from the first family reported with Ritscher-Schinzel syndrome. Functional analyses of the two missense mutations revealed impaired dendritic outgrowth processes in young developing hippocampal primary neuronal cultures. We further demonstrated that these mutations, both located in the same loop on the surface of DPYSL5 monomers and oligomers, reduced the interaction of DPYSL5 with neuronal cytoskeleton-associated proteins MAP2 and βIII-tubulin. Our findings collectively indicate that the p.Glu41Lys and p.Gly47Arg variants impair DPYSL5 function on dendritic outgrowth regulation by preventing the formation of the ternary complex with MAP2 and βIII-tubulin, ultimately leading to abnormal brain development. This study adds DPYSL5 to the list of genes implicated in brain malformation and in neurodevelopmental disorders.
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http://dx.doi.org/10.1016/j.ajhg.2021.04.004DOI Listing
May 2021

Haploinsufficiency of PRR12 causes a spectrum of neurodevelopmental, eye, and multisystem abnormalities.

Genet Med 2021 Apr 6. Epub 2021 Apr 6.

Department of Genetics, University of Groningen, University Medical Center Groningen, Groningen, Netherlands.

Purpose: Proline Rich 12 (PRR12) is a gene of unknown function with suspected DNA-binding activity, expressed in developing mice and human brains. Predicted loss-of-function variants in this gene are extremely rare, indicating high intolerance of haploinsufficiency.

Methods: Three individuals with intellectual disability and iris anomalies and truncating de novo PRR12 variants were described previously. We add 21 individuals with similar PRR12 variants identified via matchmaking platforms, bringing the total number to 24.

Results: We observed 12 frameshift, 6 nonsense, 1 splice-site, and 2 missense variants and one patient with a gross deletion involving PRR12. Three individuals had additional genetic findings, possibly confounding the phenotype. All patients had developmental impairment. Variable structural eye defects were observed in 12/24 individuals (50%) including anophthalmia, microphthalmia, colobomas, optic nerve and iris abnormalities. Additional common features included hypotonia (61%), heart defects (52%), growth failure (54%), and kidney anomalies (35%). PrediXcan analysis showed that phecodes most strongly associated with reduced predicted PRR12 expression were enriched for eye- (7/30) and kidney- (4/30) phenotypes, such as wet macular degeneration and chronic kidney disease.

Conclusion: These findings support PRR12 haploinsufficiency as a cause for a novel disorder with a wide clinical spectrum marked chiefly by neurodevelopmental and eye abnormalities.
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http://dx.doi.org/10.1038/s41436-021-01129-6DOI Listing
April 2021

Large-scale targeted sequencing identifies risk genes for neurodevelopmental disorders.

Nat Commun 2020 10 1;11(1):4932. Epub 2020 Oct 1.

Oasi Research Institute-IRCCS, Troina, Italy.

Most genes associated with neurodevelopmental disorders (NDDs) were identified with an excess of de novo mutations (DNMs) but the significance in case-control mutation burden analysis is unestablished. Here, we sequence 63 genes in 16,294 NDD cases and an additional 62 genes in 6,211 NDD cases. By combining these with published data, we assess a total of 125 genes in over 16,000 NDD cases and compare the mutation burden to nonpsychiatric controls from ExAC. We identify 48 genes (25 newly reported) showing significant burden of ultra-rare (MAF < 0.01%) gene-disruptive mutations (FDR 5%), six of which reach family-wise error rate (FWER) significance (p < 1.25E-06). Among these 125 targeted genes, we also reevaluate DNM excess in 17,426 NDD trios with 6,499 new autism trios. We identify 90 genes enriched for DNMs (FDR 5%; e.g., GABRG2 and UIMC1); of which, 61 reach FWER significance (p < 3.64E-07; e.g., CASZ1). In addition to doubling the number of patients for many NDD risk genes, we present phenotype-genotype correlations for seven risk genes (CTCF, HNRNPU, KCNQ3, ZBTB18, TCF12, SPEN, and LEO1) based on this large-scale targeted sequencing effort.
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http://dx.doi.org/10.1038/s41467-020-18723-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7530681PMC
October 2020

De novo SMARCA2 variants clustered outside the helicase domain cause a new recognizable syndrome with intellectual disability and blepharophimosis distinct from Nicolaides-Baraitser syndrome.

Genet Med 2020 11 22;22(11):1838-1850. Epub 2020 Jul 22.

Department of Genetics, Robert Debré Hospital, AP-HP, Paris, France.

Purpose: Nontruncating variants in SMARCA2, encoding a catalytic subunit of SWI/SNF chromatin remodeling complex, cause Nicolaides-Baraitser syndrome (NCBRS), a condition with intellectual disability and multiple congenital anomalies. Other disorders due to SMARCA2 are unknown.

Methods: By next-generation sequencing, we identified candidate variants in SMARCA2 in 20 individuals from 18 families with a syndromic neurodevelopmental disorder not consistent with NCBRS. To stratify variant interpretation, we functionally analyzed SMARCA2 variants in yeasts and performed transcriptomic and genome methylation analyses on blood leukocytes.

Results: Of 20 individuals, 14 showed a recognizable phenotype with recurrent features including epicanthal folds, blepharophimosis, and downturned nasal tip along with variable degree of intellectual disability (or blepharophimosis intellectual disability syndrome [BIS]). In contrast to most NCBRS variants, all SMARCA2 variants associated with BIS are localized outside the helicase domains. Yeast phenotype assays differentiated NCBRS from non-NCBRS SMARCA2 variants. Transcriptomic and DNA methylation signatures differentiated NCBRS from BIS and those with nonspecific phenotype. In the remaining six individuals with nonspecific dysmorphic features, clinical and molecular data did not permit variant reclassification.

Conclusion: We identified a novel recognizable syndrome named BIS associated with clustered de novo SMARCA2 variants outside the helicase domains, phenotypically and molecularly distinct from NCBRS.
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http://dx.doi.org/10.1038/s41436-020-0898-yDOI Listing
November 2020

Severe paroxysmal dyskinesias without epilepsy in a RHOBTB2 mutation carrier.

Parkinsonism Relat Disord 2020 08 29;77:87-88. Epub 2020 Jun 29.

Department of Neurology and Center of Clinical Neuroscience, First Faculty of Medicine, Charles University and General University Hospital in Prague, Prague, Czech Republic.

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http://dx.doi.org/10.1016/j.parkreldis.2020.06.028DOI Listing
August 2020

Teaching a difficult topic using a problem-based concept resembling a computer game: development and evaluation of an e-learning application for medical molecular genetics.

BMC Med Educ 2019 Oct 24;19(1):390. Epub 2019 Oct 24.

Department of Biology and Medical Genetics, Charles University Second Faculty of Medicine and University Hospital Motol, V Uvalu 84, 150 06, Prague, Czech Republic.

Background: Genetic testing rapidly penetrates into all medical specialties and medical students must acquire skills in this area. However, many of them consider it difficult. Furthermore, many find these topics less appealing and not connected to their future specialization in different fields of clinical medicine. Student-centred strategies such as problem-based learning, gamification and the use of real data can increase the appeal of a difficult topic such as genetic testing, a field at the crossroads of genetics, molecular biology and bioinformatics.

Methods: We designed an electronic teaching application which students registered in the undergraduate Medical Biology course can access online. A study was carried out to assess the influence of implementation of the new method. We performed pretest/posttest evaluation and analyzed the results using the sign test with median values. We also collected students' personal comments.

Results: The newly developed interactive application simulates the process of molecular genetic diagnostics of a hereditary disorder in a family. Thirteen tasks guide students through clinical and laboratory steps needed to reach the final diagnosis. Genetics and genomics are fields strongly dependent on electronic databases and computer-based data analysis tools. The tasks employ publicly available internet bioinformatic resources used routinely in medical genetics departments worldwide. Authenticity is assured by the use of modified and de-identified clinical and laboratory data from real families analyzed in our previous research projects. Each task contains links to databases and data processing tools needed to solve the task, and an answer box. If the entered answer is correct, the system allows the user to proceed to the next task. The solving of consecutive tasks arranged into a single narrative resembles a computer game, making the concept appealing. There was a statistically significant improvement of knowledge and skills after the practical class, and most comments on the application were positive. A demo version is available at https://medbio.lf2.cuni.cz/demo_m/ . Full version is available on request from the authors.

Conclusions: Our concept proved to be appealing to the students and effective in teaching medical molecular genetics. It can be modified for training in the use of electronic information resources in other medical specialties.
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http://dx.doi.org/10.1186/s12909-019-1817-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6813102PMC
October 2019

Missense Mutations in NKAP Cause a Disorder of Transcriptional Regulation Characterized by Marfanoid Habitus and Cognitive Impairment.

Am J Hum Genet 2019 11 3;105(5):987-995. Epub 2019 Oct 3.

Zebrafish Core Facility, The Children's Hospital of Philadelphia, Philadelphia, PA 19104, USA.

NKAP is a ubiquitously expressed nucleoplasmic protein that is currently known as a transcriptional regulatory molecule via its interaction with HDAC3 and spliceosomal proteins. Here, we report a disorder of transcriptional regulation due to missense mutations in the X chromosome gene, NKAP. These mutations are clustered in the C-terminal region of NKAP where NKAP interacts with HDAC3 and post-catalytic spliceosomal complex proteins. Consistent with a role for the C-terminal region of NKAP in embryogenesis, nkap mutant zebrafish with a C-terminally truncated NKAP demonstrate severe developmental defects. The clinical features of affected individuals are highly conserved and include developmental delay, hypotonia, joint contractures, behavioral abnormalities, Marfanoid habitus, and scoliosis. In affected cases, transcriptome analysis revealed the presence of a unique transcriptome signature, which is characterized by the downregulation of long genes with higher exon numbers. These observations indicate the critical role of NKAP in transcriptional regulation and demonstrate that perturbations of the C-terminal region lead to developmental defects in both humans and zebrafish.
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http://dx.doi.org/10.1016/j.ajhg.2019.09.009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6848994PMC
November 2019

Parental gonadal but not somatic mosaicism leading to de novo NFIX variants shared by two brothers with Malan syndrome.

Am J Med Genet A 2019 10 1;179(10):2119-2123. Epub 2019 Aug 1.

Department of Biology and Medical Genetics, Charles University 2nd Faculty of Medicine and University Hospital Motol, Prague, Czech Republic.

The importance of gonadal mosaicism in families with apparently de novo mutations is being increasingly recognized. We report on two affected brothers initially suggestive of X-linked or autosomal recessive inheritance. Malan syndrome due to shared NFIX variants was diagnosed in the brothers using exome sequencing. The boys shared the same paternal but not maternal haplotype around NFIX, and deep amplicon sequencing showed ~7% of the variant in paternal sperm but not in paternal blood and saliva. We performed review of previous cases of gonadal mosaicism, which suggests that the phenomenon is not uncommon. Gonadal mosaicism is often not accompanied by somatic mosaicism in tissues routinely used for testing, and if both types of mosaicism are present, the frequency of the variant in sperm is often higher than in somatic cells. In families with shared apparently de novo variants without evidence of parental somatic mosaicism, the transmitting parent may be determined through haplotyping of exome variants. Gonadal mosaicism has important consequences for recurrence risks and should be considered in genetic counseling in families with de novo variants.
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http://dx.doi.org/10.1002/ajmg.a.61302DOI Listing
October 2019

De Novo Heterozygous POLR2A Variants Cause a Neurodevelopmental Syndrome with Profound Infantile-Onset Hypotonia.

Am J Hum Genet 2019 08 25;105(2):283-301. Epub 2019 Jul 25.

Molecular Cancer Research, Center for Molecular Medicine, University Medical Center Utrecht, Utrecht University, Oncode Institute, 3584 CT Utrecht, the Netherlands.

The RNA polymerase II complex (pol II) is responsible for transcription of all ∼21,000 human protein-encoding genes. Here, we describe sixteen individuals harboring de novo heterozygous variants in POLR2A, encoding RPB1, the largest subunit of pol II. An iterative approach combining structural evaluation and mass spectrometry analyses, the use of S. cerevisiae as a model system, and the assessment of cell viability in HeLa cells allowed us to classify eleven variants as probably disease-causing and four variants as possibly disease-causing. The significance of one variant remains unresolved. By quantification of phenotypic severity, we could distinguish mild and severe phenotypic consequences of the disease-causing variants. Missense variants expected to exert only mild structural effects led to a malfunctioning pol II enzyme, thereby inducing a dominant-negative effect on gene transcription. Intriguingly, individuals carrying these variants presented with a severe phenotype dominated by profound infantile-onset hypotonia and developmental delay. Conversely, individuals carrying variants expected to result in complete loss of function, thus reduced levels of functional pol II from the normal allele, exhibited the mildest phenotypes. We conclude that subtle variants that are central in functionally important domains of POLR2A cause a neurodevelopmental syndrome characterized by profound infantile-onset hypotonia and developmental delay through a dominant-negative effect on pol-II-mediated transcription of DNA.
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http://dx.doi.org/10.1016/j.ajhg.2019.06.016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6699192PMC
August 2019

A novel variant of C12orf4 in a consanguineous Armenian family confirms the etiology of autosomal recessive intellectual disability type 66 with delineation of the phenotype.

Mol Genet Genomic Med 2019 09 23;7(9):e865. Epub 2019 Jul 23.

Department of Biology and Medical Genetics, Charles University 2nd Faculty of Medicine and University Hospital Motol, Prague, Czech Republic.

Background: Intellectual disability (ID) is a feature of many rare diseases caused by thousands of genes. This genetic heterogeneity implies that pathogenic variants in a specific gene are found only in a small number of patients, and difficulties arise in the definition of prevailing genotype and characteristic phenotype associated with that gene. One of such very rare disorders is autosomal recessive ID type 66 (OMIM #618221) caused by defects in C12orf4. Up to now, six families have been reported with mostly truncating variants. The spectrum of the clinical phenotype was not emphasized in previous reports, and detailed phenotype was not always available from previous patients, especially from large cohort studies.

Methods: Exome sequencing was performed in a consanguineous Armenian family with two affected adult brothers.

Results: The patients carry a novel homozygous nonsense C12orf4 variant. The integration of previous data and phenotyping of the brothers indicate that the clinical picture of C12orf4 defects involves hypotonia in infancy, rather severe ID, speech impairment, and behavioral problems such as aggressiveness, unstable mood, and autistic features. Several other symptoms are more variable and less consistent.

Conclusion: This rather nonsyndromic and nonspecific clinical picture implies that additional patients with C12orf4 defects will likely continue to be identified using the "genotype-first" approach, rather than based on clinical assessment. The phenotype needs further delineation in future reports.
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http://dx.doi.org/10.1002/mgg3.865DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6732288PMC
September 2019

De novo mutations in the GTP/GDP-binding region of RALA, a RAS-like small GTPase, cause intellectual disability and developmental delay.

PLoS Genet 2018 11 30;14(11):e1007671. Epub 2018 Nov 30.

HudsonAlpha Institute for Biotechnology, Huntsville, AL, United States of America.

Mutations that alter signaling of RAS/MAPK-family proteins give rise to a group of Mendelian diseases known as RASopathies. However, among RASopathies, the matrix of genotype-phenotype relationships is still incomplete, in part because there are many RAS-related proteins and in part because the phenotypic consequences may be variable and/or pleiotropic. Here, we describe a cohort of ten cases, drawn from six clinical sites and over 16,000 sequenced probands, with de novo protein-altering variation in RALA, a RAS-like small GTPase. All probands present with speech and motor delays, and most have intellectual disability, low weight, short stature, and facial dysmorphism. The observed rate of de novo RALA variants in affected probands is significantly higher (p = 4.93 x 10(-11)) than expected from the estimated random mutation rate. Further, all de novo variants described here affect residues within the GTP/GDP-binding region of RALA; in fact, six alleles arose at only two codons, Val25 and Lys128. The affected residues are highly conserved across both RAL- and RAS-family genes, are devoid of variation in large human population datasets, and several are homologous to positions at which disease-associated variants have been observed in other GTPase genes. We directly assayed GTP hydrolysis and RALA effector-protein binding of the observed variants, and found that all but one tested variant significantly reduced both activities compared to wild-type. The one exception, S157A, reduced GTP hydrolysis but significantly increased RALA-effector binding, an observation similar to that seen for oncogenic RAS variants. These results show the power of data sharing for the interpretation and analysis of rare variation, expand the spectrum of molecular causes of developmental disability to include RALA, and provide additional insight into the pathogenesis of human disease caused by mutations in small GTPases.
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http://dx.doi.org/10.1371/journal.pgen.1007671DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6291162PMC
November 2018

Correction: IQSEC2-related encephalopathy in males and females: a comparative study including 37 novel patients.

Genet Med 2019 Aug;21(8):1897-1898

APHP, Service de genetique medicale, Necker- Enfants Malades Hospital, Imagine Institute, Paris Descartes University, Paris, France.

This Article was originally published under Nature Research's License to Publish, but has now been made available under a CC BY 4.0 license. The PDF and HTML versions of the Article have been modified accordingly.
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http://dx.doi.org/10.1038/s41436-018-0327-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7608434PMC
August 2019

IQSEC2-related encephalopathy in males and females: a comparative study including 37 novel patients.

Genet Med 2019 04 12;21(4):837-849. Epub 2018 Sep 12.

APHP, Service de genetique medicale, Necker-Enfants Malades Hospital, Imagine Institute, Paris Descartes University, Paris, France.

Purpose: Variants in IQSEC2, escaping X inactivation, cause X-linked intellectual disability with frequent epilepsy in males and females. We aimed to investigate sex-specific differences.

Methods: We collected the data of 37 unpublished patients (18 males and 19 females) with IQSEC2 pathogenic variants and 5 individuals with variants of unknown significance and reviewed published variants. We compared variant types and phenotypes in males and females and performed an analysis of IQSEC2 isoforms.

Results: IQSEC2 pathogenic variants mainly led to premature truncation and were scattered throughout the longest brain-specific isoform, encoding the synaptic IQSEC2/BRAG1 protein. Variants occurred de novo in females but were either de novo (2/3) or inherited (1/3) in males, with missense variants being predominantly inherited. Developmental delay and intellectual disability were overall more severe in males than in females. Likewise, seizures were more frequently observed and intractable, and started earlier in males than in females. No correlation was observed between the age at seizure onset and severity of intellectual disability or resistance to antiepileptic treatments.

Conclusion: This study provides a comprehensive overview of IQSEC2-related encephalopathy in males and females, and suggests that an accurate dosage of IQSEC2 at the synapse is crucial during normal brain development.
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http://dx.doi.org/10.1038/s41436-018-0268-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6752297PMC
April 2019

Molecular Cytogenetic Diagnostics of Marker Chromosomes: Analysis in Four Prenatal Cases and Long-Term Clinical Evaluation of Carriers.

Cytogenet Genome Res 2018 9;154(4):187-195. Epub 2018 May 9.

Department of Biology and Medical Genetics, Charles University 2nd Faculty of Medicine and University Hospital Motol, Prague, Czech Republic.

The prenatal finding of a small supernumerary marker chromosome (sSMC) is a challenge for genetic counseling. Our analytic algorithm is based on sSMC frequencies and multicolor FISH to accelerate the procedure. The chromosomal origin, size, and degree of mosaicism of the sSMC then determine the prognosis. We illustrate the effectiveness on 4 prenatally identified de novo mosaic sSMCs derived from chromosomes 13/21, X, 3, and 17. Three sSMC carriers had a good prognosis and apparently healthy children were born, showing no abnormality till the last examination at the age of 4 years. One case had a poor prognosis, and the parents decided to terminate the pregnancy. Our work contributes to the laboratory and clinical management of prenatally detected sSMCs. FISH is a reliable method for fast sSMC evaluation and prognosis assessment; it prevents unnecessary delays and uncertainty, allows informed decision making, and reduces unnecessary pregnancy terminations.
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http://dx.doi.org/10.1159/000488790DOI Listing
July 2018

Association of 17q24.2-q24.3 deletions with recognizable phenotype and short telomeres.

Am J Med Genet A 2018 06 25;176(6):1438-1442. Epub 2018 Apr 25.

Department of Biology and Medical Genetics, 2nd Faculty of Medicine and University Hospital Motol, Charles University, Prague, Czech Republic.

Microdeletions of 17q24.2-q24.3 have been described in several patients with developmental and speech delay, growth retardation, and other features. The relatively large size and limited overlap of the deletions complicate the genotype-phenotype correlation. We identified a girl with intellectual disability, growth retardation, dysmorphic features, and a de novo 2.8 Mb long deletion of 17q24.2-q24.3. Her phenotype was strikingly similar to one previously described boy with Dubowitz syndrome (MIM 223370) and a de novo 3.9 Mb long deletion encompassing the deletion of our patient. In addition, both patients had the shortest telomeres among normal age-matched controls. Our review of all 17q24.2-q24.3 deletion patients revealed additional remarkable phenotypic features shared by the patients, some of which have consequences for their management. Proposed novel genotype-phenotype correlations based on new literature information on the region include the role of PSMD12 and BPTF, the genes recently associated with syndromic neurodevelopmental disorders, and a possible role of the complex topologically associated domain structure of the region, which may explain some of the phenotypic discrepancies observed between patients with similar but not identical deletions. Nevertheless, although different diagnoses including the Dubowitz, Nijmegen breakage (MIM 251260), Silver-Russell (MIM 180860), or Myhre (MIM 139210) syndromes were originally considered in the 17q24.2-q24.3 deletion patients, they clearly belong to one diagnostic entity defined by their deletions and characterized especially by developmental delay, specific facial dysmorphism, abnormalities of extremities and other phenotypes, and possibly also short telomere length.
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http://dx.doi.org/10.1002/ajmg.a.38711DOI Listing
June 2018

Very short DNA segments can be detected and handled by the repair machinery during germline chromothriptic chromosome reassembly.

Hum Mutat 2018 05 20;39(5):709-716. Epub 2018 Feb 20.

Department of Biology and Medical Genetics, Charles University 2nd Faculty of Medicine and University Hospital Motol, Prague, Czech Republic.

Analyses at nucleotide resolution reveal unexpected complexity of seemingly simple and balanced chromosomal rearrangements. Chromothripsis is a rare complex aberration involving local shattering of one or more chromosomes and reassembly of the resulting DNA segments. This can influence gene expression and cause abnormal phenotypes. We studied the structure and mechanism of a seemingly balanced de novo complex rearrangement of four chromosomes in a boy with developmental and growth delay. Microarray analysis revealed two paternal de novo deletions of 0.7 and 2.5 Mb at two of the breakpoints in 1q24.3 and 6q24.1-q24.2, respectively, which could explain most symptoms of the patient. Subsequent whole-genome mate-pair sequencing confirmed the chromothriptic nature of the rearrangement. The four participating chromosomes were broken into 29 segments longer than 1 kb. Sanger sequencing of all breakpoint junctions revealed additional complexity compatible with the involvement of different repair pathways. We observed translocation of a 33 bp long DNA fragment, which may have implications for the definition of the lower size limit of structural variants. Our observations and literature review indicate that even very small fragments from shattered chromosomes can be detected and handled by the repair machinery during germline chromothriptic chromosome reassembly.
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http://dx.doi.org/10.1002/humu.23408DOI Listing
May 2018

-related intellectual disability syndrome: a recognisable entity.

J Med Genet 2017 09 22;54(9):613-623. Epub 2017 Jul 22.

Laboratory of Medical Genetics, Bambino Gesù Children's Hospital, IRCCS, Rome, Italy.

Background: Mutations in forkhead box protein P1 () cause intellectual disability (ID) and specific language impairment (SLI), with or without autistic features (MIM: 613670). Despite multiple case reports no specific phenotype emerged so far.

Methods: We correlate clinical and molecular data of 25 novel and 23 previously reported patients with defects. We evaluated FOXP1 activity by an in vitro luciferase model and assessed protein stability in vitro by western blotting.

Results: Patients show ID, SLI, neuromotor delay (NMD) and recurrent facial features including a high broad forehead, bent downslanting palpebral fissures, ptosis and/or blepharophimosis and a bulbous nasal tip. Behavioural problems and autistic features are common. Brain, cardiac and urogenital malformations can be associated. More severe ID and NMD, sensorineural hearing loss and feeding difficulties are more common in patients with interstitial 3p deletions (14 patients) versus patients with monogenic defects (34 patients). Mutations result in impaired transcriptional repression and/or reduced protein stability.

Conclusions: -related ID syndrome is a recognisable entity with a wide clinical spectrum and frequent systemic involvement. Our data will be helpful to evaluate genotype-phenotype correlations when interpreting next-generation sequencing data obtained in patients with ID and/or SLI and will guide clinical management.
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http://dx.doi.org/10.1136/jmedgenet-2017-104579DOI Listing
September 2017

Analysis of 31-year-old patient with SYNGAP1 gene defect points to importance of variants in broader splice regions and reveals developmental trajectory of SYNGAP1-associated phenotype: case report.

BMC Med Genet 2017 06 2;18(1):62. Epub 2017 Jun 2.

Department of Biology and Medical Genetics, Charles University 2nd Faculty of Medicine and University Hospital Motol, Plzenska 130/221, 15000, Prague 5, Czech Republic.

Background: Whole exome sequencing is a powerful tool for the analysis of genetically heterogeneous conditions. The prioritization of variants identified often focuses on nonsense, frameshift and canonical splice site mutations, and highly deleterious missense variants, although other defects can also play a role. The definition of the phenotype range and course of rare genetic conditions requires long-term clinical follow-up of patients.

Case Presentation: We report an adult female patient with severe intellectual disability, severe speech delay, epilepsy, autistic features, aggressiveness, sleep problems, broad-based clumsy gait and constipation. Whole exome sequencing identified a de novo mutation in the SYNGAP1 gene. The variant was located in the broader splice donor region of intron 10 and replaced G by A at position +5 of the splice site. The variant was predicted in silico and shown experimentally to abolish the regular splice site and to activate a cryptic donor site within exon 10, causing frameshift and premature termination. The overall clinical picture of the patient corresponded well with the characteristic SYNGAP1-associated phenotype observed in previously reported patients. However, our patient was 31 years old which contrasted with most other published SYNGAP1 cases who were much younger. Our patient had a significant growth delay and microcephaly. Both features normalised later, although the head circumference stayed only slightly above the lower limit of the norm. The patient had a delayed puberty. Her cognitive and language performance remained at the level of a one-year-old child even in adulthood and showed a slow decline. Myopathic facial features and facial dysmorphism became more pronounced with age. Although the gait of the patient was unsteady in childhood, more severe gait problems developed in her teens. While the seizures remained well-controlled, her aggressive behaviour worsened with age and required extensive medication.

Conclusions: The finding in our patient underscores the notion that the interpretation of variants identified using whole exome sequencing should focus not only on variants in the canonical splice dinucleotides GT and AG, but also on broader splice regions. The long-term clinical follow-up of our patient contributes to the knowledge of the developmental trajectory in individuals with SYNGAP1 gene defects.
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http://dx.doi.org/10.1186/s12881-017-0425-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5457574PMC
June 2017

Dominant variants in the splicing factor PUF60 cause a recognizable syndrome with intellectual disability, heart defects and short stature.

Eur J Hum Genet 2016 01 2;25(1):43-51. Epub 2016 Nov 2.

FHU TRANSLAD, Centre de Référence Maladies Rares de l'Est, Centre de Génétique, CHU de Dijon, Dijon, France.

Verheij syndrome, also called 8q24.3 microdeletion syndrome, is a rare condition characterized by ante- and postnatal growth retardation, microcephaly, vertebral anomalies, joint laxity/dislocation, developmental delay (DD), cardiac and renal defects and dysmorphic features. Recently, PUF60 (Poly-U Binding Splicing Factor 60 kDa), which encodes a component of the spliceosome, has been discussed as the best candidate gene for the Verheij syndrome phenotype, regarding the cardiac and short stature phenotype. To date, only one patient has been reported with a de novo variant in PUF60 that probably affects function (c.505C>T leading to p.(His169Tyr)) associated with DD, microcephaly, craniofacial and cardiac defects. Additional patients were required to confirm the pathogenesis of this association and further delineate the clinical spectrum. Here we report five patients with de novo heterozygous variants in PUF60 identified using whole exome sequencing. Variants included a splice-site variant (c.24+1G>C), a frameshift variant (p.(Ile136Thrfs*31)), two nonsense variants (p.(Arg448*) and p.(Lys301*)) and a missense change (p.(Val483Ala)). All six patients with a PUF60 variant (the five patients of the present study and the unique reported patient) have the same core facial gestalt as 8q24.3 microdeletions patients, associated with DD. Other findings include feeding difficulties (3/6), cardiac defects (5/6), short stature (5/6), joint laxity and/or dislocation (5/6), vertebral anomalies (3/6), bilateral microphthalmia and irido-retinal coloboma (1/6), bilateral optic nerve hypoplasia (2/6), renal anomalies (2/6) and branchial arch defects (2/6). These results confirm that PUF60 is a major driver for the developmental, craniofacial, skeletal and cardiac phenotypes associated with the 8q24.3 microdeletion.
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http://dx.doi.org/10.1038/ejhg.2016.133DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5159768PMC
January 2016

Expanded repeats in dizygotic twins referred for diagnosis of autism versus absence of expanded repeats at screening of 330 children with autism.

Neuropsychiatr Dis Treat 2016 19;12:2367-2372. Epub 2016 Sep 19.

Department of Biology and Medical Genetics.

Myotonic dystrophy type 1 (DM1) belongs to the broad spectrum of genetic disorders associated with autism spectrum disorders (ASD). ASD were reported predominantly in congenital and early childhood forms of DM1. We describe dizygotic twin boys with ASD who were referred for routine laboratory genetic testing and in whom karyotyping, gene testing, and single nucleotide polymorphism array analysis yielded negative results. The father of the boys was later diagnosed with suspected DM1, and testing revealed characteristic gene expansions in his genome as well as in the genomes of both twins and their elder brother, who also suffered from ASD. In accord with previous reports on childhood forms of DM1, our patients showed prominent neuropsychiatric phenotypes characterized especially by hypotonia, developmental and language delay, emotional and affective lability, lowered adaptability, and social withdrawal. The experience with this family and multiple literature reports of ASD in DM1 on the one side but the lack of literature data on the frequency of gene expansions in ASD patients on the other side prompted us to screen the gene in a sample of 330 patients with ASD who were first seen by a geneticist before they were 10 years of age, before the muscular weakness, which may signal DM1, usually becomes obvious. The absence of any gene expansions in this cohort indicates that targeted gene testing can be recommended only in ASD patients with specific symptoms or family history suggestive of DM1.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5034902PMC
http://dx.doi.org/10.2147/NDT.S113917DOI Listing
September 2016

A patient showing features of both SBBYSS and GPS supports the concept of a KAT6B-related disease spectrum, with mutations in mid-exon 18 possibly leading to combined phenotypes.

Eur J Med Genet 2015 Oct 11;58(10):550-5. Epub 2015 Sep 11.

Department of Biology and Medical Genetics, Charles University 2nd Faculty of Medicine and University Hospital Motol, Prague, Czech Republic.

Genitopatellar syndrome (GPS) and Say-Barber-Biesecker-Young-Simpson syndrome (SBBYSS) are two distinct clinically overlapping syndromes caused by de novo heterozygous truncating mutations in the KAT6B gene encoding lysine acetyltransferase 6B, a part of the histone H3 acetyltransferase complex. We describe an 8-year-old girl with a KAT6B mutation and a combined GPS/SBBYSS phenotype. The comparison of this patient with 61 previously published cases with KAT6B mutations and GPS, SBBYSS or combined GPS/SBBYSS phenotypes allowed us to separate the KAT6B mutations into four groups according to their position in the gene (reflecting nonsense mediated RNA decay and protein domains) and their clinical outcome. We suggest that mutations in mid-exon 18 corresponding to the C-terminal end of the acidic (Asp/Glu-rich) domain of KAT6B may have more variable expressivity leading to GPS, SBBYSS or combined phenotypes, in contrast to defects in other regions of the gene which contribute more specifically to either GPS or SBBYSS. Notwithstanding the clinical overlap, our cluster analysis of phenotypes of all known patients with KAT6B mutations supports the existence of two clinical entities, GPS and SBBYSS, as poles within the KAT6B-related disease spectrum. The awareness of these phenomena is important for qualified genetic counselling of patients with KAT6B mutations.
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http://dx.doi.org/10.1016/j.ejmg.2015.09.004DOI Listing
October 2015

BCL11A deletions result in fetal hemoglobin persistence and neurodevelopmental alterations.

J Clin Invest 2015 Jun 4;125(6):2363-8. Epub 2015 May 4.

A transition from fetal hemoglobin (HbF) to adult hemoglobin (HbA) normally occurs within a few months after birth. Increased production of HbF after this period of infancy ameliorates clinical symptoms of the major disorders of adult β-hemoglobin: β-thalassemia and sickle cell disease. The transcription factor BCL11A silences HbF and has been an attractive therapeutic target for increasing HbF levels; however, it is not clear to what extent BCL11A inhibits HbF production or mediates other developmental functions in humans. Here, we identified and characterized 3 patients with rare microdeletions of 2p15-p16.1 who presented with an autism spectrum disorder and developmental delay. Moreover, these patients all exhibited substantial persistence of HbF but otherwise retained apparently normal hematologic and immunologic function. Of the genes within 2p15-p16.1, only BCL11A was commonly deleted in all of the patients. Evaluation of gene expression data sets from developing and adult human brains revealed that BCL11A expression patterns are similar to other genes associated with neurodevelopmental disorders. Additionally, common SNPs within the second intron of BCL11A are strongly associated with schizophrenia. Together, the study of these rare patients and orthogonal genetic data demonstrates that BCL11A plays a central role in silencing HbF in humans and implicates BCL11A as an important factor for neurodevelopment.
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http://dx.doi.org/10.1172/JCI81163DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4497765PMC
June 2015

Hypophosphatasia due to uniparental disomy.

Bone 2015 Dec 1;81:765-766. Epub 2015 May 1.

Department of Biology and Medical Genetics, Charles University 2nd Faculty of Medicine and University Hospital Motol, Prague, Czech Republic.

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http://dx.doi.org/10.1016/j.bone.2015.04.041DOI Listing
December 2015

HCFC1 loss-of-function mutations disrupt neuronal and neural progenitor cells of the developing brain.

Hum Mol Genet 2015 Jun 3;24(12):3335-47. Epub 2015 Mar 3.

School of Paediatrics and Reproductive Health, Robinson Research Institute and School of Molecular and Biomedical Sciences, University of Adelaide, Adelaide 5000, Australia,

Both gain- and loss-of-function mutations have recently implicated HCFC1 in neurodevelopmental disorders. Here, we extend our previous HCFC1 over-expression studies by employing short hairpin RNA to reduce the expression of Hcfc1 in embryonic neural cells. We show that in contrast to over-expression, loss of Hcfc1 favoured proliferation of neural progenitor cells at the expense of differentiation and promoted axonal growth of post-mitotic neurons. To further support the involvement of HCFC1 in neurological disorders, we report two novel HCFC1 missense variants found in individuals with intellectual disability (ID). One of these variants, together with three previously reported HCFC1 missense variants of unknown pathogenicity, were functionally assessed using multiple cell-based assays. We show that three out of the four variants tested result in a partial loss of HCFC1 function. While over-expression of the wild-type HCFC1 caused reduction in HEK293T cell proliferation and axonal growth of neurons, these effects were alleviated upon over-expression of three of the four HCFC1 variants tested. One of these partial loss-of-function variants disrupted a nuclear localization sequence and the resulting protein displayed reduced ability to localize to the cell nucleus. The other two variants displayed negative effects on the expression of the HCFC1 target gene MMACHC, which is responsible for the metabolism of cobalamin, suggesting that these individuals may also be susceptible to cobalamin deficiency. Together, our work identifies plausible cellular consequences of missense HCFC1 variants and identifies likely and relevant disease mechanisms that converge on embryonic stages of brain development.
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http://dx.doi.org/10.1093/hmg/ddv083DOI Listing
June 2015

Long term follow-up in a patient with a de novo microdeletion of 14q11.2 involving CHD8.

Am J Med Genet A 2015 Apr 3;167A(4):837-41. Epub 2015 Mar 3.

Department of Biology and Medical Genetics, Charles University 2nd Faculty of Medicine and University Hospital Motol, Prague, Czech Republic.

We identified a de novo deletion of 14q11.2 in a Czech patient with developmental delay, mild autistic features, macrosomy, macrocephaly, orthognathic deformities, and dysmorphic facial features. The clinical follow-up of the patient lasting 14 years documented changes in the facial dysmorphism from infancy to adolescence. The deletion affects approximately 200 kb of DNA with five protein-coding genes and two snoRNA genes. Two of the protein-coding genes, SUPT16H and CHD8, have been proposed as candidate genes for a new microdeletion syndrome. Our patient further supports the existence of this syndrome and extends its phenotypic spectrum, especially points to the possibility that orthognathic deformities may be associated with microdeletions of 14q11.2. CHD8 mutations have been found in patients with neurodevelopmental disorders and macrocephaly. The HNRNPC gene, repeatedly deleted in patients with developmental delay, is another candidate as its 5́ end is adjacent to the deletion, and the expression of this gene may be affected by position effect.
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http://dx.doi.org/10.1002/ajmg.a.36957DOI Listing
April 2015

A 15 Mb large paracentric chromosome 21 inversion identified in Czech population through a pair of flanking duplications.

Mol Cytogenet 2014 19;7:51. Epub 2014 Aug 19.

Department of Biology and Medical Genetics, Charles University 2nd Faculty of Medicine and University Hospital Motol, Prague, Czech Republic.

Background: Inversions are balanced structural chromosome rearrangements, which can influence gene expression and the risk of unbalanced chromosome constitution in offspring. Many examples of inversion polymorphisms exist in human, affecting both heterochromatic regions and euchromatin.

Results: We describe a novel, 15 Mb long paracentric inversion, inv(21)(q21.1q22.11), affecting more than a third of human 21q. Despite of its length, the inversion cannot be detected using karyotyping due to similar band patterns on the normal and inverted chromosomes, and is therefore likely to escape attention. Its identification was aided by the repeated observation of the same pair of 150 kb long duplications present in cis on chromosome 21 in three Czech families subjected to microarray analysis. The finding prompted us to hypothesise that this co-occurrence of two remote duplications could be associated with an inversion of the intervening segment, and this speculation turned out to be right. The inversion was confirmed in a series of FISH experiments which also showed that the second copy of each of the duplications was always located at the opposite end of the inversion. The presence of the same pair of duplications in additional individuals reported in public databases indicates that the inversion may also be present in other populations. Three out of the total of about 4000 chromosomes 21 examined in our sample carried the duplications and were inverted, corresponding to carrier frequency of about 1/660. Although the breakpoints affect protein-coding genes, the occurrence of the inversion in normal parents and siblings of our patients and the occurrence of the duplications in unaffected controls in databases indicate that this rare variant is rather non-pathogenic. The inverted segment carried an identical shared haplotype in the three families studied. The haplotypes, however, diverged very rapidly in the flanking regions, possibly pointing to an ancient founder event at the origin of the inversion.

Conclusions: The identification of inv(21)(q21.1q22.11) supports the notion that paracentric inversions are the most common form of chromosomal variation and that some of them may still remain undetected.
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http://dx.doi.org/10.1186/1755-8166-7-51DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4236861PMC
November 2014

Deletions of 9q21.3 including NTRK2 are associated with severe phenotype.

Am J Med Genet A 2015 Jan 27;167A(1):264-7. Epub 2014 Oct 27.

Department of Biology and Medical Genetics, Charles University 2nd Faculty of Medicine and University Hospital Motol, Prague, Czech Republic.

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http://dx.doi.org/10.1002/ajmg.a.36797DOI Listing
January 2015

Monozygotic twins with 17q21.31 microdeletion syndrome.

Twin Res Hum Genet 2014 Oct 9;17(5):405-10. Epub 2014 Jun 9.

Department of Biology and Medical Genetics,Charles University 2nd Faculty of Medicine and University Hospital Motol,Prague,Czech Republic.

Chromosome 17q21.31 microdeletion syndrome is a genomic disorder caused by a recurrent 600 kb long deletion. The deletion affects the region of a common inversion present in about 20% of Europeans. The inversion is associated with the H2 haplotype carrying additional low-copy repeats susceptible to non-allelic homologous recombination, and this haplotype is prone to deletion. No instances of 17q21.31 deletions inherited from an affected parent have been reported, and the deletions always affected a parental chromosome with the H2 haplotype. The syndrome is characterized clinically by intellectual disability, hypotonia, friendly behavior and specific facial dysmorphism with long face, large tubular or pear-shaped nose and bulbous nasal tip. We present monozygotic twin sisters showing the typical clinical picture of the syndrome. The phenotype of the sisters was very similar, with a slightly more severe presentation in Twin B. The 17q21.31 microdeletion was confirmed in both patients but in neither of their parents. Potential copy number differences between the genomes of the twins were subsequently searched using high-resolution single nucleotide polymorphism (SNP) and comparative genome hybridisation (CGH) arrays. However, these analyses identified no additional aberrations or genomic differences that could potentially be responsible for the subtle phenotypic differences. These could possibly be related to the more severe perinatal history of Twin B, or to the variable expressivity of the disorder. In accord with the expectations, one of the parents (the mother) was shown to carry the H2 haplotype, and the maternal allele of chromosome 17q21.31 was missing in the twins.
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http://dx.doi.org/10.1017/thg.2014.29DOI Listing
October 2014