Publications by authors named "Niels Tommerup"

159 Publications

Acute and persistent symptoms in non-hospitalized PCR-confirmed COVID-19 patients.

Sci Rep 2021 06 23;11(1):13153. Epub 2021 Jun 23.

Department of Medical Endocrinology and Metabolism, Copenhagen University Hospital (Rigshospitalet), Copenhagen, Denmark.

Reports of persistent symptoms after hospitalization with COVID-19 have raised concern of a "long COVID" syndrome. This study aimed at determining the prevalence of and risk factors for acute and persistent symptoms in non-hospitalized patients with polymerase chain reaction (PCR) confirmed COVID-19. We conducted a cohort study of non-hospitalized participants identified via the Danish Civil Registration System with a SARS-CoV-2-positive PCR-test and available biobank samples. Participants received a digital questionnaire on demographics and COVID-19-related symptoms. Persistent symptoms: symptoms > 4 weeks (in sensitivity analyses > 12 weeks). We included 445 participants, of whom 34% were asymptomatic. Most common acute symptoms were fatigue, headache, and sneezing, while fatigue and reduced smell and taste were most severe. Persistent symptoms, most commonly fatigue and memory and concentration difficulties, were reported by 36% of 198 symptomatic participants with follow-up > 4 weeks. Risk factors for persistent symptoms included female sex (women 44% vs. men 24%, odds ratio 2.7, 95% CI 1.4-5.1, p = 0.003) and BMI (odds ratio 1.1, 95% CI 1.0-1.2, p = 0.001). In conclusion, among non-hospitalized PCR-confirmed COVID-19 patients one third were asymptomatic while one third of symptomatic participants had persistent symptoms illustrating the heterogeneity of disease presentation. These findings should be considered in health care planning and policy making related to COVID-19.
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http://dx.doi.org/10.1038/s41598-021-92045-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8222239PMC
June 2021

A GDF5 frameshift mutation segregating with Grebe type chondrodysplasia and brachydactyly type C+ in a 6 generations family: Clinical report and mini review.

Eur J Med Genet 2021 Jul 16;64(7):104226. Epub 2021 Apr 16.

Human Molecular Genetics Laboratory, National Institute for Biotechnology and Genetic Engineering (NIBGE) College, PIEAS, Faisalabad, Pakistan. Electronic address:

Different mutations in the Growth/Differentiation Factor 5 gene (GDF5) have been associated with varying types of skeletal dysplasia, including Grebe type chondrodysplasia (GTC), Hunter-Thompson syndrome, Du Pan Syndrome and Brachydactyly type C (BDC). Heterozygous pathogenic mutations exert milder effects, whereas homozygous mutations are known to manifest more severe phenotypes. In this study, we report a GDF5 frameshift mutation (c.404delC) segregating over six generations in an extended consanguineous Pakistani family. The family confirmed that both GTC and BDC are part of the GDF5 mutational spectrum, with severe GTC associated with homozygosity, and with a wide phenotypic variability among heterozygous carriers, ranging from unaffected non-penetrant carriers, to classical BDC and to novel unclassified types of brachydactylies.
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http://dx.doi.org/10.1016/j.ejmg.2021.104226DOI Listing
July 2021

Nationwide germline whole genome sequencing of 198 consecutive pediatric cancer patients reveals a high incidence of cancer prone syndromes.

PLoS Genet 2020 12 17;16(12):e1009231. Epub 2020 Dec 17.

Department of Clinical Genetics, Copenhagen University Hospital Rigshospitalet, Copenhagen, Denmark.

Purpose: Historically, cancer predisposition syndromes (CPSs) were rarely established for children with cancer. This nationwide, population-based study investigated how frequently children with cancer had or were likely to have a CPS.

Methods: Children (0-17 years) in Denmark with newly diagnosed cancer were invited to participate in whole-genome sequencing of germline DNA. Suspicion of CPS was assessed according to Jongmans'/McGill Interactive Pediatric OncoGenetic Guidelines (MIPOGG) criteria and familial cancer diagnoses were verified using population-based registries.

Results: 198 of 235 (84.3%) eligible patients participated, of whom 94/198 (47.5%) carried pathogenic variants (PVs) in a CPS gene or had clinical features indicating CPS. Twenty-nine of 198 (14.6%) patients harbored a CPS, of whom 21/198 (10.6%) harbored a childhood-onset and 9/198 (4.5%) an adult-onset CPS. In addition, 23/198 (11.6%) patients carried a PV associated with biallelic CPS. Seven of the 54 (12.9%) patients carried two or more variants in different CPS genes. Seventy of 198 (35.4%) patients fulfilled the Jongmans' and/or MIPOGG criteria indicating an underlying CPS, including two of the 9 (22.2%) patients with an adult-onset CPS versus 18 of the 21 (85.7%) patients with a childhood-onset CPS (p = 0.0022), eight of the additional 23 (34.8%) patients with a heterozygous PV associated with biallelic CPS, and 42 patients without PVs. Children with a central nervous system (CNS) tumor had family members with CNS tumors more frequently than patients with other cancers (11/44, p = 0.04), but 42 of 44 (95.5%) cases did not have a PV in a CPS gene.

Conclusion: These results demonstrate the value of systematically screening pediatric cancer patients for CPSs and indicate that a higher proportion of childhood cancers may be linked to predisposing germline variants than previously supposed.
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http://dx.doi.org/10.1371/journal.pgen.1009231DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7787686PMC
December 2020

RRP7A links primary microcephaly to dysfunction of ribosome biogenesis, resorption of primary cilia, and neurogenesis.

Nat Commun 2020 11 16;11(1):5816. Epub 2020 Nov 16.

Department of Cellular and Molecular Medicine, University of Copenhagen, Blegdamsvej 3, DK-2200, Copenhagen, Denmark.

Primary microcephaly (MCPH) is characterized by reduced brain size and intellectual disability. The exact pathophysiological mechanism underlying MCPH remains to be elucidated, but dysfunction of neuronal progenitors in the developing neocortex plays a major role. We identified a homozygous missense mutation (p.W155C) in Ribosomal RNA Processing 7 Homolog A, RRP7A, segregating with MCPH in a consanguineous family with 10 affected individuals. RRP7A is highly expressed in neural stem cells in developing human forebrain, and targeted mutation of Rrp7a leads to defects in neurogenesis and proliferation in a mouse stem cell model. RRP7A localizes to centrosomes, cilia and nucleoli, and patient-derived fibroblasts display defects in ribosomal RNA processing, primary cilia resorption, and cell cycle progression. Analysis of zebrafish embryos supported that the patient mutation in RRP7A causes reduced brain size, impaired neurogenesis and cell proliferation, and defective ribosomal RNA processing. These findings provide novel insight into human brain development and MCPH.
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http://dx.doi.org/10.1038/s41467-020-19658-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7670429PMC
November 2020

Paroxysmal Cranial Dyskinesia and Nail-Patella Syndrome Caused by a Novel Variant in the LMX1B Gene.

Mov Disord 2020 12 19;35(12):2343-2347. Epub 2020 Sep 19.

Danish Dementia Research Centre, Department of Neurology, Rigshospitalet, University of Copenhagen, Copenhagen, Denmark.

Background: In a Danish family, multiple individuals in five generations present with early-onset paroxysmal cranial dyskinesia, musculoskeletal abnormalities, and kidney dysfunction.

Objective: To demonstrate linkage and to identify the underlying genetic cause of disease.

Methods: Genome-wide single-nucleotide polymorphisms analysis, Sequence-Tagged-Site marker analyses, exome sequencing, and Sanger sequencing were performed.

Results: Linkage analyses identified a candidate locus on chromosome 9. Exome sequencing revealed a novel variant in LMX1B present in all affected individuals, logarithm of the odds (LOD) score of z = 6.54, predicted to be damaging. Nail-patella syndrome (NPS) is caused by pathogenic variants in LMX1B encoding a transcription factor essential to cytoskeletal and kidney growth and dopaminergic and serotonergic network development. NPS is characterized by abnormal musculoskeletal features and kidney dysfunction. Movement disorders have not previously been associated with NPS.

Conclusions: Paroxysmal dyskinesia is a heretofore unrecognized feature of the NPS spectrum. The pathogenic mechanism might relate to aberrant dopaminergic circuits. © 2020 International Parkinson and Movement Disorder Society.
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http://dx.doi.org/10.1002/mds.28244DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8151874PMC
December 2020

Chromothripsis and DNA Repair Disorders.

J Clin Med 2020 Feb 25;9(3). Epub 2020 Feb 25.

Kennedy Center, Department of Clinical Genetics, Copenhagen University Hospital, Rigshospitalet, 2600 Glostrup, Denmark.

Chromothripsis is a mutational mechanism leading to complex and relatively clustered chromosomal rearrangements, resulting in diverse phenotypic outcomes depending on the involved genomic landscapes. It may occur both in the germ and the somatic cells, resulting in congenital and developmental disorders and cancer, respectively. Asymptomatic individuals may be carriers of chromotriptic rearrangements and experience recurrent reproductive failures when two or more chromosomes are involved. Several mechanisms are postulated to underlie chromothripsis. The most attractive hypothesis involves chromosome pulverization in micronuclei, followed by the incorrect reassembly of fragments through DNA repair to explain the clustered nature of the observed complex rearrangements. Moreover, exogenous or endogenous DNA damage induction and dicentric bridge formation may be involved. Chromosome instability is commonly observed in the cells of patients with DNA repair disorders, such as ataxia telangiectasia, Nijmegen breakage syndrome, and Bloom syndrome. In addition, germline variations of have been associated with chromothripsis in sonic hedgehog medulloblastoma and acute myeloid leukemia. In the present review, we focus on the underlying mechanisms of chromothripsis and the involvement of defective DNA repair genes, resulting in chromosome instability and chromothripsis-like rearrangements.
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http://dx.doi.org/10.3390/jcm9030613DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7141117PMC
February 2020

Multigenic truncation of the semaphorin-plexin pathway by a germline chromothriptic rearrangement associated with Moebius syndrome.

Hum Mutat 2019 08 14;40(8):1057-1062. Epub 2019 May 14.

Wilhelm Johannsen Center for Functional Genome Research, Department of Cellular and Molecular Medicine, University of Copenhagen, Copenhagen, Denmark.

Moebius syndrome (MBS) is a congenital disorder caused by paralysis of the facial and abducens nerves. Although a number of candidate genes have been suspected, so far only mutations in PLXND1 and REV3L are confirmed to cause MBS. Here, we fine mapped the breakpoints of a complex chromosomal rearrangement (CCR) 46,XY,t(7;8;11;13) in a patient with MBS, which revealed 41 clustered breakpoints with typical hallmarks of chromothripsis. Among 12 truncated protein-coding genes, SEMA3A is known to bind to the MBS-associated PLXND1. Intriguingly, the CCR also truncated PIK3CG, which in silico interacts with REVL3 encoded by the other known MBS-gene REV3L, and with the SEMA3A/PLXND1 complex via FLT1. Additional studies of other complex rearrangements may reveal whether the multiple breakpoints in germline chromothripsis may predispose to complex multigenic disorders.
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http://dx.doi.org/10.1002/humu.23775DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6688968PMC
August 2019

Haploinsufficiency of ARHGAP42 is associated with hypertension.

Eur J Hum Genet 2019 08 21;27(8):1296-1303. Epub 2019 Mar 21.

Department of Cellular and Molecular Medicine, University of Copenhagen, Copenhagen N, 2200, Denmark.

Family studies have established that the heritability of blood pressure is significant and genome-wide association studies (GWAS) have identified numerous susceptibility loci, including one within the non-coding part of Rho GTPase-activating protein 42 gene (ARHGAP42) on chromosome 11q22.1. Arhgap42-deficient mice have significantly elevated blood pressure, but the phenotypic effects of human variants in the coding part of the gene are unknown. In a Danish cohort of carriers with apparently balanced chromosomal rearrangements, we identified a family where a reciprocal translocation t(11;18)(q22.1;q12.2) segregated with hypertension and obesity. Clinical re-examination revealed that four carriers (age 50-77 years) have had hypertension for several years along with an increased body mass index (34-43 kg/m). A younger carrier (age 23 years) had normal blood pressure and body mass index. Mapping of the chromosomal breakpoints with mate-pair and Sanger sequencing revealed truncation of ARHGAP42. A decreased expression level of ARHGAP42 mRNA in the blood was found in the translocation carriers relative to controls and allele-specific expression analysis showed monoallelic expression in the translocation carriers, confirming that the truncated allele of ARHGAP42 was not expressed. These findings support that haploinsufficiency of ARHGAP42 leads to an age-dependent hypertension. The other breakpoint truncated a regulatory domain of the CUGBP Elav-like family member 4 (CELF4) gene on chromosome 18q12.2 that harbours several GWAS signals for obesity. We thereby provide additional support for an obesity locus in the CELF4 domain.
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http://dx.doi.org/10.1038/s41431-019-0382-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6777610PMC
August 2019

A novel in-frame mutation in leads to Juvenile neuronal ceroid lipofuscinosis in a large Pakistani family.

Int J Neurosci 2019 Sep 20;129(9):890-895. Epub 2019 Mar 20.

a Human Molecular Genetics Laboratory , National Institute for Biotechnology and Genetic Engineering (NIBGE)-PIEAS , Faisalabad , Pakistan.

Neuronal ceroid lipofuscinosis (NCLs) are the most common neurodegenerative disorders, with global incidence of 1 in 100,000 live births. NCLs affect central nervous system, primarily cerebellar and cerebral cortices. Juvenile neuronal ceroid lipofuscinosis (JNCL), also known as Batten disease, is the most common form of NCLs. JNCL is primarily caused by pathogenic mutations in gene, which encodes a transporter transmembrane protein of uncertain function. The 1.02 kb deletion is the most common mutation in that results in frame shift and a premature termination leading to nonfunctional protein. Here, we invetigated a large consanguineous family consisting of four affected individuals with clincal symptoms suggestive of Juvenile neuronal ceroid lipofuscinosis. We conducted clinial and radilogical investigation of the family and performed NGS based Gene Panel sequencing comprising of five hundred and forty five candidate genes to characterize it at genetic level. We identified a novel homozygous c.181_183delGAC mutation in the gene seggregating witht the disorder in the family. The mutation induces in-frame deletion, deleting one amino acid (p.Asp61del) in CLN3 protein. The deleted amino acid aspartic acid plays an important role as general acid in enzymes active centers as well as in maintaining the ionic character of proteins. Our finding adds to genetic variability of Juvenile neuronal ceroid lipofuscinosis associated with gene and a predicted CLN3 protein interacting domain site.
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http://dx.doi.org/10.1080/00207454.2019.1586686DOI Listing
September 2019

A splice-site variant in the lncRNA gene cosegregates in the large Volkmann cataract family.

Mol Vis 2019 20;25:1-11. Epub 2019 Jan 20.

Copenhagen Center for Glycomics, Department of Cellular and Molecular Medicine, Panum Institute, University of Copenhagen, Copenhagen N, Denmark.

Purpose: To identify the mutation for Volkmann cataract () at 1p36.33.

Methods: The genes in the candidate region 1p36.33 were Sanger and parallel deep sequenced, and informative single nucleotide polymorphisms (SNPs) were identified for linkage analysis. Expression analysis with reverse transcription polymerase chain reaction (RT-PCR) of the candidate gene was performed using RNA from different human tissues. Quantitative transcription polymerase chain reaction (qRT-PCR) analysis of the gene was performed in affected and healthy individuals. Bioinformatic analysis of the linkage regions including the candidate gene was performed.

Results: Linkage analysis of the 1p36.33 CCV locus applying new marker systems obtained with Sanger and deep sequencing reduced the candidate locus from 2.1 Mb to 0.389 Mb flanked by the markers STS-22AC and rs549772338 and resulted in an logarithm of the odds (LOD) score of Z = 21.67. The identified mutation, rs763295804, affects the donor splice site in the long non-coding RNA gene (ENSG00000231050). The gene including splice-site junctions is conserved in primates but not in other mammalian genomes, and two alternative transcripts were shown with RT-PCR. One of these transcripts represented a lens cell-specific transcript. Meta-analysis of the Cross-Linking-Immuno-Precipitation sequencing (CLIP-Seq) data suggested the RNA binding protein (RBP) eIF4AIII is an active counterpart for , and several miRNA and transcription factors binding sites were predicted in the proximity of the mutation. ENCODE DNase I hypersensitivity and histone methylation and acetylation data suggest the genomic region may have regulatory functions.

Conclusions: The mutation in suggests the long non-coding RNA as the candidate cataract gene associated with the autosomal dominant inherited congenital cataract from CCV. The mutation has the potential to destroy exon/intron splicing of both transcripts of . Sanger and massive deep resequencing of the linkage region failed to identify alternative candidates suggesting the mutation in is causative for the CCV phenotype.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6377377PMC
June 2019

Replicative and non-replicative mechanisms in the formation of clustered CNVs are indicated by whole genome characterization.

PLoS Genet 2018 11 12;14(11):e1007780. Epub 2018 Nov 12.

Department of Molecular Medicine and Surgery, Center for Molecular Medicine, Karolinska Institute, Stockholm, Sweden.

Clustered copy number variants (CNVs) as detected by chromosomal microarray analysis (CMA) are often reported as germline chromothripsis. However, such cases might need further investigations by massive parallel whole genome sequencing (WGS) in order to accurately define the underlying complex rearrangement, predict the occurrence mechanisms and identify additional complexities. Here, we utilized WGS to delineate the rearrangement structure of 21 clustered CNV carriers first investigated by CMA and identified a total of 83 breakpoint junctions (BPJs). The rearrangements were further sub-classified depending on the patterns observed: I) Cases with only deletions (n = 8) often had additional structural rearrangements, such as insertions and inversions typical to chromothripsis; II) cases with only duplications (n = 7) or III) combinations of deletions and duplications (n = 6) demonstrated mostly interspersed duplications and BPJs enriched with microhomology. In two cases the rearrangement mutational signatures indicated both a breakage-fusion-bridge cycle process and haltered formation of a ring chromosome. Finally, we observed two cases with Alu- and LINE-mediated rearrangements as well as two unrelated individuals with seemingly identical clustered CNVs on 2p25.3, possibly a rare European founder rearrangement. In conclusion, through detailed characterization of the derivative chromosomes we show that multiple mechanisms are likely involved in the formation of clustered CNVs and add further evidence for chromoanagenesis mechanisms in both "simple" and highly complex chromosomal rearrangements. Finally, WGS characterization adds positional information, important for a correct clinical interpretation and deciphering mechanisms involved in the formation of these rearrangements.
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http://dx.doi.org/10.1371/journal.pgen.1007780DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6258378PMC
November 2018

Small supernumerary marker chromosomes: A legacy of trisomy rescue?

Hum Mutat 2019 02 22;40(2):193-200. Epub 2018 Nov 22.

Department of Molecular Medicine, University of Pavia, Pavia, Italy.

We studied by a whole genomic approach and trios genotyping, 12 de novo, nonrecurrent small supernumerary marker chromosomes (sSMC), detected as mosaics during pre- or postnatal diagnosis and associated with increased maternal age. Four sSMCs contained pericentromeric portions only, whereas eight had additional non-contiguous portions of the same chromosome, assembled together in a disordered fashion by repair-based mechanisms in a chromothriptic event. Maternal hetero/isodisomy was detected with a paternal origin of the sSMC in some cases, whereas in others two maternal alleles in the sSMC region and biparental haplotypes of the homologs were detected. In other cases, the homologs were biparental while the sSMC had the same haplotype of the maternally inherited chromosome. These findings strongly suggest that most sSMCs are the result of a multiple-step mechanism, initiated by maternal meiotic nondisjunction followed by postzygotic anaphase lagging of the supernumerary chromosome and its subsequent chromothripsis.
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http://dx.doi.org/10.1002/humu.23683DOI Listing
February 2019

Position effect, cryptic complexity, and direct gene disruption as disease mechanisms in de novo apparently balanced translocation cases.

PLoS One 2018 5;13(10):e0205298. Epub 2018 Oct 5.

Department of Cytogenetics and Genomics, The Cyprus Institute of Neurology and Genetics, Nicosia, Cyprus.

The majority of apparently balanced translocation (ABT) carriers are phenotypically normal. However, several mechanisms were proposed to underlie phenotypes in affected ABT cases. In the current study, whole-genome mate-pair sequencing (WG-MPS) followed by Sanger sequencing was applied to further characterize de novo ABTs in three affected individuals. WG-MPS precisely mapped all ABT breakpoints and revealed three possible underlying molecular mechanisms. Firstly, in a t(X;1) carrier with hearing loss, a highly skewed X-inactivation pattern was observed and the der(X) breakpoint mapped ~87kb upstream an X-linked deafness gene namely POU3F4, thus suggesting an underlying long-range position effect mechanism. Secondly, cryptic complexity and a chromothripsis rearrangement was identified in a t(6;7;8;12) carrier with intellectual disability. Two translocations and a heterozygous deletion disrupted SOX5; a dominant nervous system development gene previously reported in similar patients. Finally, a direct gene disruption mechanism was proposed in a t(4;9) carrier with dysmorphic facial features and speech delay. In this case, the der(9) breakpoint directly disrupted NFIB, a gene involved in lung maturation and development of the pons with important functions in main speech processes. To conclude, in contrast to familial ABT cases with identical rearrangements and discordant phenotypes, where translocations are considered coincidental, translocations seem to be associated with phenotype presentation in affected de novo ABT cases. In addition, this study highlights the importance of investigating both coding and non-coding regions to decipher the underlying pathogenic mechanisms in these patients, and supports the potential introduction of low coverage WG-MPS in the clinical investigation of de novo ABTs.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0205298PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6173455PMC
March 2019

De novo unbalanced translocations have a complex history/aetiology.

Hum Genet 2018 Oct 1;137(10):817-829. Epub 2018 Oct 1.

Department of Molecular Medicine, University of Pavia, Pavia, Italy.

We investigated 52 cases of de novo unbalanced translocations, consisting in a terminally deleted or inverted-duplicated deleted (inv-dup del) 46th chromosome to which the distal portion of another chromosome or its opposite end was transposed. Array CGH, whole-genome sequencing, qPCR, FISH, and trio genotyping were applied. A biparental origin of the deletion and duplication was detected in 6 cases, whereas in 46, both imbalances have the same parental origin. Moreover, the duplicated region was of maternal origin in more than half of the cases, with 25% of them showing two maternal and one paternal haplotype. In all these cases, maternal age was increased. These findings indicate that the primary driver for the occurrence of the de novo unbalanced translocations is a maternal meiotic non-disjunction, followed by partial trisomy rescue of the supernumerary chromosome present in the trisomic zygote. In contrast, asymmetric breakage of a dicentric chromosome, originated either at the meiosis or postzygotically, in which the two resulting chromosomes, one being deleted and the other one inv-dup del, are repaired by telomere capture, appears at the basis of all inv-dup del translocations. Notably, this mechanism also fits with the origin of some simple translocations in which the duplicated region was of paternal origin. In all cases, the signature at the translocation junctions was that of non-homologous end joining (NHEJ) rather than non-allelic homologous recombination (NAHR). Our data imply that there is no risk of recurrence in the following pregnancies for any of the de novo unbalanced translocations we discuss here.
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http://dx.doi.org/10.1007/s00439-018-1941-9DOI Listing
October 2018

Challenges for the Sustainability of University-Run Biobanks.

Biopreserv Biobank 2018 Aug 17;16(4):312-321. Epub 2018 Jul 17.

5 Center for Information and Innovation Law (CIIR), University of Copenhagen , Copenhagen, Denmark .

Most university biobanks begin like other university research projects, that is, with an idea conceived by an individual researcher in pursuit of his/her own research interests, publications, funding, and career. Some biobanks, however, come to have scientific value that goes beyond the projects that were initially responsible for the collection of the samples and data they contain. Such value may derive from among other things the uniqueness of the samples in terms of their sheer volume, the quality of the samples, the ability to link the samples with information retrieved in disease registries, or the fact that the samples represent very rare diseases. This article focuses on biobanks of this kind, and the special obligations that publicly funded universities have to ensure the sustainability of biobanks with continued scientific value. We argue that universities should adopt policies to deal with the various, diverse issues which may arise during the lifecycle of a biobank. The policies should be flexible, accommodate the freedoms of individual researchers, and reflect the multifaceted nature of biobanks. Yet they should be specific enough to provide guidance and robust enough to safeguard legal norms and ethical values. The article sets out concrete recommendations which universities should consider and act upon.
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http://dx.doi.org/10.1089/bio.2018.0054DOI Listing
August 2018

Cryptic breakpoint identified by whole-genome mate-pair sequencing in a rare paternally inherited complex chromosomal rearrangement.

Mol Cytogenet 2018 7;11:34. Epub 2018 Jun 7.

1Department of Cytogenetics and Genomics, The Cyprus Institute of Neurology and Genetics, Nicosia, Cyprus.

Background: Precise characterization of apparently balanced complex chromosomal rearrangements in non-affected individuals is crucial as they may result in reproductive failure, recurrent miscarriages or affected offspring.

Case Presentation: We present a family, where the non-affected father and daughter were found, using FISH and karyotyping, to be carriers of a three-way complex chromosomal rearrangement [t(6;7;10)(q16.2;q34;q26.1), de novo in the father]. The family suffered from two stillbirths, one miscarriage, and has a son with severe intellectual disability. In the present study, the family was revisited using whole-genome mate-pair sequencing. Interestingly, whole-genome mate-pair sequencing revealed a cryptic breakpoint on derivative (der) chromosome 6 rendering the rearrangement even more complex. FISH using a chromosome (chr) 6 custom-designed probe and a chr10 control probe confirmed that the interstitial chr6 segment, created by the two chr6 breakpoints, was translocated onto der(10). Breakpoints were successfully validated with Sanger sequencing, and small imbalances as well as microhomology were identified. Finally, the complex chromosomal rearrangement breakpoints disrupted the , , , and genes without causing any phenotype development.

Conclusions: In contrast to the majority of maternally transmitted complex chromosomal rearrangement cases, our study investigated a rare case where a complex chromosomal rearrangement, which most probably resulted from a Type IV hexavalent during the pachytene stage of meiosis I, was stably transmitted from a fertile father to his non-affected daughter. Whole-genome mate-pair sequencing proved highly successful in identifying cryptic complexity, which consequently provided further insight into the meiotic segregation of chromosomes and the increased reproductive risk in individuals carrying the specific complex chromosomal rearrangement. We propose that such complex rearrangements should be characterized in detail using a combination of conventional cytogenetic and NGS-based approaches to aid in better prenatal preimplantation genetic diagnosis and counseling in couples with reproductive problems.
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http://dx.doi.org/10.1186/s13039-018-0384-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5991433PMC
June 2018

Risks and Recommendations in Prenatally Detected De Novo Balanced Chromosomal Rearrangements from Assessment of Long-Term Outcomes.

Am J Hum Genet 2018 06 24;102(6):1090-1103. Epub 2018 May 24.

Wilhelm Johannsen Centre for Functional Genome Research, Department of Cellular and Molecular Medicine, University of Copenhagen, 2200 Copenhagen N, Denmark; Department of Clinical Genetics, Copenhagen University Hospital, Rigshospitalet, 2100 Copenhagen O, Denmark. Electronic address:

The 6%-9% risk of an untoward outcome previously established by Warburton for prenatally detected de novo balanced chromosomal rearrangements (BCRs) does not account for long-term morbidity. We performed long-term follow-up (mean 17 years) of a registry-based nationwide cohort of 41 individuals carrying a prenatally detected de novo BCR with normal first trimester screening/ultrasound scan. We observed a significantly higher frequency of neurodevelopmental and/or neuropsychiatric disorders than in a matched control group (19.5% versus 8.3%, p = 0.04), which was increased to 26.8% upon clinical follow-up. Chromosomal microarray of 32 carriers revealed no pathogenic imbalances, illustrating a low prognostic value when fetal ultrasound scan is normal. In contrast, mate-pair sequencing revealed disrupted genes (ARID1B, NPAS3, CELF4), regulatory domains of known developmental genes (ZEB2, HOXC), and complex BCRs associated with adverse outcomes. Seven unmappable autosomal-autosomal BCRs with breakpoints involving pericentromeric/heterochromatic regions may represent a low-risk group. We performed independent phenotype-aware and blinded interpretation, which accurately predicted benign outcomes (specificity = 100%) but demonstrated relatively low sensitivity for prediction of the clinical outcome in affected carriers (sensitivity = 45%-55%). This sensitivity emphasizes the challenges associated with prenatal risk prediction for long-term morbidity in the absence of phenotypic data given the still immature annotation of the morbidity genome and poorly understood long-range regulatory mechanisms. In conclusion, we upwardly revise the previous estimates of Warburton to a morbidity risk of 27% and recommend sequencing of the chromosomal breakpoints as the first-tier diagnostic test in pregnancies with a de novo BCR.
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http://dx.doi.org/10.1016/j.ajhg.2018.04.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5992120PMC
June 2018

Congenital olfactory impairment is linked to cortical changes in prefrontal and limbic brain regions.

Brain Imaging Behav 2018 Dec;12(6):1569-1582

Danish Research Centre for Magnetic Resonance, Centre for Functional and Diagnostic Imaging and Research, Copenhagen University Hospital Hvidovre, Hvidovre, Denmark.

The human sense of smell is closely associated with morphological differences of the fronto-limbic system, specifically the piriform cortex and medial orbitofrontal cortex (mOFC). Still it is unclear whether cortical volume in the core olfactory areas and connected brain regions are shaped differently in individuals who suffer from lifelong olfactory deprivation relative to healthy normosmic individuals. To address this question, we examined if regional variations in gray matter volume were associated with smell ability in seventeen individuals with isolated congenital olfactory impairment (COI) matched with sixteen normosmic controls. All subjects underwent whole-brain magnetic resonance imaging, and voxel-based morphometry was used to estimate regional variations in grey matter volume. The analyses showed that relative to controls, COI subjects had significantly larger grey matter volumes in left middle frontal gyrus and right superior frontal sulcus (SFS). COI subjects with severe olfactory impairment (anosmia) had reduced grey matter volume in the left mOFC and increased volume in right piriform cortex and SFS. Within the COI group olfactory ability, measured with the "Sniffin' Sticks" test, was positively associated with larger grey matter volume in right posterior cingulate and parahippocampal cortices whereas the opposite relationship was observed in controls. Across COI subjects and controls, better olfactory detection threshold was associated with smaller volume in right piriform cortex, while olfactory identification was negatively associated with right SFS volume. Our findings suggest that lifelong olfactory deprivation trigger changes in the cortical volume of prefrontal and limbic brain regions previously linked to olfactory memory.
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http://dx.doi.org/10.1007/s11682-017-9817-5DOI Listing
December 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

Regulatory variants of FOXG1 in the context of its topological domain organisation.

Eur J Hum Genet 2018 02 30;26(2):186-196. Epub 2017 Dec 30.

Wilhelm Johannsen Centre for Functional Genome Research, Department of Cellular and Molecular Medicine, University of Copenhagen, Copenhagen, Denmark.

FOXG1 syndrome is caused by FOXG1 intragenic point mutations, or by long-range position effects (LRPE) of intergenic structural variants. However, the size of the FOXG1 regulatory landscape is uncertain, because the associated topologically associating domain (TAD) in fibroblasts is split into two domains in embryonic stem cells (hESC). Indeed, it has been suggested that the pathogenetic mechanism of deletions that remove the stem-cell-specific TAD boundary may be enhancer adoption due to ectopic activity of enhancer(s) located in the distal hESC-TAD. Herein we map three de novo translocation breakpoints to the proximal regulatory domain of FOXG1. The classical FOXG1 syndrome in these and in other translocation patients, and in a patient with an intergenic deletion that removes the hESC-specific TAD boundary, do not support the hypothesised enhancer adoption as a main contributor to the FOXG1 syndrome. Also, virtual 4 C and HiC-interaction data suggest that the hESC-specific TAD boundary may not be critical for FOXG1 regulation in a majority of human cells and tissues, including brain tissues and a neuronal progenitor cell line. Our data support the importance of a critical regulatory region (SRO) proximal to the hESC-specific TAD boundary. We further narrow this critical region by a deletion distal to the hESC-specific boundary, associated with a milder clinical phenotype. The distance from FOXG1 to the SRO ( > 500 kb) highlight a limitation of ENCODE DNase hypersensitivity data for functional prediction of LRPE. Moreover, the SRO has little overlap with a cluster of frequently associating regions (FIREs) located in the proximal hESC-TAD.
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http://dx.doi.org/10.1038/s41431-017-0011-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5839045PMC
February 2018

Enrichment of megabase-sized DNA molecules for single-molecule optical mapping and next-generation sequencing.

Sci Rep 2017 12 20;7(1):17893. Epub 2017 Dec 20.

Department of Cellular and Molecular Medicine, Faculty of Health and Medical Sciences, University of Copenhagen, Nørre Alle 14, Copenhagen, 2200, Denmark.

Next-generation sequencing (NGS) has caused a revolution, yet left a gap: long-range genetic information from native, non-amplified DNA fragments is unavailable. It might be obtained by optical mapping of megabase-sized DNA molecules. Frequently only a specific genomic region is of interest, so here we introduce a method for selection and enrichment of megabase-sized DNA molecules intended for single-molecule optical mapping: DNA from a human cell line is digested by the NotI rare-cutting enzyme and size-selected by pulsed-field gel electrophoresis. For demonstration, more than 600 sub-megabase- to megabase-sized DNA molecules were recovered from the gel and analysed by denaturation-renaturation optical mapping. Size-selected molecules from the same gel were sequenced by NGS. The optically mapped molecules and the NGS reads showed enrichment from regions defined by NotI restriction sites. We demonstrate that the unannotated genome can be characterized in a locus-specific manner via molecules partially overlapping with the annotated genome. The method is a promising tool for investigation of structural variants in enriched human genomic regions for both research and diagnostic purposes. Our enrichment method could potentially work with other genomes or target specified regions by applying other genomic editing tools, such as the CRISPR/Cas9 system.
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http://dx.doi.org/10.1038/s41598-017-18091-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5738345PMC
December 2017

Breakpoint mapping and haplotype analysis of translocation t(1;12)(q43;q21.1) in two apparently independent families with vascular phenotypes.

Mol Genet Genomic Med 2018 01 23;6(1):56-68. Epub 2017 Nov 23.

Department of Health, National Institute for Health and Welfare, Helsinki, Finland.

Background: The risk of serious congenital anomaly for de novo balanced translocations is estimated to be at least 6%. We identified two apparently independent families with a balanced t(1;12)(q43;q21.1) as an outcome of a "Systematic Survey of Balanced Chromosomal Rearrangements in Finns." In the first family, carriers (n = 6) manifest with learning problems in childhood, and later with unexplained neurological symptoms (chronic headache, balance problems, tremor, fatigue) and cerebral infarctions in their 50s. In the second family, two carriers suffer from tetralogy of Fallot, one from transient ischemic attack and one from migraine. The translocation cosegregates with these vascular phenotypes and neurological symptoms.

Methods And Results: We narrowed down the breakpoint regions using mate pair sequencing. We observed conserved haplotypes around the breakpoints, pointing out that this translocation has arisen only once. The chromosome 1 breakpoint truncates a CHRM3 processed transcript, and is flanked by the 5' end of CHRM3 and the 3' end of RYR2. TRHDE, KCNC2, and ATXN7L3B flank the chromosome 12 breakpoint.

Conclusions: This study demonstrates a balanced t(1;12)(q43;q21.1) with conserved haplotypes on the derived chromosomes. The translocation seems to result in vascular phenotype, with or without neurological symptoms, in at least two families. We suggest that the translocation influences the positional expression of CHRM3, RYR2, TRHDE, KCNC2, and/or ATXN7L3B.
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http://dx.doi.org/10.1002/mgg3.346DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5823676PMC
January 2018

Genome-Wide Supported Risk Variants in , , , , and Contribute to Schizophrenia Susceptibility in Pakistani Population.

Psychiatry Investig 2017 Sep 11;14(5):687-692. Epub 2017 Sep 11.

Human Molecular Genetics Laboratory, Health Biotechnology Division, National Institute for Biotechnology and Genetic Engineering (NIBGE), PIEAS, Faisalabad, Pakistan.

Objective: Schizophrenia is a chronic neuropsychiatric disease afflicting around 1.1% of the population worldwide. Recently, , , , , and have been reported as the most robustly emerging candidates involved in the etiology of schizophrenia. In this case control study, we performed an association analysis of rs1625579 (), rs1006737, rs4765905 (), rs10503253 (), rs1076560 (), rs12704290, rs6465084, and rs148754219 () in Pakistani population.

Methods: Schizophrenia was diagnosed on the basis of the Diagnostic and Statistical Manual of Mental Disorders 4th ed (DSM-IV). Detailed clinical information, family history of all patients and healthy controls were collected. RFLP based case control association study was performed in a Pakistani cohort of 508 schizophrenia patients and 300 healthy control subjects. Alleles and genotype frequencies were calculated using SPSS.

Results: A significant difference in the genotype and allele frequencies for rs4765905, rs1076560 and rs6465084 were found between the patients and controls (p=0.000).

Conclusion: This study provides substantial evidence supporting the role of , and as schizophrenia susceptibility genes in Pakistani population.
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http://dx.doi.org/10.4306/pi.2017.14.5.687DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5639139PMC
September 2017

Homozygous mutation in the NPHP3 gene causing foetal nephronophthisis.

Nephrology (Carlton) 2017 Oct;22(10):818-820

Human Molecular Genetics Laboratory, Health Biotechnology Division, National Institute for Biotechnology and Genetic Engineering (NIBGE), PIEAS, Faisalabad, Pakistan.

We present a case of a foetal sonographic finding of hyper-echogenic kidneys, which led to a strategic series of genetic tests and identified a homozygous mutation (c.424C > T, p. R142*) in the NPHP3 gene. Our study provides a rare presentation of NPHP3-related ciliopathy and adds to the mutation spectrum of the gene, being the first one from Pakistani population. With a thorough literature review, it also advocates for molecular assessment of ciliopathies to improve risk estimate for future pregnancies, and identify predisposed asymptomatic carriers.
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http://dx.doi.org/10.1111/nep.13097DOI Listing
October 2017

A novel mutation in CDK5RAP2 gene causes primary microcephaly with speech impairment and sparse eyebrows in a consanguineous Pakistani family.

Eur J Med Genet 2017 Dec 2;60(12):627-630. Epub 2017 Aug 2.

Human Molecular Genetics Laboratory, National Institute for Biotechnology and Genetic Engineering (NIBGE)-PIEAS, Faisalabad, Pakistan. Electronic address:

CDK5RAP2 gene encodes a centrosomal protein, highly expressed in fetal brain and essentially indispensable for its normal development, as biallelic mutations in it lead to primary microcephaly (MCPH). Despite being known as MCPH linked gene for more than a decade, the phenotypic spectrum of CDK5RAP2 mutations is still under explored as only eleven families have been reported worldwide. Here, we analyzed a consanguineous Pakistani MCPH family, characterized by moderate to severe intellectual disability, speech impairment, moderately short stature and sparse eyebrows. Whole exome sequencing of the proband identified a 2bp duplication in exon 34 of CDK5RAP2 that causes frame-shift, leading to a premature stop codon. The resultant transcript is resistant to nonsense mediated decay, suggesting that the mutation leads to a truncated protein lacking C-terminal domains; CDK5R1, and Cnn motif 2 (CM2), required for its localization to centrosome and Golgi Apparatus. Clinical variability observed in the family highlights the importance of further detailed clinical description of patients with CDK5RAP2 mutations.
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http://dx.doi.org/10.1016/j.ejmg.2017.07.017DOI Listing
December 2017

The identification and functional annotation of RNA structures conserved in vertebrates.

Genome Res 2017 08 9;27(8):1371-1383. Epub 2017 May 9.

Center for non-coding RNA in Technology and Health (RTH), University of Copenhagen, DK-1870 Frederiksberg, Denmark.

Structured elements of RNA molecules are essential in, e.g., RNA stabilization, localization, and protein interaction, and their conservation across species suggests a common functional role. We computationally screened vertebrate genomes for conserved RNA structures (CRSs), leveraging structure-based, rather than sequence-based, alignments. After careful correction for sequence identity and GC content, we predict ∼516,000 human genomic regions containing CRSs. We find that a substantial fraction of human-mouse CRS regions (1) colocalize consistently with binding sites of the same RNA binding proteins (RBPs) or (2) are transcribed in corresponding tissues. Additionally, a CaptureSeq experiment revealed expression of many of our CRS regions in human fetal brain, including 662 novel ones. For selected human and mouse candidate pairs, qRT-PCR and in vitro RNA structure probing supported both shared expression and shared structure despite low abundance and low sequence identity. About 30,000 CRS regions are located near coding or long noncoding RNA genes or within enhancers. Structured (CRS overlapping) enhancer RNAs and extended 3' ends have significantly increased expression levels over their nonstructured counterparts. Our findings of transcribed uncharacterized regulatory regions that contain CRSs support their RNA-mediated functionality.
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http://dx.doi.org/10.1101/gr.208652.116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5538553PMC
August 2017

Abdominal Wall Defects in Greenland 1989-2015.

Birth Defects Res 2017 Jul 2;109(11):836-842. Epub 2017 May 2.

Department of Cellular and Molecular Medicine, Panum Institute, University of Copenhagen, Copenhagen, Denmark.

Background: In the last decades, an increasing rate of gastroschisis but not of omphalocele has been reported worldwide. Greenland is the world's largest island, but 80% is covered by an ice cap, it has a small population of around 56,000 peoples (as of 2016). The occurrence of abdominal wall defects has never been investigated in Greenland.

Methods: The present study is based on data retrieved from three nationwide and two local registries in the Greenlandic health care system over 27 years (1989-2015).

Results: We identified 33 infants with abdominal wall defects born in the study time period. All cases were reclassified to 28 cases of gastroschisis, four cases of omphalocele, and there was 1 infant in the indeterminate group. The point prevalence at birth for gastroschisis increased significantly from 8 to 35 (average 10.7) per 10,000 liveborn and -stillborn infants. Mothers below 20 years of age represented 23% of all cases and the prevalence for this group was 17 per 10,000 liveborn and stillborn. Perinatal mortality for infants with gastroschisis was high (18%), and 1 year survival was 71%. For omphalocele, the prevalence varied from 8 to 11 per 10,000 liveborn and stillborn infants. There was no increasing rate in the period, further highlighting an etiological difference between gastroschisis and omphalocele.

Conclusion: This study confirms the increasing prevalence of gastroschisis in Greenland in the period from 1989 to 2015. The average was 10.7 per 10,000 liveborn and -stillborn infants and, to the best of our knowledge, this is the highest prevalence ever reported. Birth Defects Research 109:836-842, 2017. © 2017 Wiley Periodicals, Inc.
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http://dx.doi.org/10.1002/bdr2.1025DOI Listing
July 2017

Accurate Breakpoint Mapping in Apparently Balanced Translocation Families with Discordant Phenotypes Using Whole Genome Mate-Pair Sequencing.

PLoS One 2017 10;12(1):e0169935. Epub 2017 Jan 10.

Department of Cytogenetics and Genomics, The Cyprus Institute of Neurology and Genetics, Nicosia, Cyprus.

Familial apparently balanced translocations (ABTs) segregating with discordant phenotypes are extremely challenging for interpretation and counseling due to the scarcity of publications and lack of routine techniques for quick investigation. Recently, next generation sequencing has emerged as an efficacious methodology for precise detection of translocation breakpoints. However, studies so far have mainly focused on de novo translocations. The present study focuses specifically on familial cases in order to shed some light to this diagnostic dilemma. Whole-genome mate-pair sequencing (WG-MPS) was applied to map the breakpoints in nine two-way ABT carriers from four families. Translocation breakpoints and patient-specific structural variants were validated by Sanger sequencing and quantitative Real Time PCR, respectively. Identical sequencing patterns and breakpoints were identified in affected and non-affected members carrying the same translocations. PTCD1, ATP5J2-PTCD1, CADPS2, and STPG1 were disrupted by the translocations in three families, rendering them initially as possible disease candidate genes. However, subsequent mutation screening and structural variant analysis did not reveal any pathogenic mutations or unique variants in the affected individuals that could explain the phenotypic differences between carriers of the same translocations. In conclusion, we suggest that NGS-based methods, such as WG-MPS, can be successfully used for detailed mapping of translocation breakpoints, which can also be used in routine clinical investigation of ABT cases. Unlike de novo translocations, no associations were determined here between familial two-way ABTs and the phenotype of the affected members, in which the presence of cryptic imbalances and complex chromosomal rearrangements has been excluded. Future whole-exome or whole-genome sequencing will potentially reveal unidentified mutations in the patients underlying the discordant phenotypes within each family. In addition, larger studies are needed to determine the exact percentage for phenotypic risk in families with ABTs.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0169935PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5225008PMC
August 2017
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