New mutations have long been known to cause genetic disease, but their true contribution to the disease burden can only now be determined using family-based whole-genome or whole-exome sequencing ...approaches. In this Review we discuss recent findings suggesting that de novo mutations play a prominent part in rare and common forms of neurodevelopmental diseases, including intellectual disability, autism and schizophrenia. De novo mutations provide a mechanism by which early-onset reproductively lethal diseases remain frequent in the population. These mutations, although individually rare, may capture a significant part of the heritability for complex genetic diseases that is not detectable by genome-wide association studies.
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DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Abstract
Aims
Truncating titin variants (TTNtv) are the most prevalent genetic cause of dilated cardiomyopathy (DCM). We aim to study clinical parameters and long-term outcomes related to the TTNtv ...genotype and determine the related molecular changes at tissue level in TTNtv DCM patients.
Methods and results
A total of 303 consecutive and extensively phenotyped DCM patients (including cardiac imaging, Holter monitoring, and endomyocardial biopsy) underwent DNA sequencing of 47 cardiomyopathy-associated genes including TTN, yielding 38 TTNtv positive (13%) patients. At long-term follow-up (median of 45 months, up to 12 years), TTNtv DCM patients had increased ventricular arrhythmias compared to other DCM, but a similar survival. Arrhythmias are especially prominent in TTNtv patients with an additional environmental trigger (i.e. virus infection, cardiac inflammation, systemic disease, toxic exposure). Importantly, cardiac mass is reduced in TTNtv patients, despite similar cardiac function and dimensions at cardiac magnetic resonance. These enhanced life-threatening arrhythmias and decreased cardiac mass in TTNtv DCM patients go along with significant cardiac energetic and matrix alterations. All components of the mitochondrial electron transport chain are significantly upregulated in TTNtv hearts at RNA-sequencing. Also, interstitial fibrosis was augmented in TTNtv patients at histological and transcript level.
Conclusion
Truncating titin variants lead to pronounced cardiac alterations in mitochondrial function, with increased interstitial fibrosis and reduced hypertrophy. Those structural and metabolic alterations in TTNtv hearts go along with increased ventricular arrhythmias at long-term follow-up, with a similar survival and overall cardiac function.
Next generation sequencing can be used to search for Mendelian disease genes in an unbiased manner by sequencing the entire protein-coding sequence, known as the exome, or even the entire human ...genome. Identifying the pathogenic mutation amongst thousands to millions of genomic variants is a major challenge, and novel variant prioritization strategies are required. The choice of these strategies depends on the availability of well-phenotyped patients and family members, the mode of inheritance, the severity of the disease and its population frequency. In this review, we discuss the current strategies for Mendelian disease gene identification by exome resequencing. We conclude that exome strategies are successful and identify new Mendelian disease genes in approximately 60% of the projects. Improvements in bioinformatics as well as in sequencing technology will likely increase the success rate even further. Exome sequencing is likely to become the most commonly used tool for Mendelian disease gene identification for the coming years.
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DOBA, EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, IZUM, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, SIK, UILJ, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Exome sequencing is revolutionizing Mendelian disease gene identification. This results in improved clinical diagnosis, more accurate genotype-phenotype correlations and new insights into the role of ...rare genomic variation in disease.
Implementation of novel genetic diagnostic tests is generally driven by technological advances because they promise shorter turnaround times and/or higher diagnostic yields. Other aspects, including ...impact on clinical management or cost-effectiveness, are often not assessed in detail prior to implementation.
We studied the clinical utility of whole-exome sequencing (WES) in complex pediatric neurology in terms of diagnostic yield and costs. We analyzed 150 patients (and their parents) presenting with complex neurological disorders of suspected genetic origin. In a parallel study, all patients received both the standard diagnostic workup (e.g., cerebral imaging, muscle biopsies or lumbar punctures, and sequential gene-by-gene-based testing) and WES simultaneously.
Our unique study design allowed direct comparison of diagnostic yield of both trajectories and provided insight into the economic implications of implementing WES in this diagnostic trajectory. We showed that WES identified significantly more conclusive diagnoses (29.3%) than the standard care pathway (7.3%) without incurring higher costs. Exploratory analysis of WES as a first-tier diagnostic test indicates that WES may even be cost-saving, depending on the extent of other tests being omitted.
Our data support such a use of WES in pediatric neurology for disorders of presumed genetic origin.Genet Med advance online publication 23 March 2017.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Loeys-Dietz syndrome (LDS) associates with a tissue signature for high transforming growth factor (TGF)-β signaling but is often caused by heterozygous mutations in genes encoding positive effectors ...of TGF-β signaling, including either subunit of the TGF-β receptor or SMAD3, thereby engendering controversy regarding the mechanism of disease. Here, we report heterozygous mutations or deletions in the gene encoding the TGF-β2 ligand for a phenotype within the LDS spectrum and show upregulation of TGF-β signaling in aortic tissue from affected individuals. Furthermore, haploinsufficient Tgfb2(+/-) mice have aortic root aneurysm and biochemical evidence of increased canonical and noncanonical TGF-β signaling. Mice that harbor both a mutant Marfan syndrome (MFS) allele (Fbn1(C1039G/+)) and Tgfb2 haploinsufficiency show increased TGF-β signaling and phenotypic worsening in association with normalization of TGF-β2 expression and high expression of TGF-β1. Taken together, these data support the hypothesis that compensatory autocrine and/or paracrine events contribute to the pathogenesis of TGF-β-mediated vasculopathies.
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DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Intellectual disability (ID) disorders are genetically and phenotypically highly heterogeneous and present a major challenge in clinical genetics and medicine. Although many genes involved in ID have ...been identified, the etiology is unknown in most affected individuals. Moreover, the function of most genes associated with ID remains poorly characterized. Evidence is accumulating that the control of gene transcription through epigenetic modification of chromatin structure in neurons has an important role in cognitive processes and in the etiology of ID. However, our understanding of the key molecular players and mechanisms in this process is highly fragmentary. Here, we identify a chromatin-modification module that underlies a recognizable form of ID, the Kleefstra syndrome phenotypic spectrum (KSS). In a cohort of KSS individuals without mutations in EHMT1 (the only gene known to be disrupted in KSS until now), we identified de novo mutations in four genes, MBD5, MLL3, SMARCB1, and NR1I3, all of which encode epigenetic regulators. Using Drosophila, we demonstrate that MBD5, MLL3, and NR1I3 cooperate with EHMT1, whereas SMARCB1 is known to directly interact with MLL3. We propose a highly conserved epigenetic network that underlies cognition in health and disease. This network should allow the design of strategies to treat the growing group of ID pathologies that are caused by epigenetic defects.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Haploinsufficiency (HI) is the best characterized mechanism through which dominant mutations exert their effect and cause disease. Non-haploinsufficiency (NHI) mechanisms, such as gain-of-function ...and dominant-negative mechanisms, are often characterized by the spatial clustering of mutations, thereby affecting only particular regions or base pairs of a gene. Variants leading to haploinsufficency might occasionally cluster as well, for example in critical domains, but such clustering is on the whole less pronounced with mutations often spread throughout the gene. Here we exploit this property and develop a method to specifically identify genes with significant spatial clustering patterns of de novo mutations in large cohorts. We apply our method to a dataset of 4,061 de novo missense mutations from published exome studies of trios with intellectual disability and developmental disorders (ID/DD) and successfully identify 15 genes with clustering mutations, including 12 genes for which mutations are known to cause neurodevelopmental disorders. For 11 out of these 12, NHI mutation mechanisms have been reported. Additionally, we identify three candidate ID/DD-associated genes of which two have an established role in neuronal processes. We further observe a higher intolerance to normal genetic variation of the identified genes compared to known genes for which mutations lead to HI. Finally, 3D modeling of these mutations on their protein structures shows that 81% of the observed mutations are unlikely to affect the overall structural integrity and that they therefore most likely act through a mechanism other than HI.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Abstract
Aims
The dilated cardiomyopathy (DCM) phenotype is the result of combined genetic and acquired triggers. Until now, clinical decision-making in DCM has mainly been based on ejection fraction ...(EF) and NYHA classification, not considering the DCM heterogenicity. The present study aimed to identify patient subgroups by phenotypic clustering integrating aetiologies, comorbidities, and cardiac function along cardiac transcript levels, to unveil pathophysiological differences between DCM subgroups.
Methods and results
We included 795 consecutive DCM patients from the Maastricht Cardiomyopathy Registry who underwent in-depth phenotyping, comprising extensive clinical data on aetiology and comorbodities, imaging and endomyocardial biopsies. Four mutually exclusive and clinically distinct phenogroups (PG) were identified based upon unsupervised hierarchical clustering of principal components: PG1 mild systolic dysfunction, PG2 auto-immune, PG3 genetic and arrhythmias, and PG4 severe systolic dysfunction. RNA-sequencing of cardiac samples (n = 91) revealed a distinct underlying molecular profile per PG: pro-inflammatory (PG2, auto-immune), pro-fibrotic (PG3; arrhythmia), and metabolic (PG4, low EF) gene expression. Furthermore, event-free survival differed among the four phenogroups, also when corrected for well-known clinical predictors. Decision tree modelling identified four clinical parameters (auto-immune disease, EF, atrial fibrillation, and kidney function) by which every DCM patient from two independent DCM cohorts could be placed in one of the four phenogroups with corresponding outcome (n = 789; Spain, n = 352 and Italy, n = 437), showing a feasible applicability of the phenogrouping.
Conclusion
The present study identified four different DCM phenogroups associated with significant differences in clinical presentation, underlying molecular profiles and outcome, paving the way for a more personalized treatment approach.
Graphical Abstract
IMPORTANCE: Severe coronavirus disease 2019 (COVID-19) can occur in younger, predominantly male, patients without preexisting medical conditions. Some individuals may have primary immunodeficiencies ...that predispose to severe infections caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). OBJECTIVE: To explore the presence of genetic variants associated with primary immunodeficiencies among young patients with COVID-19. DESIGN, SETTING, AND PARTICIPANTS: Case series of pairs of brothers without medical history meeting the selection criteria of young (age <35 years) brother pairs admitted to the intensive care unit (ICU) due to severe COVID-19. Four men from 2 unrelated families were admitted to the ICUs of 4 hospitals in the Netherlands between March 23 and April 12, 2020. The final date of follow-up was May 16, 2020. Available family members were included for genetic variant segregation analysis and as controls for functional experiments. EXPOSURE: Severe COVID-19. MAIN OUTCOME AND MEASURES: Results of rapid clinical whole-exome sequencing, performed to identify a potential monogenic cause. Subsequently, basic genetic and immunological tests were performed in primary immune cells isolated from the patients and family members to characterize any immune defects. RESULTS: The 4 male patients had a mean age of 26 years (range, 21-32), with no history of major chronic disease. They were previously well before developing respiratory insufficiency due to severe COVID-19, requiring mechanical ventilation in the ICU. The mean duration of ventilatory support was 10 days (range, 9-11); the mean duration of ICU stay was 13 days (range, 10-16). One patient died. Rapid clinical whole-exome sequencing of the patients and segregation in available family members identified loss-of-function variants of the X-chromosomal TLR7. In members of family 1, a maternally inherited 4-nucleotide deletion was identified (c.2129_2132del; p.Gln710Argfs*18); the affected members of family 2 carried a missense variant (c.2383G>T; p.Val795Phe). In primary peripheral blood mononuclear cells from the patients, downstream type I interferon (IFN) signaling was transcriptionally downregulated, as measured by significantly decreased mRNA expression of IRF7, IFNB1, and ISG15 on stimulation with the TLR7 agonist imiquimod as compared with family members and controls. The production of IFN-γ, a type II IFN, was decreased in patients in response to stimulation with imiquimod. CONCLUSIONS AND RELEVANCE: In this case series of 4 young male patients with severe COVID-19, rare putative loss-of-function variants of X-chromosomal TLR7 were identified that were associated with impaired type I and II IFN responses. These preliminary findings provide insights into the pathogenesis of COVID-19.
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