Showing papers by "Paul Coucke published in 2013"

Deficiency for the ER-stress transducer OASIS causes severe recessive osteogenesis imperfecta in humans

Abstract: Osteogenesis imperfecta (OI) is a clinically and genetically heterogeneous brittle bone disorder. Whereas dominant OI is mostly due to heterozygous mutations in either COL1A1 or COL1A2, encoding type I procollagen, recessive OI is caused by biallelic mutations in genes encoding proteins involved in type I procollagen processing or chaperoning. Hitherto, some OI cases remain molecularly unexplained. We detected a homozygous genomic deletion of CREB3L1 in a family with severe OI. CREB3L1 encodes OASIS, an endoplasmic reticulum-stress transducer that regulates type I procollagen expression during murine bone formation. This is the first report linking CREB3L1 to human recessive OI, thereby expanding the OI gene spectrum.

Comprehensive Clinical and Molecular Analysis of 12 Families with Type 1 Recessive Cutis Laxa

Abstract: Autosomal recessive cutis laxa type I (ARCL type I) is characterized by generalized cutis laxa with pulmonary emphysema and/or vascular complications. Rarely, mutations can be identified in FBLN4 or FBLN5. Recently, LTBP4 mutations have been implicated in a similar phenotype. Studying FBLN4, FBLN5, and LTBP4 in 12 families with ARCL type I, we found bi-allelic FBLN5 mutations in two probands, whereas nine probands harbored biallelic mutations in LTBP4. FBLN5 and LTBP4 mutations cause a very similar phenotype associated with severe pulmonary emphysema, in the absence of vascular tortuosity or aneurysms. Gastrointestinal and genitourinary tract involvement seems to be more severe in patients with LTBP4 mutations. Functional studies showed that most premature termination mutations in LTBP4 result in severely reduced mRNA and protein levels. This correlated with increased transforming growth factor-beta (TGFβ) activity. However, one mutation, c.4127dupC, escaped nonsense-mediated decay. The corresponding mutant protein (p.Arg1377Alafs(*) 27) showed reduced colocalization with fibronectin, leading to an abnormal morphology of microfibrils in fibroblast cultures, while retaining normal TGFβ activity. We conclude that LTBP4 mutations cause disease through both loss of function and gain of function mechanisms.

A sensitive and specific diagnostic test for hearing loss using a microdroplet PCR-based approach and next generation sequencing.

Abstract: Implementing DNA diagnostics in clinical practice for extremely heterogeneous diseases such as hearing loss is challenging, especially when attempting to reach high sensitivity and specificity in a cost-effective fashion. Next generation sequencing has enabled the development of such a test, but the most commonly used genomic target enrichment methods such as hybridization-based capture suffer from restrictions. In this study, we have adopted a new flexible approach using microdroplet PCR-based technology for target enrichment, in combination with massive parallel sequencing to develop a DNA diagnostic test for autosomal recessive hereditary hearing loss. This approach enabled us to identify the genetic basis of hearing loss in 9 of 24 patients, a success rate of 37.5%. Our method also proved to have high sensitivity and specificity. Currently, routine molecular genetic diagnostic testing for deafness is in most cases only performed for the GJB2 gene and a positive result is typically only obtained in 10-20% of deaf children. Individuals with mutations in GJB2 had already been excluded in our selected set of 24 patients. Therefore, we anticipate that our deafness test may lead to a genetic diagnosis in roughly 50% of unscreened autosomal recessive deafness cases. We propose that this diagnostic testing approach represents a significant improvement in clinical practice as a standard diagnostic tool for children with hearing loss.

Twenty patients including 7 probands with autosomal dominant cutis laxa confirm clinical and molecular homogeneity

Abstract: Background: Elastin gene mutations have been associated with a variety of phenotypes. Autosomal dominant cutis laxa (ADCL) is a rare disorder that presents with lax skin, typical facial characteristics, inguinal hernias, aortic root dilatation and pulmonary emphysema. In most patients, frameshift mutations are found in the 3’ region of the elastin gene (exons 30-34) which result in a C-terminally extended protein, though exceptions have been reported. Methods: We clinically and molecularly characterized the thus far largest cohort of ADCL patients, consisting of 19 patients from six families and one sporadic patient. Results: Molecular analysis showed C-terminal frameshift mutations in exon 30, 32, and 34 of the elastin gene and identified a mutational hotspot in exon 32 (c.2262delA). This cohort confirms the previously reported clinical constellation of skin laxity (100%), inguinal hernias (51%), aortic root dilatation (55%) and emphysema (37%). Conclusion: ADCL is a clinically and molecularly homogeneous disorder, but intra- and interfamilial variability in the severity of organ involvement needs to be taken into account. Regular cardiovascular and pulmonary evaluations are imperative in the clinical follow-up of these patients.

Whole exome sequencing is an efficient, sensitive and specific method of mutation detection in osteogenesis imperfecta and Marfan syndrome

Abstract: Osteogenesis imperfecta (OI) and Marfan syndrome (MFS) are common Mendelian disorders. Both conditions are usually diagnosed clinically, as genetic testing is expensive due to the size and number of potentially causative genes and mutations. However, genetic testing may benefit patients, at-risk family members and individuals with borderline phenotypes, as well as improving genetic counseling and allowing critical differential diagnoses. We assessed whether whole exome sequencing (WES) is a sensitive method for mutation detection in OI and MFS. WES was performed on genomic DNA from 13 participants with OI and 10 participants with MFS who had known mutations, with exome capture followed by massive parallel sequencing of multiplexed samples. Single nucleotide polymorphisms (SNPs) and small indels were called using Genome Analysis Toolkit (GATK) and annotated with ANNOVAR. CREST, exomeCopy and exomeDepth were used for large deletion detection. Results were compared with the previous data. Specificity was calculated by screening WES data from a control population of 487 individuals for mutations in COL1A1, COL1A2 and FBN1. The target capture of five exome capture platforms was compared. All 13 mutations in the OI cohort and 9/10 in the MFS cohort were detected (sensitivity=95.6%) including non-synonymous SNPs, small indels (<10 bp), and a large UTR5/exon 1 deletion. One mutation was not detected by GATK due to strand bias. Specificity was 99.5%. Capture platforms and analysis programs differed considerably in their ability to detect mutations. Consumable costs for WES were low. WES is an efficient, sensitive, specific and cost-effective method for mutation detection in patients with OI and MFS. Careful selection of platform and analysis programs is necessary to maximize success.

Second family with the boston‐type craniosynostosis syndrome: Novel mutation and expansion of the clinical spectrum

Abstract: Craniosynostosis, caused by early fusion of one or more cranial sutures, can affect the coronal or lambdoid sutures, or include premature fusion of the sagittal (scaphocephaly) or metopic suture (trigonocephaly). Often occurring as isolated finding, their co-existence in a craniosynostosis syndrome is infrequent. Wedescribeafour-generationfamilywithvariableexpressionof a craniosynostosis phenotype with scaphocephaly and a particularly severe trigonocephaly. Molecular analysis revealed a missense mutation in the MSX2—associated with the Boston-type craniosynostosis syndrome—affecting the sameamino-acid residue as in the original Boston family. Besides unique features such as the cranial sutures involved, minor limb abnormalities and incomplete penetrance, our patients share with the original family autosomal dominant inheritance and the presence of multiple endocranial erosions on CT imaging. Though these findings appear to be important diagnostic clues for MSX2related craniosynostosis, it is noteworthy that the first affected generation in this family presented merely with isolated sagittal or unicoronal craniosynostosis and cutaneous syndactyly. Molecular analysis of MSX2 should therefore be considered in patients with isolated scaphocephaly/unicoronal synostosis, especiallyinthepresenceofafamilyhistoryforcraniosynostosisor syndactyly. 2013 Wiley Periodicals, Inc.

Dermatosparaxis (Ehlers-Danlos type VIIC): prenatal diagnosis following a previous pregnancy with unexpected skull fractures at delivery.

Abstract: Dermatosparaxis Ehlers–Danlos syndrome (or EDS VIIC), a rare autosomal recessive connective tissue disorder, is characterized by extreme skin fragility, premature rupture of membranes in pregnancy, and spontaneous rupture of internal organs Here we report a second patient with EDS VIIC presenting with congenital skull fractures and skin lacerations at birth, complications which may occur more frequently than previously thought in this condition We also discuss the role of prenatal diagnosis in the management of a subsequent normal pregnancy © 2013 Wiley Periodicals, Inc

Zebrafish Models for Ectopic Mineralization Disorders: Practical Issues from Morpholino Design to Post-Injection Observations

Abstract: Zebrafish (ZF, Danio rerio) has emerged as an important and popular model species to study different human diseases. Key regulators of skeletal development and calcium metabolism are highly conserved between mammals and ZF. The corresponding orthologs share significant sequence similarities and an overlap in expression patterns when compared to mammals, making ZF a potential model for the study of mineralization-related disorders and soft tissue mineralization. To characterize the function of early mineralization-related genes in ZF, these genes can be knocked down by injecting morpholinos into early stage embryos. Validation of the morpholino needs to be performed and the concern of aspecific effects can be addressed by applying one or more independent techniques to knock down the gene of interest. Post-injection assessment of early mineralization defects can be done using general light microscopy, calcein staining, Alizarin red staining, Alizarin red-Alcian blue double staining, and by the use of transgenic lines. Examination of general molecular defects can be done by performing protein and gene expression analysis, and more specific processes can be explored by investigating ectopic mineralization-related mechanisms such as apoptosis and mitochondrial dysfunction. In this paper, we will discuss all details about the aforementioned techniques; shared knowledge will be very useful for the future investigation of ZF models for ectopic mineralization disorders and to understand the underlying pathways involved in soft tissue calcification.

New insights into the molecular diagnosis and management of heritable thoracic aortic aneurysms and dissections.

Abstract: Since the identification of the fibrillin‑1 gene as the causal gene for Marfan syndrome, our knowledge of molecular genetics and the applicability of genetic testing for heritable thoracic aneurysms and dissections (H-TAD) in clinical practice have increased substantially. Several new syndromes related to H-TAD have been described and the list of mutated genes in syndromal and nonsyndromal H-TAD is rapidly expanding. This knowledge has led to a significant improvement of our insight into the underlying pathophysiology of H-TAD resulting in new opportunities for targeted treatment, as well as in improved risk stratification. Clinicians involved in the care for H-TAD patients require a basic knowledge of the disease entities and need to be correctly informed on the applicability of genetic testing in their patients and families. Gene‑tailored treatment and management should now be considered as part of good clinical practice. We provide a systematic overview of genetic H-TAD entities and practical recommendations for genetic testing and patient management.

Genes in Thoracic Aortic Aneurysms and Dissections - Do they Matter?: Translation and Integration of Research and Modern Genetic Techniques into Daily Clinical Practice.

Abstract: Since the identification of the fibrillin-1 gene as the causal gene for Marfan syndrome, our knowledge of molecular genetics and the applicability of genetic testing in clinical practice have expanded dramatically. Several new syndromes related to thoracic aortic aneurysms and dissections (TAAD) have been described and the list of underlying genes in syndromal and nonsyndromal TAAD already includes more than 10 different genes and is rapidly expanding. Based on this knowledge, our insights into the underlying pathophysiology of TAAD have improved significantly, and new opportunities for targeted treatment have emerged. Clinicians involved in the care of TAAD patients require a basic knowledge of the disease entities and need to be informed on the applicability of genetic testing in their patients and families. Gene-tailored treatment and management is indeed no science fiction anymore and should now be considered as part of good clinical practice. We provide a systematic overview of genetic TAAD entities and practical recommendations for genetic testing and patient management.