Detection of variants in dystroglycanopathy-associated genes through the application of targeted whole-exome sequencing analysis to a large cohort of patients with unexplained limb-girdle muscle weakness

Background: Dystroglycanopathies are a clinically and genetically heterogeneous group of disorders that are typically characterised by limb-girdle muscle weakness. Mutations in 18 different genes have been associated with dystroglycanopathies, the encoded proteins of which typically modulate the binding of alpha-dystroglycan to extracellular matrix ligands by altering its glycosylation. This results in a disruption of the structural integrity of the myocyte, ultimately leading to muscle degeneration. Methods: Deep phenotypic information was gathered using the PhenoTips online software for 1001 patients with unexplained limb-girdle muscle weakness from 43 different centres across 21 European and Middle Eastern countries. Whole-exome sequencing with at least 250 ng DNA was completed using an Illumina exome capture and a 38 Mb baited target. Genes known to be associated with dystroglycanopathies were analysed for disease-causing variants. Results: Suspected pathogenic variants were detected in DPM3, ISPD, POMT1 and FKTN in one patient each, in POMK in two patients, in GMPPB in three patients, in FKRP in eight patients and in POMT2 in ten patients. This indicated a frequency of 2.7% for the disease group within the cohort of 1001 patients with unexplained limb-girdle muscle weakness. The phenotypes of the 27 patients were highly variable, yet with a fundamental presentation of proximal muscle weakness and elevated serum creatine kinase. Conclusions: Overall, we have identified 27 patients with suspected pathogenic variants in dystroglycanopathy-associated genes. We present evidence for the genetic and phenotypic diversity of the dystroglycanopathies as a disease group, while also highlighting the advantage of incorporating next-generation sequencing into the diagnostic pathway of rare diseases.Peer reviewe

Sequential targeted exome sequencing of 1001 patients affected by unexplained limb-girdle weakness

Several hundred genetic muscle diseases have been described, all of which are rare. Their clinical and genetic heterogeneity means that a genetic diagnosis is challenging. We established an international consortium, MYO-SEQ, to aid the work-ups of muscle disease patients and to better understand disease etiology. Exome sequencing was applied to 1001 undiagnosed patients recruited from more than 40 neuromuscular disease referral centers; standardized phenotypic information was collected for each patient. Exomes were examined for variants in 429 genes associated with muscle conditions. We identified suspected pathogenic variants in 52% of patients across 87 genes. We detected 401 novel variants, 116 of which were recurrent. Variants in CAPN3, DYSF, ANO5, DMD, RYR1, TTN, COL6A2, and SGCA collectively accounted for over half of the solved cases; while variants in newer disease genes, such as BVES and POGLUT1, were also found. The remaining well-characterized unsolved patients (48%) need further investigation. Using our unique infrastructure, we developed a pathway to expedite muscle disease diagnoses. Our data suggest that exome sequencing should be used for pathogenic variant detection in patients with suspected genetic muscle diseases, focusing first on the most common disease genes described here, and subsequently in rarer and newly characterized disease genes

Detection of variants in dystroglycanopathy-associated genes through the application of targeted whole-exome sequencing analysis to a large cohort of patients with unexplained limb-girdle muscle weakness

Abstract Background Dystroglycanopathies are a clinically and genetically heterogeneous group of disorders that are typically characterised by limb-girdle muscle weakness. Mutations in 18 different genes have been associated with dystroglycanopathies, the encoded proteins of which typically modulate the binding of α-dystroglycan to extracellular matrix ligands by altering its glycosylation. This results in a disruption of the structural integrity of the myocyte, ultimately leading to muscle degeneration. Methods Deep phenotypic information was gathered using the PhenoTips online software for 1001 patients with unexplained limb-girdle muscle weakness from 43 different centres across 21 European and Middle Eastern countries. Whole-exome sequencing with at least 250 ng DNA was completed using an Illumina exome capture and a 38 Mb baited target. Genes known to be associated with dystroglycanopathies were analysed for disease-causing variants. Results Suspected pathogenic variants were detected in DPM3, ISPD, POMT1 and FKTN in one patient each, in POMK in two patients, in GMPPB in three patients, in FKRP in eight patients and in POMT2 in ten patients. This indicated a frequency of 2.7% for the disease group within the cohort of 1001 patients with unexplained limb-girdle muscle weakness. The phenotypes of the 27 patients were highly variable, yet with a fundamental presentation of proximal muscle weakness and elevated serum creatine kinase. Conclusions Overall, we have identified 27 patients with suspected pathogenic variants in dystroglycanopathy-associated genes. We present evidence for the genetic and phenotypic diversity of the dystroglycanopathies as a disease group, while also highlighting the advantage of incorporating next-generation sequencing into the diagnostic pathway of rare diseases

Biallelic and monoallelic variants in PLXNA1 are implicated in a novel neurodevelopmental disorder with variable cerebral and eye anomalies.

PURPOSE: To investigate the effect of PLXNA1 variants on the phenotype of patients with autosomal dominant and recessive inheritance patterns and to functionally characterize the zebrafish homologs plxna1a and plxna1b during development. METHODS: We assembled ten patients from seven families with biallelic or de novo PLXNA1 variants. We describe genotype-phenotype correlations, investigated the variants by structural modeling, and used Morpholino knockdown experiments in zebrafish to characterize the embryonic role of plxna1a and plxna1b. RESULTS: Shared phenotypic features among patients include global developmental delay (9/10), brain anomalies (6/10), and eye anomalies (7/10). Notably, seizures were predominantly reported in patients with monoallelic variants. Structural modeling of missense variants in PLXNA1 suggests distortion in the native protein. Our zebrafish studies enforce an embryonic role of plxna1a and plxna1b in the development of the central nervous system and the eye. CONCLUSION: We propose that different biallelic and monoallelic variants in PLXNA1 result in a novel neurodevelopmental syndrome mainly comprising developmental delay, brain, and eye anomalies. We hypothesize that biallelic variants in the extracellular Plexin-A1 domains lead to impaired dimerization or lack of receptor molecules, whereas monoallelic variants in the intracellular Plexin-A1 domains might impair downstream signaling through a dominant-negative effect

Processing top-k dominating queries in metric spaces

Top-k dominating queries combine the natural idea of selecting the k best items with a comprehensive "goodness" criterion based on dominance. A point p1 dominates p2 if p1 is as good as p2 in all attributes and is strictly better in at least one. Existing works address the problem in settings where data objects are multidimensional points. However, there are domains where we only have access to the distance between two objects. In cases like these, attributes reflect distances from a set of input objects and are dynamically generated as the input objects change. Consequently, prior works from the literature cannot be applied, despite the fact that the dominance relation is still meaningful and valid. For this reason, in this work, we present the first study for processing top-k dominating queries over distance-based dynamic attribute vectors, defined over a metric space.We propose four progressive algorithms that utilize the properties of the underlying metric space to efficiently solve the problem and present an extensive, comparative evaluation on both synthetic and real-world datasets. © 2016 ACM

Left atrial voltage mapping using a new impedance-based algorithm in patients with paroxysmal atrial fibrillation

Aims: Atrial fibrosis is associated with the pathogenesis and progression of atrial fibrillation (AF). We sought to evaluate the extent of left atrial (LA) scarring in patients with paroxysmal AF (PAF) undergoing catheter ablation using a new impedance-based algorithm. Methods: We prospectively enrolled 73 consecutive patients (43 males, 58 years) with PAF who underwent pulmonary vein antral isolation. We first performed high-density bipolar voltage mapping during sinus rhythm using Tissue Proximity Indicator (TPI), one of the features of the ConfiDense mapping module integrated in the electroanatomic mapping system. A dense LA shell was created initially without TPI (mean points 2,411) and subsequently activating TPI (mean points 1,167). Each point was classified according to the peak-to-peak bipolar voltage electrogram based on two criteria (criterion A: healthy >0.8 mV, border zone: 0.4–0.8 mV, scarred: <0.4 mV; criterion B: healthy: >0.5 mV, border zone: 0.25–0.5 mV, scarred: <0.25 mV). Results: LA voltage analysis represented significantly smaller scarred areas when mapping was performed with TPI-ON compared with TPI-OFF in both voltage criteria (average LA voltage area: 3.02 ± 5.28 cm2 vs 9.15 ± 13.11 cm2 vs in criterion A and 1.19 ± 2.54 cm2 vs 5.61 ± 9.56 cm2 in criterion B). A statistically significant voltage difference was observed in all segments of the left atrium between the two mapping protocols, particularly on the inferior wall. Conclusion: A more specific delineation of LA fibrosis may be produced using the TPI feature of the ConfiDense mapping module, through elimination of false-positive annotated mapping points due to low contact. © 2018 Wiley Periodicals, Inc

) See paragraph at end of paper for supplementary material available

The use of trimethylsilyl cyanide (TMSCN) as a reagent for the direct formation of trimethylsilyl cyanohydrin ethers 3 from ketones is reported. The advantages in using TMSCN as opposed to hydrogen cyanide are illustrated by the formation of cyanohydrin ethers of ketones that do not form stable cyanohydrins. The reduction of derivatives 3 with lithium aluminum hydride is reported to afford fi-aminomethyl alcohols 4 in good yield. The combined carbonyl derivatization-reduction sequence should afford a general synthesis of 4 useful in executing ring expansion reactions. A great deal of attention has been devoted to the conversion of ketones to p-aminomethyl alcohols 4. Interest in these derivatives has largely centered around their use in the Tiffeneau-Demjanov ring expansion of cycloalkanones.2 The major difficulty in this general homologation process has been associated with the lack of reliable routes to p-aminomethyl alcohols. The two classical methods for effecting this transformation have involved the formation and subsequent reduction of either ketone cyanohydrins3 2 or P-nitromethyl alc o h o l~.~ Both procedures have suffered from lack of generality and low overall yields for the desired transformat i~n .~ For the more widely used homologation sequence proceeding through ketone cyanohydrins, the yield of pamino alcohol 4 is directly dependent upon the stability of the cyanohydrin 2, the formation of which is highly dependent upon the steric and strain factors in the ketone.6 Recently Parham and coworkers have shown that cyanohydrin ethers can be prepared by the acid-catalyzed addition of HCN to both alkyl78 and trimethylsilyl enol ethers,7b,c and that the resultant cyanohydrin derivatives can be reduced with LiA1H4 to the desired p-amino ethers or alcohols. Although this approach results in the synthesis of derivatives of unstable cyanohydrins, the sequence requires the synthesis of the appropriate enol derivative, thus lengthening as well as restricting the homologation sequence to those systems for which enol ethers are easily prepared. In conjunction with our interests in exploring the utility of trimethylsilyl cyanide (TMSCN)g as a useful reagent in organic synthesis, we would like to report on its advantages in effecting carbonyl aminomethylation via the asilyloxy nitriles 3 (eq I). Our previous studies have shown that, in contrast to the substrate sensitivity of HCN-carbonyl addition reactions, the addition of TMSCN to both ketones and aldehydes is a general, high-yield process.8 Apparently this is a consequence of the alteration in the AH for the carbonyl addi

Recurrent Mutations in the Basic Domain of TWIST2 Cause Ablepharon Macrostomia and Barber-Say Syndromes

Ablepharon macrostomia syndrome (AMS) and Barber-Say syndrome (BSS) are rare congenital ectodermal dysplasias characterized by similar clinical features. To establish the genetic basis of AMS and BSS, we performed extensive clinical phenotyping, whole exome and candidate gene sequencing, and functional validations. We identified a recurrent de novo mutation in TWIST2 in seven independent AMS-affected families, as well as another recurrent de novo mutation affecting the same amino acid in ten independent BSS-affected families. Moreover, a genotype-phenotype correlation was observed, because the two syndromes differed based solely upon the nature of the substituting amino acid: a lysine at TWIST2 residue 75 resulted in AMS, whereas a glutamine or alanine yielded BSS. TWIST2 encodes a basic helix-loop-helix transcription factor that regulates the development of mesenchymal tissues. All identified mutations fell in the basic domain of TWIST2 and altered the DNA-binding pattern of Flag-TWIST2 in HeLa cells. Comparison of wild-type and mutant TWIST2 expressed in zebrafish identified abnormal developmental phenotypes and widespread transcriptome changes. Our results suggest that autosomal-dominant TWIST2 mutations cause AMS or BSS by inducing protean effects on the transcription factor's DNA binding