Missense variants in ANKRD11 cause KBG syndrome by impairment of stability or transcriptional activity of the encoded protein

KBG syndrome; Missense variants; Neurodevelopmental disordersSíndrome KBG; Variants de missense; Trastorns del neurodesenvolupamentSíndrome KBG; Variantes de missense; Trastornos del neurodesarrolloPurpose Although haploinsufficiency of ANKRD11 is among the most common genetic causes of neurodevelopmental disorders, the role of rare ANKRD11 missense variation remains unclear. We characterized clinical, molecular, and functional spectra of ANKRD11 missense variants. Methods We collected clinical information of individuals with ANKRD11 missense variants and evaluated phenotypic fit to KBG syndrome. We assessed pathogenicity of variants through in silico analyses and cell-based experiments. Results We identified 20 unique, mostly de novo, ANKRD11 missense variants in 29 individuals, presenting with syndromic neurodevelopmental disorders similar to KBG syndrome caused by ANKRD11 protein truncating variants or 16q24.3 microdeletions. Missense variants significantly clustered in repression domain 2 at the ANKRD11 C-terminus. Of the 10 functionally studied missense variants, 6 reduced ANKRD11 stability. One variant caused decreased proteasome degradation and loss of ANKRD11 transcriptional activity. Conclusion Our study indicates that pathogenic heterozygous ANKRD11 missense variants cause the clinically recognizable KBG syndrome. Disrupted transrepression capacity and reduced protein stability each independently lead to ANKRD11 loss-of-function, consistent with haploinsufficiency. This highlights the diagnostic relevance of ANKRD11 missense variants, but also poses diagnostic challenges because the KBG-associated phenotype may be mild and inherited pathogenic ANKRD11 (missense) variants are increasingly observed, warranting stringent variant classification and careful phenotyping

Missense variants in ANKRD11 cause KBG syndrome by impairment of stability or transcriptional activity of the encoded protein

Purpose Although haploinsufficiency of ANKRD11 is among the most common genetic causes of neurodevelopmental disorders, the role of rare ANKRD11 missense variation remains unclear. We characterized clinical, molecular, and functional spectra of ANKRD11 missense variants. Methods We collected clinical information of individuals with ANKRD11 missense variants and evaluated phenotypic fit to KBG syndrome. We assessed pathogenicity of variants through in silico analyses and cell-based experiments. Results We identified 20 unique, mostly de novo, ANKRD11 missense variants in 29 individuals, presenting with syndromic neurodevelopmental disorders similar to KBG syndrome caused by ANKRD11 protein truncating variants or 16q24.3 microdeletions. Missense variants significantly clustered in repression domain 2 at the ANKRD11 C-terminus. Of the 10 functionally studied missense variants, 6 reduced ANKRD11 stability. One variant caused decreased proteasome degradation and loss of ANKRD11 transcriptional activity. Conclusion Our study indicates that pathogenic heterozygous ANKRD11 missense variants cause the clinically recognizable KBG syndrome. Disrupted transrepression capacity and reduced protein stability each independently lead to ANKRD11 loss-of-function, consistent with haploinsufficiency. This highlights the diagnostic relevance of ANKRD11 missense variants, but also poses diagnostic challenges because the KBG-associated phenotype may be mild and inherited pathogenic ANKRD11 (missense) variants are increasingly observed, warranting stringent variant classification and careful phenotyping

Twist exome capture allows for lower average sequence coverage in clinical exome sequencing

Background Exome and genome sequencing are the predominant techniques in the diagnosis and research of genetic disorders. Sufficient, uniform and reproducible/consistent sequence coverage is a main determinant for the sensitivity to detect single-nucleotide (SNVs) and copy number variants (CNVs). Here we compared the ability to obtain comprehensive exome coverage for recent exome capture kits and genome sequencing techniques. Results We compared three different widely used enrichment kits (Agilent SureSelect Human All Exon V5, Agilent SureSelect Human All Exon V7 and Twist Bioscience) as well as short-read and long-read WGS. We show that the Twist exome capture significantly improves complete coverage and coverage uniformity across coding regions compared to other exome capture kits. Twist performance is comparable to that of both short- and long-read whole genome sequencing. Additionally, we show that even at a reduced average coverage of 70× there is only minimal loss in sensitivity for SNV and CNV detection. Conclusion We conclude that exome sequencing with Twist represents a significant improvement and could be performed at lower sequence coverage compared to other exome capture techniques

Functional genomics of developmental disorders

DNA methylation, or the epigenetic modification of primarily cytosine bases within DNA to 5-methylcytosine through the addition of a methyl group, is an epigenetic mark with a variety of biological and cellular roles. Genetic and environmental influences can perturb DNA methylation patterns in humans, and the set of differentially methylated CpG sites perturbed can be collectively called a DNA methylation signature. In this thesis, I characterise DNA methylation signatures as a diagnostic biomarker for children with rare developmental disorders in chromatin-modifying genes. I show that DNA methylation signatures are a general property of these genes, that they have substantial clinical and diagnostic utility, and that they can be used to resolve variants of uncertain significance. I also show that these signatures are robust across scientific centres and can be generated across multiple tissues. Lastly, I compare DNA methylation signatures generated from methylation microarrays to those generated from genome-wide long read sequencing data, and provide evidence that long read sequencing is a reliable and scalable method to profile 5-methylcytosine for DNA methylation signature-based classification. Overall, my work emphasises the need for scalable, cost-effective, and relatively high-throughput biomarkers in the characterisation and diagnosis of rare developmental disorder syndromes.Wellcome Sanger Institute, Hinxton, U

Differences in the number of de novo mutations between individuals are due to small family-specific effects and stochasticity

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Application of metabolite set enrichment analysis on untargeted metabolomics data prioritises relevant pathways and detects novel biomarkers for inherited metabolic disorders

Untargeted metabolomics (UM) allows for the simultaneous measurement of hundreds of metabolites in a single analytical run. The sheer amount of data generated in UM hampers its use in patient diagnostics because manual interpretation of all features is not feasible. Here, we describe the application of a pathway‐based metabolite set enrichment analysis method to prioritise relevant biological pathways in UM data. We validate our method on a set of 55 patients with a diagnosed inherited metabolic disorder (IMD) and show that it complements feature‐based prioritisation of biomarkers by placing the features in a biological context. In addition, we find that by taking enriched pathways shared across different IMDs, we can identify common drugs and compounds that could otherwise obscure genuine disease biomarkers in an enrichment method. Finally, we demonstrate the potential of this method to identify novel candidate biomarkers for known IMDs. Our results show the added value of pathway‐based interpretation of UM data in IMD diagnostics context

Twist exome capture allows for lower average sequence coverage in clinical exome sequencing.

BACKGROUND: Exome and genome sequencing are the predominant techniques in the diagnosis and research of genetic disorders. Sufficient, uniform and reproducible/consistent sequence coverage is a main determinant for the sensitivity to detect single-nucleotide (SNVs) and copy number variants (CNVs). Here we compared the ability to obtain comprehensive exome coverage for recent exome capture kits and genome sequencing techniques. RESULTS: We compared three different widely used enrichment kits (Agilent SureSelect Human All Exon V5, Agilent SureSelect Human All Exon V7 and Twist Bioscience) as well as short-read and long-read WGS. We show that the Twist exome capture significantly improves complete coverage and coverage uniformity across coding regions compared to other exome capture kits. Twist performance is comparable to that of both short- and long-read whole genome sequencing. Additionally, we show that even at a reduced average coverage of 70× there is only minimal loss in sensitivity for SNV and CNV detection. CONCLUSION: We conclude that exome sequencing with Twist represents a significant improvement and could be performed at lower sequence coverage compared to other exome capture techniques

Twist exome capture allows for lower average sequence coverage in clinical exome sequencing

Abstract Background Exome and genome sequencing are the predominant techniques in the diagnosis and research of genetic disorders. Sufficient, uniform and reproducible/consistent sequence coverage is a main determinant for the sensitivity to detect single-nucleotide (SNVs) and copy number variants (CNVs). Here we compared the ability to obtain comprehensive exome coverage for recent exome capture kits and genome sequencing techniques. Results We compared three different widely used enrichment kits (Agilent SureSelect Human All Exon V5, Agilent SureSelect Human All Exon V7 and Twist Bioscience) as well as short-read and long-read WGS. We show that the Twist exome capture significantly improves complete coverage and coverage uniformity across coding regions compared to other exome capture kits. Twist performance is comparable to that of both short- and long-read whole genome sequencing. Additionally, we show that even at a reduced average coverage of 70× there is only minimal loss in sensitivity for SNV and CNV detection. Conclusion We conclude that exome sequencing with Twist represents a significant improvement and could be performed at lower sequence coverage compared to other exome capture techniques