Mohammed Uddin

Detection of Clinically Relevant Genetic Variants in Autism Spectrum Disorder by Whole-Genome Sequencing

Autism Spectrum Disorder (ASD) demonstrates high heritability and familial clustering, yet the genetic causes remain only partially understood as a result of extensive clinical and genomic heterogeneity. Whole-genome sequencing (WGS) shows promise as a tool for identifying ASD risk genes as well as unreported mutations in known loci, but an assessment of its full utility in an ASD group has not been performed. We used WGS to examine 32 families with ASD to detect de novo or rare inherited genetic variants predicted to be deleterious (loss-of-function and damaging missense mutations). Among ASD probands, we identified deleterious de novo mutations in six of 32 (19%) families and X-linked or autosomal inherited alterations in ten of 32 (31%) families (some had combinations of mutations). The proportion of families identified with such putative mutations was larger than has been previously reported; this yield was in part due to the comprehensive and uniform coverage afforded by WGS. Deleterious variants were found in four unrecognized, nine known, and eight candidate ASD risk genes. Examples include CAPRIN1 and AFF2 (both linked to FMR1, which is involved in fragile X syndrome), VIP (involved in social-cognitive deficits), and other genes such as SCN2A and KCNQ2 (linked to epilepsy), NRXN1, and CHD7, which causes ASD-associated CHARGE syndrome. Taken together, these results suggest that WGS and thorough bioinformatic analyses for de novo and rare inherited mutations will improve the detection of genetic variants likely to be associated with ASD or its accompanying clinical symptoms.

Whole-genome sequencing of quartet families with autism spectrum disorder

Autism spectrum disorder (ASD) is genetically heterogeneous, with evidence for hundreds of susceptibility loci. Previous microarray and exome-sequencing studies have examined portions of the genome in simplex families (parents and one ASD-affected child) having presumed sporadic forms of the disorder. We used whole-genome sequencing (WGS) of 85 quartet families (parents and two ASD-affected siblings), consisting of 170 individuals with ASD, to generate a comprehensive data resource encompassing all classes of genetic variation (including noncoding variants) and accompanying phenotypes, in apparently familial forms of ASD. By examining de novo and rare inherited single-nucleotide and structural variations in genes previously reported to be associated with ASD or other neurodevelopmental disorders, we found that some (69.4%) of the affected siblings carried different ASD-relevant mutations. These siblings with discordant mutations tended to demonstrate more clinical variability than those who shared a risk variant. Our study emphasizes that substantial genetic heterogeneity exists in ASD, necessitating the use of WGS to delineate all genic and non-genic susceptibility variants in research and in clinical diagnostics.

Molecular Diagnostic Yield of Chromosomal Microarray Analysis and Whole-Exome Sequencing in Children With Autism Spectrum Disorder

IMPORTANCE The use of genome-wide tests to provide molecular diagnosis for individuals with autism spectrum disorder (ASD) requires more study. OBJECTIVE To perform chromosomal microarray analysis (CMA) and whole-exome sequencing (WES) in a heterogeneous group of children with ASD to determine the molecular diagnostic yield of these tests in a sample typical of a developmental pediatric clinic. DESIGN, SETTING, AND PARTICIPANTS The sample consisted of 258 consecutively ascertained unrelated children with ASD who underwent detailed assessments to define morphology scores based on the presence of major congenital abnormalities and minor physical anomalies. The children were recruited between 2008 and 2013 in Newfoundland and Labrador, Canada. The probands were stratified into 3 groups of increasing morphological severity: essential, equivocal, and complex (scores of 0-3, 4-5, and ≥6). EXPOSURES All probands underwent CMA, with WES performed for 95 proband-parent trios. MAIN OUTCOMES AND MEASURES The overall molecular diagnostic yield for CMA and WES in a population-based ASD sample stratified in 3 phenotypic groups. RESULTS Of 258 probands, 24 (9.3%, 95%CI, 6.1%-13.5%) received a molecular diagnosis from CMA and 8 of 95 (8.4%, 95%CI, 3.7%-15.9%) from WES. The yields were statistically different between the morphological groups. Among the children who underwent both CMA and WES testing, the estimated proportion with an identifiable genetic etiology was 15.8% (95%CI, 9.1%-24.7%; 15/95 children). This included 2 children who received molecular diagnoses from both tests. The combined yield was significantly higher in the complex group when compared with the essential group (pairwise comparison, P = .002). [table: see text]. CONCLUSIONS AND RELEVANCE Among a heterogeneous sample of children with ASD, the molecular diagnostic yields of CMA and WES were comparable, and the combined molecular diagnostic yield was higher in children with more complex morphological phenotypes in comparison with the children in the essential category. If replicated in additional populations, these findings may inform appropriate selection of molecular diagnostic testing for children affected by ASD.

Whole genome sequencing resource identifies 18 new candidate genes for autism spectrum disorder

We are performing whole-genome sequencing of families with autism spectrum disorder (ASD) to build a resource (MSSNG) for subcategorizing the phenotypes and underlying genetic factors involved. Here we report sequencing of 5,205 samples from families with ASD, accompanied by clinical information, creating a database accessible on a cloud platform and through a controlled-access internet portal. We found an average of 73.8 de novo single nucleotide variants and 12.6 de novo insertions and deletions or copy number variations per ASD subject. We identified 18 new candidate ASD-risk genes and found that participants bearing mutations in susceptibility genes had significantly lower adaptive ability (P = 6 × 10−4). In 294 of 2,620 (11.2%) of ASD cases, a molecular basis could be determined and 7.2% of these carried copy number variations and/or chromosomal abnormalities, emphasizing the importance of detecting all forms of genetic variation as diagnostic and therapeutic targets in ASD.

Brain-expressed exons under purifying selection are enriched for de novo mutations in autism spectrum disorder

A universal challenge in genetic studies of autism spectrum disorders (ASDs) is determining whether a given DNA sequence alteration will manifest as disease. Among different population controls, we observed, for specific exons, an inverse correlation between exon expression level in brain and burden of rare missense mutations. For genes that harbor de novo mutations predicted to be deleterious, we found that specific critical exons were significantly enriched in individuals with ASD relative to their siblings without ASD (P < 1.13 × 10−38; odds ratio (OR) = 2.40). Furthermore, our analysis of genes with high exonic expression in brain and low burden of rare mutations demonstrated enrichment for known ASD-associated genes (P < 3.40 × 10−11; OR = 6.08) and ASD-relevant fragile-X protein targets (P < 2.91 × 10−157; OR = 9.52). Our results suggest that brain-expressed exons under purifying selection should be prioritized in genotype-phenotype studies for ASD and related neurodevelopmental conditions.

ARTS1 polymorphisms are associated with ankylosing spondylitis in Koreans

Objective To test the association between ARTS1 polymorphisms and Koreans with ankylosing spondylitis (AS). Methods All patients and controls were Korean. 872 patients with AS fulfilling the modified New York criteria and 403 healthy controls were genotyped for five single nucleotide polymorphisms (SNPs), rs27044, rs17482078, rs10050860, rs30107 and rs2287987, known to be associated with AS in Caucasians. Results SNPs rs27044 (p=9.37 × 10−7) and rs30187 (p=7.16 × 10−6) of ARTS1 were significantly associated with AS in Koreans. There was no significant association for rs17482078, rs10050860 and rs2287987. Two four-marker haplotypes were found to be associated with AS (GCCT: p=4.71×10−7, CCCC: p=8.56×10−6). Conclusions This is first confirmation in a non-Caucasian population that genetic polymorphisms in ARTS1 are associated with AS, implicating common pathogenetic mechanisms in Korean and Caucasian patients with AS.

Genome-wide characteristics of de novo mutations in autism

De novo mutations (DNMs) are important in autism spectrum disorder (ASD), but so far analyses have mainly been on the ~1.5% of the genome encoding genes. Here, we performed whole-genome sequencing (WGS) of 200 ASD parent–child trios and characterised germline and somatic DNMs. We confirmed that the majority of germline DNMs (75.6%) originated from the father, and these increased significantly with paternal age only (P=4.2×10−10). However, when clustered DNMs (those within 20 kb) were found in ASD, not only did they mostly originate from the mother (P=7.7×10−13), but they could also be found adjacent to de novo copy number variations where the mutation rate was significantly elevated (P=2.4×10−24). By comparing with DNMs detected in controls, we found a significant enrichment of predicted damaging DNMs in ASD cases (P=8.0×10−9; odds ratio=1.84), of which 15.6% (P=4.3×10−3) and 22.5% (P=7.0×10−5) were non-coding or genic non-coding, respectively. The non-coding elements most enriched for DNM were untranslated regions of genes, regulatory sequences involved in exon-skipping and DNase I hypersensitive regions. Using microarrays and a novel outlier detection test, we also found aberrant methylation profiles in 2/185 (1.1%) of ASD cases. These same individuals carried independently identified DNMs in the ASD-risk and epigenetic genes DNMT3A and ADNP. Our data begins to characterize different genome-wide DNMs, and highlight the contribution of non-coding variants, to the aetiology of ASD.

A high-resolution copy-number variation resource for clinical and population genetics

Purpose:Chromosomal microarray analysis to assess copy-number variation has become a first-tier genetic diagnostic test for individuals with unexplained neurodevelopmental disorders or multiple congenital anomalies. More than 100 cytogenetic laboratories worldwide use the new ultra-high resolution Affymetrix CytoScan-HD array to genotype hundreds of thousands of samples per year. Our aim was to develop a copy-number variation resource from a new population sample that would enable more accurate interpretation of clinical genetics data on this microarray platform and others.Methods:Genotyping of 1,000 adult volunteers who are broadly representative of the Ontario population (as obtained from the Ontario Population Genomics Platform) was performed with the CytoScan-HD microarray system, which has 2.7 million probes. Four independent algorithms were applied to detect copy-number variations. Reproducibility and validation metrics were quantified using sample replicates and quantitative-polymerase chain reaction, respectively.Results:DNA from 873 individuals passed quality control and we identified 71,178 copy-number variations (81 copy-number variations/individual); 9.8% (6,984) of these copy-number variations were previously unreported. After applying three layers of filtering criteria, from our highest confidence copy-number variation data set we obtained >95% reproducibility and >90% validation rates (73% of these copy-number variations overlapped at least one gene).Conclusion:The genotype data and annotated copy-number variations for this largely Caucasian population will represent a valuable public resource enabling clinical genetics research and diagnostics.Genet Med 17 9, 747–752.

Copy number variation in Han Chinese individuals with autism spectrum disorder

BackgroundAutism spectrum disorders (ASDs) are a group of neurodevelopmental conditions with a demonstrated genetic etiology. Rare (<1% frequency) copy number variations (CNVs) account for a proportion of the genetic events involved, but the contribution of these events in non-European ASD populations has not been well studied. Here, we report on rare CNVs detected in a cohort of individuals with ASD of Han Chinese background.MethodsDNA samples were obtained from 104 ASD probands and their parents who were recruited from Harbin, China. Samples were genotyped on the Affymetrix CytoScan HD platform. Rare CNVs were identified by comparing data with 873 technology-matched controls from Ontario and 1,235 additional population controls of Han Chinese ethnicity.ResultsOf the probands, 8.6% had at least 1 de novo CNV (overlapping the GIGYF2, SPRY1, 16p13.3, 16p11.2, 17p13.3-17p13.2, DMD, and NAP1L6 genes/loci). Rare inherited CNVs affected other plausible neurodevelopmental candidate genes including GRID2, LINGO2, and SLC39A12. A 24-kb duplication was also identified at YWHAE, a gene previously implicated in ASD and other developmental disorders. This duplication is observed at a similar frequency in cases and in population controls and is likely a benign Asian-specific copy number polymorphism.ConclusionsOur findings help define genomic features relevant to ASD in the Han Chinese and emphasize the importance of using ancestry-matched controls in medical genetic interpretations.

Rates, distribution and implications of postzygotic mosaic mutations in autism spectrum disorder

We systematically analyzed postzygotic mutations (PZMs) in whole-exome sequences from the largest collection of trios (5,947) with autism spectrum disorder (ASD) available, including 282 unpublished trios, and performed resequencing using multiple independent technologies. We identified 7.5% of de novo mutations as PZMs, 83.3% of which were not described in previous studies. Damaging, nonsynonymous PZMs within critical exons of prenatally expressed genes were more common in ASD probands than controls (P < 1 × 10−6), and genes carrying these PZMs were enriched for expression in the amygdala (P = 5.4 × 10−3). Two genes (KLF16 and MSANTD2) were significantly enriched for PZMs genome-wide, and other PZMs involved genes (SCN2A, HNRNPU and SMARCA4) whose mutation is known to cause ASD or other neurodevelopmental disorders. PZMs constitute a significant proportion of de novo mutations and contribute importantly to ASD risk.