Genetic Diagnostic Evaluation of Trio-Based Whole Exome Sequencing Among Children With Diagnosed or Suspected Autism Spectrum Disorder.
ABSTRACT: Autism spectrum disorder (ASD) is a group of clinically and genetically heterogeneous neurodevelopmental disorders. Recent tremendous advances in the whole exome sequencing (WES) enable rapid identification of variants associated with ASD including single nucleotide variations (SNVs) and indels. To further explore genetic etiology of ASD in Chinese children with negative findings of copy number variants (CNVs), we applied WES in 80 simplex families with a single affected offspring with ASD or suspected ASD, and validated variations predicted to be damaging by Sanger sequencing. The results showed that an overall diagnostic yield of 8.8% (9.2% in the group of ASD and 6.7% in the group of suspected ASD) was observed in our cohort. Among patients with diagnosed ASD, developmental delay or intellectual disability (DD/ID) was the most common comorbidity with a diagnostic yield of 13.3%, followed by seizures (50.0%) and craniofacial anomalies (40.0%). All of identified de novo SNVs and indels among patients with ASD were loss of function (LOF) variations and were slightly more frequent among female (male vs. female: 7.3% vs. 8.5%). A total of seven presumed causative genes (CHD8, AFF2, ADNP, POGZ, SHANK3, IL1RAPL1, and PTEN) were identified in this study. In conclusion, WES is an efficient diagnostic tool for diagnosed ASD especially those with negative findings of CNVs and other neurological disorders in clinical practice, enabling early identification of disease related genes and contributing to precision and personalized medicine.
Project description:The routine assessment to determine the genetic etiology for fetal ultrasound anomalies follows a sequential approach, which usually takes about 6-8 weeks turnaround time (TAT). We evaluated the clinical utility of simultaneous detection of copy number variations (CNVs) and single nucleotide variants (SNVs)/small insertion-deletions (indels) in fetuses with a normal karyotype with ultrasound anomalies. We performed CNV detection by chromosomal microarray analysis (CMA) or low pass CNV-sequencing (CNV-seq), and in parallel SNVs/indels detection by trio-based clinical exome sequencing (CES) or whole exome sequencing (WES). Eight-three singleton pregnancies with a normal fetal karyotype were enrolled in this prospective observational study. Pathogenic or likely pathogenic variations were identified in 30 cases (CNVs in 3 cases, SNVs/indels in 27 cases), indicating an overall molecular diagnostic rate of 36.1% (30/83). Two cases had both a CNV of uncertain significance (VOUS) and likely pathogenic SNV, and one case carried both a VOUS CNV and an SNV. We demonstrated that simultaneous analysis of CNVs and SNVs/indels can improve the diagnostic yield of prenatal diagnosis with shortened reporting time, namely, 2-3 weeks. Due to the relatively long TAT for sequential procedure for prenatal genetic diagnosis, as well as recent sequencing technology advancements, it is clinically necessary to consider the simultaneous evaluation of CNVs and SNVs/indels to enhance the diagnostic yield and timely TAT, especially for cases in the late second trimester or third trimester.
Project description:Genetic variants account for more than half of the cases with congenital or prelingual onset hearing loss. Autosomal recessive nonsyndromic hearing loss (ARNSHL) is the most common subgroup. Whole-exome sequencing (WES) has been shown to be effective detecting deafness-causing single-nucleotide variants (SNVs) and insertion/deletions (INDELs). After analyzing the WES data for causative SNVs or INDELs involving previously reported deafness genes in 78 families with ARNSHL, we searched for copy number variants (CNVs) through two different tools in 24 families that remained unresolved. We detected large homozygous deletions in STRC and OTOA in single families. Thus, causative CNVs in known deafness genes explain 2 out of 78 (2.6%) families in our sample set. We conclude that CNVs can be reliably detected through WES and should be the part of pipelines used to clarify genetic basis of hearing loss.
Project description:We compared whole-exome sequencing (WES) and whole-genome sequencing (WGS) in six unrelated individuals. In the regions targeted by WES capture (81.5% of the consensus coding genome), the mean numbers of single-nucleotide variants (SNVs) and small insertions/deletions (indels) detected per sample were 84,192 and 13,325, respectively, for WES, and 84,968 and 12,702, respectively, for WGS. For both SNVs and indels, the distributions of coverage depth, genotype quality, and minor read ratio were more uniform for WGS than for WES. After filtering, a mean of 74,398 (95.3%) high-quality (HQ) SNVs and 9,033 (70.6%) HQ indels were called by both platforms. A mean of 105 coding HQ SNVs and 32 indels was identified exclusively by WES whereas 692 HQ SNVs and 105 indels were identified exclusively by WGS. We Sanger-sequenced a random selection of these exclusive variants. For SNVs, the proportion of false-positive variants was higher for WES (78%) than for WGS (17%). The estimated mean number of real coding SNVs (656 variants, ?3% of all coding HQ SNVs) identified by WGS and missed by WES was greater than the number of SNVs identified by WES and missed by WGS (26 variants). For indels, the proportions of false-positive variants were similar for WES (44%) and WGS (46%). Finally, WES was not reliable for the detection of copy-number variations, almost all of which extended beyond the targeted regions. Although currently more expensive, WGS is more powerful than WES for detecting potential disease-causing mutations within WES regions, particularly those due to SNVs.
Project description:We aimed to achieve a retrospective molecular diagnosis by applying state-of-the-art genomic sequencing methods to past patients with T-B+NK+ severe combined immunodeficiency (SCID). We included identification of copy number variations (CNVs) by whole exome sequencing (WES) using the CNV calling method ExomeDepth to detect gene alterations for which routine Sanger sequencing analysis is not suitable, such as large heterozygous deletions.Of a total of 12 undiagnosed patients with T-B+NK+ SCID, we analyzed eight probands by WES, using GATK to detect single nucleotide variants (SNVs) and small insertions and deletions (INDELs) and ExomeDepth to detect CNVs.We found heterozygous single- or multi-exon deletions in IL7R, a known disease gene for autosomal recessive T-B+NK+ SCID, in four families (seven patients). In three families (five patients), these deletions coexisted with a heterozygous splice site or nonsense mutation elsewhere in the same gene, consistent with compound heterozygosity. In our cohort, about a quarter of T-B+NK+ SCID patients (26%) had such compound heterozygous IL7R deletions.We show that heterozygous IL7R exon deletions are common in T-B+NK+ SCID and are detectable by WES. They should be considered if Sanger sequencing fails to detect homozygous or compound heterozygous IL7R SNVs or INDELs.
Project description:Whole-exome sequencing (WES) studies have demonstrated the contribution of de novo loss-of-function single-nucleotide variants (SNVs) to autism spectrum disorder (ASD). However, challenges in the reliable detection of de novo insertions and deletions (indels) have limited inclusion of these variants in prior analyses. By applying a robust indel detection method to WES data from 787 ASD families (2,963 individuals), we demonstrate that de novo frameshift indels contribute to ASD risk (OR = 1.6; 95% CI = 1.0-2.7; p = 0.03), are more common in female probands (p = 0.02), are enriched among genes encoding FMRP targets (p = 6 × 10(-9)), and arise predominantly on the paternal chromosome (p < 0.001). On the basis of mutation rates in probands versus unaffected siblings, we conclude that de novo frameshift indels contribute to risk in approximately 3% of individuals with ASD. Finally, by observing clustering of mutations in unrelated probands, we uncover two ASD-associated genes: KMT2E (MLL5), a chromatin regulator, and RIMS1, a regulator of synaptic vesicle release.
Project description:Autism spectrum disorder (ASD) is a complex neurodevelopmental disorder with strong genetic contributions. To provide a comprehensive resource for the genetic evidence of ASD, we have updated the Autism KnowledgeBase (AutismKB) to version 2.0. AutismKB 2.0 integrates multiscale genetic data on 1379 genes, 5420 copy number variations and structural variations, 11 669 single-nucleotide variations or small insertions/deletions (SNVs/indels) and 172 linkage regions. In particular, AutismKB 2.0 highlights 5669 de novo SNVs/indels due to their significant contribution to ASD genetics and includes 789 mosaic variants due to their recently discovered contributions to ASD pathogenesis. The genes and variants are annotated extensively with genetic evidence and clinical evidence. To help users fully understand the functional consequences of SNVs and small indels, we provided comprehensive predictions of pathogenicity with iFish, SIFT, Polyphen etc. To improve user experiences, the new version incorporates multiple query methods, including simple query, advanced query and batch query. It also functionally integrates two analytical tools to help users perform downstream analyses, including a gene ranking tool and an enrichment analysis tool, KOBAS. AutismKB 2.0 is freely available and can be a valuable resource for researchers.
Project description:More than 98% of the human genome is made up of non-coding DNA, but techniques to ascertain its contribution to human disease have lagged far behind our understanding of protein coding variations. Autism spectrum disorder (ASD) has been mostly associated with coding variations via de novo single nucleotide variants (SNVs), recessive/homozygous SNVs, or de novo copy number variants (CNVs); however, most ASD cases continue to lack a genetic diagnosis. We analyzed 187 consanguineous ASD families for biallelic CNVs. Recessive deletions were significantly enriched in affected individuals relative to their unaffected siblings (17% versus 4%, p?<?0.001). Only a small subset of biallelic deletions were predicted to result in coding exon disruption. In contrast, biallelic deletions in individuals with ASD were enriched for overlap with regulatory regions, with 23/28 CNVs disrupting histone peaks in ENCODE (p?<?0.009). Overlap with regulatory regions was further demonstrated by comparisons to the 127-epigenome dataset released by the Roadmap Epigenomics project, with enrichment for enhancers found in primary brain tissue and neuronal progenitor cells. Our results suggest a novel noncoding mechanism of ASD, describe a powerful method to identify important noncoding regions in the human genome, and emphasize the potential significance of gene activation and regulation in cognitive and social function.
Project description:Childhood-onset schizophrenia (COS), a very rare and severe chronic psychiatric condition, is defined by an onset of positive symptoms (delusions, hallucinations and disorganized speech or behavior) before the age of 13. COS is associated with other neurodevelopmental disorders such as autism spectrum disorder (ASD) and attention deficit and hyperactivity disorder. Copy number variations (CNVs) represent well documented neurodevelopmental disorder risk factors and, recently, de novo single nucleotide variations (SNVs) in genes involved in brain development have also been implicated in the complex genetic architecture of COS. Here, we aim to review the genetic changes (CNVs and SNVs) reported for COS, going from previous studies to the whole genome sequencing era. We carried out a systematic review search in PubMed using the keywords "childhood(early)-onset schizophrenia(psychosis)" and "genetic(s) or gene(s) or genomic(s)" without language and date limitations. The main inclusion criteria are COS (onset before 13 years old) and all changes/variations at the DNA level (CNVs or SNVs). Thirty-six studies out of 205 met the inclusion criteria. Cytogenetic abnormalities (n = 72, including 66 CNVs) were identified in 16 autosomes and 2 sex chromosomes (X, Y), some with a higher frequency and clinical significance than others (e.g., 2p16.3, 3q29, 15q13.3, 22q11.21 deletions; 2p25.3, 3p25.3 and 16p11.2 duplications). Thirty-one single nucleotide mutations in genes principally involved in brain development and/or function have been found in 12 autosomes and one sex chromosome (X). We also describe five SNVs in X-linked genes inherited from a healthy mother, arguing for the X-linked recessive inheritance hypothesis. Moreover, ATP1A3 (19q13.2) is the only gene carrying more than one SNV in more than one patient, making it a strong candidate for COS. Mutations were distributed in various chromosomes illustrating the genetic heterogeneity of COS. More than 90% of CNVs involved in COS are also involved in ASD, supporting the idea that there may be genetic overlap between these disorders. Different mutations associated with COS are probably still unknown, and pathogenesis might also be explained by the association of different genetic variations (two or more CNVs or CNVs and SNVs) as well as association with early acquired brain lesions such as infection, hypoxia, or early childhood trauma.
Project description:High-coverage whole-genome sequencing data of a single ethnicity can provide a useful catalogue of population-specific genetic variations, and provides a critical resource that can be used to more accurately identify pathogenic genetic variants. We report a comprehensive analysis of the Korean population, and present the Korean National Standard Reference Variome (KoVariome). As a part of the Korean Personal Genome Project (KPGP), we constructed the KoVariome database using 5.5 terabases of whole genome sequence data from 50 healthy Korean individuals in order to characterize the benign ethnicity-relevant genetic variation present in the Korean population. In total, KoVariome includes 12.7M single-nucleotide variants (SNVs), 1.7M short insertions and deletions (indels), 4K structural variations (SVs), and 3.6K copy number variations (CNVs). Among them, 2.4M (19%) SNVs and 0.4M (24%) indels were identified as novel. We also discovered selective enrichment of 3.8M SNVs and 0.5M indels in Korean individuals, which were used to filter out 1,271 coding-SNVs not originally removed from the 1,000 Genomes Project when prioritizing disease-causing variants. KoVariome health records were used to identify novel disease-causing variants in the Korean population, demonstrating the value of high-quality ethnic variation databases for the accurate interpretation of individual genomes and the precise characterization of genetic variations.
Project description:Autism spectrum disorder (ASD) is a complex group of clinically heterogeneous neurodevelopmental disorders with unclear etiology and pathogenesis. Genetic studies have identified numerous candidate genetic variants, including de novo mutated ASD-associated genes; however, the function of these de novo mutated genes remains unclear despite extensive bioinformatics resources. Accordingly, it is not easy to assign priorities to numerous candidate ASD-associated genes for further biological analysis. Here we developed a convenient system for identifying an experimental evidence-based annotation of candidate ASD-associated genes. We performed trio-based whole-exome sequencing in 30 sporadic cases of ASD and identified 37 genes with de novo single-nucleotide variations (SNVs). Among them, 5 of those 37 genes, POGZ, PLEKHA4, PCNX, PRKD2 and HERC1, have been previously reported as genes with de novo SNVs in ASD; and consultation with in silico databases showed that only HERC1 might be involved in neural function. To examine whether the identified gene products are involved in neural functions, we performed small hairpin RNA-based assays using neuroblastoma cell lines to assess neurite development. Knockdown of 8 out of the 14 examined genes significantly decreased neurite development (P<0.05, one-way analysis of variance), which was significantly higher than the number expected from gene ontology databases (P=0.010, Fisher's exact test). Our screening system may be valuable for identifying the neural functions of candidate ASD-associated genes for further analysis and a substantial portion of these genes with de novo SNVs might have roles in neuronal systems, although further detailed analysis might eliminate false positive genes from identified candidate ASD genes.