Cognitive phenotypes and genomic copy number variations.

The association of copy number variations (CNVs), differing numbers of copies of genetic sequence at locations in the genome, with phenotypes such as intellectual disability has been almost exclusively evaluated using clinically ascertained cohorts. The contribution of these genetic variants to cognitive phenotypes in the general population remains unclear. To investigate the clinical features conferred by CNVs associated with known syndromes in adult carriers without clinical preselection and to assess the genome-wide consequences of rare CNVs (frequency ≤0.05%; size ≥250 kilobase pairs [kb]) on carriers' educational attainment and intellectual disability prevalence in the general population. The population biobank of Estonia contains 52,000 participants enrolled from 2002 through 2010. General practitioners examined participants and filled out a questionnaire of health- and lifestyle-related questions, as well as reported diagnoses. Copy number variant analysis was conducted on a random sample of 7877 individuals and genotype-phenotype associations with education and disease traits were evaluated. Our results were replicated on a high-functioning group of 993 Estonians and 3 geographically distinct populations in the United Kingdom, the United States, and Italy. Phenotypes of genomic disorders in the general population, prevalence of autosomal CNVs, and association of these variants with educational attainment (from less than primary school through scientific degree) and prevalence of intellectual disability. Of the 7877 in the Estonian cohort, we identified 56 carriers of CNVs associated with known syndromes. Their phenotypes, including cognitive and psychiatric problems, epilepsy, neuropathies, obesity, and congenital malformations are similar to those described for carriers of identical rearrangements ascertained in clinical cohorts. A genome-wide evaluation of rare autosomal CNVs (frequency, ≤0.05%; ≥250 kb) identified 831 carriers (10.5%) of the screened general population. Eleven of 216 (5.1%) carriers of a deletion of at least 250 kb (odds ratio [OR], 3.16; 95% CI, 1.51-5.98; P = 1.5e-03) and 6 of 102 (5.9%) carriers of a duplication of at least 1 Mb (OR, 3.67; 95% CI, 1.29-8.54; P = .008) had an intellectual disability compared with 114 of 6819 (1.7%) in the Estonian cohort. The mean education attainment was 3.81 (P = 1.06e-04) among 248 (≥250 kb) deletion carriers and 3.69 (P = 5.024e-05) among 115 duplication carriers (≥1 Mb). Of the deletion carriers, 33.5% did not graduate from high school (OR, 1.48; 95% CI, 1.12-1.95; P = .005) and 39.1% of duplication carriers did not graduate high school (OR, 1.89; 95% CI, 1.27-2.8; P = 1.6e-03). Evidence for an association between rare CNVs and lower educational attainment was supported by analyses of cohorts of adults from Italy and the United States and adolescents from the United Kingdom. Known pathogenic CNVs in unselected, but assumed to be healthy, adult populations may be associated with unrecognized clinical sequelae. Additionally, individually rare but collectively common intermediate-size CNVs may be negatively associated with educational attainment. Replication of these findings in additional population groups is warranted given the potential implications of this observation for genomics research, clinical care, and public health.

Cancer is usually considered to be an acquired disease caused by the progressive accumulation of somatic deoxyribonucleic acid (DNA) changes. It is clear, however, that inherited factors may also play a significant role in the initiation of this disease. Some of these factors represent loss‐of‐function mutations in tumour suppressor genes, resulting in an increased cancer risk among carriers. Such an increased risk may also be attributed to the presence of multiple polymorphic variants such as single nucleotide polymorphisms (SNPs), each of which may convey a mild effect upon cancer susceptibility. DNA copy number variation (CNV) and other structural variations in the human genome are increasingly recognised as an alternative source of genetic variation that may influence cancer risk. Specifically, rare CNVs may affect important cancer‐associated genes and/or pathways and, thus, provide an explanation for moderate‐ to high‐risk cancer families. Therefore, it is anticipated that the identification of rare germline CNVs in unexplained familial and early onset cancer patients will contribute to our understanding of cancer predisposition and development. Key Concepts: Copy number variation (CNV) is a common form of normal variation affecting a significant proportion of the human genome. CNVs frequently encompass annotated genes. Disruption and/or silencing of critical genes located in rare CNVs may result in increased risk of disease, including cancer. Genome‐wide copy number analysis can be used as a strategy to identify novel moderate‐ to high‐penetrant cancer predisposing genes and/or mechanisms. Discovery of cancer‐predisposing CNVs may reveal new cancer syndromes that exhibit additional clinical features.

There now exist multiple lines of evidence pointing to a significant genetic component underlying the aetiology of autism spectrum disorders (ASDs). The advent of methodologies for scanning the human genome at high resolution, coupled with the recognition of copy number variation (CNV) as a prevalent source of genomic variation, has led to new strategies in the identification of clinically relevant loci. Balanced genomic changes, such as translocations and inversions, also contribute to ASD, but current studies have shown that screening with microarrays has up to fivefold increase in diagnostic yield. Recent work by our group and others has shown unbalanced genomic alterations that are likely pathogenic in upwards of 10% of cases, highlighting an important role for CNVs in the genetic aetiology of ASD. A trend in our empirical data has shifted focus for discovery of candidate loci towards individually rare but highly penetrant CNVs instead of looking for common variants of low penetrance. This strategy has proven largely successful in identifying ASD-susceptibility candidate loci, including gains and losses at 16p11.2, SHANK2, NRXN1, and PTCHD1. Another emerging and intriguing trend is the identification of the same genes implicated by rare CNVs across neurodevelopmental disorders, including schizophrenia, attention deficit hyperactivity disorder, and intellectual disability. These observations indicate that similar pathways may be involved in phenotypically distinct outcomes. Although interrogation of the genome at high resolution has led to these novel discoveries, it has also made cataloguing, characterization, and clinical interpretation of the increasing amount of CNV data difficult. Herein, we describe the history of genomic structural variation in ASD and how CNV discovery has been used to pinpoint novel ASD-susceptibility loci. We also discuss the overlap of CNVs across neurodevelopmental disorders and comment on the current challenges of understanding the relationship between CNVs and associated phenotypes in a clinical context.

Rare chromosomal deletions and duplications in attention-deficit hyperactivity disorder: a genome-wide analysis

Attention-deficit/hyperactivity disorder (ADHD) is a common, highly heritable neurodevelopmental disorder. Genetic loci have not yet been identified by genome-wide association studies. Rare copy number variations (CNVs), such as chromosomal deletions or duplications, have been implicated in ADHD and other neurodevelopmental disorders. To identify rare (frequency ⩽1%) CNVs that increase the risk of ADHD, we performed a whole-genome CNV analysis based on 489 young ADHD patients and 1285 adult population-based controls and identified one significantly associated CNV region. In tests for a global burden of large (>500 kb) rare CNVs, we observed a nonsignificant (P=0.271) 1.126-fold enriched rate of subjects carrying at least one such CNV in the group of ADHD cases. Locus-specific tests of association were used to assess if there were more rare CNVs in cases compared with controls. Detected CNVs, which were significantly enriched in the ADHD group, were validated by quantitative (q)PCR. Findings were replicated in an independent sample of 386 young patients with ADHD and 781 young population-based healthy controls. We identified rare CNVs within the parkinson protein 2 gene (PARK2) with a significantly higher prevalence in ADHD patients than in controls (P=2.8 × 10−4 after empirical correction for genome-wide testing). In total, the PARK2 locus (chr 6: 162 659 756–162 767 019) harboured three deletions and nine duplications in the ADHD patients and two deletions and two duplications in the controls. By qPCR analysis, we validated 11 of the 12 CNVs in ADHD patients (P=1.2 × 10−3 after empirical correction for genome-wide testing). In the replication sample, CNVs at the PARK2 locus were found in four additional ADHD patients and one additional control (P=4.3 × 10−2). Our results suggest that copy number variants at the PARK2 locus contribute to the genetic susceptibility of ADHD. Mutations and CNVs in PARK2 are known to be associated with Parkinson disease.

The phenotypic manifestations of rare CNVs in schizophrenia

Copy number variants (CNVs) are major variations contributing to the gene heterogeneity of congenital heart diseases (CHD). pulmonary atresia with ventricular septal defect (PA-VSD) is a rare form of cyanotic CHD characterized by complex manifestations and the genetic determinants underlying PA-VSD are still largely unknown. We investigated rare CNVs in a recruited cohort of 100 unrelated patients with PA-VSD, PA-IVS, or TOF and a population-matched control cohort of 100 healthy children using whole-exome sequencing. Comparing rare CNVs in PA-VSD cases and that in PA-IVS or TOF positive controls, we observed twenty-two rare CNVs only in PA-VSD, five rare CNVs only in PA-VSD and TOF as well as thirteen rare CNVs only in PA-VSD and PA-IVS. Six of these CNVs were considered pathogenic or potentially pathogenic to PA-VSD: 16p11.2 del (PPP4C and TBX6), 5q35.3 del (FLT4), 5p13.1 del (RICTOR), 6p21.33 dup (TNXB), 7p15.2 del (HNRNPA2B1), and 19p13.3 dup (FGF22). The gene networks showed that four putative candidate genes for PA-VSD, PPP4C, FLT4, RICTOR, and FGF22 had strong interaction with well-known cardiac genes relevant to heart or blood vessel development. Meanwhile, the analysis of transcriptome array revealed that PPP4C and RICTOR were also significantly expressed in human embryonic heart. In conclusion, three rare novel CNVs were identified only in PA-VSD: 16p11.2 del (PPP4C), 5q35.3 del (FLT4) and 5p13.1 del (RICTOR), implicating novel candidate genes of interest for PA-VSD. Our study provided new insights into understanding for the pathogenesis of PA-VSD and helped elucidate critical genes for PA-VSD.

Collaborative Genome-Wide Association and Copy Number Variation Analysis of Tourette Syndrome

This review examines the role of copy number variation in the human genome as a newly recognized determinant of lipoprotein and metabolic phenotypes. Much of the recent progress defining the molecular basis of lipoprotein disorders has been the result of studying genomic DNA at the single nucleotide level, for instance with nucleotide sequence analysis or genotyping to detect single nucleotide polymorphisms. Focus on single nucleotides, however, fails to capture the complete spectrum of potential genetic variability. Recent genome-wide mapping studies have demonstrated the surprising ubiquity of large-scale copy number variations in apparently healthy people, adding to the complexity of the 'normal' genome, but also emphasizing this form of genetic variation as a potential disease mechanism. The application of this understanding to the genetics of lipoprotein disorders has been rapid. For instance, the use of novel techniques to detect copy number variations, such as multiplex ligation-dependent probe amplification, has revealed many additional causative mutations in the low-density lipoprotein receptor gene in patients with familial hypercholesterolemia. Copy number variations thus represent a new level of genomic variation that is both an important mechanism of monogenic lipoprotein disorders and a potential contributor to common complex lipoprotein and metabolic phenotypes in the general population.

Introduction: Bicuspid Aortic Valve (BAV), the most common adult congenital heart defect, is a major cause of aortic insufficiency or stenosis requiring valve replacement and thoracic aortic aneurysms predisposing to acute aortic dissections. The spectrum of BAV ranges from early onset valve and aortic complications to sporadic late onset disease. We determined the frequency and gene content of rare genomic copy number variants (CNVs) in isolated BAV cases. Methods: We performed genome-wide SNP microarray analysis of familial BAV cases with early onset complications in the UTHealth BAV Research Registry (EBAV, n=394) and elderly probands from the International BAV Consortium (BAVCON, n=4216). CNVs were detected in Illumina genotypes using the PennCNV, QuantiSNP and cnvPartition algorithms. BAV cases were compared to 16,576 controls without known cardiovascular disease from the Database of Genotypes and Phenotypes. Only CNV calls with >6 consecutive variants, >20 kb in length, identified by at least two algorithms and intersected with known genes were included. For comparison, we assessed a cohort with left ventricular outflow tract obstructive lesions including BAV (LVOTO, n=1561) using identical methods. CNV burden and associations were tested using PLINK. Results: We identified 84 large recurrent CNVs in BAV cases that are absent or rare (<0.1%) in controls. 34 rare CNVs overlap between BAV and LVOTO cases and involve candidate genes that interact with each other during heart development. The largest and most prevalent of the recurrent CNVs are 8p23 duplications involving GATA4 (OR 340, CI: 42-2700). CNVs of genes that cause BAV when mutated were significantly enriched in cases (n=135, P =1x10 -27 ). The overall burden of rare genic CNVs, specifically large deletions, was also increased in BAV cases (empiric P < 1x10 -5 ) and was higher in younger EBAV cases than in older BAVCON cases. Conclusion: We identified likely pathogenic CNVs in more than 10% of BAV cases, implicating alterations of candidate genes at these loci in the pathogenesis of BAV.

Copy-Number Disorders Are a Common Cause of Congenital Kidney Malformations

BackgroundCopy number variation is an important class of genomic variation that has been reported in 75% of the human genome. However, it is underreported in African populations. Copy number variants (CNVs) could have important impacts on disease susceptibility and environmental adaptation. To describe CNVs and their possible impacts in Africans, we sequenced genomes of 232 individuals from three major African ethno-linguistic groups: (1) Niger Congo A from Guinea and Côte d’Ivoire, (2) Niger Congo B from Uganda and the Democratic Republic of Congo and (3) Nilo-Saharans from Uganda. We used GenomeSTRiP and cn.MOPS to identify copy number variant regions (CNVRs).ResultsWe detected 7608 CNVRs, of which 2172 were only deletions, 2384 were only insertions and 3052 had both. We detected 224 previously un-described CNVRs. The majority of novel CNVRs were present at low frequency and were not shared between populations. We tested for evidence of selection associated with CNVs and also for population structure. Signatures of selection identified previously, using SNPs from the same populations, were overrepresented in CNVRs. When CNVs were tagged with SNP haplotypes to identify SNPs that could predict the presence of CNVs, we identified haplotypes tagging 3096 CNVRs, 372 CNVRs had SNPs with evidence of selection (iHS > 3) and 222 CNVRs had both. This was more than expected (p < 0.0001) and included loci where CNVs have previously been associated with HIV, Rhesus D and preeclampsia. When integrated with 1000 Genomes CNV data, we replicated their observation of population stratification by continent but no clustering by populations within Africa, despite inclusion of Nilo-Saharans and Niger-Congo populations within our dataset.ConclusionsNovel CNVRs in the current study increase representation of African diversity in the database of genomic variants. Over-representation of CNVRs in SNP signatures of selection and an excess of SNPs that both tag CNVs and are subject to selection show that CNVs may be the actual targets of selection at some loci. However, unlike SNPs, CNVs alone do not resolve African ethno-linguistic groups. Tag haplotypes for CNVs identified may be useful in predicting African CNVs in future studies where only SNP data is available.

Rare copy number variants (CNVs)--deletions and duplications--have recently been established as important risk factors for both generalized and focal epilepsies. A systematic assessment of the role of CNVs in epileptic encephalopathies, the most devastating and often etiologically obscure group of epilepsies, has not been performed. We evaluated 315 patients with epileptic encephalopathies characterized by epilepsy and progressive cognitive impairment for rare CNVs using a high-density, exon-focused, whole-genome oligonucleotide array. We found that 25 of 315 (7.9%) of our patients carried rare CNVs that may contribute to their phenotype, with at least one-half being clearly or likely pathogenic. We identified 2 patients with overlapping deletions at 7q21 and 2 patients with identical duplications of 16p11.2. In our cohort, large deletions were enriched in affected individuals compared to controls, and 4 patients harbored 2 rare CNVs. We screened 2 novel candidate genes found within the rare CNVs in our cohort but found no mutations in our patients with epileptic encephalopathies. We highlight several additional novel candidate genes located in CNV regions. Our data highlight the significance of rare CNVs in the epileptic encephalopathies, and we suggest that CNV analysis should be considered in the genetic evaluation of these patients. Our findings also highlight novel candidate genes for further study.

Targeted Array CGH

Objective To detect the genomic copy number variations(CNVs) of mice embryonic neural tissue with neural tube defects(NTDs) induced by methotrexate(MTX), and investigate the molecular genetic mechanisms between folic acid metabolism disorders and NTDs pathogenesis. Methods C57BL/6J NTD mice model was induced by MTX on gestational day 7.5, and the maternal serum and NTD embryonic neural tissues were collected; the array-comparative genomic hybridization(array-CGH) assay was utilized to analyze the whole genomic CNVs in NTD embryonic neural tissues; reverse transcription polymerase chain reaction(RT-PCR) was used to confirm the positive results; the maternal serum concentrations of folic acid and related metabolites and dihydrofolate reductase(DHFR) activity were detected by liquid chromatography-tandem mass spectrometry and enzymatic methods, respectively. Results Array-CGH and RT-PCR results showed the 3 high confidence CNVs on XqE3, XqA1.1-qA2 and XqA1.1 in the NTD embryonic neural tissues.The NTD maternal serum concentrations of 5-methyltetrahydrofolate, 5-formyltetrahydrofolic acid, S-adenosyl methionine and DHFR activity were reduced significantly compared with the control group, and there were statistical differences(all P<0.05). Conclusions There are obvious CNVs in embryonic neural tissue of NTD mice induced by MTX and folic acid dysmetabolism might cause mice embryonic neural tube developmental disorders through CNVs. Key words: Neural tube defects; Folic acid; Copy number variations