Anna C. Need

Large recurrent microdeletions associated with schizophrenia.

Reduced fecundity, associated with severe mental disorders, places negative selection pressure on risk alleles and may explain, in part, why common variants have not been found that confer risk of disorders such as autism, schizophrenia and mental retardation. Thus, rare variants may account for a larger fraction of the overall genetic risk than previously assumed. In contrast to rare single nucleotide mutations, rare copy number variations (CNVs) can be detected using genome-wide single nucleotide polymorphism arrays. This has led to the identification of CNVs associated with mental retardation and autism. In a genome-wide search for CNVs associating with schizophrenia, we used a population-based sample to identify de novo CNVs by analysing 9,878 transmissions from parents to offspring. The 66 de novo CNVs identified were tested for association in a sample of 1,433 schizophrenia cases and 33,250 controls. Three deletions at 1q21.1, 15q11.2 and 15q13.3 showing nominal association with schizophrenia in the first sample (phase I) were followed up in a second sample of 3,285 cases and 7,951 controls (phase II). All three deletions significantly associate with schizophrenia and related psychoses in the combined sample. The identification of these rare, recurrent risk variants, having occurred independently in multiple founders and being subject to negative selection, is important in itself. CNV analysis may also point the way to the identification of additional and more prevalent risk variants in genes and pathways involved in schizophrenia.

Common variants conferring risk of schizophrenia

Schizophrenia is a complex disorder, caused by both genetic and environmental factors and their interactions. Research on pathogenesis has traditionally focused on neurotransmitter systems in the brain, particularly those involving dopamine. Schizophrenia has been considered a separate disease for over a century, but in the absence of clear biological markers, diagnosis has historically been based on signs and symptoms. A fundamental message emerging from genome-wide association studies of copy number variations (CNVs) associated with the disease is that its genetic basis does not necessarily conform to classical nosological disease boundaries. Certain CNVs confer not only high relative risk of schizophrenia but also of other psychiatric disorders. The structural variations associated with schizophrenia can involve several genes and the phenotypic syndromes, or the ‘genomic disorders’, have not yet been characterized. Single nucleotide polymorphism (SNP)-based genome-wide association studies with the potential to implicate individual genes in complex diseases may reveal underlying biological pathways. Here we combined SNP data from several large genome-wide scans and followed up the most significant association signals. We found significant association with several markers spanning the major histocompatibility complex (MHC) region on chromosome 6p21.3-22.1, a marker located upstream of the neurogranin gene (NRGN) on 11q24.2 and a marker in intron four of transcription factor 4 (TCF4) on 18q21.2. Our findings implicating the MHC region are consistent with an immune component to schizophrenia risk, whereas the association with NRGN and TCF4 points to perturbation of pathways involved in brain development, memory and cognition.

A genome-wide association study in chronic obstructive pulmonary disease (COPD): identification of two major susceptibility loci.

There is considerable variability in the susceptibility of smokers to develop chronic obstructive pulmonary disease (COPD). The only known genetic risk factor is severe deficiency of α1-antitrypsin, which is present in 1–2% of individuals with COPD. We conducted a genome-wide association study (GWAS) in a homogenous case-control cohort from Bergen, Norway (823 COPD cases and 810 smoking controls) and evaluated the top 100 single nucleotide polymorphisms (SNPs) in the family-based International COPD Genetics Network (ICGN; 1891 Caucasian individuals from 606 pedigrees) study. The polymorphisms that showed replication were further evaluated in 389 subjects from the US National Emphysema Treatment Trial (NETT) and 472 controls from the Normative Aging Study (NAS) and then in a fourth cohort of 949 individuals from 127 extended pedigrees from the Boston Early-Onset COPD population. Logistic regression models with adjustments of covariates were used to analyze the case-control populations. Family-based association analyses were conducted for a diagnosis of COPD and lung function in the family populations. Two SNPs at the α-nicotinic acetylcholine receptor (CHRNA 3/5) locus were identified in the genome-wide association study. They showed unambiguous replication in the ICGN family-based analysis and in the NETT case-control analysis with combined p-values of 1.48×10−10, (rs8034191) and 5.74×10−10 (rs1051730). Furthermore, these SNPs were significantly associated with lung function in both the ICGN and Boston Early-Onset COPD populations. The C allele of the rs8034191 SNP was estimated to have a population attributable risk for COPD of 12.2%. The association of hedgehog interacting protein (HHIP) locus on chromosome 4 was also consistently replicated, but did not reach genome-wide significance levels. Genome-wide significant association of the HHIP locus with lung function was identified in the Framingham Heart study (Wilk et al., companion article in this issue of PLoS Genetics; doi:10.1371/journal.pgen.1000429). The CHRNA 3/5 and the HHIP loci make a significant contribution to the risk of COPD. CHRNA3/5 is the same locus that has been implicated in the risk of lung cancer.

A Genome-Wide Investigation of SNPs and CNVs in Schizophrenia

We report a genome-wide assessment of single nucleotide polymorphisms (SNPs) and copy number variants (CNVs) in schizophrenia. We investigated SNPs using 871 patients and 863 controls, following up the top hits in four independent cohorts comprising 1,460 patients and 12,995 controls, all of European origin. We found no genome-wide significant associations, nor could we provide support for any previously reported candidate gene or genome-wide associations. We went on to examine CNVs using a subset of 1,013 cases and 1,084 controls of European ancestry, and a further set of 60 cases and 64 controls of African ancestry. We found that eight cases and zero controls carried deletions greater than 2 Mb, of which two, at 8p22 and 16p13.11-p12.4, are newly reported here. A further evaluation of 1,378 controls identified no deletions greater than 2 Mb, suggesting a high prior probability of disease involvement when such deletions are observed in cases. We also provide further evidence for some smaller, previously reported, schizophrenia-associated CNVs, such as those in NRXN1 and APBA2. We could not provide strong support for the hypothesis that schizophrenia patients have a significantly greater “load” of large (>100 kb), rare CNVs, nor could we find common CNVs that associate with schizophrenia. Finally, we did not provide support for the suggestion that schizophrenia-associated CNVs may preferentially disrupt genes in neurodevelopmental pathways. Collectively, these analyses provide the first integrated study of SNPs and CNVs in schizophrenia and support the emerging view that rare deleterious variants may be more important in schizophrenia predisposition than common polymorphisms. While our analyses do not suggest that implicated CNVs impinge on particular key pathways, we do support the contribution of specific genomic regions in schizophrenia, presumably due to recurrent mutation. On balance, these data suggest that very few schizophrenia patients share identical genomic causation, potentially complicating efforts to personalize treatment regimens.

Clinical application of exome sequencing in undiagnosed genetic conditions

Background There is considerable interest in the use of next-generation sequencing to help diagnose unidentified genetic conditions, but it is difficult to predict the success rate in a clinical setting that includes patients with a broad range of phenotypic presentations. Methods The authors present a pilot programme of whole-exome sequencing on 12 patients with unexplained and apparent genetic conditions, along with their unaffected parents. Unlike many previous studies, the authors did not seek patients with similar phenotypes, but rather enrolled any undiagnosed proband with an apparent genetic condition when predetermined criteria were met. Results This undertaking resulted in a likely genetic diagnosis in 6 of the 12 probands, including the identification of apparently causal mutations in four genes known to cause Mendelian disease (TCF4, EFTUD2, SCN2A and SMAD4) and one gene related to known Mendelian disease genes (NGLY1). Of particular interest is that at the time of this study, EFTUD2 was not yet known as a Mendelian disease gene but was nominated as a likely cause based on the observation of de novo mutations in two unrelated probands. In a seventh case with multiple disparate clinical features, the authors were able to identify homozygous mutations in EFEMP1 as a likely cause for macular degeneration (though likely not for other features). Conclusions This study provides evidence that next-generation sequencing can have high success rates in a clinical setting, but also highlights key challenges. It further suggests that the presentation of known Mendelian conditions may be considerably broader than currently recognised.

Rare deletions at 16p13.11 predispose to a diverse spectrum of sporadic epilepsy syndromes.

Deletions at 16p13.11 are associated with schizophrenia, mental retardation, and most recently idiopathic generalized epilepsy. To evaluate the role of 16p13.11 deletions, as well as other structural variation, in epilepsy disorders, we used genome-wide screens to identify copy number variation in 3812 patients with a diverse spectrum of epilepsy syndromes and in 1299 neurologically-normal controls. Large deletions (> 100 kb) at 16p13.11 were observed in 23 patients, whereas no control had a deletion greater than 16 kb. Patients, even those with identically sized 16p13.11 deletions, presented with highly variable epilepsy phenotypes. For a subset of patients with a 16p13.11 deletion, we show a consistent reduction of expression for included genes, suggesting that haploinsufficiency might contribute to pathogenicity. We also investigated another possible mechanism of pathogenicity by using hybridization-based capture and next-generation sequencing of the homologous chromosome for ten 16p13.11-deletion patients to look for unmasked recessive mutations. Follow-up genotyping of suggestive polymorphisms failed to identify any convincing recessive-acting mutations in the homologous interval corresponding to the deletion. The observation that two of the 16p13.11 deletions were larger than 2 Mb in size led us to screen for other large deletions. We found 12 additional genomic regions harboring deletions > 2 Mb in epilepsy patients, and none in controls. Additional evaluation is needed to characterize the role of these exceedingly large, non-locus-specific deletions in epilepsy. Collectively, these data implicate 16p13.11 and possibly other large deletions as risk factors for a wide range of epilepsy disorders, and they appear to point toward haploinsufficiency as a contributor to the pathogenicity of deletions.

WGAViewer: Software for genomic annotation of whole genome association studies

To meet the immediate need for a framework of post-whole genome association (WGA) annotation, we have developed WGAViewer, a suite of JAVA software tools that provides a user-friendly interface to automatically annotate, visualize, and interpret the set of P-values emerging from a WGA study. Most valuably, it can be used to highlight possible functional mechanisms in an automatic manner, for example, by directly or indirectly implicating a polymorphism with an apparent link to gene expression, and help to generate hypotheses concerning the possible biological bases of observed associations. The easily interpretable diagrams can then be used to identify the associations that seem most likely to be biologically relevant, and to select genomic regions that may need to be resequenced in a search for candidate causal variants. In this report, we used our recently completed study on host control of HIV-1 viral load during the asymptomatic set point period as an illustration for the heuristic annotation of this software and its contributive role in a successful WGA project.

The Characterization of Twenty Sequenced Human Genomes

We present the analysis of twenty human genomes to evaluate the prospects for identifying rare functional variants that contribute to a phenotype of interest. We sequenced at high coverage ten “case” genomes from individuals with severe hemophilia A and ten “control” genomes. We summarize the number of genetic variants emerging from a study of this magnitude, and provide a proof of concept for the identification of rare and highly-penetrant functional variants by confirming that the cause of hemophilia A is easily recognizable in this data set. We also show that the number of novel single nucleotide variants (SNVs) discovered per genome seems to stabilize at about 144,000 new variants per genome, after the first 15 individuals have been sequenced. Finally, we find that, on average, each genome carries 165 homozygous protein-truncating or stop loss variants in genes representing a diverse set of pathways.

Long-Range LD Can Confound Genome Scans in Admixed Populations

A.L.P. is supported by a Ruth Kirschstein National Research Service Award from the NIH. N.P. is supported by a K-01 career development award from the NIH. D.R. is supported by a Burroughs Wellcome Career Development Award in the Biomedical Sciences. This research was also supported by U-01 award HG004168 from the NIH (D.R. and N.P.), by NIDDK grant PO1DK46763 (J.I.R.), and by the Board of Governors Chair in Medical Genetics at Cedars-Sinai Medical Center (J.I.R.). Genotyping of the Puerto Rican samples was supported in part by grant M01-RR00425 to the Cedars-Sinai GCRC genotyping core (K.D.T.) and by NIH grant DK62413 (K.D.T).

Next generation disparities in human genomics: concerns and remedies.

Studies of human genetics, particularly genome-wide association studies (GWAS), have concentrated heavily on European populations, with individuals of African ancestry rarely represented. Reasons for this include the distribution of biomedical funding and the increased population structure and reduced linkage disequilibrium in African populations. Currently, few GWAS findings have clinical utility and, therefore, the field has not yet contributed to health-care disparities. As human genomics research progresses towards the whole-genome sequencing era, however, more clinically relevant results are likely to be discovered. As we discuss here, to avoid the genetics community contributing to healthcare disparities, it is important to adopt measures to ensure that populations of diverse ancestry are included in genomic studies, and that no major population groups are excluded.