FinnGen provides genetic insights from a well-phenotyped isolated population.

Population isolates such as those in Finland benefit genetic research because deleterious alleles are often concentrated on a small number of low-frequency variants (0.1% ≤ minor allele frequency < 5%). These variants survived the founding bottleneck rather than being distributed over a large number of ultrarare variants. Although this effect is well established in Mendelian genetics, its value in common disease genetics is less explored1,2. FinnGen aims to study the genome and national health register data of 500,000 Finnish individuals. Given the relatively high median age of participants (63 years) and the substantial fraction of hospital-based recruitment, FinnGen is enriched for disease end points. Here we analyse data from 224,737 participants from FinnGen and study 15 diseases that have previously been investigated in large genome-wide association studies (GWASs). We also include meta-analyses of biobank data from Estonia and the United Kingdom. We identified 30 new associations, primarily low-frequency variants, enriched in the Finnish population. A GWAS of 1,932 diseases also identified 2,733 genome-wide significant associations (893 phenome-wide significant (PWS), P < 2.6 × 10-11) at 2,496 (771 PWS) independent loci with 807 (247 PWS) end points. Among these, fine-mapping implicated 148 (73 PWS) coding variants associated with 83 (42 PWS) end points. Moreover, 91 (47 PWS) had an allele frequency of <5% in non-Finnish European individuals, of which 62 (32 PWS) were enriched by more than twofold in Finland. These findings demonstrate the power of bottlenecked populations to find entry points into the biology of common diseases through low-frequency, high impact variants.

Genetic associations of protein-coding variants in human disease.

Genome-wide association studies (GWAS) have identified thousands of genetic variants linked to the risk of human disease. However, GWAS have so far remained largely underpowered in relation to identifying associations in the rare and low-frequency allelic spectrum and have lacked the resolution to trace causal mechanisms to underlying genes1. Here we combined whole-exome sequencing in 392,814 UK Biobank participants with imputed genotypes from 260,405 FinnGen participants (653,219 total individuals) to conduct association meta-analyses for 744 disease endpoints across the protein-coding allelic frequency spectrum, bridging the gap between common and rare variant studies. We identified 975 associations, with more than one-third being previously unreported. We demonstrate population-level relevance for mutations previously ascribed to causing single-gene disorders, map GWAS associations to likely causal genes, explain disease mechanisms, and systematically relate disease associations to levels of 117 biomarkers and clinical-stage drug targets. Combining sequencing and genotyping in two population biobanks enabled us to benefit from increased power to detect and explain disease associations, validate findings through replication and propose medical actionability for rare genetic variants. Our study provides a compendium of protein-coding variant associations for future insights into disease biology and drug discovery.

Analysis of protein-coding genetic variation in 60,706 humans.

Large-scale reference data sets of human genetic variation are critical for the medical and functional interpretation of DNA sequence changes. Here we describe the aggregation and analysis of high-quality exome (protein-coding region) DNA sequence data for 60,706 individuals of diverse ancestries generated as part of the Exome Aggregation Consortium (ExAC). This catalogue of human genetic diversity contains an average of one variant every eight bases of the exome, and provides direct evidence for the presence of widespread mutational recurrence. We have used this catalogue to calculate objective metrics of pathogenicity for sequence variants, and to identify genes subject to strong selection against various classes of mutation; identifying 3,230 genes with near-complete depletion of predicted protein-truncating variants, with 72% of these genes having no currently established human disease phenotype. Finally, we demonstrate that these data can be used for the efficient filtering of candidate disease-causing variants, and for the discovery of human 'knockout' variants in protein-coding genes.

Haplotype Sharing Provides Insights into Fine-Scale Population History and Disease in Finland.

Finland provides unique opportunities to investigate population and medical genomics because of its adoption of unified national electronic health records, detailed historical and birth records, and serial population bottlenecks. We assembled a comprehensive view of recent population history (≤100 generations), the timespan during which most rare-disease-causing alleles arose, by comparing pairwise haplotype sharing from 43,254 Finns to that of 16,060 Swedes, Estonians, Russians, and Hungarians from geographically and linguistically adjacent countries with different population histories. We find much more extensive sharing in Finns, with at least one ≥ 5 cM tract on average between pairs of unrelated individuals. By coupling haplotype sharing with fine-scale birth records from more than 25,000 individuals, we find that although haplotype sharing broadly decays with geographical distance, there are pockets of excess haplotype sharing; individuals from northeast Finland typically share several-fold more of their genome in identity-by-descent segments than individuals from southwest regions. We estimate recent effective population-size changes through time across regions of Finland, and we find that there was more continuous gene flow as Finns migrated from southwest to northeast between the early- and late-settlement regions than was dichotomously described previously. Lastly, we show that haplotype sharing is locally enriched by an order of magnitude among pairs of individuals sharing rare alleles and especially among pairs sharing rare disease-causing variants. Our work provides a general framework for using haplotype sharing to reconstruct an integrative view of recent population history and gain insight into the evolutionary origins of rare variants contributing to disease.

Quantifying the Impact of Rare and Ultra-rare Coding Variation across the Phenotypic Spectrum.

There is a limited understanding about the impact of rare protein-truncating variants across multiple phenotypes. We explore the impact of this class of variants on 13 quantitative traits and 10 diseases using whole-exome sequencing data from 100,296 individuals. Protein-truncating variants in genes intolerant to this class of mutations increased risk of autism, schizophrenia, bipolar disorder, intellectual disability, and ADHD. In individuals without these disorders, there was an association with shorter height, lower education, increased hospitalization, and reduced age at enrollment. Gene sets implicated from GWASs did not show a significant protein-truncating variants burden beyond what was captured by established Mendelian genes. In conclusion, we provide a thorough investigation of the impact of rare deleterious coding variants on complex traits, suggesting widespread pleiotropic risk.

A multiomic approach to characterize the temporal sequence in Alzheimer's disease-related pathology.

No single-omic approach completely elucidates the multitude of alterations taking place in Alzheimer's disease (AD). Here, we coupled transcriptomic and phosphoproteomic approaches to determine the temporal sequence of changes in mRNA, protein, and phosphopeptide expression levels from human temporal cortical samples, with varying degree of AD-related pathology. This approach highlighted fluctuation in synaptic and mitochondrial function as the earliest pathological events in brain samples with AD-related pathology. Subsequently, increased expression of inflammation and extracellular matrix-associated gene products was observed. Interaction network assembly for the associated gene products, emphasized the complex interplay between these processes and the role of addressing post-translational modifications in the identification of key regulators. Additionally, we evaluate the use of decision trees and random forests in identifying potential biomarkers differentiating individuals with different degree of AD-related pathology. This multiomic and temporal sequence-based approach provides a better understanding of the sequence of events leading to AD.

Biallelic Variants in UBA5 Link Dysfunctional UFM1 Ubiquitin-like Modifier Pathway to Severe Infantile-Onset Encephalopathy.

The ubiquitin fold modifier 1 (UFM1) cascade is a recently identified evolutionarily conserved ubiquitin-like modification system whose function and link to human disease have remained largely uncharacterized. By using exome sequencing in Finnish individuals with severe epileptic syndromes, we identified pathogenic compound heterozygous variants in UBA5, encoding an activating enzyme for UFM1, in two unrelated families. Two additional individuals with biallelic UBA5 variants were identified from the UK-based Deciphering Developmental Disorders study and one from the Northern Finland Intellectual Disability cohort. The affected individuals (n = 9) presented in early infancy with severe irritability, followed by dystonia and stagnation of development. Furthermore, the majority of individuals display postnatal microcephaly and epilepsy and develop spasticity. The affected individuals were compound heterozygous for a missense substitution, c.1111G>A (p.Ala371Thr; allele frequency of 0.28% in Europeans), and a nonsense variant or c.164G>A that encodes an amino acid substitution p.Arg55His, but also affects splicing by facilitating exon 2 skipping, thus also being in effect a loss-of-function allele. Using an in vitro thioester formation assay and cellular analyses, we show that the p.Ala371Thr variant is hypomorphic with attenuated ability to transfer the activated UFM1 to UFC1. Finally, we show that the CNS-specific knockout of Ufm1 in mice causes neonatal death accompanied by microcephaly and apoptosis in specific neurons, further suggesting that the UFM1 system is essential for CNS development and function. Taken together, our data imply that the combination of a hypomorphic p.Ala371Thr variant in trans with a loss-of-function allele in UBA5 underlies a severe infantile-onset encephalopathy.

De Novo Truncating Variants in ASXL2 Are Associated with a Unique and Recognizable Clinical Phenotype.

The ASXL genes (ASXL1, ASXL2, and ASXL3) participate in body patterning during embryogenesis and encode proteins involved in epigenetic regulation and assembly of transcription factors to specific genomic loci. Germline de novo truncating variants in ASXL1 and ASXL3 have been respectively implicated in causing Bohring-Opitz and Bainbridge-Ropers syndromes, which result in overlapping features of severe intellectual disability and dysmorphic features. ASXL2 has not yet been associated with a human Mendelian disorder. In this study, we performed whole-exome sequencing in six unrelated probands with developmental delay, macrocephaly, and dysmorphic features. All six had de novo truncating variants in ASXL2. A careful review enabled the recognition of a specific phenotype consisting of macrocephaly, prominent eyes, arched eyebrows, hypertelorism, a glabellar nevus flammeus, neonatal feeding difficulties, hypotonia, and developmental disabilities. Although overlapping features with Bohring-Opitz and Bainbridge-Ropers syndromes exist, features that distinguish the ASXL2-associated condition from ASXL1- and ASXL3-related disorders are macrocephaly, absence of growth retardation, and more variability in the degree of intellectual disabilities. We were also able to demonstrate with mRNA studies that these variants are likely to exert a dominant-negative effect, given that both alleles are expressed in blood and the mutated ASXL2 transcripts escape nonsense-mediated decay. In conclusion, de novo truncating variants in ASXL2 underlie a neurodevelopmental syndrome with a clinically recognizable phenotype. This report expands the germline disorders that are linked to the ASXL genes.

Biallelic loss-of-function P4HTM gene variants cause hypotonia, hypoventilation, intellectual disability, dysautonomia, epilepsy, and eye abnormalities (HIDEA syndrome).

PURPOSE: A new syndrome with hypotonia, intellectual disability, and eye abnormalities (HIDEA) was previously described in a large consanguineous family. Linkage analysis identified the recessive disease locus, and genome sequencing yielded three candidate genes with potentially pathogenic biallelic variants: transketolase (TKT), transmembrane prolyl 4-hydroxylase (P4HTM), and ubiquitin specific peptidase 4 (USP4). However, the causative gene remained elusive. METHODS: International collaboration and exome sequencing were used to identify new patients with HIDEA and biallelic, potentially pathogenic, P4HTM variants. Segregation analysis was performed using Sanger sequencing. P4H-TM wild-type and variant constructs without the transmembrane region were overexpressed in insect cells and analyzed using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and western blot. RESULTS: Five different homozygous or compound heterozygous pathogenic P4HTM gene variants were identified in six new and six previously published patients presenting with HIDEA. Hypoventilation, obstructive and central sleep apnea, and dysautonomia were identified as novel features associated with the phenotype. Characterization of three of the P4H-TM variants demonstrated yielding insoluble protein products and, thus, loss-of-function. CONCLUSIONS: Biallelic loss-of-function P4HTM variants were shown to cause HIDEA syndrome. Our findings enable diagnosis of the condition, and highlight the importance of assessing the need for noninvasive ventilatory support in patients.

CTCF variants in 39 individuals with a variable neurodevelopmental disorder broaden the mutational and clinical spectrum.

PURPOSE: Pathogenic variants in the chromatin organizer CTCF were previously reported in seven individuals with a neurodevelopmental disorder (NDD). METHODS: Through international collaboration we collected data from 39 subjects with variants in CTCF. We performed transcriptome analysis on RNA from blood samples and utilized Drosophila melanogaster to investigate the impact of Ctcf dosage alteration on nervous system development and function. RESULTS: The individuals in our cohort carried 2 deletions, 8 likely gene-disruptive, 2 splice-site, and 20 different missense variants, most of them de novo. Two cases were familial. The associated phenotype was of variable severity extending from mild developmental delay or normal IQ to severe intellectual disability. Feeding difficulties and behavioral abnormalities were common, and variable other findings including growth restriction and cardiac defects were observed. RNA-sequencing in five individuals identified 3828 deregulated genes enriched for known NDD genes and biological processes such as transcriptional regulation. Ctcf dosage alteration in Drosophila resulted in impaired gross neurological functioning and learning and memory deficits. CONCLUSION: We significantly broaden the mutational and clinical spectrum ofCTCF-associated NDDs. Our data shed light onto the functional role of CTCF by identifying deregulated genes and show that Ctcf alterations result in nervous system defects in Drosophila.

Saccular Intracranial Aneurysms in Children When Both Parents Are Sporadic or Familial Carriers of Saccular Intracranial Aneurysms.

BACKGROUND AND PURPOSE: To study the penetrance of saccular intracranial aneurysm (IA) disease in children when both parents carry the disease. PATIENTS AND METHODS: The Kuopio IA Patient and Family Database includes all 4,411 IA patients admitted to the Kuopio University Hospital from its defined Eastern Finnish catchment population since 1980. We fused IA database with hospital diagnoses for IA patients and their 46,021 relatives from a national registry to identify couples concordant for IA disease. Penetrance of IA disease and hypertension were studied in these families. RESULTS: A total of 3,659 IA patients had 1 or more children. In total, 18 couples concordant for the IA disease with a total of 48 children, all born healthy, were identified. Hypertension was diagnosed in 23 (64%) of the 36 parents, and 7 of the 12 sporadic-sporadic couples were concordant for hypertension. Six sporadic-sporadic couples were concordant for subarachnoid haemorrhage (SAH). None of the 24 children to the 12 sporadic-sporadic couples had been diagnosed with SAH or IA disease. Instead, 11 (46%) of the 24 children to the 6 familial-sporadic couples had a diagnosed with SAH or IA disease. CONCLUSIONS: Couples concordant for IA disease are uncommon but not exceedingly rare. Biparental sporadic exposure does not seem to increase the risk of a clinically diagnosed IA disease or SAH in the offspring. IAs were common in the children with biparental sporadic-familial exposure.

Irregular Shape Identifies Ruptured Intracranial Aneurysm in Subarachnoid Hemorrhage Patients With Multiple Aneurysms.

BACKGROUND AND PURPOSE: We investigated which aneurysm-related risk factors for rupture best discriminate ruptured versus unruptured saccular intracranial aneurysms (sIAs) in subarachnoid hemorrhage patients with multiple sIAs. METHODS: We included 264 subarachnoid hemorrhage patients with a ruptured sIA and at least one additional unruptured sIA, from the Kuopio Intracranial Aneurysm database from 2003 to 2015. These patients had 268 ruptured and 445 unruptured sIAs. Angiograms of the 713 sIAs were reevaluated for multiple variables describing aneurysm shape. Multivariate generalized linear mixed models were used to calculate odds ratios with corresponding 95% confidence intervals for the independent risk factors for aneurysm rupture. RESULTS: In the multivariate analysis, only sIA size (P<0.004) and irregular shape (P<0.000) independently associated with sIA rupture. As an independent risk factor, irregular shape showed the strongest association with rupture (odds ratio 90.3; 95% confidence interval, 47.0-173.5). The sIA location, flow angles, bottleneck factor, or aspect ratio were not significantly associated with rupture. CONCLUSIONS: Irregular shape may identify the ruptured sIA better than size in patients presenting with aSAH and multiple sIAs.

High risk population isolate reveals low frequency variants predisposing to intracranial aneurysms.

3% of the population develops saccular intracranial aneurysms (sIAs), a complex trait, with a sporadic and a familial form. Subarachnoid hemorrhage from sIA (sIA-SAH) is a devastating form of stroke. Certain rare genetic variants are enriched in the Finns, a population isolate with a small founder population and bottleneck events. As the sIA-SAH incidence in Finland is >2× increased, such variants may associate with sIA in the Finnish population. We tested 9.4 million variants for association in 760 Finnish sIA patients (enriched for familial sIA), and in 2,513 matched controls with case-control status and with the number of sIAs. The most promising loci (p<5E-6) were replicated in 858 Finnish sIA patients and 4,048 controls. The frequencies and effect sizes of the replicated variants were compared to a continental European population using 717 Dutch cases and 3,004 controls. We discovered four new high-risk loci with low frequency lead variants. Three were associated with the case-control status: 2q23.3 (MAF 2.1%, OR 1.89, p 1.42×10-9); 5q31.3 (MAF 2.7%, OR 1.66, p 3.17×10-8); 6q24.2 (MAF 2.6%, OR 1.87, p 1.87×10-11) and one with the number of sIAs: 7p22.1 (MAF 3.3%, RR 1.59, p 6.08×-9). Two of the associations (5q31.3, 6q24.2) replicated in the Dutch sample. The 7p22.1 locus was strongly differentiated; the lead variant was more frequent in Finland (4.6%) than in the Netherlands (0.3%). Additionally, we replicated a previously inconclusive locus on 2q33.1 in all samples tested (OR 1.27, p 1.87×10-12). The five loci explain 2.1% of the sIA heritability in Finland, and may relate to, but not explain, the increased incidence of sIA-SAH in Finland. This study illustrates the utility of population isolates, familial enrichment, dense genotype imputation and alternate phenotyping in search for variants associated with complex diseases.

De Novo Aneurysm Formation in Carriers of Saccular Intracranial Aneurysm Disease in Eastern Finland.

BACKGROUND AND PURPOSE: Formation of new (de novo) aneurysms in patients carrying saccular intracranial aneurysm (sIA) disease has been published, but data from population-based cohorts are scarce. METHODS: Kuopio sIA database (http://www.uef.fi/ns) contains all unruptured and ruptured sIA patients admitted to Kuopio University Hospital from its Eastern Finnish catchment population. We studied the incidence and risk factors for de novo sIA formation in 1419 sIA patients with ≥5 years of angiographic follow-up, a total follow-up of 18 526 patient-years. RESULTS: There were 42 patients with a total of 56 de novo sIAs, diagnosed in a median of 11.7 years after the first sIA diagnosis. The cumulative incidence of de novo sIAs was 0.23% per patient-year and that of subarachnoid hemorrhage from a ruptured de novo sIA 0.05% per patient-year. The risk of de novo sIA discovery per patient-year increased with younger age at the first sIA diagnosis: 2.2% in the patients aged <20 years and 0.46% in the patients aged between 20 and 39 years. In Cox regression analysis, smoking history and younger age at the first sIA diagnosis significantly associated with de novo sIA formation, but female sex, multiple sIAs, and sIA family did not. CONCLUSIONS: Patients aged < 40 years at the first sIA diagnosis are in a significant risk of developing de novo sIAs, and they should be scheduled for long-term angiographic follow-up. Smoking increases the risk of de novo sIA formation, suggesting long-term follow-up for smokers. Antismoking efforts are highly recommended for sIA patients.

Antidepressant Use After Aneurysmal Subarachnoid Hemorrhage: A Population-Based Case-Control Study.

BACKGROUND AND PURPOSE: To elucidate the predictors of antidepressant use after subarachnoid hemorrhage from saccular intracranial aneurysm (sIA-SAH) in a population-based cohort with matched controls. METHODS: The Kuopio sIA database includes all unruptured and ruptured sIA cases admitted to the Kuopio University Hospital from its defined catchment population in Eastern Finland, with 3 matched controls for each patient. The use of all prescribed medicines has been fused from the Finnish national registry of prescribed medicines. In the present study, 2 or more purchases of antidepressant medication indicated antidepressant use. The risk factors of the antidepressant use were analyzed in 940 patients alive 12 months after sIA-SAH, and the classification tree analysis was used to create a predicting model for antidepressant use after sIA-SAH. RESULTS: The 940 12-month survivors of sIA-SAH had significantly more antidepressant use (odds ratio, 2.6; 95% confidence interval, 2.2-3.1) than their 2676 matched controls (29% versus 14%). Classification tree analysis, based on independent risk factors, was used for the best prediction model of antidepressant use after sIA-SAH. Modified Rankin Scale until 12 months was the most potent predictor, followed by condition (Hunt and Hess Scale) and age on admission for sIA-SAH. CONCLUSIONS: The sIA-SAH survivors use significantly more often antidepressants, indicative of depression, than their matched population controls. Even with a seemingly good recovery (modified Rankin Scale score, 0) at 12 months after sIA-SAH, there is a significant risk of depression requiring antidepressant medication.

Extending the phenotypic spectrum of RBFOX1 deletions: Sporadic focal epilepsy.

Partial deletions of the RBFOX1 gene encoding the neuronal splicing regulator have been reported in a range of neurodevelopmental diseases including idiopathic/genetic generalized epilepsy (IGE/GGE), childhood focal epilepsy, and self-limited childhood benign epilepsy with centrotemporal spikes (BECTS, rolandic epilepsy), and autism. The protein regulates alternative splicing of many neuronal transcripts involved in the homeostatic control of neuronal excitability. Herein, we examined whether structural deletions affecting RBFOX1 exons confer susceptibility to common forms of juvenile and adult focal epilepsy syndromes. We screened 807 unrelated patients with sporadic focal epilepsy, and we identified seven hemizygous exonic RBFOX1 deletions in patients with sporadic focal epilepsy (0.9%) in comparison to one deletion found in 1,502 controls. The phenotypes of the patients carrying RBFOX1 deletions comprise magnetic resonance imaging (MRI)-negative epilepsy of unknown etiology with frontal and temporal origin (n = 5) and two patients with temporal lobe epilepsy with hippocampal sclerosis. The epilepsies were largely pharmacoresistant but not associated with intellectual disability. Our study extends the phenotypic spectrum of RBFOX1 deletions as a risk factor for focal epilepsy and suggests that exonic RBFOX1 deletions are involved in the broad spectrum of focal and generalized epilepsies.

Genome-wide association study of intracranial aneurysm identifies a new association on chromosome 7.

BACKGROUND AND PURPOSE: Common variants have been identified using genome-wide association studies which contribute to intracranial aneurysms (IA) susceptibility. However, it is clear that the variants identified to date do not account for the estimated genetic contribution to disease risk. METHODS: Initial analysis was performed in a discovery sample of 2617 IA cases and 2548 controls of white ancestry. Novel chromosomal regions meeting genome-wide significance were further tested for association in 2 independent replication samples: Dutch (717 cases; 3004 controls) and Finnish (799 cases; 2317 controls). A meta-analysis was performed to combine the results from the 3 studies for key chromosomal regions of interest. RESULTS: Genome-wide evidence of association was detected in the discovery sample on chromosome 9 (CDKN2BAS; rs10733376: P<1.0×10(-11)), in a gene previously associated with IA. A novel region on chromosome 7, near HDAC9, was associated with IA (rs10230207; P=4.14×10(-8)). This association replicated in the Dutch sample (P=0.01) but failed to show association in the Finnish sample (P=0.25). Meta-analysis results of the 3 cohorts reached statistical significant (P=9.91×10(-10)). CONCLUSIONS: We detected a novel region associated with IA susceptibility that was replicated in an independent Dutch sample. This region on chromosome 7 has been previously associated with ischemic stroke and the large vessel stroke occlusive subtype (including HDAC9), suggesting a possible genetic link between this stroke subtype and IA.

Evidence for the additivity of rare and common variant burden throughout the spectrum of intellectual disability.

Intellectual disability (ID) is a common disorder, yet there is a wide spectrum of impairment from mild to profoundly affected individuals. Mild ID is seen as the low extreme of the general distribution of intelligence, while severe ID is often seen as a monogenic disorder caused by rare, pathogenic, highly penetrant variants. To investigate the genetic factors influencing mild and severe ID, we evaluated rare and common variation in the Northern Finland Intellectual Disability cohort (n = 1096 ID patients), a cohort with a high percentage of mild ID (n = 550) and from a population bottleneck enriched in rare, damaging variation. Despite this enrichment, we found only a small percentage of ID was due to recessive Finnish-enriched variants (0.5%). A larger proportion was linked to dominant variation, with a significant burden of rare, damaging variation in both mild and severe ID. This rare variant burden was enriched in more severe ID (p = 2.4e-4), patients without a relative with ID (p = 4.76e-4), and in those with features associated with monogenic disorders. We also found a significant burden of common variants associated with decreased cognitive function, with no difference between mild and more severe ID. When we included common and rare variants in a joint model, the rare and common variants had additive effects in both mild and severe ID. A multimodel inference approach also found that common and rare variants together best explained ID status (ΔAIC = 16.8, ΔBIC = 10.2). Overall, we report evidence for the additivity of rare and common variant burden throughout the spectrum of intellectual disability.