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The missing heritability of breast cancer could be partially attributed to rare variants (MAF < 0.5%). To identify breast cancer-associated rare coding variants, we conducted whole-exome sequencing (~50×) in genomic DNA samples obtained from 831 breast cancer cases and 839 controls of Chinese females. Using burden tests for each gene that included rare missense or predicted deleterious variants, we identified 29 genes showing promising associations with breast cancer risk. We replicated the association for two genes, OGDHL and BRCA2, at a Bonferroni-corrected p < 0.05, by genotyping an independent set of samples from 1,628 breast cancer cases and 1,943 controls. The association for OGDHL was primarily driven by three predicted deleterious variants (p.Val827Met, p.Pro839Leu, p.Phe836Ser; p < 0.01 for all). For BRCA2, we characterized a total of 27 disruptive variants, including 18 nonsense, six frameshift and three splicing variants, whereas they were only detected in cases, but none of the controls. All of these variants were either very rare (AF < 0.1%) or not detected in >4,500 East Asian women from the genome Aggregation database (gnomAD), providing additional support to our findings. Our study revealed a potential novel gene and multiple disruptive variants of BRCA2 for breast cancer risk, which may identify high-risk women in Chinese populations.
© 2019 UICC.

BACKGROUND - CaM (calmodulin) mutations are associated with congenital arrhythmia susceptibility (calmodulinopathy) and are most often de novo. In this report, we sought to broaden the genotype-phenotype spectrum of calmodulinopathies with 2 novel calmodulin mutations and to investigate mosaicism in 2 affected families.
METHODS - CaM mutations were identified in 4 independent cases by DNA sequencing. Biochemical and electrophysiological studies were performed to determine functional consequences of each mutation.
RESULTS - Genetic studies identified 2 novel CaM variants (-E141K in 2 cases; -E141V) and one previously reported CaM pathogenic variant (-D130G) among 4 probands with shared clinical features of prolonged QTc interval (range 505-725 ms) and documented ventricular arrhythmia. A fatal outcome occurred for 2 of the cases. The parents of all probands were asymptomatic with normal QTc duration. However, 2 of the families had multiple affected offspring or multiple occurrences of intrauterine fetal demise. The mother from the family with recurrent intrauterine fetal demise exhibited the -E141K mutant allele in 25% of next-generation sequencing reads indicating somatic mosaicism, whereas -D130G was present in 6% of captured molecules of the paternal DNA sample, also indicating mosaicism. Two novel mutations (E141K and E141V) impaired Ca binding affinity to the C-domain of CaM. Human-induced pluripotent stem cell-derived cardiomyocytes overexpressing mutant or wild-type CaM showed that both mutants impaired Ca-dependent inactivation of L-type Ca channels and prolonged action potential duration.
CONCLUSIONS - We report 2 families with somatic mosaicism associated with arrhythmogenic calmodulinopathy, and demonstrate dysregulation of L-type Ca channels by 2 novel CaM mutations affecting the same residue. Parental mosaicism should be suspected in families with unexplained fetal arrhythmia or fetal demise combined with a documented CaM mutation.

BACKGROUND - Mutations in SLC6A1 have been associated mainly with myoclonic atonic epilepsy (MAE) and intellectual disability. We identified a novel missense mutation in a patient with Lennox-Gastaut syndrome (LGS) characterized by severe seizures and developmental delay.
METHODS - Exome Sequencing was performed in an epilepsy patient cohort. The impact of the mutation was evaluated by H γ-aminobutyric acid (GABA) uptake, structural modeling, live cell microscopy, cell surface biotinylation and a high-throughput assay flow cytometry in both neurons and non neuronal cells.
RESULTS - We discovered a heterozygous missense mutation (c700G to A [pG234S) in the SLC6A1 encoding GABA transporter 1 (GAT-1). Structural modeling suggests the mutation destabilizes the global protein conformation. With transient expression of enhanced yellow fluorescence protein (YFP) tagged rat GAT-1 cDNAs, we demonstrated that the mutant GAT-1(G234S) transporter had reduced total protein expression in both rat cortical neurons and HEK 293 T cells. With a high-throughput flow cytometry assay and live cell surface biotinylation, we demonstrated that the mutant GAT-1(G234S) had reduced cell surface expression. H radioactive labeling GABA uptake assay in HeLa cells indicated a reduced function of the mutant GAT-1(G234S).
CONCLUSIONS - This mutation caused instability of the mutant transporter protein, which resulted in reduced cell surface and total protein levels. The mutation also caused reduced GABA uptake in addition to reduced protein expression, leading to reduced GABA clearance, and altered GABAergic signaling in the brain. The impaired trafficking and reduced GABA uptake function may explain the epilepsy phenotype in the patient.
Copyright © 2019. Published by Elsevier Inc.

Electrocardiographic PR interval measures atrio-ventricular depolarization and conduction, and abnormal PR interval is a risk factor for atrial fibrillation and heart block. Our genome-wide association study of over 92,000 European-descent individuals identifies 44 PR interval loci (34 novel). Examination of these loci reveals known and previously not-yet-reported biological processes involved in cardiac atrial electrical activity. Genes in these loci are over-represented in cardiac disease processes including heart block and atrial fibrillation. Variants in over half of the 44 loci were associated with atrial or blood transcript expression levels, or were in high linkage disequilibrium with missense variants. Six additional loci were identified either by meta-analysis of ~105,000 African and European-descent individuals and/or by pleiotropic analyses combining PR interval with heart rate, QRS interval, and atrial fibrillation. These findings implicate developmental pathways, and identify transcription factors, ion-channel genes, and cell-junction/cell-signaling proteins in atrio-ventricular conduction, identifying potential targets for drug development.

We conducted the largest investigation of predisposition variants in cancer to date, discovering 853 pathogenic or likely pathogenic variants in 8% of 10,389 cases from 33 cancer types. Twenty-one genes showed single or cross-cancer associations, including novel associations of SDHA in melanoma and PALB2 in stomach adenocarcinoma. The 659 predisposition variants and 18 additional large deletions in tumor suppressors, including ATM, BRCA1, and NF1, showed low gene expression and frequent (43%) loss of heterozygosity or biallelic two-hit events. We also discovered 33 such variants in oncogenes, including missenses in MET, RET, and PTPN11 associated with high gene expression. We nominated 47 additional predisposition variants from prioritized VUSs supported by multiple evidences involving case-control frequency, loss of heterozygosity, expression effect, and co-localization with mutations and modified residues. Our integrative approach links rare predisposition variants to functional consequences, informing future guidelines of variant classification and germline genetic testing in cancer.
Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

The spatial distribution of genetic variation within proteins is shaped by evolutionary constraint and provides insight into the functional importance of protein regions and the potential pathogenicity of protein alterations. Here, we comprehensively evaluate the 3D spatial patterns of human germline and somatic variation in 6,604 experimentally derived protein structures and 33,144 computationally derived homology models covering 77% of all human proteins. Using a systematic approach, we quantify differences in the spatial distributions of neutral germline variants, disease-causing germline variants, and recurrent somatic variants. Neutral missense variants exhibit a general trend toward spatial dispersion, which is driven by constraint on core residues. In contrast, germline disease-causing variants are generally clustered in protein structures and form clusters more frequently than recurrent somatic variants identified from tumor sequencing. In total, we identify 215 proteins with significant spatial constraints on the distribution of disease-causing missense variants in experimentally derived protein structures, only 65 (30%) of which have been previously reported. This analysis identifies many clusters not detectable from sequence information alone; only 12% of proteins with significant clustering in 3D were identified from similar analyses of linear protein sequence. Furthermore, spatial analyses of mutations in homology-based structural models are highly correlated with those from experimentally derived structures, supporting the use of computationally derived models. Our approach highlights significant differences in the spatial constraints on different classes of mutations in protein structure and identifies regions of potential function within individual proteins.
Copyright © 2018 American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

BACKGROUND - Next-generation sequencing of individuals with genetic diseases often detects candidate rare variants in numerous genes, but determining which are causal remains challenging. We hypothesized that the spatial distribution of missense variants in protein structures contains information about function and pathogenicity that can help prioritize variants of unknown significance (VUS) and elucidate the structural mechanisms leading to disease.
RESULTS - To illustrate this approach in a clinical application, we analyzed 13 candidate missense variants in regulator of telomere elongation helicase 1 (RTEL1) identified in patients with Familial Interstitial Pneumonia (FIP). We curated pathogenic and neutral RTEL1 variants from the literature and public databases. We then used homology modeling to construct a 3D structural model of RTEL1 and mapped known variants into this structure. We next developed a pathogenicity prediction algorithm based on proximity to known disease causing and neutral variants and evaluated its performance with leave-one-out cross-validation. We further validated our predictions with segregation analyses, telomere lengths, and mutagenesis data from the homologous XPD protein. Our algorithm for classifying RTEL1 VUS based on spatial proximity to pathogenic and neutral variation accurately distinguished 7 known pathogenic from 29 neutral variants (ROC AUC = 0.85) in the N-terminal domains of RTEL1. Pathogenic proximity scores were also significantly correlated with effects on ATPase activity (Pearson r = -0.65, p = 0.0004) in XPD, a related helicase. Applying the algorithm to 13 VUS identified from sequencing of RTEL1 from patients predicted five out of six disease-segregating VUS to be pathogenic. We provide structural hypotheses regarding how these mutations may disrupt RTEL1 ATPase and helicase function.
CONCLUSIONS - Spatial analysis of missense variation accurately classified candidate VUS in RTEL1 and suggests how such variants cause disease. Incorporating spatial proximity analyses into other pathogenicity prediction tools may improve accuracy for other genes and genetic diseases.

BRAF and MEK inhibitors have improved clinical outcomes in advanced, BRAF -mutated melanomas. Acquired resistance occurs in most patients, with numerous and diverse drivers. We obtained pretreatment and progression biopsies from a patient who progressed on dabrafenib and trametinib. In addition to a preserved BRAF mutation, an internal deletion (rearrangement) of BRAF was observed in the progression sample. This deletion involved exons 2-8, which includes the Ras-binding domain, and is analogous to previously documented BRAF fusions and splice variants known to reactivate RAS-RAF-MEK-ERK signaling. In a large cohort of melanomas, 10 additional internal deletions were identified (0.4% of all melanomas; nine of which had concurrent BRAF mutations), as well as sporadically in other tumor types. Thus, we describe a novel mechanism of resistance to BRAF and MEK inhibition.
© 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

BACKGROUND - Genome-wide association studies (GWAS) have yielded variants at >30 loci that associate with atrial fibrillation (AF), including rare coding mutations in the sarcomere genes MYH6 and MYL4.
OBJECTIVES - The aim of this study was to search for novel AF associations and in doing so gain insights into the mechanisms whereby variants affect AF risk, using electrocardiogram (ECG) measurements.
METHODS - The authors performed a GWAS of 14,255 AF cases and 374,939 controls, using whole-genome sequence data from the Icelandic population, and tested novel signals in 2,002 non-Icelandic cases and 12,324 controls. They then tested the AF variants for effect on cardiac electrical function by using measurements in 289,297 ECGs from 62,974 individuals.
RESULTS - The authors discovered 2 novel AF variants, the intergenic variant rs72700114, between the genes LINC01142 and METTL11B (risk allele frequency = 8.1%; odds ratio [OR]: 1.26; p = 3.1 × 10), and the missense variant p.Gly4098Ser in PLEC (frequency = 1.2%; OR: 1.55; p = 8.0 × 10), encoding plectin, a cytoskeletal cross-linking protein that contributes to integrity of cardiac tissue. The authors also confirmed 29 reported variants. p.Gly4098Ser in PLEC significantly affects various ECG measurements in the absence of AF. Other AF variants have diverse effects on the conduction system, ranging from none to extensive.
CONCLUSIONS - The discovery of a missense variant in PLEC affecting AF combined with recent discoveries of variants in the sarcomere genes MYH6 and MYL4 points to an important role of myocardial structure in the pathogenesis of the disease. The diverse associations between AF variants and ECG measurements suggest fundamentally different categories of mechanisms contributing to the development of AF.
Copyright © 2017 American College of Cardiology Foundation. Published by Elsevier Inc. All rights reserved.

BACKGROUND AND PURPOSE - Apoptosis-inducing factor mitochondrion-associated-1 (AIFM1) in mitochondria has captured a great deal of attention due to its well-described function in apoptosis. Mutations in AIFM1 have resulted in multiple clinical phenotypes, including X-linked Charcot-Marie-Tooth disease type 4. These syndromes usually involve multiple locations within the nervous system and/or multiple organs. This study describes a novel missense mutation in AIFM1 and its associated peripheral nerve disease.
METHODS - Patients with AIFM1 mutation were characterized clinically, electrophysiologically, genetically and by magnetic resonance imaging. The fibroblasts were isolated from the patients to study mitochondrial OXPHOS complexes.
RESULTS - We identified a family with a novel missense mutation (Phe210Leu) in AIFM1 who developed an isolated late-onset axonal polyneuropathy in which the central nervous system and other organs were spared. Interestingly, this Phe210Leu mutation resulted in abnormal assembly of mitochondrial complex I and III, and failed to disrupt AIFM1 binding with mitochondrial intermembrane space import and assembly protein 40 (MIA40) in the patients' cells. Deficiency of either AIFM1 or MIA40 is known to impair the assembly of mitochondrial complex I and IV. However, levels of both AIFM1 and MIA40 were unchanged.
CONCLUSIONS - Phe210Leu mutation in AIFM1 induces an axonal polyneuropathy that might be contributed by the misassembly of mitochondrial complex I and III. This misassembly appears to be independent of the traditional mechanism via AIFM1/MIA40 deficiency.
© 2017 EAN.