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Germline DDX41 variants are the most common mutations predisposing to acute myeloid leukemia (AML)/myelodysplastic syndrome (MDS) in adults, but the causal variant (CV) landscape and clinical spectrum of hematologic malignancies (HMs) remain unexplored. Here, we analyzed the genomic profiles of 176 patients with HM carrying 82 distinct presumably germline DDX41 variants among a group of 9821 unrelated patients. Using our proposed DDX41-specific variant classification, we identified features distinguishing 116 patients with HM with CV from 60 patients with HM with variant of uncertain significance (VUS): an older age (median 69 years), male predominance (74% in CV vs 60% in VUS, P = .03), frequent concurrent somatic DDX41 variants (79% in CV vs 5% in VUS, P < .0001), a lower somatic mutation burden (1.4 ± 0.1 in CV vs 2.9 ± 0.04 in VUS, P = .012), near exclusion of canonical recurrent genetic abnormalities including mutations in NPM1, CEBPA, and FLT3 in AML, and favorable overall survival (OS) in patients with AML/MDS. This superior OS was determined independent of blast count, abnormal karyotypes, and concurrent variants, including TP53 in patients with AML/MDS, regardless of patient’s sex, age, or specific germline CV, suggesting that germline DDX41 variants define a distinct clinical entity. Furthermore, unrelated patients with myeloproliferative neoplasm and B-cell lymphoma were linked by DDX41 CV, thus expanding the known disease spectrum. This study outlines the CV landscape, expands the phenotypic spectrum in unrelated DDX41-mutated patients, and underscores the urgent need for gene-specific diagnostic and clinical management guidelines.

While molecular testing of hematologic malignancies is now standard of care, there is variability in practice and testing capabilities between different academic laboratories, with common questions arising on how to best meet clinical expectations. A survey was sent to hematopathology subgroup members of the Genomics Organization for Academic Laboratories (GOAL) consortium to assess current and future practice and potentially establish a reference for peer institutions. Responses were received from eighteen academic tertiary-care laboratories regarding next-generation sequencing (NGS) panel design, sequencing protocols and metrics, assay characteristics, laboratory operations, case reimbursement, and development plans. Differences in NGS panel size, use, and gene content were reported. Gene content for myeloid processes was reported to be generally excellent while genes for lymphoid processes were less well covered. Turnaround time (TAT) for acute cases including acute myeloid leukemia was reported to range from 2-7 to 15-21 calendar days with different approaches to achieving rapid TAT described. To help guide NGS panel design and standardize gene content, consensus gene lists based on current and future NGS panels in development were generated. Most survey respondents expected molecular testing at academic laboratories to continue to be viable in the future with rapid TAT for acute cases likely to remain an important factor. Molecular testing reimbursement was reported to be a major concern. The results of this survey and subsequent discussions improve the shared understanding of differences in testing practices for hematologic malignancies between institutions and will help provide a more consistent level of patient care.

Genetic disorders presenting in the neonatal period can have a significant impact on morbidity and mortality. Early diagnosis can facilitate timely prognostic counseling to families and possibility of precision care, which could improve outcome. As availability of diagnostic testing expands, the required knowledge base of the neonatologist must also expand to include proper application and understanding of genetic testing modalities, especially where availability of clinical genetics consultation is limited. Herein, we review genetic tests utilized in the neonatal intensive care unit (NICU) providing background on the technology, clinical indications, advantages, and limitations of the tests. This review will span from classic cytogenetics to the evolving role of next generation sequencing and its impact on the management of neonatal disease.

Hundreds of genes are implicated in autism spectrum disorder (ASD) but the mechanisms through which they contribute to ASD pathophysiology remain elusive. Here, we analyzed leukocyte transcriptomics from 1-4 year-old male toddlers with ASD or typical development from the general population. We discovered a perturbed gene network that includes genes that are highly expressed during fetal brain development and which is dysregulated in hiPSC-derived neuron models of ASD. High-confidence ASD risk genes emerge as upstream regulators of the network, and many risk genes may impact the network by modulating RAS/ERK, PI3K/AKT, and WNT/β-catenin signaling pathways. We found that the degree of dysregulation in this network correlated with the severity of ASD symptoms in the toddlers. These results demonstrate how the heterogeneous genetics of ASD may dysregulate a core network to influence brain development at prenatal and very early postnatal ages and, thereby, the severity of later ASD symptoms.

Background Both RNA-Seq and sample freeze-thaw are ubiquitous. However, knowledge about the impact of freeze-thaw on downstream analyses is limited. The lack of common quality metrics that are sufficiently sensitive to freeze-thaw and RNA degradation, e.g. the RNA Integrity Score, makes such assessments challenging. Results Here we quantify the impact of repeated freeze-thaw cycles on the reliability of RNA-Seq by examining poly(A)-enriched and ribosomal RNA depleted RNA-seq from frozen leukocytes drawn from a toddler Autism cohort. To do so, we estimate the relative noise, or percentage of random counts, separating technical replicates. Using this approach we measured noise associated with RIN and freeze-thaw cycles. As expected, RIN does not fully capture sample degradation due to freeze-thaw. We further examined differential expression results and found that three freeze-thaws should extinguish the differential expression reproducibility of similar experiments. Freeze-thaw also resulted in a 3′ shift in the read coverage distribution along the gene body of poly(A)-enriched samples compared to ribosomal RNA depleted samples, suggesting that library preparation may exacerbate freeze-thaw-induced sample degradation. Conclusion The use of poly(A)-enrichment for RNA sequencing is pervasive in library preparation of frozen tissue, and thus, it is important during experimental design and data analysis to consider the impact of repeated freeze-thaw cycles on reproducibility. Graphical abstract

BackgroundTumor mutation burden (TMB) is a biomarker frequently reported by clinical laboratories, which is derived by quantifying of the number of single nucleotide or indel variants (mutations) identified by next-generation sequencing of tumors. TMB values can inform prognosis or predict the response of a patient’s tumor to immune checkpoint inhibitor therapy. Methods for the calculation of TMB are not standardized between laboratories, with significant variables being the gene content of the panels sequenced and the inclusion or exclusion of synonymous variants in the calculations. The impact of these methodological differences has not been investigated and the concordance of reported TMB values between laboratories is unknown.MethodsSequence variant lists from more than 9000 tumors of various types were downloaded from The Cancer Genome Atlas. Variant lists were filtered to include only appropriate variant types (ie, non-synonymous only or synonymous and non-synonymous variants) within the genes found in five commonly used targeted solid tumor gene panels as well as an in-house gene panel. Calculated TMB was paired with corresponding overall survival (OS) data of each patient.ResultsRegression analysis indicates high concordance of TMB as derived from the examined panels. TMB derived from panels was consistently and significantly lower than that derived from a whole exome. TMB, as derived from whole exome or the examined panels, showed a significant correlation with OS in the examined data.ConclusionsTMB derived from the examined gene panels was analytically equivalent between panels, but not between panels and whole-exome sequencing. Correlation between TMB and OS is significant if TMB method-specific cut-offs are used. These results suggest that TMB values, as derived from the gene panels examined, are analytically and prognostically equivalent.

The co-occurrence of HIV and alcohol use disorder (AUD) amplifies risk for neural injury and neurocognitive deficits. However, the substantial neurocognitive heterogeneity across HIV+/AUD+ individuals suggests inter-individual differences in vulnerability to the neurotoxicity of comorbid HIV/AUD. Genetic variation in alcohol dehydrogenase (ADH), which metabolizes ethanol, may contribute to inter-individual neurocognitive variability. We evaluated associations between five ADH single-nucleotide polymorphisms (SNPs) and neurocognition in men stratified by HIV and lifetime AUD status. Neurobehavioral assessments were administered to 153 men. Three-way ANOVAs examined the interaction of HIV, AUD, and ADH SNPs on global and domain-specific demographically corrected T scores. Follow-up ANCOVAs adjusted for age, estimated verbal IQ, depression, and remote non-alcohol substance use disorders. HIV/AUD groups differed globally and for verbal fluency, working memory, executive function, and processing speed T scores specifically, with HIV+/AUD+ exhibiting the poorest performance. ADH4 (rs1126671) was associated with large effects on working memory (d = - 1.16, p = .001) and executive function (d = - 0.77, p = .028) selectively in HIV+/AUD+, which remained significant in ANCOVA models. ADH1A (rs3819197) moderated the deleterious effects of HIV+/AUD+ on processing speed such that HIV+/AUD+ related to slower information processing in A allele carriers but not GG homozygotes (ps

This multi-institutional study was undertaken to evaluate interrater reliability of the 2017 Association for Molecular Pathology/American Society of Clinical Oncology/College of American Pathologists guidelines for interpretation and reporting of oncology sequence variants and to assess current practices and perceptions surrounding these guidelines. Fifty-one variants were distributed to 20 participants from 10 institutions for classification using the new guidelines. Agreement was assessed using chance-corrected agreement (Cohen κ). κ was 0.35. To evaluate if data sharing could help resolve disagreements, a summary of variant classifications and additional information about each variant were distributed to all participants. κ improved to 0.7 after the original classifications were revised. Participants were invited to take a web-based survey regarding their perceptions of the guidelines. Only 20% (n = 3) of the survey respondents had prior experience with the guidelines in clinical practice. The main perceived barriers to guideline implementation included the complexity of the guidelines, discordance between clinical actionability and pathobiologic relevance, lack of familiarity with the new classifications, and uncertainty when applying criteria to potential germline variants. This study demonstrates noteworthy discordances between pathologists for variant classification in solid tumors when using the 2017 Association for Molecular Pathology/American Society of Clinical Oncology/College of American Pathologists guidelines. These findings highlight potential areas for clarification/refinement before mainstream clinical adoption.

The use of next generation sequencing (NGS) to profile tumor genomes for the presence of diagnostic, prognostic, or therapeutically targetable variants is revolutionizing the practice of oncology and is increasingly utilized in clinical laboratory settings. Beginning with the isolation of DNA of sufficient quality and quantity from a tumor specimen, the creation of a library of genomic fragments representing the portion of the genome of interest, ranging from a few genes to the entire exome, is the first step required in the sequencing process. Fixed tumor tissue in the form of a tissue block is the most commonly encountered specimen for analysis in a clinical setting. Special precautions must be employed to ensure that material isolated from these specimens is suitable for use. Once DNA is obtained, one of the most commonly used methods for library preparation involves fluid phase hybridization-based capture of the genomic regions to be interrogated. This multistep process involves fragmentation of the DNA to a uniform size distribution, ligating adapter molecules which are labeled with specific barcodes to enable downstream sequencing and sample identification, and the use of a multiplexed pool of biotinylated single stranded RNA or DNA hybridization probes to recognize and capture the targeted genomic regions. Fragments which are not specifically captured during the hybridization process are removed via a series of wash steps, and a final low cycle amplification is used to prepare the library of captured fragments for sequencing. In this chapter, we provide a step-by-step guide to the preparation of fixed tissue-derived DNA libraries for sequencing via the Illumina process and highlight some of the precautions necessary when working with these types of specimens.

The use of next-generation sequencing technologies has enabled the analysis of a wide spectrum of somatic mutations in tumors. This analysis can be carried out using various strategies including the use of small panels of focused, clinically actionable genes, large panels of cancer-related genes, whole exomes, and the entire genome. One of the main goals in these analyses is to identify key mutations in these tumors that drive the oncogenic process. Depending on the gene, mutations can have altering effects, such as loss of function mutations in tumor suppressor genes, to mutations that activate genes such as kinases involved with cell cycle progression or proliferation. Once the sequencing process is complete, and the alignment of the large collection of reads to the reference genome and variant calling has been carried out, one is left with a large collection of variants. The challenge then becomes assigning where the variant resides in the genome with respect to coding regions, splice site regions, regulatory regions, and what potential functional effect these variants may have on the resulting protein. Other helpful information includes determining if the variant has been identified before, and if so, the tumor type associated with the variant. In addition, if the tumor profiling experiment is not conducted with a matched specimen representing the inherited genome, various tools are helpful to determine if the variant is likely to be an inherited polymorphism or a somatic event. In this chapter, we review the various tools available for annotating variants to assist in filtering down and prioritizing the hundreds to thousands of variants down to the key variants likely to be driver mutations and relevant to the tumor being profiled.

The use of next-generation sequencing and hybridization-based capture for target enrichment have enabled the interrogation of coding regions of several clinically significant cancer genes in tumor specimens using both targeted panels of a few to hundreds of genes, to whole-exome panels encompassing coding regions of all genes in the genome. Next-generation sequencing (NGS) technologies produce millions of relatively short segments of sequences or reads that require bioinformatics tools to map reads back to a reference genome using various read alignment tools, as well as to determine differences between single bases (single nucleotide variants or SNVs) or multiple bases (insertions and deletions or indels) between the aligned reads and the reference genome to call variants. In addition to single nucleotide changes or small insertions and deletions, high copy gains and losses can also be gleaned from NGS data to call gene amplifications and deletions. Throughout these processes, numerous quality control metrics can be assessed at each step to ensure that the resulting called variants are of high quality and are accurate. In this chapter we review common tools used to generate reads from Illumina-derived sequence data, align reads, and call variants from hybridization-based targeted NGS panel data generated from tumor FFPE-derived DNA specimens as well as basic quality metrics to assess for each assayed specimen.

The use of next-generation sequencing technologies has enabled the analysis of a wide spectrum of somatic mutations in tumors. This analysis can be carried out using various strategies including the use of small panels of focused, clinically actionable genes, large panels of cancer-related genes, whole exomes, and the entire genome. One of the main goals in these analyses is to identify key mutations in these tumors that drive the oncogenic process. Depending on the gene, mutations can have altering effects, such as loss of function mutations in tumor suppressor genes, to mutations that activate genes such as kinases involved with cell cycle progression or proliferation. Once the sequencing process is complete, and the alignment of the large collection of reads to the reference genome and variant calling has been carried out, one is left with a large collection of variants. The challenge then becomes assigning where the variant resides in the genome with respect to coding regions, splice site regions, regulatory regions, and what potential functional effect these variants may have on the resulting protein. Other helpful information includes determining if the variant has been identified before, and if so, the tumor type associated with the variant. In addition, if the tumor profiling experiment is not conducted with a matched specimen representing the inherited genome, various tools are helpful to determine if the variant is likely to be an inherited polymorphism or a somatic event. In this chapter, we review the various tools available for annotating variants to assist in filtering down and prioritizing the hundreds to thousands of variants down to the key variants likely to be driver mutations and relevant to the tumor being profiled.

The use of next generation sequencing (NGS) to profile tumor genomes for the presence of diagnostic, prognostic, or therapeutically targetable variants is revolutionizing the practice of oncology and is increasingly utilized in clinical laboratory settings. Beginning with the isolation of DNA of sufficient quality and quantity from a tumor specimen, the creation of a library of genomic fragments representing the portion of the genome of interest, ranging from a few genes to the entire exome, is the first step required in the sequencing process. Fixed tumor tissue in the form of a tissue block is the most commonly encountered specimen for analysis in a clinical setting. Special precautions must be employed to ensure that material isolated from these specimens is suitable for use. Once DNA is obtained, one of the most commonly used methods for library preparation involves fluid phase hybridization-based capture of the genomic regions to be interrogated. This multistep process involves fragmentation of the DNA to a uniform size distribution, ligating adapter molecules which are labeled with specific barcodes to enable downstream sequencing and sample identification, and the use of a multiplexed pool of biotinylated single stranded RNA or DNA hybridization probes to recognize and capture the targeted genomic regions. Fragments which are not specifically captured during the hybridization process are removed via a series of wash steps, and a final low cycle amplification is used to prepare the library of captured fragments for sequencing. In this chapter, we provide a step-by-step guide to the preparation of fixed tissue-derived DNA libraries for sequencing via the Illumina process and highlight some of the precautions necessary when working with these types of specimens.

The use of next-generation sequencing and hybridization-based capture for target enrichment have enabled the interrogation of coding regions of several clinically significant cancer genes in tumor specimens using both targeted panels of a few to hundreds of genes, to whole-exome panels encompassing coding regions of all genes in the genome. Next-generation sequencing (NGS) technologies produce millions of relatively short segments of sequences or reads that require bioinformatics tools to map reads back to a reference genome using various read alignment tools, as well as to determine differences between single bases (single nucleotide variants or SNVs) or multiple bases (insertions and deletions or indels) between the aligned reads and the reference genome to call variants. In addition to single nucleotide changes or small insertions and deletions, high copy gains and losses can also be gleaned from NGS data to call gene amplifications and deletions. Throughout these processes, numerous quality control metrics can be assessed at each step to ensure that the resulting called variants are of high quality and are accurate. In this chapter we review common tools used to generate reads from Illumina-derived sequence data, align reads, and call variants from hybridization-based targeted NGS panel data generated from tumor FFPE-derived DNA specimens as well as basic quality metrics to assess for each assayed specimen.

International audience; C-reactive protein (CRP) is a sensitive biomarker of chronic low-grade inflammation and is associated with multiple complex diseases. The genetic determinants of chronic inflammation remain largely unknown, and the causal role of CRP in several clinical outcomes is debated. We performed two genome-wide association studies (GWASs), on HapMap and 1000 Genomes imputed data, of circulating amounts of CRP by using data from 88 studies comprising 204,402 European individuals. Additionally, we performed in silico functional analyses and Mendelian randomization analyses with several clinical outcomes. The GWAS meta-analyses of CRP revealed 58 distinct genetic loci (p < 5 × 10-8). After adjustment for body mass index in the regression analysis, the associations at all except three loci remained. The lead variants at the distinct loci explained up to 7.0% of the variance in circulating amounts of CRP. We identified 66 gene sets that were organized in two substantially correlated clusters, one mainly composed of immune pathways and the other characterized by metabolic pathways in the liver. Mendelian randomization analyses revealed a causal protective effect of CRP on schizophrenia and a risk-increasing effect on bipolar disorder. Our findings provide further insights into the biology of inflammation and could lead to interventions for treating inflammation and its clinical consequences.Copyright © 2018 American Society of Human Genetics. All rights reserved.

INTRODUCTION: The Val allele of the Val158Met single-nucleotide polymorphism of the catechol-o-methyltransferase gene (COMT) confers greater catabolism of dopamine (DA) in the prefrontal cortex (PFC) than the Met allele. Met/Met homozygotes typically outperform Val-carriers on tests of executive function (EF), perhaps resulting from increased DA bioavailability. Methamphetamine (METH) causes large releases of DA, which is associated with neurotoxicity and executive dysfunction in chronic METH users. We hypothesized that, contrary to its effect in non-METH-using populations, slower DA clearance conferred by Met/Met will relate to worse EF in METH users. METHODS: 149 non-Hispanic White men, stratified by METH dependence (METH+/−) and COMT (Val/Val, Val/Met, Met/Met), completed three tests of EF: Wisconsin Card Sorting Test (WCST), Stroop Color-Word Test (Stroop), and Trail Making Test Part B (Trails B). Demographically-adjusted test scores were averaged to create an EF composite T-score. We examined the interaction of METH and COMT on the EF composite and individual test T-scores, controlling for premorbid functioning and alcohol use. RESULTS: METH group differences in EF were evident only among Met/Met carriers (beta = −9.36, p

Genome-wide association studies of case-control status have advanced the understanding of the genetic basis of psychiatric disorders. Further progress may be gained by increasing sample size but also by new analysis strategies that advance the exploitation of existing data, especially for clinically important quantitative phenotypes. The functionally-informed efficient region-based test strategy (FIERS) introduced herein uses prior knowledge on biological function and dependence of genotypes within a powerful statistical framework with improved sensitivity and specificity for detecting consistent genetic effects across studies. As proof of concept, FIERS was used for the first genome-wide single nucleotide polymorphism (SNP)-based investigation on bipolar disorder (BD) that focuses on an important aspect of disease course, the functional outcome. FIERS identified a significantly associated locus on chromosome 15 (hg38: chr15:48965004 - 49464789 bp) with consistent effect strength between two independent studies (GAIN/TGen: European Americans, BOMA: Germans; n = 1592 BD patients in total). Protective and risk haplotypes were found on the most strongly associated SNPs. They contain a CTCF binding site (rs586758); CTCF sites are known to regulate sets of genes within a chromatin domain. The rs586758 - rs2086256 - rs1904317 haplotype is located in the promoter flanking region of the COPS2 gene, close to microRNA4716, and the EID1, SHC4, DTWD1 genes as plausible biological candidates. While implication with BD is novel, COPS2, EID1, and SHC4 are known to be relevant for neuronal differentiation and function and DTWD1 for psychopharmacological side effects. The test strategy FIERS that enabled this discovery is equally applicable for tag SNPs and sequence data. peerReviewed

© 2015 Nature America, Inc. All rights reserved. Socioeconomic disparities are associated with differences in cognitive development. The extent to which this translates to disparities in brain structure is unclear. We investigated relationships between socioeconomic factors and brain morphometry, independently of genetic ancestry, among a cohort of 1,099 typically developing individuals between 3 and 20 years of age. Income was logarithmically associated with brain surface area. Among children from lower income families, small differences in income were associated with relatively large differences in surface area, whereas, among children from higher income families, similar income increments were associated with smaller differences in surface area. These relationships were most prominent in regions supporting language, reading, executive functions and spatial skills; surface area mediated socioeconomic differences in certain neurocognitive abilities. These data imply that income relates most strongly to brain structure among the most disadvantaged children.

© 2014 The Authors. Objective: The diagnostic boundaries of sleep disorders are under considerable debate. The main sleep disorders are partly heritable therefore, defining heritable pathophysiologic mechanisms could delineate diagnoses and suggest treatment. We collected clinical data and DNA from consenting patients scheduled to undergo clinical polysomnograms, to expand our understanding of the polymorphisms associated with the phenotypes of particular sleep disorders. Methods: Patients at least 21 years of age were recruited to contribute research questionnaires, and to provide access to their medical records, saliva for deoxyribonucleic acid (DNA), and polysomnographic data. From these complex data, 38 partly overlapping phenotypes were derived indicating complaints, subjective and objective sleep timing, and polysomnographic disturbances. A custom chip was used to genotype 768 single-nucleotide polymorphisms (SNPs). Additional assays derived ancestry-informative markers (eg, 751 participants of European ancestry). Linear regressions controlling for age, gender, and ancestry were used to assess the associations of each phenotype with each of the SNPs, highlighting those with Bonferroni-corrected significance. Results: In peroxisome proliferator-activated receptor gamma, coactivator 1 beta (PPARGC1B), rs6888451 was associated with several markers of obstructive sleep apnea. In aryl hydrocarbon receptor nuclear translocator-like (ARNTL), rs10766071 was associated with decreased polysomnographic sleep duration. The association of rs3923809 in BTBD9 with periodic limb movements in sleep was confirmed. SNPs in casein kinase 1 delta (CSNK1D rs11552085), cryptochrome 1 (CRY1 rs4964515), and retinoic acid receptor-related orphan receptor A (RORA rs11071547) were less persuasively associated with sleep latency and time of falling asleep. Conclusions: SNPs associated with several sleep phenotypes were suggested, but due to risks of false discovery, independent replications are needed before the importance of these associations can be assessed, followed by investigation of molecular mechanisms.

Background Complex biological networks of atherosclerosis are largely unknown. Objective The main objective of the Genetic Loci and the Burden of Atherosclerotic Lesions study is to assemble comprehensive biological networks of atherosclerosis using advanced cardiovascular imaging for phenotyping, a panomic approach to identify underlying genomic, proteomic, metabolomic, and lipidomic underpinnings, analyzed by systems biology–driven bioinformatics. Methods By design, this is a hypothesis-free unbiased discovery study collecting a large number of biologically related factors to examine biological associations between genomic, proteomic, metabolomic, lipidomic, and phenotypic factors of atherosclerosis. The Genetic Loci and the Burden of Atherosclerotic Lesions study ( NCT01738828 ) is a prospective, multicenter, international observational study of atherosclerotic coronary artery disease. Approximately 7500 patients are enrolled and undergo non–contrast-enhanced coronary calcium scanning by CT for the detection and quantification of coronary artery calcium, as well as coronary artery CT angiography for the detection and quantification of plaque, stenosis, and overall coronary artery disease burden. In addition, patients undergo whole genome sequencing, DNA methylation, whole blood–based transcriptome sequencing, unbiased proteomics based on mass spectrometry, as well as metabolomics and lipidomics on a mass spectrometry platform. The study is analyzed in 3 subsequent phases, and each phase consists of a discovery cohort and an independent validation cohort. For the primary analysis, the primary phenotype will be the presence of any atherosclerotic plaque, as detected by cardiac CT. Additional phenotypic analyses will include per patient maximal luminal stenosis defined as 50% and 70% diameter stenosis. Single-omic and multi-omic associations will be examined for each phenotype; putative biomarkers will be assessed for association, calibration, discrimination, and reclassification.

Waist circumference (WC) and waist-to-hip ratio (WHR) are surrogate measures of central adiposity that are associated with adverse cardiovascular events, type 2 diabetes and cancer independent of body mass index (BMI). WC and WHR are highly heritable with multiple susceptibility loci identified to date. We assessed the association between SNPs and BMI-adjusted WC and WHR and unadjusted WC in up to 57 412 individuals of European descent from 22 cohorts collaborating with the NHLBI's Candidate Gene Association Resource (CARe) project. The study population consisted of women and men aged 20-80 years. Study participants were genotyped using the ITMAT/Broad/CARE array, which includes ∼50 000 cosmopolitan tagged SNPs across ∼2100 cardiovascular-related genes. Each trait was modeled as a function of age, study site and principal components to control for population stratification, and we conducted a fixed-effects meta-analysis. No new loci for WC were observed. For WHR analyses, three novel loci were significantly associated (P < 2.4 × 10(-6)). Previously unreported rs2811337-G near TMCC1 was associated with increased WHR (β ± SE, 0.048 ± 0.008, P = 7.7 × 10(-9)) as was rs7302703-G in HOXC10 (β = 0.044 ± 0.008, P = 2.9 × 10(-7)) and rs936108-C in PEMT (β = 0.035 ± 0.007, P = 1.9 × 10(-6)). Sex-stratified analyses revealed two additional novel signals among females only, rs12076073-A in SHC1 (β = 0.10 ± 0.02, P = 1.9 × 10(-6)) and rs1037575-A in ATBDB4 (β = 0.046 ± 0.01, P = 2.2 × 10(-6)), supporting an already established sexual dimorphism of central adiposity-related genetic variants. Functional analysis using ENCODE and eQTL databases revealed that several of these loci are in regulatory regions or regions with differential expression in adipose tissue.

Blood pressure (BP) is a heritable risk factor for cardiovascular disease. To investigate genetic associations with systolic BP (SBP), diastolic BP (DBP), mean arterial pressure (MAP), and pulse pressure (PP), we genotyped ∼50,000 SNPs in up to 87,736 individuals of European ancestry and combined these in a meta-analysis. We replicated findings in an independent set of 68,368 individuals of European ancestry. Our analyses identified 11 previously undescribed associations in independent loci containing 31 genes including PDE1A, HLA-DQB1, CDK6, PRKAG2, VCL, H19, NUCB2, RELA, HOXC@ complex, FBN1, and NFAT5 at the Bonferroni-corrected array-wide significance threshold (p < 6 × 10-7) and confirmed 27 previously reported associations. Bioinformatic analysis of the 11 loci provided support for a putative role in hypertension of several genes, such as CDK6 and NUCB2. Analysis of potential pharmacological targets in databases of small molecules showed that ten of the genes are predicted to be a target for small molecules. In summary, we identified previously unknown loci associated with BP. Our findings extend our understanding of genes involved in BP regulation, which may provide new targets for therapeutic intervention or drug response stratification. © 2014 The American Society of Human Genetics.

Philadelphia chromosome negative myeloproliferative neoplasms (MPNs) are characterized by the overproduction of mature blood cells and variable bone marrow fibrosis. MPNs attributed to dysregulation of the Janus kinase 2 (JAK2) pathway include polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis (PMF). Somatic mutations in JAK2, thrombopoietin receptor (MPL), and calreticulin (CALR) have been identified as driver mutations with direct or upstream upregulation of JAK2. CALR mutations are the most recently described, with the two most common mutations being a 52-base pair (bp) deletion (type 1) or 5 bp insertion (type2) in exon 9. Studies have shown a prognostic advantage to type1/type 1 like CALR driven MPNs over JAK2, MPL, and type2 CALR driven MPNs. Rarer CALR exon 9 mutations have also been identified in presumed MPN patients negative for JAK2 and MPL mutations. In these cases variable predicted changes to the CALR protein have resulted in speculative interpretations as to their relevance in the diagnosis of a suspected MPN. Here we report a patient with a longstanding history of myelofibrosis, thrombocytosis, and anemia, eventually determined to have an in-frame presumed germline (due to variant fraction and identification in the patient's child) CALR mutation downstream of a somatic type 1 CALR mutation. The overall compounded alterations generate a type 1 like mutation previously not described (to the best of our knowledge) in the literature. The patient is a 70-year old female noted to be persistently anemic all her life. While a bone marrow assessment was recommended early in life, the patient declined workup until a marrow biopsy was eventually performed at age 50. The biopsy reportedly showed mild marrow fibrosis and the patient was trailed on erythropoietin for her anemia with little benefit. At age 59 the patient was noted with thrombocytosis (478 X 109/L) and mild splenomegaly. Repeat marrow biopsy showed hypercellular marrow, marked fibrosis (WHO grade 3/3), and megakaryocyte clustering. JAK2 was noted to not be mutated. Over the next decade the patient developed symptomatic splenomegly and continued to be anemic with eventual intermittent transfusion requirement at age 65, pushing her risk to DIPPS-plus intermediate-2. During this period successive treatments included Revlimid, trial sonic hedgehog (shh) inhibitor, and JAK2 inhibitors with intervening multiyear long spans without treatment. Marrow fibrosis over this time was predominantly unchanged on the various therapies but symptomatic splenomegaly decreased on shh and JAK2 inhibitors. At present the patient requires intermittent transfusions and is on a JAK2 inhibitor. Recent NGS testing of marrow aspirate identified an in-frame presumed germline CALR mutation downstream of a compound somatic type 1 CALR mutation. A high molecular risk ASXL1 mutation was also identified. The in-frame CALR mutation results in a 9bp in frame deletion (c.1191_1199del, p.398_400delGluGluAsp), which has been reported at least 10 times in the literature. The various reports have one event mentioned as being presumably germline and non-pathogenic, while the other reports are equivocal to presumed pathogenic in light of negative JAK2 and MPL mutations in patients with clinical suspicion for a MPN. At our institution we have identified 4 instances of this 9bp deletion, 3 show allele fractions suggestive of being germline and 1 case with an allele fraction consistent with being a somatic mutation. In one germline case the patient also had a JAK2 V617F mutation and a diagnosis of a MPN. The other presumed germline case was found in an offspring of the patient described in this report and currently shows no signs of a MPN. The presumed somatic 9bp deletion was seen in patient with a hypocellular marrow myelodysplastic syndrome, found to also have a TET2 mutation and normal karyotype. While in-frame CALR exon 9 mutations are rare and predominantly considered germline non-pathogenic polymorphisms, there may be value in reporting such events in the context of patients with myeloid neoplasms as to not miss possible disease modifying mutations which may become apparent when aggregating multi-institution data sets. The patient highlighted in this report exhibits a strikingly long and relatively indolent disease course, notably despite an adverse prognostic risk category and high molecular risk mutation. Disclosures No relevant conflicts of interest to declare.

Written and verbal languages are neurobehavioral traits vital to the development of communication skills. Unfortunately, disorders involving these traits-specifically reading disability (RD) and language impairment (LI)-are common and prevent affected individuals from developing adequate communication skills, leaving them at risk for adverse academic, socioeconomic and psychiatric outcomes. Both RD and LI are complex traits that frequently co-occur, leading us to hypothesize that these disorders share genetic etiologies. To test this, we performed a genome-wide association study on individuals affected with both RD and LI in the Avon Longitudinal Study of Parents and Children. The strongest associations were seen with markers in ZNF385D (OR = 1.81, P = 5.45 × 10(-7) ) and COL4A2 (OR = 1.71, P = 7.59 × 10(-7) ). Markers within NDST4 showed the strongest associations with LI individually (OR = 1.827, P = 1.40 × 10(-7) ). We replicated association of ZNF385D using receptive vocabulary measures in the Pediatric Imaging Neurocognitive Genetics study (P = 0.00245). We then used diffusion tensor imaging fiber tract volume data on 16 fiber tracts to examine the implications of replicated markers. ZNF385D was a predictor of overall fiber tract volumes in both hemispheres, as well as global brain volume. Here, we present evidence for ZNF385D as a candidate gene for RD and LI. The implication of transcription factor ZNF385D in RD and LI underscores the importance of transcriptional regulation in the development of higher order neurocognitive traits. Further study is necessary to discern target genes of ZNF385D and how it functions within neural development of fluent language.

Background Genome-wide association studies have identified several genetic loci associated with variation in resting heart rate in European and Asian populations. No study has evaluated genetic variants associated with heart rate in African Americans. Objective To identify novel genetic variants associated with resting heart rate in African Americans. Methods Ten cohort studies participating in the Candidate-gene Association Resource and Continental Origins and Genetic Epidemiology Network consortia performed genome-wide genotyping of single nucleotide polymorphisms (SNPs) and imputed 2,954,965 SNPs using HapMap YRI and CEU panels in 13,372 participants of African ancestry. Each study measured the RR interval (ms) from 10-second resting 12-lead electrocardiograms and estimated RR-SNP associations using covariate-adjusted linear regression. Random-effects meta-analysis was used to combine cohort-specific measures of association and identify genome-wide significant loci ( P ≤2.5×10 −8 ). Results Fourteen SNPs on chromosome 6q22 exceeded the genome-wide significance threshold. The most significant association was for rs9320841 (+13 ms per minor allele; P = 4.98×10 −15 ). This SNP was approximately 350 kb downstream of GJA1 , a locus previously identified as harboring SNPs associated with heart rate in Europeans. Adjustment for rs9320841 also attenuated the association between the remaining 13 SNPs in this region and heart rate. In addition, SNPs in MYH6 , which have been identified in European genome-wide association study, were associated with similar changes in the resting heart rate as this population of African Americans. Conclusions An intergenic region downstream of GJA1 (the gene encoding connexin 43, the major protein of the human myocardial gap junction) and an intragenic region within MYH6 are associated with variation in resting heart rate in African Americans as well as in populations of European and Asian origin.

Blood pressure (BP) is a heritable determinant of risk for cardiovascular disease (CVD). To investigate genetic associations with systolic BP (SBP), diastolic BP (DBP), mean arterial pressure (MAP) and pulse pressure (PP), we genotyped ∼50 000 single-nucleotide polymorphisms (SNPs) that capture variation in ∼2100 candidate genes for cardiovascular phenotypes in 61 619 individuals of European ancestry from cohort studies in the USA and Europe. We identified novel associations between rs347591 and SBP (chromosome 3p25.3, in an intron of HRH1) and between rs2169137 and DBP (chromosome1q32.1 in an intron of MDM4) and between rs2014408 and SBP (chromosome 11p15 in an intron of SOX6), previously reported to be associated with MAP. We also confirmed 10 previously known loci associated with SBP, DBP, MAP or PP (ADRB1, ATP2B1, SH2B3/ATXN2, CSK, CYP17A1, FURIN, HFE, LSP1, MTHFR, SOX6) at array-wide significance (P < 2.4 × 10(-6)). We then replicated these associations in an independent set of 65 886 individuals of European ancestry. The findings from expression QTL (eQTL) analysis showed associations of SNPs in the MDM4 region with MDM4 expression. We did not find any evidence of association of the two novel SNPs in MDM4 and HRH1 with sequelae of high BP including coronary artery disease (CAD), left ventricular hypertrophy (LVH) or stroke. In summary, we identified two novel loci associated with BP and confirmed multiple previously reported associations. Our findings extend our understanding of genes involved in BP regulation, some of which may eventually provide new targets for therapeutic intervention.

Background— The PR interval, as measured by the resting, standard 12-lead ECG, reflects the duration of atrial/atrioventricular nodal depolarization. Substantial evidence exists for a genetic contribution to PR, including genome-wide association studies that have identified common genetic variants at 9 loci influencing PR in populations of European and Asian descent. However, few studies have examined loci associated with PR in African Americans. Methods and Results— We present results from the largest genome-wide association study to date of PR in 13 415 adults of African descent from 10 cohorts. We tested for association between PR (ms) and ≈2.8 million genotyped and imputed single-nucleotide polymorphisms. Imputation was performed using HapMap 2 YRI and CEU panels. Study-specific results, adjusted for global ancestry and clinical correlates of PR, were meta-analyzed using the inverse variance method. Variation in genome-wide test statistic distributions was noted within studies (λ range: 0.9–1.1), although not after genomic control correction was applied to the overall meta-analysis (λ: 1.008). In addition to generalizing previously reported associations with MEIS1, SCN5A, ARHGAP24, CAV1 , and TBX5 to African American populations at the genome-wide significance level ( P −8 ), we also identified a novel locus: ITGA9 , located in a region previously implicated in SCN5A expression. The 3p21 region harboring SCN5A also contained 2 additional independent secondary signals influencing PR ( P −8 ). Conclusions— This study demonstrates the ability to map novel loci in African Americans as well as the generalizability of loci associated with PR across populations of African, European, and Asian descent.

Background— Ethnic differences in cardiac arrhythmia incidence have been reported, with a particularly high incidence of sudden cardiac death and low incidence of atrial fibrillation in individuals of African ancestry. We tested the hypotheses that African ancestry and common genetic variants are associated with prolonged duration of cardiac repolarization, a central pathophysiological determinant of arrhythmia, as measured by the electrocardiographic QT interval. Methods and Results— First, individual estimates of African and European ancestry were inferred from genome-wide single-nucleotide polymorphism (SNP) data in 7 population-based cohorts of African Americans (n=12 097) and regressed on measured QT interval from ECGs. Second, imputation was performed for 2.8 million SNPs, and a genome-wide association study of QT interval was performed in 10 cohorts (n=13 105). There was no evidence of association between genetic ancestry and QT interval ( P =0.94). Genome-wide significant associations ( P –8 ) were identified with SNPs at 2 loci, upstream of the genes NOS1AP (rs12143842, P =2×10 –15 ) and ATP1B1 (rs1320976, P =2×10 –10 ). The most significant SNP in NOS1AP was the same as the strongest SNP previously associated with QT interval in individuals of European ancestry. Low probability values ( P –5 ) were observed for SNPs at several other loci previously identified in genome-wide association studies in individuals of European ancestry, including KCNQ1 , KCNH2 , LITAF , and PLN . Conclusions— We observed no difference in duration of cardiac repolarization with global genetic indices of African American ancestry. In addition, our genome-wide association study extends the association of polymorphisms at several loci associated with repolarization in individuals of European ancestry to include individuals of African ancestry.

© The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com. The electrocardiographic QRS duration, a measure of ventricular depolarization and conduction, is associated with cardiovascular mortality. While single nucleotide polymorphisms (SNPs) associated with QRS duration have been identified at 22 loci in populations of European descent, the genetic architecture of QRS duration in non-European populations is largely unknown. We therefore performed a genome-wide association study (GWAS) meta-analysis of QRS duration in 13,031 African Americans from ten cohorts and a transethnic GWAS meta-analysis with additional results from populations of European descent. In the African American GWAS, a single genome-wide significant SNP association was identified (rs3922844, P = 4 × 10-14) in intron 16 of SCN5A, a voltage-gated cardiac sodium channel gene. The QRS-prolonging rs3922844 C allele was also associated with decreased SCN5A RNA expression in human atrial tissue (P = 1.1 × 10-4). High density genotyping revealed that the SCN5A association region in African Americans was confined to intron 16. Transethnic GWAS meta-analysis identified novel SNP associations on chromosome 18 in MYL12A (rs1662342, P = 4.9 × 10-8) and chromosome 1 near CD1E and SPTA1 (rs7547997, P = 7.9 × 10-9). The 22 QRS loci previously identified in populations of European descent were enriched for significant SNP associations with QRS duration in African Americans (P = 9.9 × 10-7), and index SNP associations in or near SCN5A, SCN10A, CDKN1A, NFIA, HAND1, TBX5 and SETBP1 replicated in African Americans. In summary, rs3922844 was associated with QRS duration and SCN5A expression, two novel QRS loci were identified using transethnic meta-analysis, and a significant proportion of QRS-SNP associations discovered in populations of European descent were transferable to African Americans when adequate power was achieved.

A wealth of genetic associations for cardiovascular and metabolic phenotypes in humans has been accumulating over the last decade, in particular a large number of loci derived from recent genome wide association studies (GWAS). True complex disease-associated loci often exert modest effects, so their delineation currently requires integration of diverse phenotypic data from large studies to ensure robust meta-analyses. We have designed a gene-centric 50 K single nucleotide polymorphism (SNP) array to assess potentially relevant loci across a range of cardiovascular, metabolic and inflammatory syndromes. The array utilizes a "cosmopolitan" tagging approach to capture the genetic diversity across approximately 2,000 loci in populations represented in the HapMap and SeattleSNPs projects. The array content is informed by GWAS of vascular and inflammatory disease, expression quantitative trait loci implicated in atherosclerosis, pathway based approaches and comprehensive literature searching. The custom flexibility of the array platform facilitated interrogation of loci at differing stringencies, according to a gene prioritization strategy that allows saturation of high priority loci with a greater density of markers than the existing GWAS tools, particularly in African HapMap samples. We also demonstrate that the IBC array can be used to complement GWAS, increasing coverage in high priority CVD-related loci across all major HapMap populations. DNA from over 200,000 extensively phenotyped individuals will be genotyped with this array with a significant portion of the generated data being released into the academic domain facilitating in silico replication attempts, analyses of rare variants and cross-cohort meta-analyses in diverse populations. These datasets will also facilitate more robust secondary analyses, such as explorations with alternative genetic models, epistasis and gene-environment interactions.

The goal of the International HapMap Project is to determine the common patterns of DNA sequence variation in the human genome and to make this information freely available in the public domain. An international consortium is developing a map of these patterns across the genome by determining the genotypes of one million or more sequence variants, their frequencies and the degree of association between them, in DNA samples from populations with ancestry from parts of Africa, Asia and Europe. The HapMap will allow the discovery of sequence variants that affect common disease, will facilitate development of diagnostic tools, and will enhance our ability to choose targets for therapeutic intervention. © 2003 Nature Publishing Group.

Genome-wide association studies (GWASs) have identified many SNPs underlying variations in plasma-lipid levels. We explore whether additional loci associated with plasma-lipid phenotypes, such as high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), total cholesterol (TC), and triglycerides (TGs), can be identified by a dense gene-centric approach. Our meta-analysis of 32 studies in 66,240 individuals of European ancestry was based on the custom ∼50,000 SNP genotyping array (the ITMAT-Broad-CARe array) covering ∼2,000 candidate genes. SNP-lipid associations were replicated either in a cohort comprising an additional 24,736 samples or within the Global Lipid Genetic Consortium. We identified four, six, ten, and four unreported SNPs in established lipid genes for HDL-C, LDL-C, TC, and TGs, respectively. We also identified several lipid-related SNPs in previously unreported genes: DGAT2, HCAR2, GPIHBP1, PPARG, and FTO for HDL-C; SOCS3, APOH, SPTY2D1, BRCA2, and VLDLR for LDL-C; SOCS3, UGT1A1, BRCA2, UBE3B, FCGR2A, CHUK, and INSIG2 for TC; and SERPINF2, C4B, GCK, GATA4, INSR, and LPAL2 for TGs. The proportion of explained phenotypic variance in the subset of studies providing individual-level data was 9.9% for HDL-C, 9.5% for LDL-C, 10.3% for TC, and 8.0% for TGs. This large meta-analysis of lipid phenotypes with the use of a dense gene-centric approach identified multiple SNPs not previously described in established lipid genes and several previously unknown loci. The explained phenotypic variance from this approach was comparable to that from a meta-analysis of GWAS data, suggesting that a focused genotyping approach can further increase the understanding of heritability of plasma lipids. © 2012 The American Society of Human Genetics.

© 2016 American Academy of Neurology. Objective: The aims of the current study were to determine whether children with the 6 different APOE ϵ genotypes show differences in gray matter maturation, particularly for those with ϵ4 and ϵ2 alleles, which are associated with poorer outcomes in many neurologic disorders. Methods: A total of 1,187 healthy children (aged 3-20 years, 52.1% boys, 47.9% girls) with acceptable data from the cross-sectional Pediatric Imaging Neurocognition and Genetics Study were evaluated for the effects of 6 APOE ϵ genotypes on macroscopic and microscopic cortical and subcortical gray matter structures (measured with 3-tesla MRI and FreeSurfer for automated morphometry) and on cognition (NIH Toolbox). Results: Among APOE ϵ4 carriers, age-related changes in brain structures and cognition varied depending on genotype, with the smallest hippocampi in ϵ2ϵ4 children, the lowest hippocampal fractional anisotropy in younger ϵ4ϵ4 children, the largest medial orbitofrontal cortical areas in ϵ3ϵ4 children, and age-dependent thinning of the entorhinal cortex in ϵ4ϵ4 children. Younger ϵ4ϵ4 children had the lowest scores on executive function and working memory, while younger ϵ2ϵ4 children performed worse on attention tasks. Larger parietal gyri in the younger ϵ2ϵ4 children, and thinner temporal and cingulate isthmus cortices or smaller hippocampi in the younger ϵ4ϵ4 children, predicted poorer performance on attention or working memory. Conclusions: Our findings validated and extended prior smaller studies that showed altered brain development in APOE ϵ4-carrier children. The ϵ4ϵ4 and ϵ2ϵ4 genotypes may negatively influence brain development and brain aging at the extremes of age. Studying APOE ϵ polymorphisms in young children may provide the earliest indicators for individuals who might benefit from early interventions or preventive measures for future brain injuries and dementia.

© 2015 Elsevier Inc. The main objective of the multi-site Pediatric Imaging, Neurocognition, and Genetics (PING) study was to create a large repository of standardized measurements of behavioral and imaging phenotypes accompanied by whole genome genotyping acquired from typically-developing children varying widely in age (3 to 20 years). This cross-sectional study produced sharable data from 1493 children, and these data have been described in several publications focusing on brain and cognitive development. Researchers may gain access to these data by applying for an account on the PING portal and filing a data use agreement. Here we describe the recruiting and screening of the children and give a brief overview of the assessments performed, the imaging methods applied, the genetic data produced, and the numbers of cases for whom different data types are available. We also cite sources of more detailed information about the methods and data. Finally we describe the procedures for accessing the data and for using the PING data exploration portal.

FTO affects changes in BMI during both childhood and adulthood. However, its effect on onset age of overweight in adulthood is not known. To address this question, we conducted a study to examine effects of FTO tag SNPs on censored age of overweight in the longitudinal Bogalusa Heart Study (BHS) cohort, which began in 1973-1974. Of participating subjects, 658 whites (308 males and 350 females) with genotype data were selected for the study. The FTO gene was examined by a survival analysis of 30 tag SNPs regarding their association with left, interval and right-censored adult overweight. After adjustment for birth weight and sex, SNP rs9939609 has a small nominal p value of 0.004 for the association with onset age, which has an expected proportion of false positives of 9.6 % after adjusting for multiple tests. It was estimated that genotypes AA, AT and TT have onset age (standard error) of 22.82 (1.07), 28.96 (1.04) and 27.76 (1.04) years, respectively, for a 50 % cumulative proportion of overweight in the population. Genotypes AA, AT and TT, respectively, have estimated survival probability of 65.8, 78.7 and 76.8 % at the age of 18; and survival probability of 6.5, 11.8 and 10.7 % at the age of 60. The odds ratios of survival beyond age ≥18 years are 0.52 for AA versus AT and 0.58 for AA versus TT. We thus concluded that risk genetic variants at FTO gene can accelerate the onset age and influence the survival odds of overweight in younger adults.

Objectives This study sought to evaluate the influence of single nucleotide polymorphisms (SNPs) on the pharmacodynamic effect of high- or standard-dose clopidogrel after percutaneous coronary intervention (PCI). Background There is a lack of prospective, multicenter data regarding the effect of different genetic variants on clopidogrel pharmacodynamics over time in patients undergoing PCI. Methods The GRAVITAS (Gauging Responsiveness with A VerifyNow assay–Impact on Thrombosis And Safety) trial screened patients with platelet function testing after PCI and randomly assigned those with high on-treatment reactivity (OTR) to either high- or standard-dose clopidogrel; a cohort of patients without high OTR were also followed. DNA samples obtained from 1,028 patients were genotyped for 41 SNPs in 17 genes related to platelet reactivity. After adjusting for clinical characteristics, the associations between the SNPs and OTR using linear regression were evaluated. Results CYP2C19*2 was significantly associated with OTR at 12 to 24 h (R2 = 0.07, p = 2.2 × 10−15), 30 days (R2 = 0.10, p = 1.3 × 10−7), and 6 months after PCI (R2 = 0.07, p = 1.9 × 10−11), whereas PON1, ABCB1 3435 C→T, and other candidate SNPs were not. Carriers of 1 and 2 reduced-function CYP2C19 alleles were significantly more likely to display persistently high OTR at 30 days and 6 months, irrespective of treatment assignment. The portion of the risk of persistently high OTR at 30 days attributable to reduced-function CYP2C19 allele carriage was 5.2% in the patients randomly assigned to high-dose clopidogrel. Conclusions CYP2C19, but not PON1 or ABCB1, is a significant determinant of the pharmacodynamic effects of clopidogrel, both early and late after PCI. In patients with high OTR identified by platelet function testing, the CYP2C19 genotype provides limited incremental information regarding the risk of persistently high reactivity with clopidogrel 150-mg maintenance dosing. (Genotype Information and Functional Testing Study [GIFT]; NCT00992420)

At late stage, nearly all breast cancers are heterogeneous and refractory to treatment, like metaplastic breast cancer is at an early stage. These rare carcinomas are highly aggressive and de-differentiated. They are enriched for mesenchymal and stem cell features and essentially fail current therapies. As metaplastic tumors provide a time-compressed picture of breast cancer progression early on, understanding these tumors will yield insight into mechanisms that drive breast cancer into advanced stages and treatment resistance. To investigate a genetic basis for heterogeneity in metaplastic breast cancer, we established a progression model comprising three cell lines. The cell lines were derived from a primary tumor, a local recurrence and a pleural effusion of a 40-year old patient. The primary tumor was a stage III invasive metaplastic, triple negative, inflammatory breast cancer, resected after neoadjuvant chemotherapy (capecitabine and taxotere, then adriamycin and one cycle of bevacizumab). The local recurrence, biopsied seven months post mastectomy, developed after the patient received adjuvant carboplatin and gemcitabine for 3 cycles and then radiation to the chest wall. At this time, the patient had lung metastases and was treated with taxol and bevacizumab yielding a mixed response. Local invasive growth continued and a malignant pleural effusion developed four months later. Analyzing the genetic and molecular characteristics of this progression model in vitro, its tumorigenicity and metastasis in vivo, and interrogating lead findings in a growing collection of metaplastic tumors helps us to dissect the genetic heterogeneity in breast cancer, and potentially to identify the cell types that drive disease progression and treatment resistance. Our gene expression analyses and genomic evaluations identified epithelial to mesenchymal transition (EMT) as a key characteristic in the progression and treatment resistance of this cancer. Major changes in cytoskeletal genes, chemokines and their receptors, amplification of drug transporter proteins, metalloproteinases and matrix proteins seen with increasing motility and invasiveness along with recruitment of host inflammatory responses in the in vivo model, loss of chromosomal regions harboring known and putative tumor suppressors, and deletions of genes encoding proteins for metabolic inactivation of sex hormones in the breast tissue, along with specific loss of clusters of desmosomal genes are guiding our understanding of metaplastic breast cancer progression. The results provide insight into the development, the extremely invasive nature, and treatment resistance of these tumors. Our collaborative network of clinicians, pathologists, translational genomic researchers and bioinformatics specialists will enable us to identify and prioritize genetic events as disease drivers, prognostic biomarkers of disease progression, and determinants of treatment resistance. Our goal is to identify molecular and functional targets for effective therapy and evaluate them in the clinic. Lessons learned from metaplastic breast cancer will improve our understanding of breast cancer progression in general, and could translate into effective treatments for advanced breast cancer where current standard of care is failing. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr PD03-07.

Clinical relevance of a genetic predisposition to elevated blood pressure was quantified during the transition from childhood to adulthood in a population-based Finnish cohort (N=2357). Blood pressure was measured at baseline in 1980 (age 3–18 years) and in follow-ups in 1983, 1986, 2001, and 2007. Thirteen single nucleotide polymorphisms associated with blood pressure were genotyped, and 3 genetic risk scores associated with systolic and diastolic blood pressures and their combination were derived for all of the participants. Effects of the genetic risk score were 0.47 mm Hg for systolic and 0.53 mm Hg for diastolic blood pressures (both P P =0.015). Replications in 1194 participants of the Bogalusa Heart Study showed essentially similar results. The participants in the highest quintile of the combination genetic risk score had a 1.82-fold risk of hypertension in adulthood ( P

Genes affecting glutamate neurotransmission featured prominently in associations between 94 genes and 12 inherited physiological or cognitive characteristics of schizophrenia. Of 16,620 possible SNP-trait associations, 47 showed strong significance.

ObjectivesThe objective of this study was to evaluate the antiplatelet effect of clopidogrel 150 mg/day in patients with high on-treatment reactivity (OTR) and to further assess this effect according to CYP2C19 genotype.BackgroundHigh OTR is associated with ischemic events in clopidogrel-treated patients after percutaneous coronary intervention. Alternative dosing regimens might enhance platelet inhibition.MethodsPatients with high OTR receiving a standard clopidogrel regimen were identified with the VerifyNow P2Y12 assay and administered clopidogrel 150 mg daily for 7 days, after which OTR was reassessed. Comprehensive CYP2C19 genotyping was performed with the BeadXpress platform (Illumina, San Diego, California) for the *2, *3, *4, *5, *6, *7, *8, and *17 variants.ResultsA total of 41 subjects were enrolled, 20 of whom were carriers of a CYP2C19 loss-of-function (LoF) allele. High-dose clopidogrel significantly reduced OTR from 285 ± 47 P2Y12 reaction units (PRU) to 220 ± 91 PRU (p < 0.001). There were no significant differences in antiplatelet effect according to CYP2C19 status, although the reduction in reactivity was minimal in the small number of patients homozygous for LoF alleles (n = 3, 28 ± 31 PRU, p = NS). Increasing body mass index was independently and negatively associated with the reduction in OTR (p = 0.009).ConclusionsIn patients with high OTR, clopidogrel 150 mg/day results in a significant reduction in platelet reactivity. Carriage of an LoF CYP2C19 polymorphism does not seem to have a major influence on dose effect. The observed lack of effect in patients with 2 copies of a CYP2C19 LoF allele must be confirmed by larger studies.

Telomeres are engaged in a host of cellular functions, and their length is regulated by multiple genes. Telomere shortening, in the course of somatic cell replication, ultimately leads to replicative senescence. In humans, rare mutations in genes that regulate telomere length have been identified in monogenic diseases such as dyskeratosis congenita and idiopathic pulmonary fibrosis, which are associated with shortened leukocyte telomere length (LTL) and increased risk for aplastic anemia. Shortened LTL is observed in a host of aging-related complex genetic diseases and is associated with diminished survival in the elderly. We report results of a genome-wide association study of LTL in a consortium of four observational studies ( n = 3,417 participants with LTL and genome-wide genotyping). SNPs in the regions of the oligonucleotide/oligosaccharide-binding folds containing one gene ( OBFC1 ; rs4387287; P = 3.9 × 10 −9 ) and chemokine (C-X-C motif) receptor 4 gene (CXCR4 ; rs4452212; P = 2.9 × 10 −8 ) were associated with LTL at a genome-wide significance level ( P < 5 × 10 −8 ). We attempted replication of the top SNPs at these loci through de novo genotyping of 1,893 additional individuals and in silico lookup in another observational study ( n = 2,876), and we confirmed the association findings for OBFC1 but not CXCR4 . In addition, we confirmed the telomerase RNA component (TERC) as a gene associated with LTL ( P = 1.1 × 10 −5 ). The identification of OBFC1 through genome-wide association as a locus for interindividual variation in LTL in the general population advances the understanding of telomere biology in humans and may provide insights into aging-related disorders linked to altered LTL dynamics.

To identify bipolar disorder (BD) genetic susceptibility factors, we conducted two genome-wide association (GWA) studies: one involving a sample of individuals of European ancestry (EA; n=1001 cases; n=1033 controls), and one involving a sample of individuals of African ancestry (AA; n=345 cases; n=670 controls). For the EA sample, single-nucleotide polymorphisms (SNPs) with the strongest statistical evidence for association included rs5907577 in an intergenic region at Xq27.1 (P=1.6 x 10(-6)) and rs10193871 in NAP5 at 2q21.2 (P=9.8 x 10(-6)). For the AA sample, SNPs with the strongest statistical evidence for association included rs2111504 in DPY19L3 at 19q13.11 (P=1.5 x 10(-6)) and rs2769605 in NTRK2 at 9q21.33 (P=4.5 x 10(-5)). We also investigated whether we could provide support for three regions previously associated with BD, and we showed that the ANK3 region replicates in our sample, along with some support for C15Orf53; other evidence implicates BD candidate genes such as SLITRK2. We also tested the hypothesis that BD susceptibility variants exhibit genetic background-dependent effects. SNPs with the strongest statistical evidence for genetic background effects included rs11208285 in ROR1 at 1p31.3 (P=1.4 x 10(-6)), rs4657247 in RGS5 at 1q23.3 (P=4.1 x 10(-6)), and rs7078071 in BTBD16 at 10q26.13 (P=4.5 x 10(-6)). This study is the first to conduct GWA of BD in individuals of AA and suggests that genetic variations that contribute to BD may vary as a function of ancestry.

The last few years have seen extensive efforts to catalogue human genetic variation and correlate it with phenotypic differences. Most common SNPs have now been assessed in genome-wide studies for statistical associations with many complex traits, including many important common diseases. Although these studies have provided new biological insights, only a limited amount of the heritable component of any complex trait has been identified and it remains a challenge to elucidate the functional link between associated variants and phenotypic traits. Technological advances, such as the ability to detect rare and structural variants, and a clear understanding of the challenges in linking different types of variation with phenotype, will be essential for future progress.

We have completed a second-generation linkage map that incorporates sequence-based positional information. This new map, the Rutgers Map v.2, includes 28,121 polymorphic markers with physical positions corroborated by recombination-based data. Sex-averaged and sex-specific linkage map distances, along with confidence intervals, have been estimated for all map intervals. In addition, a regression-based smoothed map is provided that facilitates interpolation of positions of unmapped markers on this map. With nearly twice as many markers as our first-generation map, the Rutgers Map continues to be a unique and comprehensive resource for obtaining genetic map information for large sets of polymorphic markers.

Infections of unknown origin and an altered immune response have been hypothesized to play a role in the pathogenesis of schizophrenia. We have previously identified two single nucleotide polymorphisms (SNPs) of the IL-10 receptor 1 (IL-10R1) causing a substitution of glycine 330 to arginine (G330R) and of serine 138 to glycine (S138G). A possible association between these IL-10R1 variants and schizophrenia has been investigated in the present study. DNA of 101 unrelated Austrian patients with a DSM-III-R (Diagnostic and Statistical Manual of Mental Disorders) consensus diagnosis of schizophrenia (n = 70) or schizoaffective disorder (n = 31) and DNA of 121 German schizophrenic patients (DSM-III-R) was analyzed for the presence of S138G and G330R by allele-specific multiplex PCRs. Data from patients were compared with 250 unrelated, psychiatric healthy controls. No difference in allele frequency was detected between patients and controls (G330R: 34.0% vs. 30.0%, P = 0.208; S138G: 19.7% vs. 16.6%, P = 0.235; by Fisher's exact test). However, there was a significant difference in genotype distribution (wt/wt, wt/mut, mut/mut) for G330R between patients (46.8%, 38.3%, 14.9%) and controls (47.6%, 44.8%, 7.6%; Fisher's test P = 0.032). No such difference was seen for S138G. Our results suggest that homozygosity of the IL-10R1 G330R allele is associated with schizophrenia and may contribute to the expression of disease phenotype in susceptible individuals.

© 2015, Springer-Verlag Berlin Heidelberg. Anxiety is a risk factor for many adverse neuropsychiatric and socioeconomic outcomes, and has been linked to functional and structural changes in the ventromedial prefrontal cortex (VMPFC). However, the nature of these differences, as well as how they develop in children and adolescents, remains poorly understood. More effective interventions to minimize the negative consequences of anxiety require better understanding of its neurobiology in children. Recent research suggests that structural imaging studies may benefit from clearly delineating between cortical surface area and thickness when examining these associations, as these distinct cortical phenotypes are influenced by different cellular mechanisms and genetic factors. The present study examined relationships between cortical surface area and thickness of the VMPFC and a self-report measure of anxiety (SCARED-R) in 287 youths aged 7–20 years from the Pediatric Imaging, Neurocognition, and Genetics (PING) study. Age and gender interactions were examined for significant associations in order to test for developmental differences. Cortical surface area and thickness were also examined simultaneously to determine whether they contribute independently to the prediction of anxiety. Anxiety was negatively associated with relative cortical surface area of the VMPFC as well as with global cortical thickness, but these associations diminished with age. The two cortical phenotypes contributed additively to the prediction of anxiety. These findings suggest that higher anxiety in children may be characterized by both delayed expansion of the VMPFC and an altered trajectory of global cortical thinning. Further longitudinal studies will be needed to confirm these findings.

The proteins encoded by the classical HLA class I and class II genes in the major histocompatibility complex (MHC) are highly polymorphic and play an essential role in self/non-self immune recognition. HLA variation is a crucial determinant of transplant rejection and susceptibility to a large number of infectious and autoimmune disease1. Yet identification of causal variants is problematic due to linkage disequilibrium (LD) that extends across multiple HLA and non-HLA genes in the MHC2,3. We therefore set out to characterize the LD patterns between the highly polymorphic HLA genes and background variation by typing the classical HLA genes and >7,500 common single nucleotide polymorphisms (SNPs) and deletion/insertion polymorphisms (DIPs) across four population samples. The analysis provides informative tag SNPs that capture some of the variation in the MHC region and that could be used in initial disease association studies, and provides new insight into the evolutionary dynamics and ancestral origins of the HLA loci and their haplotypes.