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• Add Health: Longitudinal Study of a Nationally Representative Sample of Adolescents in Grades 7-12 in the United States during the 1994-95 School Year, Followed into Adulthood with Five Interviews/Surveys in 1995, 1996, 2001-02, 2008, and 2016-18

  The National Longitudinal Study of Adolescent to Adult Health [Add Health] is an ongoing longitudinal study of a nationally representative U.S. cohort of more than 20,000 adolescents in grades 7-12 (aged 12-19 years) in 1994 followed into adulthood with five interviews/surveys in 1995, 1996, 2001-02, 2008, and 2016-18. Add Health was designed to understand how social environments and behaviors in adolescence are linked to health and achievement outcomes in young adulthood. Add Health contains unprecedented environmental, behavioral, psychosocial, biological, and genetic data from early adolescence and into adulthood on a large, nationally representative cohort with unprecedented racial, ethnic, socioeconomic, and geographic diversity. Add Health has a large, multidisciplinary user base of over 50,000 researchers around the world who have published over 3,400 research articles. Add Health is housed at the Carolina Population Center of the University of North Carolina at Chapel Hill. Add Health datasets are distributed according to a tiered data disclosure plan designed to protect the data from the risk of direct and indirect disclosure of respondent identity. Add Health's large sample size, population diversity and rich longitudinal database of psychosocial, physical, and contextual data will permit investigation of an exceptionally broad range of phenotypes with known genetic variation. Prospective longitudinal measures are available to document change over time in each of these phenotypes, as well as change in the social environment and life experiences, making the Add Health sample ideal for understanding genetic linkages with health and behavior across the life course. The original design of Add Health included important features for understanding biological processes in health and developmental trajectories across the life course of young people, including an embedded genetic sample with more than 3,000 pairs of adolescents with varying biological resemblance (e.g., twins, full sibs, half sibs, and adolescents who grew up in the same household but have no biological relationship), testing of saliva and urine for sexually transmitted infections and HIV, and biomarkers of cardiovascular health, metabolic processes, immune function, renal function, and inflammation. Add Health therefore has critical objective indicators of health status and disease markers in young adulthood, well before chronic illness or its complications emerge in later adulthood. Because DNA has been collected on the full sample at Wave IV, it is possible to link genetic profiles with social, behavioral, and biological measures over time from adolescence into adulthood. Add Health sampled the multiple environments in which young people live their lives, including the family, peers, school, neighborhood, community, and relationship dyads, and provides independent and direct measurement of these environments over time. Add Health contains extensive longitudinal information on health-related behavior, including life histories of physical activity, involvement in risk behavior, substance use, sexual behavior, civic engagement, education, and multiple indicators of health status based on self-report (e.g., general health, chronic illness), direct measurement (e.g., overweight status and obesity), and biomarkers. No other data resource with this expanse of genotype and phenotype data on a large nationally representative longitudinal sample with race, ethnic, socioeconomic, and geographic diversity exists. A complete reference guide on study design and accomplishments can be found on the Add Health website: https://cdr.lib.unc.edu/concern/articles/6t053j27s.

  Study phs001367

• Heart Failure Network - Phosphodiesterase-5 Inhibition to Improve Clinical Status and Exercise Capacity in Diastolic Heart Failure (HFN RELAX-BioLINCC)

  Data Access NOTE: Please refer to the “Authorized Access” section below for information about how access to the data from this accession differs from many other dbGaP accessions. Related StudyEcho images are available through HFN RELAX-ImagingObjectivesThe RELAX trial tested the hypothesis that chronic phosphodiesterase type-5 inhibitor therapy with sildenafil would improve exercise capacity and clinical status in heart failure patients with normal ejection fraction, as compared to placebo. Background Heart failure (HF) with preserved ejection fraction (HFpEF) is a common and highly morbid condition that is characterized by chronic exercise intolerance, progressive functional decline and a high rate of readmission. At the time of the RELAX trial, clinical trials of renin-angiotensin system antagonists had not demonstrated improvement in outcomes or clinical status in HFpEF, and effective therapies were needed. Phosphodiesterase type-5 (PDE-5) metabolizes the nitric oxide (NO) and natriuretic peptide (NP) systems' second messenger cyclic guanosine monophosphate (cGMP), and thus may limit beneficial NO and NP actions in the heart, vasculature and kidney. Pre-clinical studies suggest that inhibition of PDE-5 reverses adverse cardiac structural and functional remodeling and enhances vascular, neuroendocrine and renal function. In clinical studies, PDE-5 inhibitor therapy improved exercise tolerance and clinical status in patients with idiopathic pulmonary arterial hypertension and in patients with HF and reduced ejection fraction (HFrEF). A small, single-center study in HFpEF observed improved hemodynamics, left ventricular (LV) diastolic function, right ventricular (RV) systolic function, LV hypertrophy and lung function with chronic PDE-5 inhibition as compared to placebo. In aggregate, these studies suggested the potential for PDE-5 inhibition to ameliorate several key pathophysiological perturbations in HFpEF, and thus improve exercise capacity and clinical status. ParticipantsA total of 216 participants were enrolled in the trial with 113 in the Sildenafil group and 103 in the placebo group. Design Participants who met screening criteria underwent baseline studies including a history and physical examination, cardiopulmonary exercise test (CPXT), six-minute walk distance, Minnesota Living with Heart Failure Questionnaire (MLWHFQ), echocardiography, cardiac magnetic resonance imaging, and phlebotomy for biomarkers. Subjects were then randomly assigned, in a 1:1 ratio, to either the sildenafil or placebo intervention group. The study drug was administered orally at 20 mg three times daily (TID) for 12 weeks. If the dose was well tolerated at 12 weeks, it was increased to 60 mg TID for another 12 weeks. If side effects developed, study staff could recommend discontinuation or return to a lower or previously tolerated dose of study drug. Sildenafil levels 2 hours after a scheduled dose of study drug were obtained at 12 and 24 weeks. The primary endpoint was exercise capacity determined by change in peak oxygen consumption during the CPXT after 24 weeks of therapy. Secondary endpoints included change in six-minute walk distance at 12 and 24 weeks, change in peak oxygen consumption at 12 weeks, and a three tier score reflective of clinical status where patients were ranked based on time to death (lowest tier), time to cardiovascular or cardiorenal hospitalization (middle tier), and change in the MLWHFQ for patients alive without cardiovascular or cardiorenal hospitalization after 24 weeks (highest tier). Conclusions Chronic phosphodiesterase type-5 inhibitor therapy with sildenafil for 24 weeks did not alter exercise capacity or clinical status compared to placebo in patients with heart failure and preserved ejection fraction.

  Study phs003565

• Common Fund (CF) Genotype-Tissue Expression Project (GTEx)

  Lay Description The aim of the Genotype-Tissue Expression (GTEx) Project is to increase our understanding of how changes in our genes affect human health and disease with the ultimate goal of improving health care for future generations. GTEx will create a database that researchers can use to study how inherited changes in genes lead to common diseases. GTEx researchers are studying genes in different tissues obtained from many different people. The GTEx project also includes a study of the GTEx donor consent process - this study will help ensure that the consent process and other aspects of the project effectively address the concerns and expectations of participants in the study. GTEx is a pioneering project that uses state-of-the-art protocols for obtaining and storing a large range of organs and tissues, and for testing them in the lab. Until now, no project has analyzed genetic variation and expression in as many tissues from the same person in such a large population as planned for GTEx. Scientific Description Understanding the role of variation in the human genome is crucial to elucidating genetic contributions to human health and disease. Despite the results of genome-wide association studies (GWAS) documenting strong statistical associations between genetic variation and human traits, the functional role for most of these variants is largely unexplained. Nearly 90% of these GWAS-implicated sites lie outside of protein-coding sequences, suggesting that these variants might regulate gene expression. The goal of the Genotype-Tissue Expression (GTEx) project is to establish a resource database and tissue biobank in which to study the relationship between genetic variation and gene expression and other molecular phenotypes in reference/non-diseased tissues. The ultimate resource will include approximately 960 post-mortem donors with several dozen tissues from each, a resource large enough to study both cis- and trans- gene expression quantitative trait loci (eQTLs). Some tissue will also be banked for additional molecular analyses. To request biosamples, please see the Biobank page for more information. GTEx was initially funded as a 2-year pilot project by the NIH Common Fund (CF) in 2010, and has been scaled up after demonstration of feasibility. The project will collect and analyze RNA levels in many different human tissues and each donor will be characterized for germline genetic variation through dense genotyping arrays and sequencing of either whole exomes or whole genomes. By treating RNA expression levels as quantitative traits, eQTLs will be identified as sites containing genetic variation that correlate with changes in RNA expression. Such eQTLs have been associated with 4%-12% of expressed human genes, and with common complex human diseases, including obesity, atherosclerosis, type 2 diabetes, Crohn's disease, and asthma. Additionally, few studies have examined the tissue specificity of eQTLs. A subset of banked tissue samples will also be analyzed for other molecular phenotypes, such as DNA methylation, DNaseI hypersensitivity sites, and proteomics. The GTEx project will thus serve as a resource database and tissue bank for many future studies, especially for understanding the functional basis of inherited susceptibility to disease. All GTEx releases since Version 5 follow the NIH Genomic Data Sharing (GDS) Policy whereby there are no restrictions on use or publication after release. Additional information about the GTEx Data Release and Publication policy can be found on the GTEx Data Portal at www.gtexportal.org.

  Study phs000424

• Resuscitation Outcomes Consortium (ROC) Prehospital Resuscitation on Helicopter Study (PROHS) (ROC-PROHS-BioLINCC)

  Accessing Data: Please refer to the "Authorized Access" section below regarding accessing data through the BioData Catalyst ecosystem. The data from this accession is not available for download through dbGaP.Objective: To observe if patients with severe traumatic injuries evacuated to level 1 trauma centers on air ambulances who received prehospital red blood cells and/or plasma had decreased in-hospital mortality compared to patients who received only crystalloids.Background: Injury is the leading cause of death in adults and children between the ages of 1 and 44 years. However, approximately 40% of in-hospital deaths among injured patients involve massive truncal hemorrhage that is considered potentially salvageable. Multiple retrospective military and civilian studies have reported that transfusion involving blood component ratios approaching whole blood are associated with significant decreases in 24-hour and 30-day mortality among injured patients. Furthermore, prehospital transfusion (PHT) studies from military and civilian hospitals have shown that prehospital transfusion of plasma and red blood cells (RBCs) is not only feasible, but associated with improved coagulation status on arrival. Prehospital resuscitation practices in the US differ significantly in approach, with most systems using crystalloids while a few offer RBCs or a combination of plasma and RBCs. No large multicenter civilian studies have evaluated the use of prehospital plasma and RBCs in severely injured patients compared to crystalloids.Participants: A total of 25,118 trauma patients were admitted during the 10 month enrollment period to the nine participating centers, of which 2341 arrived by helicopter, and 1058 met the highest risk criteria. Of the high risk sub-set, 585 arrived on helicopters with blood products available and 473 patients arrived via helicopters without blood available. One hundred forty two patients (24%) of those transported on helicopters with blood products available actually received PHT and 916 patients did not.Design: PROHS was a multicenter, prospective pragmatic, observational study of prehospital resuscitation approaches. There were no study guidelines dictating resuscitation practice (i.e., use of blood products or end of resuscitation). Of the 9 participating Level I trauma centers, 5 had helicopter systems with plasma and/or RBCs while the other 4 had helicopter systems that used only crystalloid resuscitation. All patients arriving to the participating center's trauma emergency department via helicopter directly from the scene were screened and had initial data collected. Of that group, all patients who met the highest risk category or received blood during transport were followed by direct observation during the initial resuscitation period and then indirectly through medical chart review until hospital discharge or 30 days after admission (whichever occurred first).The primary outcome was in-hospital mortality at 3 hours, 24 hours, and 30 days. Other outcome measures included length of hospital stay, number of ICU days, number of ventilator days, blood product usage, GOSE score at discharge, number of patients with complications, and number of patients who required hemostatic devices. Conclusions:Because of the unexpected imbalance in injury severity between systems with and without blood products on helicopters, all analyses were inconclusive. With few units transfused to each patient and small outcome differences between groups, large randomized studies will be required to detect significant survival differences in this important population.

  Study phs003826

• Evolution of an adenomacarcinoma in response to selection by targeted kinase inhibitors

  Background - Adenocarcinomas of the tongue are rare and represent the minority (20-25%) of salivary gland tumors affecting the tongue. We investigated the utility of massively parallel sequencing to characterize an adenocarcinoma of the tongue, before and after treatment. Results - In the pre-treatment tumor we identified 7,629 genes within regions of copy number gain. 1,078 genes exhibited increased expression relative to the blood and unrelated tumors and four genes contained somatic protein-coding mutations. Our analysis suggested the tumor cells were driven by the RET oncogene. Genes whose protein products are targeted by the RET inhibitors sunitinib and sorafenib correlated with being amplified and or highly expressed. Consistent with our observations administration of sunitinib was associated with stable disease lasting 4 months, after which the lung lesions began to grow. Administration of sorafenib and sulindac provided disease stabilization for an additional 3 months after which the cancer progressed and new lesions appeared. A recurring metastasis possessed 7,288 genes within copy number amplicons, 385 genes exhibiting increased expression relative to other tumours and 9 new somatic protein coding mutations. The observed mutations and amplif ications were consistent with therapeutic resistance arising through activation of the MAPK and AKT pathways. Conclusions - We conclude that complete genomic characterization of a rare tumour has the potential to aid in clinical decision making and identifying therapeutic approaches where no established treatment protocols exist. These results also provide direct in-vivo genomic evidence for mutational evolution within a tumor under drug selection and potential mechanisms of drug resistance accrual.

  Study EGAS00000000074

• Angelman, Rett, Prader-Willi Syndromes Consortium (ARP) Angelman Syndrome Natural History Protocol

  This study is a longitudinal multidisciplinary investigation on the natural history, morbidity and mortality of Angelman Syndrome (AS). We will collect detailed longitudinal data on a cohort of AS individuals to gain a better understanding of the disease progression, and to follow the natural history of the clinical features of this cohort including assessment of quality of life and longevity. The participants to be recruited for the study will include 1) patients who have a documented molecular diagnosis of AS and 2) patients with a clear clinical diagnosis of Angelman Syndrome as determined by the Principal Investigator (PI) and the Co-investigators in this study but who do not have a known molecular defect. One of the goals of the natural history study will be to characterize the phenotypic differences between patients with Class I deletions and those with Class II deletions, particularly with respect to the issue of autism. A blood sample may be collected on the participants in order to create a DNA repository, and in some cases, to establish cell lines if further material is required for molecular studies. Alternatively, DNA may be obtained by buccal mucosa swabs/brushing. In those AS patients with known deletions involving the 15q11-q13 regions, a blood sample will be collected to perform comparative genome hybridization (CGH) microarray studies to characterize the extent of the deletion. No drugs or treatments will be administered through this protocol. In rare instances, a skin biopsy to establish a fibroblast cell line may be requested (using separate consent).

  Study phs000576

• Innate Immune Anti-Viral Deaminase Deregulation Fuels Pre-Leukemia Stem Cell Evolution

  This study evaluates the effect of Rebecsinib treatment on gene expression, isoform splicing, and ADAR1 mediated RNA editing in cells from an in vivo humanized mouse model which have been engrafted with human secondary AML cells. For analysis of cells from Rebesinib-treated animals, CD34+ cells were obtained from bone marrow and spleens of sAML-engrafted mice following once weekly treatment with 0 (Vehicle) or 5 or 10 mg/kg Rebecsinib. For analysis of cells from serial transplant recipients of cells harvested from Rebecsinib-treated mice, CD34+ cells were obtained from bone marrow and spleens of sAML-engrafted mice that were transplanted with cells from animals that received twice weekly treatment with 0 (Vehicle) or 10 mg/kg Rebecsinib. No further treatment was given to serial transplant recipients, thus allowing the analysis of molecular changes that are sustained following serial transplantation. Whole transcriptome sequencing was performed at The Scripps Research Institute Next Generation Sequencing Core on Illumina NextSeq 500 sequencers with 150bp paired-end reads.These data revealed a uniquely expressed ADAR1 splice isoform (with a retrained intronic region), global downregulation of normalized RNA editing events, and isoform switching from MCL1-L to MCL1-S in Rebecsinib-treated sAML-engrafted mice. These results demonstrate the anti-survival and anti-self-renewal effects of Rebecsinib treatment on the leukemia stem cell (LSC) population in sAML-engrafted mice as well as the reduction in the inflammation-induced, hyper-editing p150 isoform of ADAR1. Thus, Rebecsinib treatment has promise for reducing the LSC population in myeloproliferative disorders and associated cancers.

  Study phs002228

• Genome, epigenome and RNA sequences of monozygotic twins discordant for multiple sclerosis

  Monozygotic (MZ) twins have been widely employed for dissection of the relative contributions of genetics and environment in disease. In multiple sclerosis (MS), an autoimmune demyelinating disease that commonly causes neurodegeneration and disability in young adults, disease discordance in MZ twins has been interpreted to indicate environmental importance in pathogenesis. However, genetic and epigenetic differences between MZ twins have been described, challenging the accepted experimental paradigm in disambiguating effects of nature and nurture. Here, we report the genome sequences of one MS-discordant MZ twin pair and messenger RNA (mRNA) transcriptome and epigenome sequences of CD4+ lymphocytes from three MS-discordant, MZ twin pairs. No reproducible differences were detected between co-twins among ~3.6 million single nucleotide polymorphisms (SNPs) or ~0.2 million insertion-deletion polymorphisms (indels). Nor were any reproducible differences observed between siblings of the three twin pairs in HLA haplotypes, confirmed MS-susceptibility SNPs, copy number variations, mRNA and genomic SNP and indel genotypes, or expression of ~19,000 genes in CD4+ T cells. Only two to 176 differences in methylation of ~2 million CpG sites were detected between siblings of the three twin pairs, in contrast to ~800 differences in methylation between T cells of unrelated individuals and several thousand differences between tissues or normal and cancerous tissues. In the first systematic effort to estimate sequence variation among MZ co-twins, we did not find evidence for genetic, epigenetic or transcriptome differences that explained disease discordance. These are the first female, twin and autoimmune disease genome sequences reported.

  Study phs000239

• DNA Methylomic Profiling of Preeclampsia Across Pregnancy

  Preeclampsia (PE) is a hypertensive, multi-system disorder of pregnancy that significantly impacts maternal and infant morbidity/mortality across the globe, as it increases risk of cardiovascular disease and remains a leading killer of women and babies. Despite PE's significant impact on morbidity/mortality, there are no clinically reliable biomarkers that predict PE. DNA methylation, a dynamic regulator of gene expression, represents a mechanism that is known to be impacted by the environment. Because PE stems from a dysfunctional placenta that releases debris into the maternal circulation, we hypothesized that the in-vivo environment created by the dysfunctional placenta will impact DNA methylation in the maternal circulation, and that these blood-based methylation profiles would serve as a systemic biomarker of the maternal response to placental dysfunction. Our overall objective of this pilot study was to longitudinally characterize DNA methylation profiles across the three trimesters of pregnancy in the maternal blood at time points before and after clinically overt PE using a targeted (endoglin and endoglin-related genes) and a discovery-based approach. For this pilot study, 28 normotensive control participants and 28 PE case participants enrolled in the NICHD funded pregnancy study entitled Prenatal Exposures and Preeclampsia Prevention: Mechanisms of Preeclampsia and the Impact of Obesity (PEPP3; P01HD30367) were 1:1 frequency matched on self-reported race, pre-pregnancy BMI, smoking history, and gestational age at sample collection. Methylation data were collected with the Infinium® MethylationEPIC Beadchip. Methylation assay data collection were carried out at Johns Hopkins University Genetic Resources Core Facility, The SNP Center, Baltimore, MD, USA.

  Study phs001937

• National Eye Institute (NEI) Primary Open-Angle African American Glaucoma Genetics (POAAGG) Study

  The purpose of the POAAGG study is to investigate the genetic architecture of primary open-angle glaucoma (POAG) in African Americans. This population is up to eight times more likely to develop glaucoma than non-Hispanic whites, yet no large-scale genetics studies have specifically addressed this ethnic group. The POAAGG study aims to enroll 7765 patients, consisting of a discovery cohort of 5500 subjects (2000 cases and 3500 controls) and a validation cohort of 2265 subjects (1000 cases and 1265 controls). The study population consists of self-identified blacks (African Americans, African descent, or African Caribbean), 35 years or older, identified from the Philadelphia region. Subjects are recruited from the Scheie Eye Institute and its research sites in Philadelphia, PA, as well as from the Penn Medicine Biobank. Fellowship-trained glaucoma specialists classify each subject as a case, control, or glaucoma suspect based on specified criteria. As of 02/01/2017, a total of 8187 subjects have been recruited to the POAAGG study, including 2522 cases, 4545 controls, and 1120 suspects. A comprehensive genetic analysis of POAG, including a genome-wide association analysis, is currently underway. The GWAS was conducted on the Illumina Infinium Multi Ethnic Genotyping Array (MEGA), which contained custom content tailored to POAG and African Americans. A comprehensive genetic analysis of quantitative traits associated with POAG is also in progress. Over 90% of cases have full phenotypic information, including intraocular pressure, cup-to-disc ratio, central corneal thickness, retinal nerve fiber layer thickness, visual acuity, and visual fields.

  Study phs001312

• Genomic profiling for metastatic uveal melanoma from a phase I study of the protein kinase C inhibitor AEB071

  Up to 50% of patients with uveal melanoma (UM) develop metastatic disease, for which there is no effective systemic treatment. This study aimed to evaluate the safety and efficacy of the orally available protein kinase C inhibitor, AEB071, in patients with metastatic UM, and to perform genomic profiling of metastatic tumor samples, with the aim to propose combination therapies. Metastatic UM patients (n=153) were treated with AEB071 in a Phase I, single-arm study. Patients received total daily doses of AEB071 ranging from 450 to 1400 mg. First-cycle dose-limiting toxicities (DLTs) were observed in 13 patients (13%). These were most commonly gastrointestinal system toxicities and were dose related, occurring at doses > 700 mg/day. Preliminary clinical activity was observed, with 3% of patients achieving a partial response and 50% with stable disease (median duration 15 weeks). High-depth, targeted next-generation DNA sequencing (NGS) was performed on 89 metastatic tumor biopsy samples. Mutations previously identified in UM were observed, including mutations in GNAQ, GNA11, BAP1, SF3B1, PLCB4, and amplification of chromosome arm 8q. GNAQ/GNA11 mutations were observed at a similar frequency (93%) as previously reported, confirming a therapeutic window for inhibition of the downstream effector PKC in metastatic UM. In conclusion, the protein kinase C inhibitor AEB071 was well tolerated and modest clinical activity was observed in metastatic UM. The genomic findings were consistent with previous reports in primary UM. Together, our data allow envisaging combination therapies of PKC inhibitors with other compounds in metastatic UM.

  Study phs001953

• Sporadic autism exomes reveal a highly interconnected protein network of de novo mutations

  It is well established that autism spectrum disorders (ASD) have a strong genetic component; however, for at least 70% of cases, the underlying genetic cause is unknown. Under the hypothesis that de novo mutations underlie a substantial fraction of the risk for developing ASD in families with no previous history of ASD or related phenotypes-so-called sporadic or simplex families-we sequenced all coding regions of the genome (the exome) for parent-child trios exhibiting sporadic ASD, including 189 new trios and 20 that were previously reported. Additionally, we also sequenced the exomes of 50 unaffected siblings corresponding to these new (n = 31) and previously reported trios (n = 19), for a total of 617 individual exomes from 209 families deposited in dbGaP. Here we show that de novo point mutations are overwhelmingly paternal in origin (4:1 bias) and positively correlated with paternal age, consistent with the modest increased risk for children of older fathers to develop ASD. Moreover, 39% (49 of 126) of the most severe or disruptive de novo mutations map to a highly interconnected beta-catenin/chromatin remodelling protein network ranked significantly for autism candidate genes. In proband exomes, recurrent protein-altering mutations were observed in two genes: CHD8 and NTNG1. Mutation screening of six candidate genes in 1,703 ASD probands identified additional de novo, protein-altering mutations in GRIN2B, LAMC3 and SCN1A. Combined with copy number variant (CNV) data, these results indicate extreme locus heterogeneity but also provide a target for future discovery, diagnostics and therapeutics.

  Study phs000482

• Norepinephrine Transporter Blockade as a Pathophysiological Biomarker in Neurogenic Orthostatic Hypotension

  The clinical picture of autonomic failure is dominated by disabling orthostatic hypotension. Recent developments in the understanding of the underlying pathophysiology of the discrete clinical forms of autonomic failure have revealed that participants with multiple system atrophy (MSA) are characterized by impairment of central autonomic pathways crucial for autonomic cardiovascular control, but have intact peripheral postganglionic noradrenergic fibers. Participants with MSA have peripheral residual sympathetic tone that is no longer modulated by central autonomic centers and is not under baroreflex control and, therefore, cannot be harnessed to improve their orthostatic hypotension. On the other hand, participants with pure autonomic failure (PAF) and with Parkinson' s disease (PD) are characterized by neurodegeneration and loss of peripheral noradrenergic fibers, as evidenced by low levels of plasma norepinephrine and absent cardiac uptake of labeled catechols. Pharmacological inhibition of the norepinephrine transporter (NET) is an example of an approach that can take advantage of the participants' own residual sympathetic tone. NET inhibition will increase synaptic norepinephrine that is tonically released in MSA participants by preventing its reuptake. This should result in an increase in blood pressure. This is a longitudinal study of 50 participants, age 18-80 years with neurogenic orthostatic hypotension associated with impaired reflexes. The purpose of this study is to determine if the magnitude of the pressor response to atomoxetine at study entry, will be useful as a biomarker to differentiate those participants that will ultimately be shown to have pre-ganglionic (central) lesions (MSA) with those shown to have post ganglionic (peripheral) lesions (PAF and PD).

  Study phs001595

• National Heart Lung and Blood Institute Exome sequencing in SCID

  Analysis of the molecular etiologies of severe combined immunodeficiency (SCID) has led to important insights into the control of immune cell development. Most cases of SCID result from either X-linked or autosomal recessive inheritance of mutations in a known causative gene. However, in some cases, the molecular etiology remains unclear. To identify the cause of SCID in a patient known to lack the protein tyrosine phosphatase CD45, we utilized single nucleotide polymorphisms (SNP) arrays and whole exome sequencing. The patient's mother was heterozygous for an inactivating mutation in CD45, while the paternal alleles lacked mutations. The patient exhibited a single CD45 mutation identical to the maternal allele. Patient SNP array analysis revealed no change in copy number but loss of heterozygosity for the entire length of chromosome 1 (Chr1), indicating that disease was caused by uniparental disomy (UPD) with isodisomy of the entire maternal Chr1 bearing the CD45 mutation. Non-lymphoid blood cells and other mesoderm and ectoderm-derived tissues retained UPD of the entire maternal Chr1 in this patient who had undergone successful bone marrow transplantation. Exome sequencing revealed mutations in 7 additional genes bearing nonsynonymous SNPs predicted to have deleterious effects. These findings represent the first reported case of SCID caused by UPD and suggest UPD should be considered in SCID and other recessive disorders, especially when the patient appears homozygous for an abnormal gene found in only one parent. Evaluation for alterations in other genes affected by UPD should also be considered in such cases.

  Study phs000479

• Transposable Elements in FTLD-TDP and ALS-TDP

  Amyotrophic Lateral Sclerosis (ALS) is a devastating neurodegenerative disease involving the rapid and progressive loss of motor neurons. While a number of inherited mutations have been identified as causing ALS, the vast majority of cases are sporadic with no family history of disease. This has led to the model that ALS may be a complex syndrome with an unknown number of causal molecular pathways. However, nearly all ALS patient tissues display aggregates of the RNA binding protein TDP-43 (TARDBP/TAR DNA-binding protein), and mutations in the gene encoding this protein are known to cause ALS. We show that one of the normal functions of TDP-43 is to bind and regulate retrotransposons, repetitive sequences in the genome that are normally silent, due in part to the function of TDP-43. Furthermore, we demonstrate that many ALS patients with TDP-43 pathology also display de-silencing of retrotransposon RNAs, consistent with a loss of normal TDP-43 function. Those ALS patients that exhibit high levels of retrotransposon expression form a distinct subset of ALS subjects that also show other correlated alterations to the transcriptome. Using unsupervised machine learning algorithms, we identified three distinct molecular subtypes from the whole genomes and transcriptomes of a large cohort of 148 ALS patient samples. The retrotransposon re-activated subset formed 20% of ALS samples; two additional groups displayed signatures of oxidative stress (61%), and glial dysfunction (19%), respectively. Together these results demonstrate that retrotransposon re-activation is common in a subset of ALS patients with TDP-43 pathology.

  Study phs001889

• ILyAD (Indolent Lymphoma And vitamin D)

  Indolent non-Hodgkin lymphomas are incurable diseases, and require intermittent and often morbid and expensive therapy during their prolonged natural history. Lower intensity and better-tolerated, cost-effective treatment strategies are needed for these patients. Low vitamin D levels at diagnosis of indolent lymphomas are strongly associated with inferior outcomes to treatment. Standard therapy for low tumor burden, indolent lymphoma includes a single agent monoclonal antibody (rituximab). In our double-blind, placebo-controlled, randomized trial we evaluated if vitamin D supplementation with 2000IU oral cholecalciferol daily for three years improved the progression-free survival of patients with indolent lymphoma treated with single agent rituximab. Stratification factors for randomization included histology and Follicular Lymphoma International Prognostic Index (FL-IPI) prognostic score. The primary endpoint of the trial was progression-free survival over three years; secondary endpoints included response at 13 weeks, and overall survival. Eligible participants were adults with biopsy proven follicular lymphoma (grades 1-3a), small lymphocytic lymphoma, marginal zone lymphoma and mucosal-associated lymphoid tissue with no prior anti-lymphoma systemic therapy. Participants had Ann Arbor stage II or greater, measurable disease defined by Lugano criteria. Participants with osteoporosis requiring prescription treatment, symptomatic primary hyperthyroidism, hypercalcemia, creatinine greater than 1.5 times the upper limit of normal or a history of calcium-related kidney stones were excluded. Germline DNA was isolated from saliva samples obtained from participants at baseline. Whole exome sequencing was performed and SNPs were analyzed using the Agilent SureSelectXT Human All Exon V6 +UTR array. SNP data from 185 participants are available through dbGaP.

  Study phs003503

• Sequence analysis of colorectal serrated lesions

  Recent studies have shown that colorectal serrated lesions, which include sessile serrated adenomas (SSAs) and traditional serrated adenomas (TSAs), are precursors of colorectal cancer. However, the molecular mechanisms underlying the carcinogenesis, particularly in TSAs, remain largely uncharacterized. To clarify their molecular and clinicopathological characteristics, we performed mutation and methylation analyses of cancer-associated genes in 78 serrated lesions, including TSAs, SSAs and microvesicular hyperplastic polyps. Target exon sequence analysis was performed with 39 genes, including genes known to be frequently mutated in colorectal cancers and/or serrated lesions. We also used bisulfite pyrosequencing to assess the methylation status of various cancer-associated genes and marker genes of the CpG island methylator phenotype (CIMP). The prevalence of mutations in genes associated with Wnt signaling was significantly higher in TSAs than SSAs (65% vs. 28%, p < 0.01). Among those, RNF43 mutations were observed in 38% of TSAs and 17% of SSAs. In immunohistochemical studies of 39 serrated lesions, the prevalence of abnormal nuclear ?-catenin accumulation was significantly higher in TSAs (57%) than SSAs (8%) (P = 0.01). SMOC1 methylation was detected in 54% of TSAs but in no SSAs (p < 0.01). Additionally, SMOC1 methylation was more prevalent among TSAs with KRAS mutation (82%) than with BRAF mutation (38%, p = 0.03). Lesions with CIMP-high or RNF43 mutations were detected only in TSAs with BRAF mutation, suggesting two distinct carcinogenic pathways in TSAs. Mutations in genes associated with Wnt signaling play a greater role in the carcinogenesis of TSAs than SSAs.

  Study JGAS000217

• Oncogenic fate conversion by PRDM16s causes acute myeloid leukemia

  Oncogenic mutations confer aberrant replicative capacity to cells with little or no replicative capacity, generating cancer stem cells that perpetuate the tumor through extensive self-replication and differentiation blockade. However, whether oncogenes disrupt the cellular identity ofcancer stem cell by altering the developmental potential is unknown. Fate conversion has been demonstrated by ectopic expression of master transcriptional regulators, such as PU.1 and C/EBP alpha that confers myeloid cell fate to other cell types, and PRDM16 that confers brown adipose fate to white adipocytes or myoblasts. Here we show that a transcriptional regulator overexpressed in human myeloid malignancies, PRDM16s, causes oncogenic fate conversion, by transforming cells fated to form platelets and erythrocytes into myeloid leukemia-initiating cells (LICs). Prdm16s expression in hematopoietic progenitor cells caused a myelodysplastic syndrome (MDS)-like disease that progressed to acute myelogenous leukemia (AML). The myeloid diseases caused by Prdm16s exhibited expansion of megakaryocyte-erythroid progenitors (MEPs) but not granulocyte-macrophage progenitors. MEPs from Prdm16s-induced leukemia possessed LIC potential, and expression of Prdm16s in normal MEPs was sufficient to convert them to myeloid LICs and blocked megakaryocytic/erythroid potential. Prdm16s induced the expression of myeloid master regulators, including PU.1 and C/EBP alpha, by interacting with their super enhancers. Ablation of myeloid master regulators attenuated the myeloid potential and reinstalled the megakaryocytic/erythroid potential of leukemic-MEPs in mouse models and in human AML with PRDM16 rearrangement. Our study demonstrates that oncogenic Prdm16 expression converts the fate of MEPs to a malignant myeloid fate by activating myeloid master transcription factors.

  Study EGAS00001003235

• Search for new loci and low-frequency variants influencing glioma risk by exome-array analysis

  To identify protein altering variants (PAVs) for glioma we analysed Illumina HumanExome BeadChip exome array data on 1,882 glioma cases and 8,079 controls from three independent European populations. In addition to single variant tests we incorporated information on the predicted functional consequences of PAVs and analysed sets of genes with a higher likelihood of having a role in glioma on the basis of the profile of somatic mutations documented by large-scale sequencing initiatives. Globally tThere was a strong relationship between effect size and SNPs predicted to be damaging (P=2.29x10-49); however, these variants which are most likely to impact on risk, are rare (MAF&lt;5%). While no single variant showed an association which was statistically significant at the genomewide threshold a number of represented promising Notable associations - were seen with the BRCA2:c.9976A&gt;T, p.(Lys3326Ter)BRCA2 p.Lys332X variant, which has been shown documented to influence breast and lung cancer risks (OR=2.3, P=4.00x10-4 for glioblastoma [GBM]) and IDH2:c.782G&gt;A, p.(Arg261His)IDH2 p.Arg261His (OR=3.21, P=7.67x10-3, for non-GBM). Additionally, gene burden tests revealed a statistically significant association for HARS2 and risk of GBM (P=2.20x10-6). Genome scans of low frequency PAVs represent a complementary strategy to identify disease-causing variants compared with scans based on tagSNPs. Strategies to lessen the multiple testing burden by restricting analysis to PAVs with higher priors affords an opportunity to maximise study power.

  Study EGAS00001001258

• Benchmarking for alignment and variant calling

  Background: Lung cancer is a heterogeneous disease and the primary cause of cancer-related mortality worldwide. Somatic mutations, including large structural variants, are important biomarkers in lung cancer for selecting targeted therapy. Genomic studies in lung cancer have been conducted using short-read sequencing. Emerging long-read sequencing technologies are a promising alternative to study somatic structural variants, however there is no current consensus on how to process data and call somatic events. In this study, we preformed whole genome sequencing of lung cancer and matched non-tumour samples using long and short read sequencing to comprehensively benchmark three sequence aligners and six structural variant callers comprised of four generic callers (Sniffles2, cuteSV, SVIM and DELLY in generic mode) and two somatic callers (nanomonsv and DELLY in somatic modes). Results: Different combinations of aligners and variant callers influenced somatic structural variant detection. The choice of caller had a significant influence on somatic structural variant detection in terms of variant type, size, sensitivity, and accuracy. The performance of each variant caller was assessed by comparing to somatic structural variants identified by short read sequencing. When compared to somatic structural variants detected with short read sequencing, more events were detected with long read sequencing. The mean recall of somatic variant events identified by short read sequencing was higher for the somatic callers (64%) than generic callers (52%), with nanomonsv (69%) showing the best performance. Conclusion: Long read sequencing can identify somatic structural variants. The longer reads have the potential to improve our understanding of cancer development and inform personalized cancer treatment.

  Study EGAS00001007819

• Exome Sequencing Of 75 Individuals From Multiply Affected Coeliac Families

  Coeliac disease is a highly heritable common autoimmune disease involving chronic small intestinal inflammation in response to dietary wheat. The human leukocyte antigen (HLA) region, and 40 newer regions identified by genome wide association studies (GWAS) and dense fine mapping, account for ~40% of the disease heritability. We hypothesized that in pedigrees with multiple individuals with coeliac disease rare [minor allele frequency (MAF) &lt;0.5%] mutations of larger effect size (odds ratios of ~ 2 – 5) might exist. We sequenced the exomes of 75 coeliac individuals of European ancestry from 55 multiply affected families. We selected interesting variants and genes for further follow up using a combination of: an assessment of shared variants between related subjects, a model-free linkage test, and gene burden tests for multiple, potentially causal, variants. We next performed highly multiplexed amplicon resequencing of all RefSeq exons from 24 candidate genes selected on the basis of the exome sequencing data in 2,248 unrelated coeliac cases and 2,230 controls. 1,335 variants with a 99.9% genotyping call rate were observed in 4,478 samples, of which 939 were present in coding regions of 24 genes (Ti/Tv 2.99). 91.7% of coding variants were rare (MAF &lt;0.5%) and 60% were novel. Gene burden tests performed on rare functional variants identified no significant associations (P&lt;1x10-3) in the resequenced candidate genes. Our strategy of sequencing multiply affected families with deep follow up of candidate genes has not identified any new coeliac disease risk mutations.

  Study EGAS00001001093

• Comparison of the diagnostic yield of aCGH and NGS across different neurodevelopmental disorders

  Current recommendations for the genetic diagnosis of neurodevelopmental disorders (NDDs) do not include the use of next generation sequencing (NGS) and are still based on chromosomal microarrays, such as comparative genomic hybridization array (aCGH). The present study compares the diagnostic yield obtained by aCGH and clinical exome sequencing in NDD globally and its spectrum of disorders. To that end, 1412 patients clinically diagnosed with NDDs and studied with aCGH were classified into phenotype categories: Global developmental delay/intellectual disability (GDD/ID); autism spectrum disorder (ASD); and Other NDDs. These categories were further subclassified into subcategories, based on the most frequent accompanying signs and symptoms: isolated forms, forms with epilepsy; forms with micro or macrocephaly and syndromic forms. A subset of 245 patients of the 1412 were subjected to clinical exome sequencing. Diagnostic yield of aCGH and clinical exome sequencing, expressed as the number of solved cases, was compared for each phenotype category and subcategory. Clinical exome sequencing was superior than aCGH for all cases except for isolated ASD, with no cases solved by NGS. Globally, clinical exome sequencing solved 20% of cases (versus 5.7% by aCGH) and the diagnostic yield was highest for all forms of GDD/ID and lowest for Other NDDs (7.1% versus 1.4% by aCGH) and ASD (6.1% versus 3% by aCGH). In the majority of cases, diagnostic yield was higher in the phenotype subcategories than in the mother category. These results support the use of NGS as the first-tier test in the diagnostic algorithm of all NDDs followed by aCGH when necessary.

  Study EGAS00001004949

• Prediction of pigmentation phenotypes by SNP typing in a Northern German population

  Human pigmentation traits are of great interest to many research areas, from ancient DNA analysis to forensic science. We aimed to develop a gene-based predictive model for pigmentation phenotypes in a realistic target population for forensic case work from Northern Germany. Our aim was to determine whether better prediction accuracy can be achieved, or fewer genetic markers may suffice, than in previously studied, genetically more heterogeneous populations. We investigated the association between eye, hair and skin colour, and 12 candidate single nucleotide polymorphisms (SNPs) from six genes. Our study comprised two samples of 300 and 100 individuals from Northern Germany who were carefully characterized with regard to pigmentation phenotypes. The first sample was used to select trait-associated SNPs whereas the second sample served to estimate odds ratios (ORs) and to quantify the predictive capability of the respective SNP genotypes. SNP rs12913832 in HERC2 was found to be strongly associated with blue eye colour (OR=15.6, p&lt;1.2•10-4) and to yield reasonable predictive power (90% sensitivity, 63% specificity). SNP associations with hair and skin colour were weaker and genotypes less predictive. A comparison to two recently published sets of markers to predict eye and hair colour revealed that the consideration of additional SNPs with weak to moderate effect increases the predictive power in Northern Germans for eye colour, but not for hair colour. In addition, fine phenotyping and differentiation of hair colour (light / dark and red tint / no red tint) were found to increase the number of significant genotype-phenotype associations.

  Study EGAS00001001174

• Associations between APOE status and cognitive ability across the lifecourse

  The association between APOE genotype and cognitive function suggests a positive role for the e2 allele and a negative role for the e4 allele. Both alleles have relatively low frequencies in the general population, hence meta-analyses have been based on many small, heterogeneous studies. Here we report the APOE-cognition associations in the largest single analysis to date. APOE status and cognitive ability were measured in 18,337 participants from the Generation Scotland study between 2006 and 2011. The age range was 18-94 years with a mean of 47 (SD 15). Four cognitive domains were assessed: verbal declarative memory (paragraph recall), processing speed (digit symbol substitution), verbal fluency (phonemic verbal fluency), and vocabulary (Mill Hill synonyms). Linear regression was used to assess the associations between APOE genetic status and cognition. Possession of the e4 allele was associated with lower scores on the measures of memory and processing speed in subjects aged &gt;60. Across all age ranges, the e4 allele was linked to better verbal fluency scores. In younger subjects (≤60 years) the e4 allele was linked to higher vocabulary scores. There were no associations between the e2 allele and cognitive ability. As seen in previous meta-analyses, the APOE e4 allele is linked to poorer cognitive performance in the domains of memory and processing speed. By contrast, positive associations were seen between the e4 allele and measures of verbal fluency and vocabulary. All associations were relatively small and, in many cases, nominally significant despite the very large sample size.

  Study EGAS00001001235

• Exome sequencing

  Human telomere biology disorders (TBD), a heterogeneous group of disorders characterized by telomere attrition and premature aging phenotypes, are caused by inherited loss-of-function mutations in telomere-associated genes. Here, we identify three germline heterozygous missense variants in RPA1 gene in four unrelated probands presenting with short telomeres and varying clinical features of TBD including bone marrow failure, myelodysplastic syndrome, T- and B-cell lymphopenia, pulmonary fibrosis, and skin manifestations. All variants cluster to DNA binding domain A of RPA1 protein. RPA1 is an essential single-strand DNA-binding protein essential for DNA replication and repair. RPA1 has been implicated in telomere maintenance but the mechanism remains elusive. Thus far, no genetic disorder had been linked to RPA1. Using FRET assays, we showed that RPA1E240K and RPA1V227A exhibit increased binding to single-strand and telomeric DNA, implying a gain in DNA-binding function while RPA1T270A has binding properties similar to wild type. To study the mutational effect, RPA1E240K, with the highest binding capacity, was knocked-in iPSCs using CRISPR/Cas9, which showed severe telomere shortening and impaired hematopoietic differentiation. Furthermore, in the RPA1E240K patient, stable blood counts over two decades due coincided with clonal rescue hematopoiesis with an RNA-degrading truncating RPA1 mutation and a uniparental isodisomy 17p with loss of RPA1E240K allele. Using single cell DNA sequencing, the two somatic genetic rescue events were proven to be independently acquired in hematopoietic stem cells. In summary, we describe the first human disease caused by germline RPA1 variants in individuals with TBD.

  Study EGAS00001005761

• Analysis of the Elements Involved in the Enrichment of a Panel of Genomic Regions by Nanopore Sequencing Using Adaptive Sampling

  DNA sequencing using nanopore technologies with the affordable MinION device is useful for the identification and characterization of structural variants, long haplotypes, sequencing of repetitive regions and identification of epigenetic modifications. The main limitation of this approach is the low coverage obtained, which might be avoided by adaptive sampling, a computationally controlled method of enrichment for targeted genomic regions. This study dissects the factors involved in the enrichment by adaptive sampling of a panel of 18 human genome regions containing 20 genes implicated in breast cancer sequenced in 16 patients with familial breast cancer negative for NGS screening. An average coverage of 2.0x was obtained for the whole genome and 5.1x for selected regions. Sequencing time was the main factor improving coverage. The selection of long reads (&amp;gt;1 Kb) did not improve the enrichment. The length of the selected region, which in our study ranged from 126 to 565 Kb, did not play a significant role in enrichment. However, the region containing PMS2 showed significantly lower coverage, which could be explained by the high number of PMS2 pseudogenes (N = 14), which were also enriched. Our study shows new evidence of enrichment obtained by adaptive sampling in a panel of genomic regions and shows parameters, the relevance of sequencing time and the role of pseudogenes, that improve the enrichment yield with no library reloading or GPU use, data useful for a more efficient application of this procedure in future studies.

  Study EGAS00001007375

• Efficacy of nivolumab in pediatric cancers with high mutation burden and mismatch-repair deficiency

  Purpose: Checkpoint-inhibitors have limited efficacy for children with unselected solid and brain tumors. We report the first prospective pediatric trial (NCT02992964) using nivolumab exclusively for refractory non-hematological cancers harboring tumor mutation burden (TMB) ≥5 mutations/megabase (mut/Mb) and/or mismatch-repair deficiency (MMRD). Patients and methods: Twenty patients were screened, and ten ultimately included in the response cohort of whom nine had TMB &gt;10mut/Mb (three initially eligible based on MMRD) and one patient had TMB between 5-10 mut/Mb. Results: Delayed immune responses contributed to best overall response of 50%, improving on initial objective responses (20%) and leading to 2-year overall survival (OS) of 50% (95% CI; 27, 93). Four children, including three with refractory malignant gliomas are in complete remission at a median follow-up of 37-months (range: 32.4-60), culminating in 2-year OS of 43% (95% CI; 18.2, 100). Biomarker analyses confirmed benefit in children with germline MMRD, microsatellite instability, higher activated and lower regulatory circulating T-cells. Stochastic mutation accumulation driven by underlying germline MMRD impacted the tumor microenvironment, contributing to delayed responses. No benefit was observed in the single patient with a MMR-proficient tumour and TMB 7.4 mut/Mb. Conclusions: Nivolumab resulted in durable responses and prolonged survival for the first time in a pediatric trial of refractory hypermutated cancers including malignant gliomas. Novel biomarkers identified here need to be translated rapidly to clinical care to identify children who can benefit from checkpoint-inhibitors, including for upfront management of their cancers.

  Study EGAS00001007393

• Peripheral blood DNA methylome in adalimumab-treated patients with rheumatoid arthritis

  Background and aims Rheumatoid arthritis (RA) patients are currently treated with biological agents mostly aimed at cytokine blockade such as anti-TNFα therapy. Currently, there are no biomarkers to predict therapy response to these agents. Here, we aimed to predict response to adalimumab (ADA) treatment in RA patients using DNA methylation in peripheral blood Methods DNA methylation profiling on whole peripheral blood from 92 RA patients before the start of adalimumab (ADA) was determined using Illumina HumanMethylationEPIC BeadChip Array. After 6 months, treatment response was assessed according to the Alliance of Associations for Rheumatology (EULAR) criteria for disease response. Patients were classified as responders (DAS28&lt;3.2 or decrease of 1.2 points) or as non-responders (DAS28&gt;5.1 or decrease of less than 0.6 points). Machine learning models were built with gradient boosting and stability selection models to predict response prior to ADA treatment with predictor DNA methylation markers. Results We demonstrated a 27-feature panel of DNA methylation markers or CpG classifiers that predict response in RA patients treated with ADA. Forty-nine patients were assigned as responder and 43 patients assigned as non-responders. We differentiate responders from non-responders with a high sensitivity (AUC 0.76). The predictor CpGs annotated to genes involved in immunological- and pathophysiological pathways related to RA such as T-cell signaling, B-cell pathology and angiogenesis. Conclusion Our findings indicate that DNA methylation signatures discriminate responders and non-responders to ADA treatment and can serve as a tool for therapy prediction.

  Study EGAS00001007578

• NHLBI TOPMed: The Genetic Epidemiology of Asthma in Costa Rica

  This administrative supplement to the project, "The Genetic Epidemiology of Asthma in Costa Rica" (R37 HL066289) is in response to NOT-HL-14-029 to perform whole genome sequencing (WGS) on existing NHLBI populations. We focus on asthma because of its public health significance. Asthma affects 26 million U.S. children and adults, remains a major cause of morbidity (one-half million hospitalizations a year) and is the most common cause of school and work days lost. Asthma-related costs are estimated to be over $12.7 billion annually. The Asthma Probands for both the extended pedigrees and the trios utilized in this study were selected on the basis of a physician diagnosis of asthma; a history of recurrent asthma attacks or at least 2 respiratory symptoms; and either airway hyperresponsiveness to methacholine or significant response to bronchodilator (Albuterol) administration. These criteria are identical to the criteria used in the Childhood Asthma Management Program (CAMP). The three primary goals of this project are to: (1) identify common and rare genetic variants that determine asthma and its associated phenotypes (height, weight, IgE level, lung function, bronchodilator response, steroid treatment response) through whole genome sequencing (WGS); (2) perform novel family based association analysis of our WGS data to identify novel genes for asthma; and (3) integrate epigenomic and transcriptomic data with our WGS data and determine the epistatic interactions present using systems genomics approaches. Identification of the molecular determinants of asthma remains an important priority in translational science. Genome-wide association studies (GWAS) have been successful in this regard, identifying at least 10 novel susceptibility genes for asthma. However, as with most complex traits, the variants identified by GWAS explain only a fraction of the estimated heritability of this disorder. Herein, we propose a novel family-based study design and state-of-the-art genome sequencing techniques to map a set of sequence variants for asthma and its associated phenotypes and assess the interrelationships of the identified genes and variants using systems genomics methods. We have assembled a team of investigators highly-skilled and expert in whole genome sequencing (Drs. Michael Cho and Benjamin Raby), genetic association analysis (Drs. Scott T. Weiss, Jessica Lasky-Su and Christoph Lange), integrative genomics (Drs. Raby, Kelan Tantisira, Augusto Litonjua and Dawn DeMeo), and systems genomics (Drs. Weiss, Amitabh Sharma, Lange and Raby) to address this important problem with both a novel study design and data set.

  Study phs000988

• Virginia PrIMeD Study

  The goal of this study was to determine whether screening children (2-16 yrs) in pediatric waiting rooms for genetic risk of type 1 diabetes (T1D) could be applied to diverse (by geography, genetic ancestry) communities, and for those at "high genetic risk," if barriers prevent subsequent screening for presence of islet autoantibodies. A total of 3,818 children were recruited from eight general pediatric and specialty clinics across Virginia. Clinical Research Coordinators were stationed at each clinic and obtained informed consent/assent, a brief medical history, and a saliva sample for DNA extraction and determination of their genetic risk using a T1D Genetic Risk Score (T1D GRS). Children were present in the clinic for a "healthy" visit, although children with and without a history of T1D were recruited into the study. Age, sex, self-reported ancestry/race, T1D history, and other medical history information was obtained. DNA was extracted from the saliva samples and genotyping with a T1D-focused array using 74 SNPs (including 26 SNPs in the HLA region) provided data to generate a T1D GRS for each individual. Of the 3,818 children recruited, there was a slight excess of males (1,959, 51.3%), a majority of white and Hispanic/Latino ancestry (82.8%), and 91 (2.4%) with prevalent T1D. A total of 542 (14.2%) had "high T1D genetic risk" (T1D GRS > 5). Although the T1D GRS included non-HLA genetic variants, 80% of those with "high genetic risk" would have been classified using only those SNPs in the HLA region alone. Of children with "high genetic risk" and without pre-existing T1D (n=494), 7.0% (34/494) consented for islet autoantibody screening. Among children with pre-existing T1D (n=91), 52% (n=48) also had a "high genetic risk." The HLA-focused T1D GRS was not significantly associated with age at onset of T1D. Of those at high genetic risk that consented for autoantibody testing (n=34) and completed the blood collection (n=28), two (2/28, 7.1%) tested positive for multiple autoantibodies and were at significant risk of developing T1D. Follow up of 55% of parents/guardians identified 2 (of 2,096, 0.095%) participants who developed T1D, one at "high genetic risk" and one with a first-degree relative with T1D. A major factor in obtaining islet autoantibody testing was concern over SARS-Cov-2 exposure and an independent non-scheduled blood collection.Individual-level data at each SNP (genotyped and imputed) for T1D genetic risk and affiliated data are provided in dbGaP.

  Study phs003609

• RNA-seq of longitudinal human blood and nose swab samples from a controlled human infection experiment with SARS-CoV-2 virus

  Controlled human infection experiments enable longitudinal profiling of immune responses to a pathogen. 36 healthy volunteers aged 18-29 years, with no evidence of previous infection or vaccination, were inoculated with SARS-CoV-2 virus and quarantined for 14 days. Blood samples (n=374) for RNA sequencing were collected into PAXgene tubes before virus challenge, 6 hours after challenge, daily thereafter for 14 days and on day 28. Mid-turbinate nose swabs (n=96) for RNA sequencing were collected before virus challenge, and on days 1, 3, 5, 7, 10 and 14 after challenge, preserved in RNAprotect. 18 of 36 participants developed a replicative SARS-CoV-2 infection as evidenced by consecutive PCR-positive swabs for the virus. For every participant, blood RNA from selected days were extracted and depleted for genomic DNA and globin mRNA, before cDNA libraries were constructed using KAPA RNA HyperPrep with RiboErase kits. Libraries were sequenced on the Illumina NovaSeq 6000 platform using NovaSeq 6000 S4 Reagent Kits (200 cycles). Nose swab RNA samples were extracted and depleted for genomic DNA before cDNA libraries were constructed using KAPA mRNA HyperPrep Kits. Libraries were sequenced on the Illumina NextSeq platform the using the NextSeq 500/550 High Output Kit (75 cycles).

  Dataset EGAD50000000942

• Somatic mutation rates scale with lifespan across mammals

  The rates and patterns of somatic mutation in normal tissues are largely unknown outside of humans. Comparative analyses can shed light on the diversity of mutagenesis across species and on long-standing hypotheses regarding the evolution of somatic mutation rates and their role in cancer and ageing. Here, we used whole-genome sequencing of 208 intestinal crypts from 56 individuals to study the landscape of somatic mutation across 16 mammalian species. We found somatic mutagenesis to be dominated by seemingly endogenous mutational processes in all species, including 5-methylcytosine deamination and oxidative damage. With some differences, mutational signatures in other species resembled those described in humans, although the relative contribution of each signature varied across species. Remarkably, the somatic mutation rate per year varied greatly across species and exhibited a strong inverse relationship with species lifespan, with no other life-history trait studied displaying a comparable association. Despite widely different life histories among the species surveyed, including ~30-fold variation in lifespan and ~40,000-fold variation in body mass, the somatic mutation burden at the end of lifespan varied only by a factor of ~3. These data unveil common mutational processes across mammals and suggest that somatic mutation rates are evolutionarily constrained and may be a determinant of lifespan.

  Dataset EGAD00001008032

• scRNA-seq of HSPC treated with gemcitabine and carboplatin

  4 samples of 2000000 HSPCs in 2 ml media each were then prepared and treated for 24 hours: carboplatin-high: 150 µg/ml carboplatin carboplatin-low: 18.75 µg/ml carboplatin gemcitabine: 25 ng/ml gemcitabine control: no drug Single-cells were subsequently extracted using the ddSEQ™ Single-Cell Isolator (Bio-Rad) and later sequenced using the SureCell™ Whole Transcriptome Analysis 3' Library Prep Kit (Illumina) on the NextSeq 500 System (Illumina) using the NextSeq 500/550 High Output Kit v2.5 150 Cycles (Illumina) all following the manufacturer's instructions. The raw FASTQ-files are available as read 1 and read 2 files from 4 lanes for each sample giving a total of 16 FASTQ-files. The FASTQ-files were also processed following the ddSeeker (Romagnoli, D., et al., BMC Genomics 2018, doi:10.1186/s12864-018-5249-x) and Drop-seq (Macosko, E.Z., et al., Cell 2015, doi:10.1016/j.cell.2015.05.002) protocols for processing scRNA-seq data to yield the final digital gene expression (dge) for each cell of each sample. This has resulted in one dge text file and one dge summary text file per sample. These 8 text-files with dge data are found in the zip-compressed analysis data-file.

  Dataset EGAD00001006080

• Targeted replication of LVOTO genes

  Non-syndromic cases of congenital heart defects (CHD) exhibit variable modes of inheritance (Mendelian and non-Mendelian). Several studies have identified strong candidates in humans by taking a candidate gene approach as well as by using whole exome next generation sequencing (NGS). So far these studies could only explain a minor fraction of the observed phenotype in humans, most of them in syndromic cases and no single study has focused on the subset of cases with left ventricular outflow tract obstruction (LVOTO). To discover novel disease-causing genes a large cohort of patients with LVOTO, approximately 100 cases, 25 families and 100 trios have been exome sequenced. This study based on NGS sequencing data yielded several known and novel compelling candidate genes, such as MYH6, NR2F2 and MYH11, but also novel ones, such as ITGB4. To evaluate the significance of our findings in a replication cohort we assembled another 1614 cases with an LVOTO phenotype from our collaborators in Toronto, Berlin and Amsterdam. Targeted resequencing in this additional cohort will help to find additional cases with mutations in the identified candidate genes to strengthen genotype-phenotype association. We will use control data from the INTERVAL project for case/control analyses The pulldowns will be performed as 24-plex ISC with 192 or greater indexes, and the sequencing will be performed with 192 samples per lane, requiring 9 lanes of sequencing.

  Dataset EGAD00001002212

• Targeted de novo phasing and long-range assembly by template mutagenesis

  Long-range sequencing with low error rate has been challenging. Sequence assembly and phasing usually require a high-quality reference genome for mapping, so working on highly-variable genomic regions or regions with no reference genome information would be difficult. In this study, we describe novel bench protocols and algorithms to obtain ultra-low-error-rate haplotype-phased sequence assemblies of regions 10 KB in length using a short-read sequencing platform that simultaneously solves the above two problems. We accomplish this by imprinting each template strand from a target region with a dense and unique mutation pattern. The mutation process randomly and independently converts ~50% of cytosines to uracils. Short-read sequencing libraries are made from both mutated and unmutated templates. A conservative de Bruijn graph approach seeds an assembly of the mutated templates, which we then extend by mapping paired-end reads. We next partition the template assemblies into two or more haplotypes after using the unmutated sequence library to recover almost all of the mutated bases. The final haplotype is assembled and corrected for residual template mutations and PCR errors. We obtain per-base-error rates below 10 9. We apply this method to a human family, correctly assembling and phasing three genomic intervals, including the highly polymorphic HLA-B gene.

  Dataset EGAD00001008444

• Targeted Myeloid DNA-Panelsequencing, MLL

  Sequencing data from a targeted myeloid DNA-Panelsequnencing at the MLL Dx, Munich lab. Targeted sequencing was performed using the Nextera DNA Flex library preparation kit, starting with 100ng of genomic DNA (Illumina, San Diego, CA, USA). The target regions were enriched by a custom xGen Lockdown panel using a hybridization capture workflow (IDT Integrated DNA Technologies, Coralville, IA, USA). All libraries were sequenced with 100bp paired-end reads on a NovaSeq6000 (Illumina) with a mean coverage of 3206x. Somatic variant calling was performed with Pisces and a sensitivity cut off of 2%. Large deletions and medium-sized insertions, as they are for example found in CALR and FLT3, were called with Pindel. Variant annotation considered the publicly available data bases Cosmic (v91), ClinVar (2020-03), gnomAd (non-cancer, v2.1.1), dbNSFP (v3.5) and UMD TP53 (2017_R2). Variants that are described as somatic, protein truncating or affecting splice sites were considered as mutations while variants with no or discrepant data base information were considered as variant of uncertain significance.

  Dataset EGAD00001008485

• EGA submission account terms

  Submission account terms Each EGA submission account has a dedicated submission box. To ensure continued service to all submitters EGA submission boxes should not exceed 8Tb in size, and cannot exceed 12Tb. Those that are near to or exceeding this value will be monitored on a weekly basis by the helpdesk team and the submitter will be contacted and guidance provided accordingly on completing their metadata registration. You will be able to register your metadata associated with the files in your submission account via EGA Submitter Portal, Webin or the EGA REST API. EGA also requests that metadata is submitted within 60 days of upload data to a submission box. When we detect files that are over 60 days old in a submission box we will notify the submitter requesting that metadata be submitted or the files removed from the submission box. It is important to note that data is not archived at the EGA until the metadata describing the files has been associated and the archival process has completed. It is only when associated metadata has been submitted that the archival process into EGA is fully triggered. EGA removes files from the submission box once they have been successfully archived, so no action is required by a submitter to remove successfully archived files. Files over 90 days old will be deleted from the submission box unless EGA Helpdesk has been contacted and confirmed an exemption with the submitter.

  Documentation submission/terms

• CIDR: The Role of Rare Coding Variation in Prostate Cancer in Men of African Ancestry - RESPOND Project 2

  In RESPOND Project 2, we seek to identify rare genetic factors that are associated with prostate cancer (PCa) risk and aggressiveness in men of African ancestry (AA). We will conduct exome sequencing of 15,000 prostate cancer cases and 5,000 controls from the RESPOND cohort and the African Ancestry Prostate Cancer Consortium (AAPC) with cases selected based on risk categories: high-risk (stage T3/T4 or Gleason 8+ or PSA&gt;20 ng/ml), intermediate-risk (stage T2b/T2c or Gleason 7 or PSA 10-20 ng/ml) and low-risk disease (stage T1/T2a and Gleason &#8804; 6 and PSA&lt;10 ng/ml). We expect the findings from this Project to significantly advance knowledge of susceptibility to aggressive PCa and racial/ethnic disparities in PCa risk, and to guide the development of future preventive, early detection and prognostic measures for AA men. The first phase of the study will include exome sequence data for approximately 7,500 cases and 5,000 controls from AAPC. The second phase of the study will include exome data for 5,000 cases from RESPOND and AAPC.ACKNOWLEDGMENTS and CONTRIBUTING SITES Multiethnic Cohort (MEC): The MEC and the genotyping in this study were supported by National Institutes of Health (NIH) grants CA63464, CA54281, CA1326792, CA148085, and HG004726. Cancer incidence data for the MEC and Los Angeles Study of Aggressive Prostate Cancer (LAAPC) studies have been collected by the Los Angeles Cancer Surveillance Program of the University of Southern California with federal funds from the National Cancer Institute (NCI)/NIH/Department of Health and Human Services (DHHS) under Contract No. N01-PC-35139, and the California Department of Health Services as part of the state-wide cancer reporting program mandated by California Health and Safety Code Section 103885, and grant 1U58DP000807-3 from the Centers for Disease Control and Prevention (CDC). Ghana Prostate Study (GPS): The Ghana Prostate Study was funded by the Intramural Program of the NCI/NIH/DHHS under Contract No. HHSN261200800001E. Men of African Descent and Carcinoma of the Prostate (MADCaP): We thank all MADCaP study participants. This work is a product of the MADCaP network. This work was supported by NCI/NIH grant U01CA184374 to Timothy Rebbeck and National Institute of General Medical Sciences (NIGMS) MIRA grant R35GM133727 to Joseph Lachance. Additional funding includes a seed grant from the Integrated Cancer Research Center at Georgia Institute of Technology. UGANDA: The UGANDA study was supported by NIH grant R01CA165862. Southern Community Cohort (SCCS) is funded by NIH grant CA092447. SCCS sample preparation was conducted at the Epidemiology Biospecimen Core Lab that is supported in part by the Vanderbilt Ingram Cancer Center (CA68485). Data on SCCS cancer cases were provided by Alabama Statewide Cancer Registry, Kentucky Cancer Registry, Office of Cancer Surveillance at Tennessee Department of Health, Florida Cancer Data System, Central Cancer Registry at North Carolina Division of Public Health, Georgia Comprehensive Cancer Registry, Louisiana Tumor Registry, Mississippi Cancer Registry, South Carolina Central Cancer Registry, Virginia Cancer Registry at Virginia Department of Health, and Cancer Registry at Arkansas Department of Health. The Arkansas Central Cancer Registry is fully funded by a grant from National Program of Cancer Registries (NPCR)/Centers for Disease Control and Prevention (CDC). Data on SCCS cancer cases from Mississippi were collected by the Mississippi Cancer Registry which participates in the NPCR/CDC. The contents of this publication are solely the responsibility of the authors and do not necessarily represent the official views of the CDC or the Mississippi Cancer Registry. Karuprostate: The Karuprostate study was supported by the French National Health Directorate and by the Association pour la Recherche sur les Tumeurs de la Prostate. S&#233;verine Ferdinand, Marc Romana. The North Carolina - Louisiana Prostate Cancer Project (PCaP) is carried out as a collaborative study supported by the Department of Defense contract DAMD 17-03-2-0052. The authors thank the staff, advisory committees and research subjects participating in the PCaP study for their important contributions. We would also like to acknowledge the UNC BioSpecimen Facility and the LSUHSC Pathology Lab for our DNA extractions, blood processing, storage and sample disbursement (https://genome.unc.edu/bsp). Gene-Environment Interaction in Prostate Study (GECAP) was supported by NIH grant ES011126. King County Prostate Cancer Study (KCPCS) was supported by NIH grants CA056678, CA082664, and CA092579, with additional support from the Fred Hutchinson Cancer Research Center. We thank the participants in these studies, and Ms. Suzanne Kolb for help with study management. The Los Angeles Study of Aggressive Prostate Cancer (LAAPC) was funded by grant 99-00524V-10258 from the Cancer Research Fund, under Interagency Agreement #97-12013 (University of California contract #98-00924V) with the Department of Health Services Cancer Research Program. Cancer incidence data for the MEC and LAAPC studies have been collected by the Los Angeles Cancer Surveillance Program of the University of Southern California with Federal funds from the NCI/NIH/DHHS under Contract No. N01-PC-35139, and the California Department of Health Services as part of the state-wide cancer reporting program mandated by California Health and Safety Code Section 103885, and grant 1U58DP000807-3 from the CDC. Prostate Cancer Studies at MD Anderson (MDA) was supported by grants CA68578, ES007784, DAMD W81XWH-07-1-0645, and CA140388.

  Study phs002637

• Whole_genome_sequencing_of_100_DDD_trios_with_suspected_noncoding_causal_mutations

  We would like to whole-genome sequence 100 trios (300 individuals) from the DDD project in which we suspect the proband has a pathogenic noncoding mutation, to 30X coverage. These have been carefully selected to maximise the chances of finding pathogenic noncoding mutations. Specifically, we have selected probands who have no likely causal mutation within the coding region but who have a heterozygous loss-of-function mutation inherited from one parent in a known recessive developmental disorder gene, and whose phenotype is a good match for the typical phenotype caused by recessive variants in that gene. We suspect that these probands may have a heterozygous noncoding mutation on the other allele which reduces expression of the gene so that, together, these two mutations are effectively equivalent to a homozygous loss-of-function mutation. About a quarter of the probands have affected siblings and, if those siblings have been exome sequenced, we have checked that they are identical-by-descent in the region of the gene. We propose to search in the promoter region, intronic and annotated regulatory regions around these genes of interest for rare variants inherited from the other parent (i.e. not the one who transmitted the loss-of-function variant), or de novo variants. If we find such variants, and demonstrate an enrichment of this compound heterozygous LoF noncoding mechanism above that expected by chance, we intend to follow up specific candidates by RNAseq and potentially CRISPR/Cas9 experiments. This data is part of a pre-publication release. For information on the proper use of pre-publication data shared by the Wellcome Trust Sanger Institute please see http://www.sanger.ac.uk/datasharing/

  Study EGAS00001002452

• Bladder Cancer Organoids as a Functional System to Model Different Disease Stages and Therapy Response

  The study 'Bladder Cancer Organoids as a Functional System to Model Different Disease Stages and Therapy Response' reports the genomic characterization at the single allele level upon Whole Exome Sequencing of Bladder Cancer (BLCa) Patient Tumor (PT) samples obtained from 18 patients and corresponding Patient Derived Organoids (PDOs), implementing the SPICE pipeline (Ciani Y et al). Our cohort included 21 PTs and 22 PDOs. In this study, we generated a unique biobank resource of BLCa organoids that encompasses a broad spectrum of disease stages. We demonstrated that PDOs retain cancer heterogeneity and mutational burden. Furthermore, we observed that BLCa PDO can have different morphologies which may reflect tumor-specific features. Indeed, a solid morphology may be associated with more advanced tumors, as supported by the high PDO genomic burden and high primary tumor stage. Moreover, with our drug screening pipeline, we demonstrated the clinical relevance and feasibility of using a BLCa PDO-based drug assay for comparing the standard of care (SOC) drugs with compounds not currently in use for BLCa. Response to SOC was highly heterogeneous with approximately 50% of non-muscle invasive bladder cancer and 33% of muscle invasive bladder cancer PDOs showing no or minimal response. The most effective targeted therapy was lapatinib. We used an integrative analysis of drug response profiles with matched PDO genomic analysis to determine enrichment thresholds for candidate markers of therapy resistance and sensitivity. Importantly, by assessing the clinical history of longitudinally sampled cases, the clonal evolution of the disease could be determined and matched with PDO drug response profiles.

  Study phs003149

• Mechanism of Action of Vitamin E in NAFLD

  We conducted a single-center prospective mechanistic randomized trial (clinicaltrials.gov NCT01792115). Adult patients with NAFLD were enrolled between May 2013 and November 2016. All patients underwent a 12 weeks run-in period of lifestyle intervention. Following the run-in phase, patients underwent a baseline evaluation including blood tests, glucose tolerance testing, measurement of liver fat content by 1H-magnetic resonance spectroscopy (MRS) and a percutaneous liver biopsy. Patients with no evidence of NAFLD on the entry liver biopsy ended their participation in the study at that point. Patients with histological confirmation of NAFLD (either NAFL or NASH) on the baseline biopsy were randomized at a 1:1:1 ratio to receive one of 3 doses of natural vitamin E (RRR-alpha-tocopherol, Pharmavite), 200, 400 or 800 IU/d, for 24 weeks, and were followed at a 1-8 week intervals in a parallel assigned design. While on vitamin E treatment, patients were encouraged to continue and adhere to the lifestyle and dietary guidelines of the run-in phase. On week 4 of vitamin E treatment, a repeat MRS and liver biopsy were performed. On week 24, a final MRS was performed to assess treatment response. The primary endpoint was the reduction of liver fat content by MRS from baseline (immediately prior to starting vitamin E) to week 24 of treatment, and a response was defined as a relative reduction of &#8805;25% in liver fat content by MRS between baseline and week 24. Early changes in the hepatic transcriptome were investigated at week 4 comparing non-responders to responder in a nested design.

  Study phs001930

• PROstate Cancer Medically Optimized Genome Enhanced ThErapy (PROMOTE) of Castration Resistant Prostate Cancer (CRPC) Patients Treated with Abiraterone Acetate

  The Prostate Cancer Medically Optimized Genome-Enhanced Therapy (PROMOTE) study uses genetic clues in castration-resistant prostate cancer that may identify an individualized treatment approach for men with the disease. Understanding the molecular biology behind castration-resistant prostate cancer has led to more treatment options, but there are still no definite conclusions about which specific drug best treats patients - maximum suppression of cancer growth while minimizing side effects. The PROMOTE study explores the genetic characteristics of each tumor to predict these treatment paradigms for the future, resulting in more effective and less toxic options for patients. Our long-term goal is to improve treatments for men with advanced prostate cancer by using genomic sequencing to increase life span and quality of life. We also will uncover novel vulnerable targets in the cancer genome that may provide new drug therapies. PARTICIPATION Eligible participants are men: With castration-resistant prostate cancer or prostate cancer not responding to hormone treatments About to begin abiraterone acetate therapy Agreeable to undergoing two tumor biopsies During the study, participants travel to Mayo Clinic for an initial biopsy (before beginning abiraterone acetate) and a second biopsy approximately three months later. The cell tissue collected is analyzed to identify gene alterations in the tumor that could eventually be targeted with treatments. Tissue is preserved for future research. Participants can continue to be treated by their local cancer care team during this period and beyond. In addition, the Mayo team carefully monitors participants' cancer via follow-up studies and the genetic signature of tumors that were biopsied so that patients may benefit from future treatments.

  Study phs001141

• Mitochondrial DNA deletion detection in POLG patients

  Accumulation of deletions in the mitochondrial genome is a pathological hallmark of both aging and a broad range of neurometabolic disorders. Prior mtDNA deletion studies have been limited by reliance upon either long range PCR or NGS strategies that target specific regions of the mitochondrial genome. We developed an ultrasensitive high throughput sequencing process (LostArc) to identify and quantify all deletions in circular DNA molecules while minimizing interference from linear DNA and PCR artefacts. We applied this process to skeletal muscle biopsies from 19 unaffected individuals (aged 17 to 93 years) and from 22 patients with heritable mitochondrial disease (17 to 80 years) bearing pathogenic variants in POLG, the nuclear gene encoding the catalytic subunit of the mitochondrial replicative DNA polymerase &#611; (Pol &#611;). To our astonishment, LostArc revealed 35 million mtDNA deletions with ~470,000 unique deletion spans. This unprecedented volume of data enabled detailed and highly informative bioinformatics analyses. We show that ablation of mtDNA is sufficient to explain skeletal muscle phenotypes associated with aging and POLG-derived disease, with a clear threshold separating symptomatic and asymptomatic individuals. Unsupervised hierarchical clustering and principle component analysis reveal distinct age- and disease-correlated deletion patterns that implicate DNA replication by Pol &#611; in the formation of mtDNA deletions. Deletion patterns also point to active mtDNA replication but little or no purifying selection against deleted mtDNA by mitophagy in postmitotic muscle fibers. Monte Carlo simulations of varied hypotheses for deletion formation, based in part on competing models of mtDNA replication, suggest the observed patterns are most consistent with deletions initiated by replication fork stalling during strand displacement mtDNA synthesis. Note that sequence data for HEK293 can be found under BioProject PRJNA642884.

  Study phs002052

• Resistance to Checkpoint Blockade Therapy Through Inactivation of Antigen Presentation

  Although immune checkpoint blockade (CPB) leads to prolonged responses in 15-40% of patients with metastatic melanoma, treatment refractory disease and progression after initial response remain major causes of mortality. While predictors of response have been reported, the common mechanisms of both primary and acquired resistance are poorly understood. To identify mechanisms of resistance and examine the evolving landscape in response to CPB, we performed whole exome sequencing (WES), immunohistochemistry (IHC), and RNA-sequencing (RNAseq) of longitudinal tumor biopsies from 17 metastatic melanoma patients treated with various CPB therapies. We found no significant changes in both mutational and neoantigen loads over time between responders and nonresponders. However, we identified abnormalities in one gene, beta-2-microglobulin (B2M), an essential component of MHC Class I antigen presentation, that were present in samples during disease progression but not regression. In total, we identified B2M aberrations in 29.4% of patients, including multiple early frameshift mutations, loss of heterozygosity (LOH) overlapping B2M, and absence of tumor-specific B2M protein expression. Additional defects in the antigen presentation and IFN&#947; pathways were identified but were not restricted to progressing lesions in our cohort. In two independent cohorts of 105 and 38 melanoma patients treated with ipilimumab (anti-CTLA4) and pembrolizumab (anti-PD1) respectively, we found that B2M LOH was enriched 3-fold in nonresponders (~30%) vs. responders (~10%) and associated with poorer overall survival (log-rank p=0.01, p=0.006). Loss of both copies of B2M was found only in nonresponders. We also found evidence for association of LOH overlapping IFNGR1 with poorer overall survival exclusively in the anti-PD1 cohort. Thus, B2M loss is likely a common mechanism of primary and acquired resistance to therapies targeting CTLA4 or PD-1.

  Study phs001427

• Gene mutation and rescue in congenital diaphragmatic hernia

  Congenital Diaphragmatic Hernia (CDH) is a common and life-threatening malformation. The most common phenotype is left-sided posterolateral (Bochdalek-type) hernia accompanied by pulmonary hypoplasia. Due to the high mortality, most of the cases have no family history for CDH. The rare presumed Mendelian instances of CDH have the potential to be extremely informative about the molecular mechanisms generating this phenotype. Three multiplex families were deemed meritorious for study by whole exome sequencing: Parents who were 1st cousins gave birth to 2 children with CDH (one of whom also had cleft lip and palate), both now deceased. A DNA sample was available from one child. We proposed to perform a SNP array to identify regions of homozygosity by descent and intersect these regions with variants detected on exome sequencing. This effort has led to the identification of 3 confirmed variants. We are determining which if any are expressed in the primordial developing diaphragm and are also using bioinformatic approaches to determine which of these candidates interact with known CDH genes. This non-consanguineous couple gave birth to 2 affected boys with an identical and lethal CDH phenotype (consisting of diaphragmatic hernia, epigastric hernia, and facial dysmorphism) and one healthy boy. We predicted X-linked inheritance and findings from X chromosome microsatellite marker analysis are consistent with this. Accordingly, we are focusing on sequence variants mapping to the X chromosome. We have identified a single candidate gene and confirmed that the variant follows an X-linked pattern of inheritance in all family members. Expression studies are underway. These affected males are second cousins. By virtue of their position in the pedigree, we predicted an inheritance pattern consistent with X-linked inheritance. Data from WES and microsatellite marker analysis are currently being integrated.

  Study phs000485

• National Cancer Institute Clinical and Laboratory Analysis of Familial Cancer

  The purpose of this study is to continue the analysis begun on 09C0079 which was focused on identification of the genetic mutation associated with a new gastric polyposis syndrome, Gastric Adenocarcinoma and Proximal Polyposis of the Stomach (GAPPS). GAPPS, first described in 2012, is an autosomal dominant gastric polyposis syndrome that confers a substantial risk for gastric adenocarcinoma. The GAPPS phenotype consists of a carpeting of greater than 100 fundic gland polyps (FGPs) in the oxyntic mucosa of the gastric body and fundus, with antral sparing with some FGPs displaying high-grade dysplasia. This is in contrast to sporadic FGPs that are benign, most often are not associated with high-grade dysplasia though low-grade dysplasia has been reported, and are fewer in number. Gastric FGPs have been found to be associated with Familial Adenomatous Polyposis (FAP) and attenuated FAP, which is an autosomal dominant condition associated with germ line mutations in APC which confers a phenotype consisting of multiple (&gt;100) adenomatous polyps in the colon and rectum developing after the first decade of life. FGPs are reported to occur in 12-84% of patients with FAP, whereas sporadic FGPs are identified in ~5% of patients undergoing upper gastrointestinal endoscopy. Gastric adenocarcinoma arising from FGPs in individuals with FAP have been reported. Other manifestations of FAP and AFAP include polyps in the upper gastrointestinal tract, congenital hypertrophy of retinal pigment epithelium, osteomas and epidermoid cysts, supernumerary teeth, desmoid formation, and other malignancies such as thyroid, small bowel cancer, hepatoblastoma, and medulloblastoma. Thus far there are no reports in GAPPS of polyposis arising in the esophagus, gastric antrum, pylorus, small intestine, or colon.

  Study phs001935

• Exome Sequencing for Diseases of the Immune System: X-linked Immunodeficiency with Magnesium Defect, EBV Infection, and Neoplasia (XMEN)

  The etiologies of primary immunodeficiencies often yield novel insights about the immune system. Although a genetic etiology has been suspected for patients with abnormally low CD4+ T cells in the absence of HIV infection or any known causes of lymphopenia, no genetic mutation has been described to date for any case of primary CD4 lymphopenia. In this study, we characterized a non-consanguineous family with two non-HIV infected boys exhibiting an inverted CD4 to CD8 T cell ratio and a history of recurrent chronic viral infections since birth. Consistent with a decreased thymic output of CD4+ T cells, the percentage of CD31+ cells in the CD4+ naive population of these patients was decreased. In addition, the activation of T cells was significantly impaired in the patient upon TCR stimulation. Given the mother&#39;s T cells show completely skewed X chromosome inactivation, we suspected that the nature of this disease is X-linked. We performed X-chromosome exon-capture targeted single-end Solexa sequencing on two brothers and the mother and found a 10 base pair deletion at an intron-exon junction of Magnesium Transporter 1 (MAGT1), a Mg2+ specific transporter. We confirmed that this deletion leads to altered splicing, frameshift, early termination of the mRNA, and deficient protein expression in the lymphocytes of the two patients. Moreover, knockdown of this transporter in T cells isolated from healthy donors can recapitulate the observed T cell activation defect while its ectopic expression in the patients&#39; lymphocytes can restore T cell stimulation. Our discovery highlights the significance of this transporter to T cell function.

  Study phs000365

• Genetic Analysis of Parkinson's Disease

  A portion of the sample includes multiplex PD families recruited as part of a previously funded study (R01NS037167; PI: Tatiana Foroud). Families were ascertained initially through a living affected sibling pair. Subsequently, ascertainment was loosened to include PD probands having a positive family history of PD in a first degree relative, who was not required to be part of the study. All individuals completed an in-person evaluation at one of the &#62; 65 Parkinson Study Group sites participating in this study. Individuals completed a detailed evaluation that included the Unified Parkinson Disease Rating Scale (UPDRS), Mini-Mental State Examination (MMSE), Geriatric Depression Scale (GDS) and a diagnostic checklist that implemented the UK PD Brain Bank inclusion and exclusion criteria. An autopsy program was established in this study and brain tissue for confirmation of diagnosis and further experimental study are available from over 100 participants. Autopsy confirmation of the PD diagnosis was nearly 100% for those who met UK PD Brain Bank criteria. In addition, because this was a cross sectional study and family members were often seen very early in disease course, among those who did not meet UK PD Brain Bank criteria but had a subsequent autopsy, a significant subset (&#62;1/3) had autopsy confirmation of PD. The second source of samples is from the NINDS Human Genetics DNA and Cell Line Repository located at the Coriell Institute. Currently DNA from 4,509 PD patients is available from the Coriell Institute. We restricted the selected sample to include only Caucasians and ensured there was no overlap with samples contributed by the PROGENI Study.

  Study phs001004

• Glial Cell Line-Derived Neurotrophic Factor (GDNF) Polymorphisms and Anxiety, Depression

  GDNF gene variants were studied as possible risk factors of depression or anxiety on a young sample. The association study involved eight (rs1981844, rs3812047, rs3096140, rs2973041, rs2910702, rs1549250, rs2973050 and rs11111) GDNF single nucleotide polymorphisms and anxiety and depression scores measured by the Hospital Anxiety and Depression Scale (HADS) on 708 Caucasian young adults with no psychiatric history. Results provided significant effects of two single nucleotide polymorphisms on anxiety scores following the Bonferroni correction for multiple testing (p=0.00070 and p=0.00138 for rs3812047 and rs3096140, respectively). Haplotype analysis confirmed the role of these SNPs (p=0.00029). A significant sex-gene interaction was also observed since the effect of the rs3812047 A allele as a risk factor of anxiety was more pronounced in males. This is the first demonstration of a significant association between the GDNF gene and mood characteristics demonstrated by the association of two SNPs of the GDNF gene (rs3812047 and rs3096140) and individual variability of anxiety using self-report data from a non-clinical sample. Reprinted from Kotyuk et. al., 2013 (Kotyuk, E., Keszler, G., Nemeth, N., Ronai, Z., Sasvari-Szekely, M., and Szekely, A. (2013). Glial Cell Line-Derived Neurotrophic Factor (GDNF) as a Novel Candidate Gene of Anxiety. PLoS One,8, (12) PMID: 24324616), with permission from Publisher (All content of articles published in PLOS journals is open access. You can read about our open access license here: http://www.plos.org/about/open-access/. To summarize, this license allows you to download, reuse, reprint, modify, distribute, and/or copy articles or images in PLOS journals, so long as the original creators are credited (e.g., including the article&#39;s citation and/or the image credit); Laura Perry; Staff EO;PLOS ONE)

  Study phs000713

• Lung Cancer Genetic Study Among Asian Never Smokers

  The Lung Cancer Genetic Study Among Asian Never Smokers includes data from four population-based studies conducted among Asian women: the Shanghai Women's Health Study (SWHS), the Hwasun Cancer Epidemiology Study (HCES1 and HCES2), the Female Lung Cancer Consortium in Asia (FLCCA, including sFLCA), and the Guangzhou Lung Cancer Study (GLCS). Shanghai Women's Health Study (SWHS): The SWHS is a population-based prospective cohort study of approximately 75,000 adult women who were recruited between 1997 and 2000. The cohort has been followed by a combination of record linkage and active follow-ups. Lung cancer patients identified in the SWHS and non-cases were included in the current study.Hwasun Cancer Epidemiology Study-Lung Cancer (HCES1 and HCES2): The HCES is a hospital-based case-control study whose goal is to identify factors of the cancer development and clinical progression in a Korean population. Included in this project were 299 female lung cancer cases from HCES2, who were diagnosed between April 2004 and February 2013 at Chonnam National University Hwasun Hospital, a cancer specified hospital in Jeollanam-do province, South Korea. Controls were randomly selected from women with no previous cancer diagnosis at enrollment in the Namwon Study and the Dong-gu study, ongoing community-based cohort studies in South Korea. A total of 1,103 controls (273 from HCES1 and 830 from HCES2) were genotyped by MEGA. Guangzhou Lung Cancer Study (GLCS): The GLCS is a case-control study including never-smoking women lung cancer cases and controls from Guangzhou, China.In this study, the DNA samples of 1,195 subjects from SWHS and GLCS were genotyped using whole exome sequencing (WES), and the DNA samples of 7,174 subjects from SWHS, HCES1, HCES2, FLCCA, and sFLCA were genotyped using the Multi-Ethnic Global Array (MEGA).

  Study phs002366