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• PGRN-RIKEN: Genetic Determinants of Clinical Cardiovascular Events in Patients Receiving Statins

  Coronary heart disease (CHD) is an important public health problem in developed countries. Statins are effective in the prevention and treatment of CHD; nevertheless, many patients receiving statins still suffer cardiovascular events (CV) such as heart attack. Identifying genetic variants responsible for differential clinical responses to statins will not only allow individual patients at high residual risk to be targeted for additional therapies, but also will define new biologic pathways contributing to statin response, and thus new targets for future therapies. Accordingly, the goal of this study is to identify genetic variants associated with clinical CV in patients receiving statins. Subjects identified for study are of European descent and include 1718 subjects with CV while on statins (cases) and 4172 subjects without CV while on statins (controls). Key research resources utilized in this effort include VanderbiltD's BioVU DNA databank and associated Synthetic Derivative database of clinical information, and software tools developed to identify drugs and clinical events using Electronic Health Record-derived structured and unstructured ("free text") data. Most cases and controls identified include three data types: ICD-9 codes, medication regimens, and medical test results. Genotyping, using IlluminaD's Infinium HumanOmniExpressExome BeadChip (OmniExpressExome), was performed by the RIKEN Integrative Medical Sciences Center (IMS) and supported by the Pharmacogenomics Research Network (PGRN)-RIKEN IMS Global Alliance.

  Study phs000963

• The National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) Genetics of Genetic Epidemiology of Metabolic Syndrome in an Island Population

  The major objective of this study was to conduct a systematic genetic study of metabolic traits involved in metabolic syndrome through collection and analysis of epidemiological, demographic, environmental, and relevant biological and clinical data from a relatively isolated island population of the eastern Adriatic coast of Croatia. The population was chosen for the following reasons: 1) in spite of practicing a largely traditional life style and dietary habits, high rates of obesity, arterial hypertension, dyslipidemia and related metabolic abnormalities were found in previous studies; 2) the population was established by a relatively small number of founders, predominantly of Slavic descent from the mainland during 15th to 18th century AD, a genetically homogeneous population living in a homogeneous environment; 3) sharing a common European ancestry, a relevant population for study in the context of the general US population; 4) Croatian collaborators have been conducting anthropological and genetic studies in these communities for over three decades. There were two major aims of the study: 1) to recruit ~1200 adult participants and collect blood samples together with demographic, anthropometric, environmental and clinical data from the island of Hvar; to perform biochemical tests to measure glucose, insulin, uric acid and lipid levels; 2) conduct a genome-wide association analysis of metabolic traits and phenotypes using genome-wide SNP arrays (Affymetrix Genome-Wide Human SNP Array 5.0).

  Study phs000737

• A Genome-Wide Association Study of Peripheral Arterial Disease

  The Electronic Medical Record Phenotypes and Community-engaged Genomic Association Study aims to identify genetic variants mediating susceptibility to peripheral arterial disease (PAD). The study leverages a biorepository of blood samples of 1688 PAD cases and 1649 controls, and the electronic medical record (EMR) to annotate the biorepository. PAD cases were identified from the vascular laboratory database as having an ankle brachial index (ABI) <0.9 at rest or after exercise or having non-compressible vessels. Controls were without prior history of atherosclerotic vascular disease and when tested, no evidence of ischemia on a stress test. Phenotypes and environmental exposures including age, ethnicity, demographic and anthropometric data are derived from the Mayo Electronic Medical Record (EMR). Comorbidities were determined using algorithms for diabetes and hypertension based on ICD-9 codes and medication use. Relevant laboratory data, including lipid levels, fasting blood sugar and serum creatinine at index date or within a 1-year window of the index date were extracted. Medication classes at index date were identified using Mayo's Natural Language Processing-based system with RxNorm codification and NDF-RT terminologies mapping. Smoking status was confirmed by natural language processing of clinical notes. Genotyping of ~600,000 SNPs across the genome is being conducted at the Broad Institute using the Illumina 660W platform. Statistical analyses will be conducted to identify genetic variants associated with susceptibility to PAD.

  Study phs000203

• NIH Division of Intramural Research Multiomic Monogenic Disease Study

  This cross-sectional, observational study utilized high-dimensional approaches, including transcriptomic analysis, serum protein profiling, and peripheral blood frequency assessments, to compare 228 adult and pediatric patients diagnosed with 22 monogenic immune-mediated conditions impacting various key immunological pathways, with 42 age- and sex-matched healthy controls. Participants were enrolled in clinical protocols at the National Institutes of Health in Bethesda, Maryland, USA. The primary objectives of the study were twofold: first, to identify disease-specific and shared "pan-disease" signatures using the collected data, and second, to develop a Random Forest classifier integrating these datasets to differentiate between age- and sex-matched healthy individuals and monogenic patients. This classifier produced a metric known as the "Immune Health Metric" (IHM). Subsequent analysis demonstrated that the IHM effectively distinguished healthy subjects from individuals with multiple polygenic autoimmune and inflammatory diseases in independent datasets. Moreover, it exhibited correlations with healthy aging and tracked disease activities and treatment responses across both immunological and non-immunological conditions. Additionally, the IHM accurately predicted age-dependent antibody responses to various vaccines, including influenza, Hepatitis A/B, Yellow Fever, Varicella Zoster, and Meningococcal vaccines.The microarray-based transcriptomic data, aptamer-based serum protein concentrations (Somalogic platform), peripheral blood frequency data, and clinical information will be made accessible through the database of Genotypes and Phenotypes (dbGaP).

  Study phs002732

• Phase 2 Study of Nivolumab and Entinostat in Unresectable or Metastatic Cholangiocarcinoma and Pancreatic Adenocarcinoma

  The study was a single-center, open-label, phase II trial (registration no. NCT03250273) designed to evaluate the efficacy of entinostat in combination with nivolumab in patients with advanced pancreatic ductal adenocarcinoma (PDAC). The aim was to assess the objective response rate using RECIST v1.1 criteria as the primary endpoint. A total of 27 patients with advanced PDA were enrolled from November 2017 to November 2020. Their treatment involved a 14-day lead-in with entinostat monotherapy, followed by a combination with nivolumab. The study utilized several molecular technologies to assess immune profiles and tumor microenvironment (TME) changes: high-dimensional mass cytometry by time of flight (CyTOF), Luminex for chemokine analysis in plasma, multiplexed immunohistochemistry (mIHC), image cytometry-based quantification, bulk RNA sequencing analysis. The objective response rate (ORR) was 11% with three patients showing a partial response (PR). The median duration of response was 10.2 months. Grade ≥ 3 treatment-related adverse events (TRAEs) occurred in 63% of patients, with the most common being decreased lymphocyte count, anemia, hypoalbuminemia, and hyponatremia. Gene expression analysis showed an enrichment in inflammatory response signaling pathways with the combination treatment. RNA sequencing data with clinical and demographic patient information will be available through dbGaP. For expression summaries, please refer to GEO submission GSE248014.

  Study phs003615

• NHLBI TOPMed: Genomic Activities such as Whole Genome Sequencing and Related Phenotypes in the Framingham Heart Study

  The Framingham Heart Study (FHS) is a prospective cohort study of 3 generations of subjects who have been followed up to 65 years to evaluate risk factors for cardiovascular disease. Its large sample of ~15,000 men and women who have been extensively phenotyped with repeated examinations make it ideal for the study of genetic associations with cardiovascular disease risk factors and outcomes. DNA samples have been collected and immortalized since the mid-1990s and are available on ~8000 study participants in 1037 families. These samples have been used for collection of GWAS array data and exome chip data in nearly all with DNA samples, and for targeted sequencing, deep exome sequencing and light coverage whole genome sequencing in limited numbers. Additionally, mRNA and miRNA expression data, DNA methylation data, metabolomics and other 'omics data are available on a sizable portion of study participants. This project will focus on deep whole genome sequencing (mean 30X coverage) in ~4100 participants and imputed to all with GWAS array data to more fully understand the genetic contributions to cardiovascular, lung, blood and sleep disorders. Also available are aptamer proteomic profiling, RNAseq and 850K array DNA methylation data that predominantly overlap with participants with WGS data. Comprehensive phenotypic and pedigree data for study participants are available through dbGaP phs000007.

  Study phs000974

• A comprehensive DNA methylation landscape of human and mouse cell lines derived from hematological malignancies

  Human cancer cell lines constitute useful models to study the primary disease and many relevant findings in tumor biology have been originated from them since the first immortalized cell line (HeLa) was obtained. The easy experimental intervention, the purity of the transformed cells and their versatility to undergo high-throughput screenings represent advantageous features of the established cancer cell lines. In recent years, major efforts have characterized in detail the multiomics make-up of hundreds of cancer cell lines and studied their association with sensitivity to anticancer drugs. Most of these studies have been genetic-centric and have not characterized in detail the epigenetic profiles underlying the characteristics of these cells or their impact on the efficacy of antitumoral compounds. In this regard, a past version of a DNA methylation microarray or a less accessible and more time-consuming such as reduced representation bisulfite sequencing have been used in those attempts to interrogate the epigenetic setting. Herein, we have obtained the DNA methylation profiles of 210 cell lines derived from hematological malignancies utilizing comprehensive DNA methylation microarrays that interrogates more than 850,000 and 285,000 CpG sites from human and mouse genomes, respectively. Importantly, we also provide a pharmacoepigenetic example of the potential use of this resource by showing how DNA methylation can predict response to nucleoside analogues.

  Study EGAS50000000627

• Germline Genetics of Myelodysplastic Syndromes (MDS) and Acute Myeloid Leukemia (AML)

  Myelodysplastic syndromes (MDS) are part of a heterogeneous and overlapping group of clonal diseases that arise in the hematopoietic stem or progenitor cells and also include acute myeloid leukemia (AML), myeloproliferative neoplasms (MPN), and the hybrid MDS/MPN entities. Individuals with MDS have a high risk of progressing to leukemia, with approximately 30% expected to develop AML. This study evaluates genetic susceptibility to myeloid malignancy using data from population-based case control studies of myelodysplastic syndromes and acute myeloid leukemia in adults (ages 18-85 years) and pediatric MDS cases with banked samples at the Center for International Blood and Marrow Transplant Research (CIBMTR). Adult cases were identified through the Minnesota Cancer Reporting System and controls were recruited from the State Driver's License/Identification Card list. DNA samples were collected via mouthwash collection kits (AML cases and controls) or Oragene saliva collection kits (adult MDS cases and controls). Pediatric MDS samples were banked at CIBMTR prior to receipt of a transplant and DNA was extracted from isolated T cells or whole blood samples. Genotyping data were generated using the Illumina Global Diversity array and quality control and genotype calling was conducted using established best practices. Our primary analyses investigated the contribution of genetic variation to MDS risk and evaluated the role of the killer cell immunoglobulin receptors (KIR) on incidence and survival.

  Study phs002962

• Collection: Hispanic Community Health Study/Study of Latinos (HCHS/SOL)

  The Hispanic Community Health Study / Study of Latinos (HCHS/SOL) is a multi-center epidemiologic study in Hispanic/Latino populations to determine the role of acculturation in the prevalence and development of disease, and to identify risk factors playing a protective or harmful role in Hispanics/Latinos. The study is sponsored by the National Heart, Lung, and Blood Institute (NHLBI) and six other institutes, centers, and offices of the National Institutes of Health (NIH). The goals of the HCHS/SOL include studying the prevalence and development of disease in Hispanics/Latinos, including the role of acculturation, and identifying disease risk factors that play protective or harmful roles in Hispanics/Latinos. A total of 16,415 persons of Cuban, Dominican, Mexican, Puerto Rican, Central American, and South American backgrounds were recruited through four Field Centers affiliated with San Diego State University, Northwestern University in Chicago, Albert Einstein College of Medicine in the Bronx area of New York, and the University of Miami. Seven additional academic centers serve as scientific and logistical support centers. Study participants aged 18-74 years took part in an extensive clinic exam and assessments to ascertain socio-demographic, cultural, environmental and biomedical characteristics. Annual follow-up interviews are conducted to determine a range of health outcomes.To request access to this collection, select phs003650 in the dbGaP when submitting a data access request.

  Study phs003650

• MRCA and MRCE SNP genotypes

  Gene expression levels can be an important link DNA between variation and phenotypic manifestations. Our previous map of global gene expression based on approximately 400K SNPs and 50K transcripts in 400 sib pairs from the MRCA family panel has been widely used to interpret the results of GWAS. Here, we more than double the size of our initial dataset with expression data on 550 additional individuals from the MRCE family panel using the Illumina whole genome expression array. We have used new statistical methods for dimension reduction to account for non-genetic effects in estimates of expression levels and we have also included SNPs imputed from the 1000 Genomes Project. Our methods reduced false discovery rates and increased the number of eQTLs mapped either locally or at a distance (i.e. in cis or trans) from 1,534 in the MRCA dataset to 4,452 (with <5% FDR). Imputation of 1000 Genomes SNPs further increased the number of eQTLs to 7,302. Using the same methods and imputed SNPs in the newly acquired MRCE dataset we identified eQTLs for 9000 genes. The combined results identify strong local and distant effects for transcripts from 14,177 genes. Our eQTL database (http://www.hsph.harvard.edu/faculty/liming-liang/software/eqtl/) based on these results is freely available to help define the function of disease-associated variants.

  Study EGAS00000000137

• Organoid BulkRNAseq

  Here, we aim to understand early determinants of brain organoid quality and generated a set of brain organoids from 12 different hPSC lines using an adaptation of the unguided Lancaster protocol (Lancaster & Knoblich, 2014). All cell lines were obtained from healthy donors. 12 donor hPSC lines were differentiated into brain organoids and a set of 72 individual organoids (6 per line) were randomly selected morphologically diverse organoids and subjected to bulk RNA sequencing for gene expression analysis. RNA was extracted using the RNeasy kit (Qiagen, Germany) according to the manufacturer’s instructions. A total of 1000 ng per sample was sent for RNA sequencing to Azenta Life Sciences (Genewiz Leipzig, Germany) for sequencing. Library preparation was performed using the NEBNext Ultra II RNA Library Prep Kit for Illumina following manufacturer’s instructions (NEB, Ipswich, MA, USA). mRNAs were first enriched with Oligo(dT) beads. The samples were sequenced using a 2x150 Pair-End (PE) configuration v1.5 at a depth of >20 million reads in each sample. Raw sequence data (.bcl files) generated from Illumina NovaSeq6000 was converted into fastq files and de-multiplexed using Illumina bcl2fastq program version 2.20. The raw sequence reads were aligned to the human reference genome GRCh38 using the STAR aligner (Dobin et al., 2013). Gene expression levels were quantified using the FeatureCounts tool (Liao et al., 2014).

  Study EGAS50000000659

• Nala TAS-LRS PGx Study

  Pharmacogenomics (PGx) testing improves medication safety and efficacy by identifying genetic variants that affect drug response. However, current technologies often fail to resolve complex loci, detect structural variants, or phase alleles accurately. Here, we present an end-to-end PGx workflow based on Targeted Adaptive Sampling-Long Read Sequencing (TAS-LRS), integrating a streamlined laboratory protocol with a bioinformatics pipeline that includes a novel CYP2D6 caller. Using 1,000 ng of DNA and three-sample multiplexing on a single PromethION flow cell, the assay achieves consistent on-target (25x) and off-target (3x) coverage, enabling accurate, haplotype-resolved testing of 35 pharmacogenes alongside genome-wide genotyping from off-target reads. We further developed the workflow into a clinically ready service and validated its performance across 17 reference and clinical samples. The assay demonstrated high concordance for small variants (99.9%) and structural variants (>95%), with phased diplotypes and metabolizer phenotypes reaching 97.7% and 98.0% concordance, respectively. Improved calls were observed in 12 genes due to enhanced genotyping, phasing, or novel allele detection. In addition, off-target reads supported accurate genome-wide imputation, comparable to short-read sequencing and superior to microarrays. These results establish the feasibility of long-read sequencing for clinical PGx testing and position TAS-LRS as a scalable solution combining both targeted and genome-wide utility.

  Study EGAS50000001116

• Identification of RNA biomarkers in Parkinson's disease iPSC-derived neuronal cells

  Parkinson?s disease (PD) is an age-related, chronic and progressive neurodegenerative disorder characterized by a loss of multifocal neurons and subsequent motor symptoms. These overt motor symptoms are often preceded by prodromal non-motor symptoms. Though a number of genetic and environmental factors have been identified to play a role in PD, more exact methods for both diagnosing and assessing prognosis are yet to be discovered. Probing the transcriptomes of control and PD cells can give some interesting insight into changes caused by the disease (in both coding and non-coding gene expression), highlighting potential RNA biomarkers that may be used for PD diagnosis and as new drug targets. The study consists of two main sources of data: matched neural stem cell (NSC) and fully differentiated dopaminergic neurons derived from iPS cells. Both sets of data are made up of three samples from a control cell line, and five samples carrying a mutation in one of several genes known to be linked to heritable PD: PARK2, PARK22 or PARK9. All transcriptome libraries were synthesized and sequenced using no-amplification non-tagging cap analysis of gene expression (nAnT-iCAGE) on the Illumina HiSeq 2500 platform. Resulting data was mapped to the human genome annotation (hg38) and processed CAGE tags were clustered ready for differential expression analyses. The 16 samples here described were sequenced across two lanes.

  Study JGAS000142

• Identification of RNA biomarkers in Parkinson's disease iPSC-derived neuronal cells

  Parkinson���s disease (PD) is an age-related, chronic and progressive neurodegenerative disorder characterized by a loss of multifocal neurons and subsequent motor symptoms. These overt motor symptoms are often preceded by prodromal non-motor symptoms. Though a number of genetic and environmental factors have been identified to play a role in PD, more exact methods for both diagnosing and assessing prognosis are yet to be discovered. Probing the transcriptomes of control and PD cells can give some interesting insight into changes caused by the disease (in both coding and non-coding gene expression), highlighting potential RNA biomarkers that may be used for PD diagnosis and as new drug targets. The study consists of two main sources of data: matched neural stem cell (NSC) and fully differentiated dopaminergic neurons derived from iPS cells. Both sets of data are made up of three samples from a control cell line, and five samples carrying a mutation in one of several genes known to be linked to heritable PD: PARK2, PARK22 or PARK9. All transcriptome libraries were synthesized and sequenced using no-amplification non-tagging cap analysis of gene expression (nAnT-iCAGE) on the Illumina HiSeq 2500 platform. Resulting data was mapped to the human genome annotation (hg38) and processed CAGE tags were clustered ready for differential expression analyses. The 16 samples here described were sequenced across two lanes.

  Study JGAS000318

• Neoadjuvant immune checkpoint blockade in mismatch-repair proficient colon cancers

  Immune checkpoint blockade has led to paradigm shifts in the treatment of various tumour types, yet limited efficacy has been observed in patients with metastatic mismatch-repair proficient (pMMR) colorectal cancer. Here we report an in-depth analysis of patients with early-stage pMMR colon cancer from the phase II NICHE study (ClinicalTrials.gov: NCT03026140). A total of 31 patients received neoadjuvant treatment of nivolumab plus ipilimumab followed by surgery. The response rate was 26% and included six patients with a major pathological response (≤10% residual viable tumour). One patient with an ongoing clinical complete response did not undergo surgery. Responses were observed despite a low tumour mutational burden in all tumours, while chromosomal genomic instability scores were significantly higher in responders compared to non-responders. Responding tumours were more likely to be TP53 mutant, KRASG12 wildtype and had significantly higher baseline expression of proliferation signatures and TCF1. Imaging mass cytometry also revealed a higher percentage of Ki-67+ cancer and Ki-67+ CD8+ T cells in responders compared to non-responders. Meanwhile, in non-responders, we observed an enrichment for an inflamed-fibrotic microenvironment associated with TGF-β expression, hinting towards possible resistance mechanisms. These results provide a comprehensive analysis of response to neoadjuvant ICB in early-stage pMMR colon cancers and identify potential biomarkers for patient selection.

  Study EGAS50000000856

• Phenotypic characterization and prognostic impact of CD103+ tissue-resident memory T cells in diffuse large B cell lymphoma

  Tissue-resident memory T (TRM) cells are a population of memory T cells that stably occupy tissues and play a key role in immunosurveillance, having been linked to favorable survival outcomes in various solid tumors. While TRM cells have been identified in lymph nodes, their phenotype and prognostic significance in B-cell non-Hodgkin lymphoma (B-NHL), including diffuse large B-cell lymphoma (DLBCL), remains poorly characterized. The frequency of CD103 expressing T cells in patient samples with DLCBL was quantified by immunofluorescence (IF) staining of tissue biopsies (n=306) and flow cytometry (n=252) of cell disaggregates, and linked with clinical outcome. The phenotype of TRM cells was characterized by spectral flow cytometry (n=10 DLBCL and 2 reactive lymph node (rLN) samples) and single-cell RNA sequencing (scRNAseq) (n=12 aggressive B cell lymphomas and 4 rLN samples). An additional 51 samples from an external dataset were included to verify the single cell transcriptome findings. The flow cytometry and scRNAseq findings in DLBCL were extrapolated to additional B-NHL entities. Through analysis of these datasets we were able to characterize CD103+ TRM-like cells in DLBCL and other B-NHL entities and identified them to represent a prognostically favorable population with an activated/cytotoxic T cell phenotype.

  Study EGAS50000000943

• RBMX functional retrocopy safeguards brain development

  Retrotransposition has generated thousands of intronless gene copies in mammalian genomes, yet their contribution to brain development and evolution remains largely unexplored. We investigated the role for RBMX retrocopy in shaping neurodevelopment and modulating disease. First, we reported 8 distinct pathogenic variants in RBMX, an X-linked splicing regulator, in 11 males presenting with intellectual disability, microcephaly, and cortical malformations. Then, we combined transcriptomic profiling, protein–protein and protein–RNA interaction studies both in human cellular models and mouse embryonic cortices to assess the functional redundancy between RBMX and its retrocopy RBMXL1. Finally, we use mouse genetics to dissect RBMX function and its compensation by RBMXL1 in corticogenesis. We show that RBMX pathogenic variants disrupt cortical development through both loss- and gain-of-function mechanisms. Surprisingly, despite severe phenotypes in humans, Rbmx-deficient mice display only mild cortical abnormalities, a discrepancy likely due to compensation by Rbmxl1, a retrocopy that arose independently in mice and humans. We demonstrate that RBMX and RBMXL1 share protein and RNA partners and act redundantly in brain development, with RBMXL1 buffering the impact of RBMX deficiency. Together, these findings establish that RBMXL1 functions as a functional paralog of RBMX safeguarding neurodevelopment by buffering deleterious variation. More broadly, these results identify retrocopies as active contributors to neurodevelopmental robustness and suggest that functional retrocopies may have facilitated the evolutionary diversification of the mammalian brain.

  Study EGAS50000001650

• Whole genome sequencing of matched primary and metastatic acral melanomas

  Next generation sequencing has enabled systematic discovery of mutational spectra in cancer samples. Here we used whole genome sequencing to characterise somatic mutations and structural variation in a primary acral melanoma and its lymph node metastasis. Our data show that the somatic mutational rates in this acral melanoma sample pair was more comparable to the rate reported in cancer genomes not associated with mutagenic exposure than in the genome of a melanoma cell line or the transcriptome of melanoma short-term cultures. Despite the perception that acral skin is sun-protected, the dominant mutational signature in these samples is compatible with damage due to ultraviolet light exposure. A nonsense mutation in ERCC5 discovered in both the primary and metastatic tumours, could also have contributed to the mutational signature through accumulation of unrepaired dipyrimidine lesions. However, evidence of transcription-coupled repair was suggested by the lower mutational rate in the transcribed regions and expressed genes. The primary and the metastasis are highly similar at the level of global gene copy number alterations, loss of heterozygosity and single nucleotide variation (SNV). Furthermore, the majority of the SNVs in the primary tumour were propagated in the metastasis and one non-synonymous coding SNV and one splice site mutation appeared to arise de novo in the metastatic lesion.

  Study EGAS00001000169

• A Cancer Cell-Line Titration Series for Evaluating Somatic Classification

  Accurate detection of somatic single nucleotide variants and small insertions and deletions from DNA sequencing experiments of tumour-normal pairs is a challenging task. Tumour samples are often contaminated with normal cells confounding the available evidence for the somatic variants. Furthermore, tumours are heterogeneous so sub-clonal variants are observed at reduced allele frequencies. We present here a cell-line titration series dataset that can be used to evaluate somatic variant calling pipelines with the goal of reliably calling true somatic mutations at low allele frequencies. Cell-line DNA was mixed with matched normal DNA at 8 different ratios to generate samples with known tumour cellularities, and exome sequenced on Illumina HiSeq to depths of >300x. The data was processed with several different variant calling pipelines and verification experiments were performed to assay >1,500 somatic variant candidates using Ion Torrent PGM as an orthogonal technology. By examining the variants called at varying cellularities and depths of coverage, we show that the best performing pipelines are able to maintain a high level of precision at any cellularity. In addition, we estimate the number of true somatic variants undetected as cellularity and coverage decrease. Our cell-line titration series dataset, along with the associated verification results, is effective for this evaluation and will serve as a valuable dataset for future somatic calling algorithm development.

  Study EGAS00001001016

• TRACERx 100: metastatic samples

  Whole exome sequencing of 10 metastatic biopsies from four TRACERx100 patients (see EGA dataset EGAS00001002247), collected either after relapse or death. The data from these samples are initially published with Abbosh, C. et al. Phylogenetic ctDNA analysis depicts early stage lung cancer evolution. Nature http://dx.doi.org/10.1038/nature22364 (2017). Abstract: Earlier detection of relapse following primary surgery for non-small cell lung cancer and the characterization of emerging subclones seeding metastatic sites might offer new therapeutic approaches to limit tumor recurrence. The potential to non-invasively track tumor evolutionary dynamics in ctDNA of early-stage lung cancer is not established. Here we conduct a patient-specific approach to ctDNA profiling in the first 100 lung TRACERx (TRAcking Cancer Evolution through therapy (Rx)) study participants, including one patient co-recruited to the PEACE (Posthumous Evaluation of Advanced Cancer Environment) post-mortem study. We identify independent predictors of ctDNA release in early-stage non-small cell lung cancer and perform tumor volume limit of detection analyses. Through blinded profiling of post-operative plasma, we observe evidence of adjuvant chemotherapy resistance and identify patients destined to experience recurrence of their lung cancer. Finally, we show that phylogenetic ctDNA profiling tracks the subclonal nature of lung cancer relapse and metastases, providing a new approach for ctDNA driven therapeutic studies.

  Study EGAS00001002415

• UK10K NEURO ASD MGAS

  In the UK10K project we propose a series of complementary genetic approaches to find new low frequency/rare variants contributing to disease phenotypes. These will be based on obtaining the genome wide sequence of 4000 samples from the TwinsUK and ALSPAC cohorts (at 6x sequence coverage), and the exome sequence (protein coding regions and related conserved sequence) of 6000 samples selected for extreme phenotypes. Our studies will focus primarily on cardiovascular-related quantitative traits, obesity and related metabolic traits, neurodevelopmental disorders and a limited number of extreme clinical phenotypes that will provide proof-of-concept for future familial trait sequencing. We will analyse directly quantitative traits in the cohorts and the selected traits in the extreme samples, and also use imputation down to 0.1% allele frequency to extend the analyses to further sample sets with genome wide genotype data. In each case we will investigate indels and larger structural variants as well as SNPs, and use statistical methods that combine rare variants in a locus or pathway as well as single-variant approaches. The MGAS (Molecular Genetics of Autism Study) samples are from a clinical sample seen by specialists at the Maudsley hospital and who have had detailed phenotypic assessments with ADI-R and ADOS.For further information on this cohort please contact Patrick Bolton (patrick.bolton@kcl.ac.uk).

  Study EGAS00001000113

• Evolution of DNA repair defects during malignant progression of low-grade gliomas after temozolomide treatment.

  Temozolomide (TMZ) increases the overall survival of patients with glioblastoma (GBM), but its role in the clinical management of diffuse low-grade gliomas (LGG) is still being defined. DNA hypermethylation of the O 6 -methylguanine-DNA methyltransferase (MGMT) promoter is associated with an improved response to TMZ treatment, while inactivation of the DNA mismatch repair (MMR) pathway is associated with therapeutic resistance and TMZ-induced mutagenesis. We previously demonstrated that TMZ treatment of LGG induces driver mutations in the RB and AKT-mTOR pathways, which may drive malignant progression to secondary GBM. To better understand the mechanisms underlying TMZ-induced mutagenesis and malignant progression, we explored the evolution of MGMT methylation and genetic alterations affecting MMR genes in a cohort of 34 treatment-naïve LGGs and their recurrences. Recurrences with TMZ-associated hypermutation had increased MGMT methylation compared to their untreated initial tumors and higher overall MGMT methylation compared to TMZ-treated non-hypermutated recurrences. A TMZ-associated mutation in one or more MMR genes was observed in five out of six TMZ-treated hypermutated recurrences. In two cases, pre-existing heterozygous deletions encompassing MGMT, or an MMR gene, were followed by TMZ-associated mutations in one of the genes of interest. These results suggest that tumor cells with methylated MGMT may undergo positive selection during TMZ treatment in the context of MMR deficiency.

  Study EGAS00001001179

• Illumina HumanOmniExpress genotyping data from the TEENAGE (TEENs of Attica: Genes and Environment) study.

  The TEENAGE study is a cross-sectional study comprising healthy Greek adolescents. The study was conducted according to the guidelines specified in the Declaration of Helsinki and all procedures involving human subjects were approved by the Institutional Review Board of Harokopio University and the Greek Ministry of Education, Lifelong Learning and Religious Affairs. The TEENAGE study target population comprised adolescent students aged 13 to 15 years attending the first three classes of public secondary schools located in the wider Athens area of Attica. The regions across Attica from which students were recruited were physically similar and all urban in nature. Sampling of regions and schools selected to participate was random. Students were enrolled from the wider urban region to ensure that they were coming from families with varying educational background and socio-economic status. Moreover, in order to exclude any vast economic disparities, all participating schools were exclusively public schools, which are free of charge. A total of twenty-four schools participated in the study out of 285 junior secondary public schools in Attica. Anthropometric, medical and dietary information was obtained. DNA samples were genotyped using Illumina HumanOmniExpress BeadChips (Illumina, San Diego, CA, USA) at the Wellcome Trust Sanger Institute, Hinxton, UK. Genotype calling algorithm used was Illuminus (Teo et al., 2007).

  Study EGAS00001000996

• PD-L1 blockade immunotherapy rewires cancer-induced emergency myelopoiesis

  Introduction Immune checkpoint blockade (ICB) immunotherapy has revolutionized cancer treatment, demonstrating exceptional clinical responses in a wide range of cancers. Despite the success, a significant proportion of patients still fail to respond, highlighting the existence of unappreciated mechanisms of immunotherapy resistance. Delineating such mechanisms is paramount to minimize immunotherapy failures and optimize the clinical benefit. Methods In this study, we treated tumour-bearing mice with PD-L1 blockage antibody (aPD-L1) immunotherapy, to investigate its effects on cancer-induced emergency myelopoiesis, focusing on bone marrow HSPCs. We examined the impact of aPD-L1 treatment on HSPC quiescence, proliferation, transcriptomic profile, and functionality. Results Herein, we reveal that aPD-L1 in tumour-bearing mice targets the hematopoietic stem and progenitor cells (HSPCs) in the bone marrow (ΒΜ), mediating their exit from quiescence and promoting their proliferation. Notably, disruption of the PDL1/PD1 axis induces transcriptomic reprogramming in HSPCs, observed in both individuals with Hodgkin lymphoma (HL) and tumour-bearing mice, shifting towards an inflammatory state. Furthermore, HSPCs from aPDL1-treated mice demonstrated resistance to cancer-induced emergency myelopoiesis, evidenced by a lower generation of MDSCs compared to control-treated mice. Discussion: Our findings shed light on unrecognized mechanisms of action of ICB immunotherapy in cancer, which involves targeting of BM-driven HSPCs and reprogramming of cancer-induced emergency myelopoiesis.

  Study EGAS00001007873

• Coding and small non-coding transcriptional landscape of tuberous sclerosis complex cortical tubers: implications for pathophysiology and treatment

  Tuberous Sclerosis Complex (TSC) is a rare genetic disorder that results from a mutation in the TSC1 or TSC2 genes leading to constitutive activation of the mechanistic target of rapamycin complex 1 (mTORC1). TSC is associated with autism, intellectual disability and severe epilepsy. Cortical tubers are believed to represent the neuropathological substrates of these disabling manifestations in TSC. In the presented study we used high-throughput RNA sequencing in combination with systems-based computational approaches to investigate the complexity of the TSC molecular network. Overall we detected 438 differentially expressed genes and 991 differentially expressed small non-coding RNAs in cortical tubers compared to autopsy control brain tissue. We observed increased expression of genes associated with inflammatory, innate and adaptive immune responses. In contrast, we observed a down-regulation of genes associated with neurogenesis and glutamate receptor signaling. MicroRNAs represented the largest class of over-expressed small non-coding RNA species in tubers. In particular, our analysis revealed that the miR-34 family (including miR-34a, miR-34b and miR-34c) was significantly over-expressed. Functional studies demonstrated the ability of miR-34b to modulate neurite outgrowth in mouse primary hippocampal neuronal cultures. This study provides new insights into the TSC transcriptomic network along with the identification of potential new treatment targets.

  Study EGAS00001002485

• UK10K NEURO ASD TAMPERE

  In the UK10K project we propose a series of complementary genetic approaches to find new low frequency/rare variants contributing to disease phenotypes. These will be based on obtaining the genome wide sequence of 4000 samples from the TwinsUK and ALSPAC cohorts (at 6x sequence coverage), and the exome sequence (protein coding regions and related conserved sequence) of 6000 samples selected for extreme phenotypes. Our studies will focus primarily on cardiovascular-related quantitative traits, obesity and related metabolic traits, neurodevelopmental disorders and a limited number of extreme clinical phenotypes that will provide proof-of-concept for future familial trait sequencing. We will analyse directly quantitative traits in the cohorts and the selected traits in the extreme samples, and also use imputation down to 0.1% allele frequency to extend the analyses to further sample sets with genome wide genotype data. In each case we will investigate indels and larger structural variants as well as SNPs, and use statistical methods that combine rare variants in a locus or pathway as well as single-variant approaches. The Tampere Autism sample set consists of samples from Finnish subjects with ASD (autism spectrum disorders) with IQs over 70 recruited from a clinical centre for the diagnosis and treatment of children with ASD. For further information on this cohort please contact either Terho Lehtimaki (terho.lehtimaki@uta.fi) or Kaija Puura (kaija.puura@pshp.fi).

  Study EGAS00001000115

• Homozygous loss-of-function variants in European cosmopolitan and isolate populations

  Homozygous loss of function (HLOF) variants provide a valuable window on gene function in humans, as well as an inventory of the human genes that are not essential for survival and reproduction. All humans carry at least a few HLOF variants, but the exact number of inactivated genes that can be tolerated is currently unknown-as are the phenotypic effects of losing function for most human genes. Here, we make use of 1432 whole exome sequences from five European populations to expand the catalogue of known human HLOF mutations; after stringent filtering of variants in our dataset, we identify a total of 173 HLOF mutations, 76 (44%) of which have not been observed previously. We find that population isolates are particularly well suited to surveys of novel HLOF genes because individuals in such populations carry extensive runs of homozygosity, which we show are enriched for novel, rare HLOF variants. Further, we make use of extensive phenotypic data to show that most HLOFs, ascertained in population-based samples, appear to have little detectable effect on the phenotype. On the contrary, we document several genes directly implicated in disease that seem to tolerate HLOF variants. Overall HLOF genes are enriched for olfactory receptor function and are expressed in testes more often than expected, consistent with reduced purifying selection and incipient pseudogenisation.

  Study EGAS00001001606

• Bulk-tissue RNA-sequencing of anterior cingulate cortex samples derived from Lewy body disease patients

  Parkinson’s disease (PD), Parkinson’s disease with dementia (PDD) and dementia with Lewy bodies (DLB) are three clinically, genetically and neuropathologically overlapping neurodegenerative diseases collectively known as the Lewy body diseases (LBDs). A variety of molecular mechanisms have been implicated in PD pathogenesis, but the mechanisms underlying PDD and DLB remain largely unknown, a knowledge gap that presents an impediment to the discovery of disease-modifying therapies. Transcriptomic profiling can contribute to addressing this gap, but remains limited in the LBDs. Here, we applied paired bulk-tissue and single-nucleus RNA-sequencing to anterior cingulate cortex samples derived from 28 individuals, including healthy controls, PD, PDD and DLB cases (n = 7 per group), to transcriptomically profile the LBDs. Using this approach, we (i) found transcriptional alterations in multiple cell types across the LBDs; (ii) discovered evidence for widespread dysregulation of RNA splicing, particularly in PDD and DLB; (iii) identified potential splicing factors, with links to other dementia-related neurodegenerative diseases, coordinating this dysregulation; and (iv) identified transcriptomic commonalities and distinctions between the LBDs that inform understanding of the relationships between these three clinical disorders. Together, these findings have important implications for the design of RNA-targeted therapies for these diseases and highlight a potential molecular “window” of therapeutic opportunity between the initial onset of PD and subsequent development of Lewy body dementia.

  Study EGAS00001005305

• scRNA-seq of HGSC tumor and ascites

  Immune regulatory metabolites are key features of the tumor microenvironment (TME), yet with a few notable exceptions, their identities remain largely unknown. We uncovered the immune regulatory metabolic states and metabolomes of sorted tumor and stromal, CD4+, and CD8+ cells from the tumor and ascites of patients with high-grade serous ovarian cancer (HGSC) using high-dimensional flow cytometry and metabolomics supplemented with single cell RNA sequencing. Flow cytometry revealed that tumor cells show a consistently greater uptake of glucose than T cells, but similar mitochondrial activity. Cells within the ascites and tumor had pervasive metabolite differences, with a striking enrichment in 1-methylnicotinamide (MNA) in T cells infiltrating the tumor compared to ascites. Despite the elevated levels of MNA in T cells, the expression of nicotinamide N-methyltransferase, the gene encoding the enzyme that catalyses the transfer of a methyl group from S-adenosylmethionine to nicotinamide, was restricted to fibroblasts and tumor cells. Treatment of T cells with MNA resulted in an increase in T cell-mediated secretion of the tumor promoting cytokine tumor necrosis factor alpha. Thus, the TME-derived metabolite MNA contributes to an alternative and non-cell autonomous mechanism of immune modulation of T cells in HGSC. Collectively, uncovering the tumor-T cell metabolome may reveal metabolic vulnerabilities that can be exploited using T cell-based immunotherapies to treat human cancer.

  Study EGAS00001004829

• Targeted de-methylation of the FOXP3-TSDR

  CD4+ regulatory T cells (Tregs) are key mediators of immunological tolerance and promising effector cells for immuno-suppressive adoptive cellular therapy to fight autoimmunity and chronic inflammation. Their functional stability is critical for their clinical utility and has been correlated to the demethylated state of the TSDR/CNS2 enhancer element in the Treg lineage transcription factor FOXP3. However, proof for a causal contribution of the TSDR de-methylation to FOXP3 stability and Treg induction is so far lacking. We here established a powerful transient-transfection CRISPR-Cas9-based epigenetic-editing method for the selective de-methylation of the TSDR within the endogenous chromatin environment of a living cell. The induced de-methylated state was stable over weeks in clonal T cell proliferation cultures even after expression of the editing complex had ceased. Epigenetic editing of the TSDR resulted in FOXP3 expression, even in its physiological isoform distribution, proving a causal role for the de-methylated TSDR in FOXP3 regulation. However, successful FOXP3 induction was not associated with a switch towards a functional Treg phenotype, in contrast to what has been reported from FOXP3 overexpression approaches. Thus, TSDR de-methylation is required, but not sufficient for a stable Treg phenotype induction. Therefore, targeted demethylation of the TSDR may be a critical addition to published in vitro Treg induction protocols which so far lack FOXP3 stability.

  Study EGAS00001004867

• 5- FU treated organoids

  5-Fluorouracil (5-FU) is a chemotherapeutic drug component that is commonly used for the treatment of solid cancers. The anticancer properties of 5-FU have been attributed to interference with nucleotide synthesis and through direct incorporation into the DNA as being a pyrimidine analog. As both mechanisms of action may have a mutational impact on surviving tumor cells, we performed three independent analyses to characterize the genomic consequences of 5-FU treatment: i) in vitro treatment of intestinal organoids with 5-FU followed by whole genome sequencing, ii) genome-wide mutation analyses in tumor samples from 5-FU treated patients with breast or colorectal cancer, and iii) analysis of paired biopsies from patients treated with 5-FU between biopsies. Our results demonstrate that both in vitro and in vivo 5-FU causes a mutational pattern that is dominated by T&gt;G substitutions in a CTT context and with strong resemblance to COSMIC signature 17. Interestingly, this signature is also found in non 5-FU treated patients, most prominently in esophageal tumors, indicating that distinct endogenous and exogenous triggers can converge into highly similar mutational signatures. Furthermore, our results suggest that 5-FU may have adverse mutagenic effects on healthy cells and contributes to genetic variability in surviving cancer cells thereby contributing to tumor evolution.

  Study EGAS00001003592

• Uncovering tumor intrinsic and extrinsic factors that regulate hepatocellular carcinoma growth using patient derived xenograft assays

  Several major factors limit our understanding of hepatocellular carcinoma (HCC). First, human HCCs are infrequently biopsied for diagnosis and thus are not often biologically interrogated. Second, HCC initiation and progression are strongly influenced by the cirrhotic microenvironment, and the exact contributions of intrinsic and extrinsic tumor factors are unclear. A powerful approach to examine the personalized biology of liver cancers and the influence of host tissues is with patient derived xenograft (PDX) models. In Asia, HCCs from hepatitis B virus patients have been efficiently converted into PDXs, but few parallel efforts from the west have been reported. In a large-scale analysis, we implanted 93 HCCs and 8 cholangiocarcinomas (CCAs) to systematically analyze host factors and to define an optimized platform for PDX development from both surgical and biopsy samples. NSG mice that had undergone partial hepatectomy (PHx) represented the best combination of engraftability, growth, and passageability, but overall rates were low and indicative of a unique intrinsic biology for HCCs in the US. PDX models preserved the histology and genetic features of parental tumors, and ultimately, 9 new models were usable for pre-clinical studies. Intriguingly, HCC PDXs were differentially sensitive to regorafenib and sorafenib and CCA PDXs were also highly sensitive to regorafenib. PDX models functionalize early and advanced stage HCCs and revealed unique biological features of liver cancers from the US.

  Study EGAS00001004020

• eMERGE Network Study of the Genetic Determinants of Resistant Hypertension

  The electronic Medical Records and Genomics (eMERGE) Network is a consortium of five participating sites (Group Health Seattle, Marshfield Clinic, Mayo Clinic, Northwestern University, and Vanderbilt University) funded by the NHGRI to investigate the use of electronic medical record systems for genomic research. The goal of eMERGE is to conduct genome-wide association studies in approximately 19,000 individuals using EMR-derived phenotypes and DNA from linked Biorepositories. Using electronic phenotyping methods, the consortium used DNA samples from all participating sites to explore the genetic determinants of resistant hypertension. Treatment resistant hypertension is a common health problem in the clinical setting. A basic definition of resistant hypertension included subjects with uncontrolled blood pressure despite use of three antihypertensive medications or subjects requiring four or more medications to maintain control. Other conditions, such as secondary causes of hypertension, were exclusions. Site and participants include: Vanderbilt University: BioVU, Vanderbilt&#39;s DNA databank, is an enabling resource for exploration of the relationships among genetic variation, disease susceptibility, and variable drug responses, and represents a key first step in moving the emerging sciences of genomics and pharmacogenomics from research tools to clinical practice. BioVU acquires DNA from discarded blood samples collected from routine patient care. The biobank is linked to de-identified clinical data extracted from Vanderbilt&#39;s EMR, which forms the basis for phenotype definitions used in genotype-phenotype correlations. Marshfield Clinic: The Marshfield Clinic Personalized Medicine Research Project is a population-based biobank in central Wisconsin with more than 20,000 adult subjects who provided written, informed consent to access their medical records and provided a blood sample from which DNA was extracted and plasma and serum stored. In addition to an average of 30 years of medical history data, a questionnaire about environmental exposures, including a detailed food frequency questionnaire, is available to facilitate gene/environment studies. Northwestern University: The NUgene Project is a repository with longitudinal medical information from participating patients at affiliated hospitals and outpatient clinics from the Northwestern University Medical Center. Participants&#39; DNA samples are coupled with data from a self-reported questionnaire and continuously updated data from our Electronic Medical Record (EMR) representing actual clinical care events. Northwestern has a state-of-the art, comprehensive inpatient and outpatient EMR system of over 2 million patients. NUgene has broad access to participant data for all outpatient visits as well as inpatient data via a consolidated data warehouse. NUgene participants consent to distribution and use of their coded DNA samples and data for a broad range of genetic research by third-party investigators. Group Health(GH)/University of Washington (UW): Aging and Dementia eMERGE study biorepository leverages rich population-based longitudinal data from both electronic medical records and in-depth research data to explore genome wide associations. Participants include Seattle-area members of GH (a large integrated health care system in Washington State) consented and enrolled in 1) the UW Alzheimer&#39;s Disease Patient Registry (ADPR) and 2) the Adult Changes in Thought (ACT) study. The ADPR (PI: Eric B. Larson; NIH/NIA U01 AG 006781) is a population-based registry of incident dementia cases designed to identify all new Alzheimer&#39;s Disease cases within GH from 1987 to 1996. Medical history, physical, laboratory testing, and neuropsychological testing were performed on all consenting potential cases for determination of dementia status by a consensus conference. The study base of the ADPR population was stable with an attrition rate of less than 1%/year. The ACT study (PI: Eric B. Larson; NIH/NIA U01 AG 006781) is an ongoing community-based cohort study of aging and dementia. The original cohort of 2,581 randomly selected dementia-free members age 65 and older was enrolled in 1994-1996 and expanded by 811 in 2000-2002. Continuous enrollment to maintain a cohort of 2,000 dementia free persons began in 2005. Participants receive biennial assessment including cognitive status determination. The ACT sub-sample is stable; for the original cohort, median enrollment in GH was 19 years prior to joining the ACT study, and 85% of the cohort has &#8805; 10 years of GH enrollment. DNA for the ADPR participants were obtained through a companion study, Genetic Differences in Cases and Controls (PI: Walter Kukull; NIH/NIA R01 AG007584). DNA obtained through both studies were extracted from blood using Gentra Systems Puregene methods. DNA concentration is determined by UV optical density. All samples are checked for quality by 260/280 ratio. For long-term storage, samples are aliquoted and stored at -70&#176;C. Mayo Clinic: The Mayo biobank is a disease-specific biobank for vascular diseases including peripheral arterial disease (PAD). PAD patients were identified from individuals referred to the non-invasive vascular laboratory for lower extremity arterial evaluation. Since 1997, laboratory findings have been recorded into an electronic database employing an in-house software package for data archiving and retrieval; this data becomes part of the Mayo EMR. Patients referred to the center with suspected PAD undergo a comprehensive non-invasive evaluation including the ankle-brachial index (ABI) - the ratio of blood pressure measured in the upper arms divided by blood pressure measured at the ankles. Controls subjects are identified from patients referred to the Cardiovascular Health Clinic for stress ECG. The prevalence of PAD in patients with normal exercise capacity who do not have inducible ischemia on the stress ECG, was &lt;1%. Data regarding risk factors for atherosclerosis such as diabetes, dyslipidemia, hypertension, and smoking are ascertained from the EMR.

  Study phs000297

• The sanger result of LAM disease

  Tumor biopsies from LAM disease were analyzed by sanger to characterize the alterations in patients ,to elucidate the landscape of genetic/genomic alterations. The dataset include 21 samples.

  Dataset EGAD00010001761

• NHGRI Genome Integrity of iPSCs Study

  The purpose of this study is to compare mutation load in induced pluripotent stem cells derived from skin fibroblasts.

  Study phs001277

• AmsterdamUMC Data Access Committee for the EPIC-CD study

  The current data access committee reviews and assesses data requests of researchers that seek to gain access to data acquired as part of the EPIC-CD project as performed at the AmsterdamUMC, Amsterdam, Netherlands.

  Dac EGAC50000000097

• Clinical data and mapping file

  Patient demographic characteristics, treatment, and survival data for IMbrave151 (N = 162), with one patient per line. Mapping file to map anonymized subject IDs to RNAseq IDs

  Dataset EGAD50000000569

• Bulk transcriptomics of Human High-Grade-B-Cell-Lymphomas differentiated according to IGH status

  GCB DLBCL MB2 DE and HGBCL-DH-BCL2(-BCL6) cases classified according to IGH status in IGH+ or IGH-undetectable

  Dataset EGAD50000001524

• Long-read RNA sequencing of human normal liver organoid with or without SF3B4 overexpression

  Project aims to find alternative splicing events and resulting transcript isoforms modulated by SF3B4

  Study EGAS50000000849

• WES dataset of a TIL-ACT metastatic melanoma cohort

  Whole-exome sequencing (WES) data from 13 patients with metastatic melanoma in a TIL-ACT cohort, generated to characterize tumor genomics and neoantigen landscapes relevant to adoptive T cell therapy.

  Dataset EGAD50000001731

• Star-based RNA count files

  To ellucidate gene activity in mRCC, RNA Sequencing data was generated as part of the EuroTarget study. Star aligner 2.7 was used to generate count files

  Dataset EGAD50000002296

• DNA methylation array study for 7 pairs of primary NSCLCs and distant metastases

  This study is to compare DNA methylation profiles between primary NSCLCs and paired distant metastases.

  Study EGAS00001004079

• Mutational_Screening_of_Human_Acute_Myleloid_Leukaemia_Samples

  Characterization of somatic mutations in Acute Myeloid Leukaemia samples to identify whether they can be classified into three mutually exclusive groups.

  Study EGAS00001000046

• PREDICT___Whole_Genomes

  PREDICT/Predicting individual response and resistance to VEGFR/mTOR pathway therapeutic intervention using biomarkers discovered through tumour functional genomics.

  Study EGAS00001000934

• ChIP-seq in colorectal cancer and paired adjacent normal mucosa

  Chip-seq samples for 20 colorectal patients with paired adjacent normal mucosa to characterize H3K27ac and H3K4me1.

  Study EGAS00001005303

• Melanoma Germlines subset for BAP1

  These samples comprise both melanoma cases and controls sequenced for a selection of loci linked to disease susceptibility. These bams are a subset of the sequencing restricted specifically to the GRCh37 coding areas of the BAP1 gene.

  Dataset EGAD00001002743

• Breast cancer topographs

  Whole genome sequencing to identify subclonal variants for subsequent mapping back to fixed tissue specimens. . This dataset contains all the data available for this study on 2023-03-09.

  Dataset EGAD00001010124

• Whole genome sequencing of 8 HER2-Positive Breast Cancer (in complement to EGAD00001001844)

  Whole genome sequencing of 8 HER2-Positive Breast Cancer (in complement to EGAD00001001844)

  Dataset EGAD00001003189

• Single Cell Targeted Sequence Capture

  This study is to ascertain whether it is feasible to extract single cell from a tumour, perform amplification, generate a library and sequence a targeted pulldown.

  Dataset EGAD00001001450

• Genetic_variation_in_Kuusamo

  The aim of this study is to characterize as completely as possible the genetic variation and structure of Kuusamo and surrounding regions. We will perform low coverage, whole genome sequencing in a relatively limited number (approximately 200) of individuals from the FINRISK study both of whose parents are born in Kuusamo, Posio or Taivalkoski. Sequencing will be performed at the Wellcome Trust Sanger Institute (WTSI). We have previously shown using simulations and genome wide SNP data that the genetic characteristics agree with known genealogical data, and that approximately 200 individuals suffice to capture the majority of the genetic variation in this population.We have also developed methods for long-range haplotyping and long-range imputation, which are more accurate than traditional methods. This will allow us to identify a majority of the founder haplotypes in this population. The information from the haplotypes identified in this study can be used in other studies to impute from genome-wide SNP data the full genome sequence of samples ascertained for specific phenotypes.

  Study EGAS00001000020

• University of North Carolina at Chapel Hill (UNC) Hepatocellular Carcinoma Study by Exome Sequencing (HCCSES)

  Genetic alterations in specific driver genes lead to disruption of cellular pathways and are critical events in the instigation and progression of hepatocellular carcinoma. As a prerequisite for individualized cancer treatment, we sought to characterize the landscape of recurrent somatic mutations in hepatocellular carcinoma. We performed whole exome sequencing on 87 hepatocellular carcinomas and matched normal adjacent tissues to an average coverage of 59x. The overall mutation rate was roughly 2 mutations per Mb, with a median of 45 non-synonymous mutations that altered the amino acid sequence (range 2 to 381). We found recurrent mutations in several genes with high transcript levels: TP53 (18%), CTNNB1 (10%), KEAP1 (8%), C16orf62 (8%), MLL4 (7%) and RAC2 (5%). Significantly affected gene families include the nucleotide-binding domain and leucine rich repeat containing family, calcium channel subunits, and histone methyltransferases. In particular, the MLL family methyltransferases for histone H3 lysine 4 were mutated in 20% of tumors. Conclusion: The NFE2L2-KEAP1 and MLL pathways are recurrently mutated in multiple cohorts of hepatocellular carcinoma.

  Study phs000627

• Establishing Reliability for Quantitative EEG, Transcranial Doppler, Behavioral Outcomes and Optical Coherence Tomography in SWS: The Next Step toward Biomarker Development

  Sturge-Weber syndrome (SWS) is a rare, congenital syndrome with vascular malformations of the brain, skin and eye. The underlying cause of SWS is unknown and the extent of involvement varies greatly from patient to patient. Those with brain involvement present with seizures, stroke-like episodes and neurologic deficits, usually in infancy. Those with eye involvement frequently develop vision threatening glaucoma and retinal injury. Treatment has been largely symptomatic and hampered by delayed diagnosis and the rarity of the condition. The purpose of this study is to determine the reliability of non-invasive measures to be developed as biomarkers for SWS and to determine inter-rater reliability where appropriate. This includes measuring reliability for: Power asymmetry on Quantitative Electroencephalogram Asymmetry of Transcranial Doppler velocity and pulsatility Behavioral outcome measures (PEDI, Erhardt, Modified Rankin Scale) Optical Coherence Tomography Approximately 40 subjects will participate in this observational, cross-sectional study. The first version study release included data of the first n=25 subjects.

  Study phs001281

• The Dynamic Landscape of Open Chromatin During Human Cortical Neurogenesis

  Non-coding regions comprise most of the human genome and harbor a significant fraction of risk alleles for neuropsychiatric diseases, yet their functions remain poorly defined. We created a high-resolution map of non-coding elements involved in human cortical neurogenesis by contrasting chromatin accessibility and gene expression in the germinal zone and cortical plate of the developing cerebral cortex. To obtain a high resolution depiction of chromatin structure and gene expression in developing human fetal cortex, we dissected the post-conception week (PCW) 15-17 human neocortex into two major anatomical divisions to distinguish between proliferating neural progenitors and post mitotic neurons: (1) GZ: the neural progenitor-enriched region encompassing the ventricular zone (VZ), subventricular zone (SVZ), and intermediate zone (IZ) and (2) CP: the neuron-enriched region containing the subplate (SP), cortical plate (CP), and marginal zone (MZ). Tissues were obtained from three independent donors and three to four technical replicates from each tissue were processed for ATAC-seq to define the landscape of accessible chromatin and RNA-seq for genome-wide gene expression profiling.

  Study phs001438

• Kids First: Genomics of Orofacial Clefts in the Philippines

  Orofacial clefts (OFCs) are the most common craniofacial structural birth defect in humans caused by incomplete formation of the upper lip and/or the palate. Over 4,000 children in the Philippines are born with an OFC every year, and the Filipino OFC birth prevalence rate of 1 in 500 live births is one of the highest in the world. Genome-wide association studies support a multifactorial etiology for OFCs, but common variants only account for up to ~25% of the heritable risk in any one population. Sequencing studies in diverse populations is essential to fully understand the genetic architecture of OFCs. The study goals of the Kids First project are to sequence a collection of case-parent trios from the Philippines, in order to identify de novo mutations and inherited rare variants, and understand the population differences in common variant signals in the Philippines versus other populations. Approximately 373 trios from the Philippines were selected for whole genome sequencing (WGS) as part of this Gabriella Miller Kids First project.

  Study phs002595

• Whole-Genome Sequencing Analysis of Extrachromosomal DNA with Amplicon Architect

  The study is related to our manuscript entitled "Single-cell genetic heterogeneity linked to immune infiltration in glioblastoma". Using multiplexed FISH analysis of 3-4 tumor cores from 17 glioblastoma formalin-fixed paraffin-embedded (FFPE) tumors, we identified two classes of tumors based on single cell co-occurrence of EGFR and CDK4 amplification. Tumors with a low odds ratio for finding EGFR and CDK4 amplified in the same cell (ORlow tumors) had higher EGFR copy number and higher diversity of copy number in cells harboring EGFR amplification. This suggested that in ORlow tumors EGFR is likely present on extrachromosomal DNA. To test this hypothesis, we performed whole genome sequencing on DNA extracted from FFPE slides from 3 ORlow samples and 3 ORhigh samples and used AmpliconArchitect to investigate the structure of the EGFR amplicon. We found multiple breakpoints and high complexity structure in the 3 tested ORlow samples, but not in the ORhigh samples, supporting the possibility of an extrachromosomal nature of EGFR amplification in the first group.

  Study phs003100

• Tumor Fraction Guided Cell-Free DNA Profiling in Metastatic Cancer Patients

  Cell-free DNA (cfDNA) profiling is increasingly used to guide cancer care, yet mutations are not always identified. The ability to detect somatic mutations in plasma depends on both assay sensitivity and the fraction of circulating DNA in plasma that is tumor-derived (i.e cfDNA tumor fraction). We hypothesized that cfDNA tumor fraction could inform the interpretation of negative cfDNA results and guide the choice of subsequent assays of greater genomic breadth or depth. Plasma samples collected from 118 metastatic cancer patients were analyzed with cf-IMPACT, a modified version of the FDA-authorized MSK-IMPACT tumor test that can detect genomic alterations in 410 cancer-associated genes. Shallow whole genome sequencing (sWGS) was also performed in the same samples to estimate cfDNA tumor fraction based on genome-wide copy number alterations using z-score statistics. The results show that cfDNA tumor fraction can inform the interpretation of negative cfDNA results and guide the selection of subsequent sequencing platforms that are most likely to identify clinically-relevant genomic alterations.

  Study phs002290

• Adolescent Idiopathic Scoliosis (AIS) 1000 Exomes Study

  Adolescent idiopathic scoliosis (AIS) is the most common pediatric spinal deformity. Although the Bracing in AIS Trial (BrAIST) recently demonstrated the effectiveness of bracing for preventing scoliosis progression in some patients, more than 20,000 children undergo major spinal fusion surgery at an annual cost of $3 billion. Spinal fusion surgery is a major operation with considerable risks and complications. Accurate methods of predicting curve progression are needed to develop personalized prevention strategies for those at high risk and to eliminate screening and treatment of those at low risk of progression. Previously identified risk factors for scoliosis curve progression include sex, age of onset, curve type, and presence of an underlying disorder. However, currently available algorithms for predicting AIS curve progression are inaccurate, possibly because the role of genetic factors has been largely unexplored. Because there is little a priori knowledge of the genetic variants involved in AIS pathology, an unbiased genome-wide approach is likely to provide the best opportunity to comprehensively identify disease-associated genes. This is a multicenter exome sequencing study of extreme cases with severe scoliosis.

  Study phs001677

• The National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS) Consortium for the Longitudinal Evaluation of African-Americans with Early Rheumatoid Arthritis (CLEAR)

  The Consortium for the Longitudinal Evaluation of African-Americans with Early Rheumatoid Arthritis (CLEAR) Registry and Repository, supported by the National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS), was established in 2000 to provide a resource for the scientific community to explore genetic and non-genetic factors affecting rheumatoid arthritis (RA) occurrence and outcomes in African Americans. The long term objective is a database and a repository of 1,100 RA and 550 matched healthy African-American subjects. This CLEAR Registry and Repository has two arms: a longitudinal arm for subjects with early RA (enrollment from 2000 to 2005) and a cross-sectional arm for subjects with any disease duration (enrollment from 2006 to 2012). CLEAR has two components: a database and a repository. The database contains extensive demographic, socioeconomic, clinical and radiographic (radiographs of hands and feet) information and bone mineral density data from DEXA scans. The repository contains genomic DNA, plasma and serum on most of the participants. Participants in CLEAR II had RNA isolated from peripheral blood cells.

  Study phs001360

• National Heart, Lung, and Blood Institute (NHLBI) Data Coordinating Center, Microbiome of the Lung and Respiratory Track, Lung HIV Microbiome Project (LHMP)

  Humans interact with countless microorganisms in multiple environments. The Lung HIV Microbiome Project (LHMP) characterized the microbiome of the lung and respiratory tract. This effort provided initial data to develop further hypotheses addressing differences between HIV-infected and HIV-uninfected individuals. The project involved multiple studies, either single center or multiple-center studies. The research design of each study varies with the objective of that study. The Lung HIV Microbiome Project (LHMP) worked collaboratively to ensure the clinical and scientific integrity and success of this project. The LHMP characterized the microbiome of the lung alone or in combination with the oropharyngeal cavities in HIV-infected individuals and HIV-uninfected controls using molecular techniques to identify bacteria. Knowledge of the lung microbiome and that of other components of the respiratory tract in healthy and diseased states may lead to the identification of predictors of disease progression and therapeutic targets for translation into better preventive and treatment strategies. Study datasets and biospecimens are available for request from https://biolincc.nhlbi.nih.gov/studies/lhmp/.

  Study phs000769

• A Pharmcogenetic Study of Bipolar Disorder in a Taiwanese Han Chinese Population (TWBP)

  Objective: To identify genetic variants associated with response to lithium prophylaxis treatment. Materials: 294 bipolar I (BPI) patients receiving lithium treatment, a subset of 1761 bipolar I (BPI) patients recruited by the Taiwan Bipolar Consortium. Lithium response was assessed based on the Alda Scale. Methods: Responders to lithium treatment were defined if the patients&#39; Alda Scale &gt; 4, 5, 6, and 7. Genome-wide association (GWA) analysis was carried out on the 294 BPI patients for the 4 definitions of responders. The SNPs showing strongest association in the GWAS were then tested for association in a replication sample of 100 patients and further tested in a prospective follow-up series of 24 patients. Results: Two SNPs on GADL1 showed the strongest associations in the four GWA analyses and in the replication set of 100 patients. These two SNPs had a sensitivity of 0.93 for predicting lithium response and differentiated between the good and poor responders in the follow-up cohort. Summary: These genetic variants are associated with response to lithium prophylaxis treatment for BPI.

  Study phs000692

• Single-Cell RNA-Sequencing of Human Prostatectomy Tissue

  The tissue microenvironment in prostate cancer is profoundly altered. How prostate cancer cells and their precursors mediate those changes is unclear, in part due to the inability to longitudinally study the disease evolution in human tissues. To overcome this limitation, we performed extensive single-cell RNA-sequencing (scRNA-seq) to assess the transcriptional profiles of the tissue microenvironment in prostate tissues from prostatectomies from men diagnosed with prostate cancer. For each subject, benign-enriched and tumor-enriched tissues were collected from the peripheral zone of the prostate. Our studies of human tissues revealed that cancer cell-intrinsic activation of MYC signaling was the top up-regulated pathway in human cancers, representing a common denominator across the well-known molecular and pathological heterogeneity of human prostate cancer. Numerous non-malignant cell states in the tumor microenvironment (TME), including non-cancerous epithelial, immune, and fibroblast cell compartments, were conserved across individuals, suggesting that these cell types may be a sequelae of the convergent MYC activation in the cancer cells.

  Study phs003480

• ScRNA-seq of human kidney immune cells of patients with ANCA-associated glomerulonephritis, Lupus Nephritis against a "healthy" nephrectomy control

  ANCA-associated glomerulonephritis (AGN) associates with a high risk of end-stage kidney disease. The role of kidney immune cells in local inflammation remains unclear. Here, we investigate kidney immune cell diversity and function. Kidney tissue from AGN patients (n=5) and a lupus nephritis (LN) patient (n=1) were aquired during a biopsy procedure for a clinical indication. Needle-core biopsies were obtained for histopathological examination, and an additional pass was performed to retrieve kidney tissue for scRNA-seq. Healthy kidney tissue (n=1) was obtained from a kidney that was surgically removed do tue due to a (non-invasive) papillary urothelial carcinoma. Immediately after collection, kidney tissue was processed into a single-cell suspension and sorted using a 4-color flow cytometry panel to isolate living, CD45+ immune cells. To aid in the multi-omic characterization, surface markers and T and B cell repertoires were sequenced in 2 samples (1 AGN patient and the nephrectomy control). These samples were incubated with an oligo-antibody TotalSeq-C cocktail containing 130 unique cell surface antigens.

  Study EGAS50000000159

• WES in HCCOs with varying Doxorubicin resistance

  We aimed to identify somatic mutations that could explain the resistance to Doxorubicin. Therefor, we purified Exomes from liver biopsies and HCC organoids using Agilent SureSelectXT (clinical research exome, HS2 V8, or Human all Exon V6 Cosmic) and performed WES. Matched non-tumoral liver biopsies served as the control for germline mutations. 12 of these samples were previously reported. Sequence reads were aligned to the reference human genome GRCh38 using Burrows-Wheeler Aligner (BWA, v0.7.17). Duplicate removal, base quality adjustment, mutation calling (Mutect2, intervals: SureSelectXT_HS_V8_S33266340, germline-recourse: gnomad.exomes.r2.1.1.sites.liftover_grch38, Panel-of-Normal: 1000g_pon.hg38), filtering of Mutect calls, and functional annotation (Funcotator, funcotator_dataSources.v1.7.20200521g) were performed using the Genome Analysis Toolkit (GATK, v4.4) according to the Broad Institute best practices in Somatic short variant discovery workflows. Mutations with a functional annotation of Deletion, Insertion, Non-stop, Splice-Site, Start-Codon and Missense mutations were kept. A list of 43 commonly mutated genes in HCC was analyzed in the HCC organoids [TCGA, Comprehensive and Integrative Genomic Characterization of Hepatocellular Carcinoma. Cell, 2017. 169(7): p. 1327-1341.e23].

  Study EGAS50000000043

• Genomic Alterations in Normal Breast Tissues Preceding Breast Cancer Diagnosis (GANPBC)

  This study was designed to examine the genomic alterations in histologically normal breast tissues of women who subsequently developed breast cancer and compare it to breast tissues from women who did not. Breast core needle biopsy tissues from 160 healthy women at the Susan G. Komen Tissue Bank were requested for this study and sequenced using whole exome sequencing. The samples included tissues from 79 participants who, during the annual follow-up with Susan G. Komen self-reported a subsequent diagnosis of invasive or non-invasive breast cancer after tissue submission; these cases were classified as pre-diagnosis normal tissues. The remaining 81 samples were from women who had no cancer diagnosis at their last follow-up and were classified as healthy control samples. Among the pre-diagnosis samples, the average time from tissue donation to breast cancer diagnosis was 4.7 years (range: 0-11). For the healthy controls, the average time from tissue donation to the last follow up was 6.1 years (range: 0-14).

  Study phs003822

• Transcriptome profiling of patient derived neural stem cells highlights the importance of CTNND2 and WNT signaling in early neuralization

  Loss of CTNND2/δ-catenin function is implicated in neurodevelopmental disorders, including Cri-du-chat syndrome, autism spectrum disorder and attention deficit hyperactivity disorder. CTNND2, the encoded protein, is crucial for the cadherin-catenin cell adhesion complex in the brain. However, the specific impact on neurodevelopment remains unclear. To investigate, we used neuroepithelial stem cells (NESCs) derived from patient-specific induced pluripotent stem cells (iPSCs) with loss of CTNND2. In monolayer cultures, we observed abnormal neural rosette formation and failure to establish stable neural stem cells. 3D organoid cultures confirmed delayed neurogenesis and slower growth in patient-derived iPSC lines. We have built a transcriptomic catalog and observed a dysregulation of the WNT signaling pathway linked to our observed cellular phenotypes. To validate we excluded the WNT activator CHIR99021 from the media, which partially rescued the transcriptomic and morphological phenotype of the patient neural induction. This study enhances our understanding of how CTNND2 loss affects early neurogenesis and neuronal maturation in patient specific cells.

  Study EGAS50000000323

• Acute Respiratory Distress Network (ARDSNet) Study 02 Late Steroid Rescue Study (LaSRS) (ARDSNet-LaSRS-BioLINCC)

  Data Access NOTE: Please refer to the &#8220;Authorized Access&#8221; section below for information about how access to the data from this accession differs from many other dbGaP accessions. Biospecimens: Access to Biospecimens is through the NHLBI Biologic Specimen and Data Repository Information Coordinating Center (BioLINCC). Biospecimens from ARDSNet-LaSRS include BAL Slides, Bronchial Lavage, and Plasma. Please note that use of biospecimens in genetic research is subject to a tiered consent.Objectives: Since previous reports suggested that corticosteroids may improve survival, this project was developed as a multicenter, randomized controlled trial of corticosteroids in patients with persistent ARDS.Background: Persistent acute respiratory distress syndrome (ARDS) is characterized by excessive fibroproliferation, ongoing inflammation, prolonged mechanical ventilation, and a substantial risk of death.Participants: There were 180 participants.Conclusions: The results do not support the routine use of methylprednisolone for persistent ARDS despite the improvement in cardiopulmonary physiology. In addition, starting methylprednisolone therapy more than two weeks after the onset of ARDS may increase the risk of death. (NEJM April 20, 2006; Vol 354, No. 16, pp 1671-84).

  Study phs003769

• Relation between transcriptome, karyotype and age in cases of sex chromosome aneuploidies.

  Why women live longer than men is still an open question in human biology. Sex chromosomes have been proposed to play a role in the observed sex gap in longevity, and the Y male chromosome has been suspected of having a potential toxic genomic impact on male longevity. It has been hypothesized that transposable element (TE) repression declines with age, potentially leading to detrimental effects such as somatic mutations and disrupted gene expression, which may accelerate the aging process. Given that the Y chromosome is rich in repeats, age-related increases in TE expression could be more pronounced in males, likely contributing to their reduced longevity compared to females. In this work, we first studied whether TE expression is associated with the number of sex chromosomes in humans. We analyzed blood transcriptomic data obtained from individuals of different karyotype compositions: 46,XX females (normal female karyotype), 46,XY males (normal male karyotype), as well as males with abnormal karyotypes, such as 47,XXY, and 47,XYY.

  Study EGAS50000001009

• 10x Multiome from Human Fetal Heart

  We hope to uncover the genetic basis of congenital heart disease by examining how genetic variants, especially those in non-coding regions, influence heart development. To achieve this goal, our research team created a single-cell atlas of human fetal heart development. The study population consists of 41 structurally normal fetal hearts spanning 6 to 22 weeks post-conception. We profiled close to 750,000 using single-cell RNA sequencing and ATAC sequencing technologies via the 10x Genomics Multiome platform.The molecular technologies employed allowed for simultaneous analysis of gene expression and chromatin accessibility at the single-cell level. This approach identified numerous distinct cell types and states within the developing heart, offering detailed insights into gene activity and enhancer usage during critical stages of heart development.Principal findings include the characterization of diverse cell populations and the identification of specific gene expression patterns and regulatory elements active during heart formation. This high-resolution atlas provides valuable insights into how genetic variants may disrupt normal heart development, particularly those in non-coding regions that affect gene regulation.

  Study phs003778

• Ancient tree-topologies and gene-flow processes among human lineages in Africa

  The deep history of human evolution in Africa remain intensely debated. We produced 73 novel high-quality whole genomes from 14 Central and Southern African populations with diverse cultural practices. With extensive simulations and machine-learning Approximate Bayesian Computation inferences, we reconstruct jointly their complex history of divergences and migrations. We find vast genome-wide diversity within and among populations, often uncorrelated with geography and cultural practices. This demonstrates the necessity to explicitly consider local genomic patterns, without merging samples indiscriminately into larger a priori categories, to reconstruct human evolutionary histories. We find that tree-like evolutions with long periods of drift between short periods of unidirectional gene-flow best explain the data, and that the lineage ancestral to Khoe-San populations diverged around 300,000 years ago from a lineage ancestral to rainforest hunter-gatherers and neighboring agriculturalists. We also find that short periods of ancient and recent gene-flow coincide with epochs of major ecological and cultural changes in Sub-Saharan Africa.

  Study EGAS50000001072

• Single Cell RNA-Sequencing of BCG Naive and Recurrent Non-Muscle Invasive Bladder Cancer Reveals a CD6/ALCAM-Mediated Immune-Suppressive Pathway

  Non-muscle invasive bladder cancer (NMIBC) represents 70&#8211;80% patients with newly diagnosed bladder cancer, and Bacillus Calmette-Guerin (BCG) remains a cornerstone treatment for intermediate- and high-risk NMIBC to prevent disease recurrence and progression. However, a significant number of patients experience recurrence after their first course of BCG, posing significant challenges in the management of the disease. We conducted single cell RNA sequencing (scRNA-Seq) on freshly collected NMIBC samples, distinguishing between those naive to BCG treatment and those that recurred post-BCG treatment. We observed a clear activation of inflammatory pathways across cell types during recurrence, but these were not associated with canonical immune checkpoint or T cell exhaustion phenotypes. Analysis of cell-to-cell communication revealed enhanced interactions between T cells and urothelial cells in BCG-recurrence, predominantly modulated by CD6 and activated leukocyte cell adhesion molecule (ALCAM). Furthermore, we found CD6hi T cells to be immunosuppressed and enriched in recurrent samples, suggesting a potential role for CD6 as an immune evasion signal in NMIBC.

  Study phs003742

• CNA differences between RNA-based subtypes of PDAC

  Pancreatic ductal adenocarcinomas (PDAC) have a dismal prognosis with a 5-year survival of <5%. Chemoresistance and recurrences, seen in approximately 70% of PDAC patients, may partly be explained by intratumor heterogeneity (ITH) through which tumors may develop ‘escape routes’. In the Pancreas Multibiopsy (PM)-project we have obtained multiple fragments from pancreatic and other peri-ampullary carcinomas in order to reveal potential ITH and identify genetic aberrations that predict outcome after surgery and response to chemotherapy. CNA-profiles were used to complement the characterization of liabilities in PDAC that we identified previously using RNAseq in the form of epithelial, secretory, compound pancreatic, and mesenchymal subgroups that showed survival outcomes. Comparison of CNA-profiles between these transcriptomics-based subgroups (with 8, 3, 6, and 6 tumors respectively per group) showed differences, ranging from ‘quiet’(i.e. hardly any CNA) to ‘wild’(i.e. lots of CNA), but these were not statistically significant (Chi2 0.09). When differentiating between the classical and basal-like molecular subtypes of PDAC differences in CNA (high or low) were even less statistically significant (Chi2 0.292).

  Study EGAS50000001218

• Systemic Inflammation and Lymphocyte Activation Precede Rheumatoid Arthritis

  The Allen Institute for Immunology-University of California, San Diego-University of Colorado Transition to Rheumatoid Arthritis (ALTRA) project is a prospective longitudinal cohort study to understand the pre-clinical development of rheumatoid arthritis (RA). We performed cross-sectional and longitudinal multi-omics analyses of peripheral immunity in the anti-citrullinated protein autoantibody-positive (ACPA+) at-risk for RA period. We provide FASTQ files from (1) single-cell RNA sequencing (scRNA-seq) of ACPA+ at-risk (n=45), ACPA+ early RA (n=11), and site-matched ACPA- controls (n=38); (2) scRNA-seq of longitudinal samples from ACPA+ at-risk who developed clinically active RA during our study (n=16 participants, n=75 samples) and ACPA- controls (n=29 subjects, n=89 samples); (3) trimodal TEA-seq of a subset of ACPA+ at-risk (n=7) and ACPA- controls (n=5). Using these data, we characterize pathogenesis prior to clinical RA diagnosis and suggest the at-risk state exhibits substantial immune alterations that could potentially be targeted for early intervention to delay or prevent autoimmunity.

  Study phs003944

• Human liver cholangiocyte organoids capture the heterogeneity of in vivo human liver ductal epithelium

  Human liver ductal epithelium is morphologically, functionally and transcriptionally heterogeneous. However, understanding the dynamics within human biliary/ductal epithelium has been hampered by the absence of an in vitro system that fully mimics its complex cellular heterogeneity. Here, we found that the human liver cholangiocyte (ductal-cell) organoids we previously published (Huch et al., 2015) are fairly homogenous and do not retain the cellular heterogeneity of the in vivo human biliary/ductal epithelium. Inspired by the knowledge of the in vivo niche, we refined our previous organoid culture medium to fully capture the cellular heterogeneity of the human ductal epithelium. We employed this refined system to analyse the interactions and relationships between the different human biliary epithelial cell states and discovered that acquiring a bipotent ductal cell state is necessary for differentiating the human ductal epithelium into functional hepatocyte-like cells. Our improved cholangiocyte organoid model represents a new platform to investigate cell plasticity and duct-to-hepatocyte differentiation in human liver.

  Study EGAS50000001000

• Genomic Copy-Number Variants Drive Apoptotic Resistance and Relapses on Immune Checkpoint Inhibitors

  We investigated the genomic mechanisms underlying acquired resistance (AR) to immune checkpoint inhibitors (ICIs) across tumor types, specifically by examining how copy number variant (CNV)-driven subclonal evolution contributes to clinical relapses in melanoma patients. We analyzed longitudinal clinical melanoma samples from patients who experienced relapses on ICI therapy, complemented by isogenic human melanoma cell line models (M486) and murine melanoma tumor systems. Single-cell whole-genome sequencing (scWGS) was employed to resolve subclonal CNV patterns in AR tumors. The scWGS data revealed that AR-dominant subclones systematically deleted pro-apoptotic genes while amplifying anti-apoptotic genes and innate resistance pathway components. Notably, shared and private AR subclonal CNVs preexisted selective pressure, with convergent evolution targeting apoptosis-regulatory pathways and resulting in downregulated apoptotic/mitochondrial priming that enables ICI resistance. scWGS datasets from isogenic human melanoma cell lines, patient-matched tumor samples, and murine melanoma models will be made available through dbGaP, enabling researchers to access high-resolution subclonal CNV profiles that define the genomic landscape of acquired ICI resistance evolution.

  Study phs004102

• Reading about genomic analysis of pan-neuroblastoma

  There are virtually no research fields that do not benefit from the large-scale genomic analysis that are so popular nowadays. For research into diseases with highly diverse clinical behavior like pediatric neuroblastoma, nevertheless, this type of approach has been a game-changer. Samuel W. Brady and colleagues, from St. Jude Children’s Research Hospital in Memphis, have gathered data from 702 neuroblastomas which were profiled by whole genome sequencing (WGS, n = 205), whole exome sequencing (WES, n = 539), and/or RNA-Seq. Less than half of the samples were generated during this study, while the others were obtained from previous work. Part of the WES data was indeed deposited in EGA in 2012 (EGA study EGAS00001000213). A scratchy summary about the huge work from Samuel W. Brady and colleagues, published in Nature Communications this October 2020 “These findings suggest that the sympathetic nervous system, the tissue from which neuroblastoma arises, is susceptible to different oncogenic insults at different times during development, which could be explored in future investigations using animal models.” Conclude the authors, highlighting the importance of their findings. Pediatric neuroblastomas are classified according to clinical and molecular features into low, intermediate, and high risk. Age plays a critical role in the classification, with younger children having an overall better prognosis. The samples included in this study belonged to all ages and risk-groups: the aim of such set up is the identification of age-associated molecular aberrations. The authors defined a mutational landscape of pediatric neuroblastoma including rare driver events, impossible to pick with a smaller data set. Interestingly, most driver mutations' frequency was found to be age-related. Those include alterations on MYCN, TERT, PTPRD, and Ras pathway, in addition to the previously reported ATRX. In addition, the large data set allowed the authors to define co-existence relations between the mutations (mutual exclusion or co-occurrence) which is a crucial puzzle piece to unravel the mechanisms driving the pathogenesis. Enjoy this really interesting article, open source on Pubmed.

  Blog genomic-analysis-of-pan-neuroblastoma

• The genetic structure of Norway

  The aim of the present study was to describe the genetic structure of the Norwegian population using genotypes from 6369 unrelated individuals with detailed information about places of residence. Using standard single marker- and haplotype-based approaches, we report evidence of two regions with distinctive patterns of genetic variation, one in the far northeast, and another in the south of Norway, as indicated by fixation indices, haplotype sharing, homozygosity and effective population size. We detect and quantify a component of Uralic Sami ancestry that is enriched in the North. On a finer scale, we find that rates of migration have been affected by topography like mountain ridges. In the broader Scandinavian context, we detect elevated relatedness between the mid- and northern border areas towards Sweden. The main finding of this study is that despite Norway’s long maritime history and as a former Danish territory, the region closest to mainland Europe in the south appears to have been the most isolated region in Norway, highlighting the open sea as a barrier to gene flow into Norway.

  Study EGAS00001004826

• Pancreatic islets PISA RNA-seq samples

  GWAS have identified more than 700 genetic signals associated with type 2 diabetes (T2D). To gain insight into the underlying molecular mechanisms, we created the Translational human pancreatic Islet Genotype tissue-Expression Resource (TIGER), aggregating &gt;500 human islet RNA-seq and genotyping datasets. We imputed genotypes using 4 reference panels and meta-analyzed cohorts to improve coverage of expression quantitative trait loci (eQTL) and developed a method to combine allele-specific expression across samples (cASE). We identified &gt;1 million islet eQTLs (56% novel), of which 53 colocalize with T2D signals (60% novel). Among them, a low-frequency allele that reduces T2D risk by half increases CCND2 expression. We identified 8 novel cASE colocalizations, among which an SLC30A8 T2D associated variant. We make all the data available through the open-access TIGER portal (http://tiger.bsc.es), which represents a comprehensive human islet genomic data resource to elucidate how genetic variation affects islet function and translate this into therapeutic insight and precision medicine for T2D.

  Study EGAS00001005535

• Long-read sequencing of diagnosis and post-therapy medulloblastoma reveals complex rearrangement patterns and epigenetic signatures

  Cancer genomes harbor a broad spectrum of structural variants (SV) driving tumorigenesis, a relevant subset of which are likely to escape discovery in short reads. We employed Oxford Nanopore Technologies (ONT) sequencing in a paired diagnostic and post-therapy medulloblastoma to unravel the haplotype-resolved somatic genetic and epigenetic landscape. We assemble complex rearrangements and such associated with telomeric sequences, including a 1.55 Megabasepair chromothripsis event. We uncover a complex SV pattern termed "templated insertion thread", characterized by short (mostly less than 1kb) insertions showing prevalent self-concatenation into highly amplified structures of up to 50kbp in size. Templated insertion threads occur in 3% of cancers, with a prevalence ranging to 74% in liposarcoma, and frequent colocalization with chromothripsis. We also perform long-read based methylome profiling and discover allele-specific methylation (ASM) effects, complex rearrangements exhibiting differential methylation, and differential promoter methylation in seven cancer-driver genes. Our study shows the potential of long-read sequencing in cancer.

  Study EGAS00001006576

• Recursive splicing in long vertebrate genes

  Recent studies have shown that expression of many genes with exceptionally long introns in the mammalian brain can be perturbed by regulatory factors linked to neurodevelopmental or neurodegenerative disorders1-3, suggesting unique regulatory mechanisms. Here we identify functional recursive splice sites (RSS) in long introns of genes expressed in the brain. These RSS are highly conserved in genes with extreme length across diverse vertebrate species and permit step-wise removal of long introns via recursive splicing. Recursive splicing requires initial definition of a “recursive exon” that is located downstream of RSS, and most often contains premature stop codons. Moreover, we show that RSS create a splicing switch driven by splice site competition in order to distinguish primary mRNA isoforms from alternative isoforms that are prevalent in long genes. The recursive exon is not detectable in the dominant mRNA isoform due to recursive splicing, but is completely included when cryptic promoters or other cryptic exons are used. Thus, by coupling inclusion of recursive exons with the use of cryptic elements, RSS act to distinguish new mRNA isoforms emerging from long genes.

  Study EGAS00001001170

• Study of pediatric hepatocellular carcinoma caused by bile salt export pump deficiency

  Hepatocellular carcinoma (HCC) is almost invariably associated with an underlying inflammatory state, whose direct contribution to the acquisition of critical genomic changes is unclear. We mapped the acquired genomic alterations in human and mouse HCCs induced by defects in hepatocyte biliary transporters, which expose hepatocytes to bile salts and cause the onset of chronic inflammation that develops into cancer. In both human and mouse cancer genomes we found few somatic point mutations with no impairment of cancer genes, but instead massive gene amplification and rearrangements. This genomic landscape differs greatly from that of virus- and alcohol-associated liver cancer. Copy number gains preferentially occurred at late stages of cancer development and frequently targeted the MAPK signaling pathway, and in particular direct regulators of JNK. The pharmacological inhibition of JNK retarded cancer progression in the mouse. Our study demonstrates that intrahepatic cholestasis leading to hepatocyte exposure to bile acids and inflammation promotes cancer through genomic modifications that can be clearly distinguished from those determined by other etiological factors.

  Study EGAS00001000749

• Sequencing_Acute_Myeloid_Leukaemia_

  This project will aim at sequencing and analysing three samples from a patient with acute myeloid leukaemia (AML) with t(15;17) translocation (acute promyelocytic leukaemia). The patient was treated with standard chemotherapy and samples (blood and bone marrow) taken for minimal residual disease measured during and after chemotherapy. 2-3years later the patient presented a second AML containing inv (16).In this project we will sequence both AML and normal genomes to find associated mutations This will allow unprecedented insights into the dynamics of appearance of secondary AML, since we will be able to use the MRD samples to track the emergence of the inv(16) leukaemia. We will also be able to distinguish whether the 2 leukaemias are clonally related or not. This project will entail sequencing and analysis of three samples (whole genome 30x coverage) using Hi-Seq. The funds from this grant will cover the cost of sequencing 9 lanes from the AML t(15;17) sample, 9 lanes from AML inv(16) sample and 8 lanes from the normal genome.

  Study EGAS00001000035

• APCDR Uganda GWAS: Genome-wide sequence variation and susceptibility loci for cardiometabolic traits in a sub-Saharan African population (UGWAS component)

  Genomic studies in African populations provide unique opportunities to understand disease aetiology, human genetic diversity and population history in a regional and a global context. To leverage the relative benefits of different strategies, we undertook a combined approach of genotyping and whole-genome sequencing (WGS) in a population-based study of 6,400 individuals from a geographically defined rural community in South-West Uganda. We present data from 4,778 individuals with genotypes for ~2.2 million SNPs from the Uganda GWAS resource (UGWAS), and sequence data on up to 1,978 individuals spanning 41.5M SNPs and 4.5M indels (UG2G); 343 individuals overlap between the two datasets. We highlight the value of the largest sequence panel from Africa to date as a global resource for variant discovery, imputation and understanding the mutational spectrum and its clinical relevance in African populations. Alongside phenotype data, we provide a rich new genomic resource for researchers in Africa and globally.

  Study EGAS00001001558

• This study explored and validated the clinical application of targeted NGS of circulating tumor DNA in identifying tumor-specific mutations and uncovering clinical actionable targets in a variety of solid tumors in large patient cohorts.

  Cancer is a disease of genetic alterations. Comprehensive genetic diagnosis is needed to match each patient to appropriate cancer therapy. However, acquisition of representative tumor samples is invasive and often infeasible. Circulating tumor DNA (ctDNA) is a promising non-invasive biomarker for cancer mutational profiling. Here we implemented targeted next generation sequencing (NGS) with a customized pan-cancer gene panel on 605 clinical ctDNA samples in multiple cancer types. Overall, tumor-specific mutations were identified in 87% of ctDNA samples, with mutation spectra highly concordant with their matched tumor tissues. 71% of patients have at least one clinical actionable mutation with 76% of which have suggested drugs approved or in clinical trials. In particular, our study reveals a unique mutation spectrum in Chinese lung cancer patients, which could be used to guide treatment decision and monitor the appearance of drug-resistant mutations. Taken together, our study demonstrated the clinical utility of target NGS-based ctDNA mutational profiling to guide cancer treatment decision.

  Study EGAS00001002251

• Pilot_experiment_on_functional_genomics_in_osteoarthritis

  The aim of this work is to apply an integrated systems approach to understand the biological underpinnings of large joint (hip and knee) osteoarthritis which culminates in the need for total joint replacement (TJR). In this pilot we will assess the feasibility of the approach in the relevant tissue. We will obtain diseased and non-diseased tissue (cartilage and endochondral bone) following TJR, coupled with a blood sample, from 12 patients. We will generate genotype data on the HumanCoreExome array and will characterise the 12 pairs of diseased and non-diseased tissue samples in terms of methylation (Illumina 450k array), transcription (RNASeq) and expression (quantitative proteomics). The pilot will help assess the feasibility of isolating sufficient levels of starting material for the different approaches, and will instigate the development of analytical approaches to synthesising the resulting data. If successful, it is envisaged that we will proceed with expanding the work to 500 patients in the first year, with a target total sample size of 3,000 in the longer term. The key collaborators in this work are Mark Wilkinson (Sheffield) and Jyoti Choudhary

  Study EGAS00001001212

• Interactions between the tumor and the systemic response of breast cancer patients

  Breast cancer (BC) research has largely focused on the molecular properties of the tumor proper. However, to understand how cancer progresses and ultimately modulates patient outcome, we also require an understanding of the molecular changes in the patient systemic response (SR). Toward this end, we generated and analyzed RNA profiles from tumor and matched blood samples in 173 BC patients. We designed a system (MIxT) to explore and link tightly co-expressed gene sets (modules) across matched tissues. The gene composition of modules, and their expression, largely vary across tissues. Distinct patterns of expression in the SR are predominantly detected in highly immunogenic BC subtypes. We also find that the expression pattern of a module in one tissue is correlated to the expression pattern of a module in the other tissue in a fashion that depends on subtype. For example, systemic immunosuppression is detected in basalL patients in a fashion that is proportional to the level of expression of several tumor modules highlighting immune evasion mechanisms active in these particular cancers.

  Study EGAS00001001804

• Identification_of_the_underlying_causal_variant_in_a_multi_generational_family_with_autosomal_dominant_common_variable_immunodeficiency

  Common variable immunodeficiency (CVID) is the most common form of primary immunodeficiency with an estimated incidence of 1:10,000. It has been apparent for many years that CVID has a genetic component, occurs frequently in families and can have both a recessive or dominant mode of inheritance. In recent years, 4 genes underlying CVID have been identified; however, mutations within in them are estimated to account for no more than 10% of all cases of CVID.We have identified a multi-generational family with autosomal dominant CVID. Genome-wide linkage analysis has mapped the locus underlying CVID in this family to an approximately 9.2 Mb interval on chromosome 3q27.3-q29, between the markers D3S3570 and D3S1265. This locus is distinct from any of the previously mapped susceptibility loci suggesting a novel genetic variant is responsible for disease in this family. The aim of this study is to use exome sequencing of affected (n = 4) and unaffected (n = 4) individuals, in tandem with the available genetic mapping data, to identify the causal variant underlying CVID in this family.

  Study EGAS00001000269

• BLUEPRINT DNA methylation profiles of graft donors in allogenic hematopoietic stem cell transplantation

  Allogenic hematopoietic stem cell transplantation (HSCT) is a curative treatment for many hematological conditions. Acute graft-versus-host disease (aGVHD) is a prevalent immune-mediated complication following HSCT. Current diagnostic biomarkers that correlate with aGVHD severity, progression, and therapy response in graft recipients are insufficient. We investigated whether epigenetic marks measured in peripheral blood of healthy graft donors stratify aGVHD severity in HLA-matched sibling recipients prior to HSCT. We measured genome-wide DNA methylation levels in peripheral blood of 91 HSCT donors using Illumina Infinium HumanMethylation450 BeadChips. We showed that epigenetic signatures underlying aGVHD severity in recipients correspond to immune pathways relevant to aGVHD etiology. We identified distinct DNA methylation marks in graft donors that are associated with aGVHD severity status in recipients. Together, our findings suggest that ‘epigenetic matching’ of HSCT donor-recipient pairs in a clinical setting may be used in conjunction with genetic matching to inform both donor selection and transplantation strategy, with the ultimate aim of improving patient outcome.

  Study EGAS00001001287

• Genotype and exome data for an Australian Aboriginal population: a reference panel for health-based research

  Genetic analyses, including genome-wide association studies and whole exome sequencing (WES), provide powerful tools for the analysis of complex and rare genetic diseases. To date there are no reference data for Aboriginal Australians to underpin the translation of health-based genomic research. Here we provide a catalog of variants called after sequencing the exomes of 72 Aboriginal individuals to a depth of 20X coverage in 80% of the sequenced nucleotides. We determined 320,976 single nucleotide variants (SNVs) and 47,313 insertions/deletions using the Genome Analysis Toolkit. We had previously genotyped a subset of the Aboriginal individuals (70/72) using the Illumina Omni2.5 BeadChip platform and found 99% concordance at overlapping sites, which suggests high quality genotyping. Finally, we compared our SNVs to six publicly available variant databases, such as dbSNP and the Exome Sequencing Project, and 70,115 of our SNVs did not overlap any of the single nucleotide polymorphic sites in all the databases. Our data set provides a useful reference point for genomic studies on Aboriginal Australians.

  Study EGAS00001001585

• Genome-wide mutational consequences of nucleotide excision repair-deficiency through XPC deletion in a human adult stem cell culture

  Nucleotide excision repair (NER) is one of the main DNA repair pathways that protect cells against genomic damage. Deficiency in this pathway can contribute to the development of cancer and accelerate aging. NER-deficiency is an important determinant for cancer treatment outcome, as NER-deficient tumors are selectively sensitive to cisplatin treatment. While NER-deficiency has been linked to mutational Signature 5, not all NER-deficient tumors are characterized by a high Signature 5 contribution, illustrating the importance to further characterize the mutational consequences of NER-deficiency. Here, we analyzed the mutational profile of a human adult stem cell (organoid) culture that is deficient in NER through XPC deletion by CRISPR-Cas9 gene-editing, and subsequent whole-genome sequencing analysis of a clonally derived organoid. We found that XPC deletion results in an increase in base substitution load and specifically induces Signature 8 mutations, a mutational signature with previously unknown etiology. Presence of Signature 8 may, therefore, serve as a novel biomarker for NER-deficient tumors and could improve personalized treatment strategies.

  Study EGAS00001002681

• Sequencing_component_for_the_whole_genome_methylation_analysis_in_PBMCs_and_cell_subsets__pilot_study_

  DNA methylation has been shown to play a major role in determining cellular phenotype by regulating gene expression. Moreover, dysregulation of differentially methylated genes has been implicated in disease pathogenesis of various conditions including cancer development as well as autoimmune diseases such as systemic Lupus erythematosus and rheumatoid arthritis. Evidence is rapidly accumulating for a role of DNA methylation in regulating immune responses in health and disease. However, the exact mechanisms remain unknown. The overall aim of the project is to investigate the role of epigenetic mechanisms in regulating immunity and their impact on autoimmune disease pathogenesis.The aim of this pilot study is to perform whole genome methylation analysis in peripheral blood mononuclear cells (PBMCs) and cell subsets (CD4, CD8, CD14, CD19, CD16 and whole PBMCs) obtained from 6 healthy volunteers. Whole genome methylation analysis will be performed using two methodological approaches, the Infinium Methylation Bead Array K450 (Illumina) and MeDIP-seq. mRNA expression arrays will also be performed in order to correlate DNA methylation with gene expression as well as genotyping on the Illumina OmniExpress chip

  Study EGAS00001000490

• RNA Editing in breast cancer

  High-throughput sequencing screens suggest that RNA editing, which consists in the substitution of adenosine with inosine by the RNA-specific adenosine deaminase (ADAR) enzyme, occurs at several thousand positions across the human genome. Recent evidences have shown that RNA-editing could promote proliferation and carcinogenesis; however, the general principles of ADAR activity on the transcriptome and how ADAR is controlled in cancers remain to be established. The main aim of this project was to investigate the phenomenon of RNA editing in breast and other cancers. The frequency of A-to-I editing was evaluated in 58 breast cancers equally distributed among the different molecular subtypes and 10 normal breast tissues. The analysis was focused on defining: the relationship between the global amount of editing and ADAR expression; the ability to predict the level of editability of specific sites; the distribution of editing in normal and tumour samples and among different breast cancer subtypes; and the clinical, pathological and genomic factors affecting editing.

  Study EGAS00001000495

• Circulating Tumor DNA Analysis Detects Minimal Residual Disease and Predicts Recurrence in Patients with Stage II Colon Cancer

  Detection of circulating tumor DNA (ctDNA) after resection of stage II colon cancer may identify those at the highest risk of recurrence and help inform adjuvant treatment decisions. We used massively-parallel-sequencing–based assays to evaluate the ability of ctDNA to detect minimal residual disease in 1046 plasma samples from a prospective cohort of 230 patients with resected stage II colon cancer. In patients not treated with adjuvant chemotherapy, ctDNA was detected post-operatively in 14 of 178 (7.9%) patients, 11 (79%) of whom had recurred at a median follow-up of 27 months; recurrence occurred in only 16 (9.8 %) of the 164 patients with negative ctDNA (HR, 18; 95% confidence interval [CI], 7.9 to 36; P&lt;0.001). In patients treated with chemotherapy, the presence of ctDNA following completion of chemotherapy was also associated with an inferior recurrence-free survival (HR, 11; 95% CI, 1.8 to 68; P=0.001). ctDNA detection after stage II colon cancer resection provides direct evidence of residual disease and identifies patients at very high risk of recurrence.

  Study EGAS00001001839

• Pilot_experiment_on_functional_genomics_in_osteoarthritis

  The aim of this work is to apply an integrated systems approach to understand the biological underpinnings of large joint (hip and knee) osteoarthritis which culminates in the need for total joint replacement (TJR). In this pilot we will assess the feasibility of the approach in the relevant tissue. We will obtain diseased and non-diseased tissue (cartilage and endochondral bone) following TJR, coupled with a blood sample, from 12 patients. We will generate genotype data on the HumanCoreExome array and will characterise the 12 pairs of diseased and non-diseased tissue samples in terms of methylation (Illumina 450k array), transcription (RNASeq) and expression (quantitative proteomics). The pilot will help assess the feasibility of isolating sufficient levels of starting material for the different approaches, and will instigate the development of analytical approaches to synthesising the resulting data. If successful, it is envisaged that we will proceed with expanding the work to 500 patients in the first year, with a target total sample size of 3,000 in the longer term. The key collaborators in this work are Mark Wilkinson (Sheffield) and Jyoti Choudhary

  Study EGAS00001001213

• Cell-free DNA TAPS for early cancer detection

  Multimodal, genome-wide characterization of epigenetic and genetic information in circulating cell-free DNA (cfDNA) could enable more sensitive early cancer detection. However, due to technological challenges associated with DNA methylation sequencing in low input cfDNA samples, most studies have been limited by DNA damage caused by bisulfite sequencing, or the qualitative nature of enrichment-based sequencing. Recently, we developed TET-assisted Pyridine Borane Sequencing (TAPS), which is a mild, bisulfite-free method for base-resolution direct DNA methylation sequencing. Here we optimized TAPS for cfDNA (cfTAPS) to provide high-quality and high-depth whole-genome cell-free methylomes. We applied cfTAPS to 85 cfDNA samples from patients with hepatocellular carcinoma (HCC) or pancreatic ductal adenocarcinoma (PDAC) and non-cancer controls. From just 10 ng cfDNA (1-3 mL of plasma), we generated the most comprehensive cfDNA methylome to date. We demonstrated that cfTAPS provides multimodal information about cfDNA characteristics, including DNA methylation, tissue of origin, and DNA fragmentation. Integrated analysis of these epigenetic and genetic features enables accurate identification of early HCC and PDAC.

  Study EGAS00001004962

• Mutations in SPINK2 induce azoospermia

  Azoospermia, characterized by the absence of spermatozoa in the ejaculate is a common cause of male infertility with a poorly characterized etiology. Exome sequencing analysis of two azoospermic brothers allowed the identification of a homozygous splice mutation in SPINK2, encoding a serine protease inhibitor believed to target acrosin, the main sperm acrosomal protease. In accord with these findings we observed that homozygous Spink2 KO male mice had azoospermia. Moreover, despite normal fertility, heterozygous male mice had a high rate of morphologically abnormal spermatozoa and a reduced sperm motility. Further analysis demonstrated that in the absence of Spink2, protease-induced stress initiates Golgi fragmentation and prevents acrosome biogenesis leading to spermatid differentiation arrest. We also observed a deleterious effect of acrosin overexpression in HEK cells, effect that was alleviated by SPINK2 coexpression confirming its role as acrosin inhibitor. These results demonstrate that SPINK2 is necessary to neutralize proteases during their cellular transit towards the acrosome and that its deficiency induces a pathological continuum ranging from oligoasthenoteratozoospermia in heterozygotes to azoospermia in homozygotes.

  Study EGAS00001002450

• Somatic_mutation_profiling_of_intestinal_crypts_from_IBD

  In this project we want to investigate the role that somatic mutations in intestinal stem cells play in IBD. We will use laser capture microdissection to ascertain epithelial cells from an individual crypts/vilus. We will extract DNA from these and use a new low-input material library prep methods to make accurate libraries for sequencing. In this pilot phase of the project we will perform 15X whole-genome sequencing of a small number of crypts biopsied from four patients. Biopsies have been taken from inflammed and noninflammed patches to compare mutation rates and mutational signatures. We will also test if crypts from inflammed patches of a single patient are clonal (either within a given inflammed patch or across inflammed patches). We will quantify the rate of cancer driver mutations in inflammed and non-inflammed crypts. 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 (including details of any publication moratoria), please see http://www.sanger.ac.uk/datasharing/

  Study EGAS00001002896

• Multimodal single-cell and bulk glioma analyses

  Glioma intratumoral heterogeneity enables adaptation to challenging microenvironments and contributes to therapeutic resistance. We integrated 914 single-cell DNA methylomes, 55,284 single-cell transcriptomes, and bulk multi-omic profiles across 11 adult IDH-mutant or IDH-wild-type gliomas to delineate sources of intratumoral heterogeneity. We show that local DNA methylation disorder associates with cell-to-cell DNA methylation differences, is elevated in more aggressive tumors, links with transcriptional disruption, and is altered in environmental stress response. Glioma cells under in vitro hypoxic and irradiation stress increased local DNA methylation disorder and shifted cell states. We identified a positive association between genetic and epigenetic instability that was supported in bulk longitudinally collected DNA methylation data. Increased DNA methylation disorder associated with accelerated disease progression, and recurrently selected DNA methylation changes were enriched for environmental stress response pathways. Our work identifies an epigenetically facilitated adaptive stress response process and highlights the importance of epigenetic heterogeneity in shaping therapeutic outcomes.

  Study EGAS00001005300

• Comprehensive pharmacogenomic characterization of gastric cancer

  Gastric cancer is among the most lethal human malignancies. Previous studies have identified molecular aberrations that constitute dynamic biological networks and genomic complexities of gastric tumors. However, the clinical translation of molecular-guided targeted therapy is hampered by challenges. Notably, solid tumors often harbor multiple genetic alterations, complicating the development of effective treatments. To address such challenges, we established a comprehensive dataset of molecularly annotated patient derivatives coupled with pharmacological profiles for 60 targeted agents to explore dynamic pharmacogenomic interactions in gastric cancers. We identified lineage-specific drug sensitivities based on histopathological and molecular subclassification, including substantial sensitivities toward VEGFR and EGFR inhibition therapies in diffuse- and signet ring-type gastric tumors, respectively. We identified potential therapeutic opportunities for WNT pathway inhibitors in ALK-mutant tumors, a significant association between PIK3CA-E542K mutation and AZD5363 response, and transcriptome expression of RNF11 as a potential predictor of response to gefitinib. Collectively, our results demonstrate the feasibility of drug screening combined with tumor molecular characterization to facilitate personalized therapeutic regimens for gastric tumors.

  Study EGAS00001004106

• Cancer immune control needs senescence induction by Stat1 dependent cell cycle regulator pathways in tumours

  Immune checkpoint bloackade (ICB)-based or natural cancer immune responses largely eliminate tumours. Yet, they require additional mechanisms to arrest those cancer cells that are not rejected. Cytokine-induced senescence (CIS) can stably arrest cancer cells, suggesting that interferon-dependent induction of senescence-inducing cell cycle regulators is needed to control those cancer cells that escape from killing. Here we report in two different cancers sensitive to T cell-mediated rejection, we show that deletion of the senescence-inducing cell cycle regulators p16Ink4a/p19Arf (Cdkn2a) or p21Cip1 (Cdkn1a) in the tumour cells abrogated both, the natural and the ICB-induced cancer immune control. Also in humans, melanoma metastases that progressed rapidly during ICB have losses of senescence-inducing genes and amplifications of senescence inhibitors. Metastatic cells also resist CIS. Such genetic and functional alterations are infrequent in metastatic melanomas regressing during ICB. Thus, activation of tumour-intrinsic, senescence-inducing cell cycle regulators is required to stably arrest those cancer cells that escape from eradication.

  Study EGAS00001004151

• Epigenomic priming of immune genes implicates oligodendroglia in multiple sclerosissusceptibility

  Multiple sclerosis (MS) is characterized by a targeted attack on oligodendroglia (OLG)and associated myelin by immune cells, which are thought to be the main drivers of MS susceptibility. Assessing chromatin accessibility and the transcriptome simultaneously at the single cell level, we found that immune genes exhibit a primedchromatin state in mouse and human OLG in a non-disease context, compatible withrapid transitions to immune-competent states in MS. We identified transcription factors as BACH1 and STAT1 involved in immune gene regulation in oligodendrocyteprecursor cells (OPCs). A subset of immune genes present bivalency of H3K4me3/H3K27me3 in OPCs, with Polycomb inhibition leading to their increased activation upon interferon-gamma (IFNg) treatment. Some MS susceptibility single-nucleotide polymorphisms (SNPs) overlap with these regulatory regions in mouse andhuman OLG, and treatment of mouse OPCs with IFNg leads to chromatin architecture remodeling at these loci and altered expression of interacting genes. Our data indicates that susceptibility for MS may involve OLG, which therefore constitute novel targets for immunological-based therapies for MS.

  Study EGAS00001005911