Search Results - EGA European Genome-Phenome Archive

502 present-day genotypes included in the '137 ancient human genomes from across the Eurasian steppe' publication

For thousands of years, the Eurasian steppe has been a centre for human migrations and cultural changes. To understand its population history following the Bronze Age migrations, 137 ancient humans were sequenced. These ancient DNA sequences were notably compared to the genetic data of present-day populations from Eurasia. Besides already published data, 502 individuals currently living in Inner Asia and Jordania were sampled and newly genotyped on diverse DNA-arrays. These new data, merged as a single dataset of 242,406 autosomal SNPs, are included in the present ENA study.

Study EGAS00001002926

Bulk RNA-seq of monocytes and in vitro cultured monocyte-derived macrophages of ANCA-associated vasculitis patients with active and stable disease and healthy controls

To study monocyte and macrophage activation in ANCA-associtated vasculitis (AAV), we performed bulk RNA sequencing of bead-selected monocytes and in vitro cultured monocyte-derived macrophages from AAV patients and healthy controls. Overview patients included for sequencing monocytes: - AAV active disease, n=4, MPO-AAV=4 - AAV remission, n=10, PR3-AAV=5, MPO-AAV=5 - Healthy controls, n=6 Overview patients included for sequencing monocyte-derived macrophages: - AAV active, n=1, PR3-AAV=1 - AAV remission, n=3, PR3-AAV=3 - Healthy controls, n=3

Study EGAS50000000311

human CMV-specific CD8+ T cells

In this study we established a comprehensive workflow to collect multi-omics single-cell data using a commercially available micro-well based platform. This included whole transcriptome, cell surface markers (targeted sequencing-based cell surface proteomics), T cell specificities, adaptive immune receptor repertoire (AIRR) profiles and sample multiplexing. With this technique we identified novel paired T cell receptor sequences for three prominent human CMV epitopes. In addition, we review the ability of dCODE dextramers to detect antigenspecific T cells at low frequencies by estimating sensitivities and specificities when used as reagents for single-cell multi-omics.

Study EGAS50000000633

Unraveling metastatic progression of breast cancer

To reconstruct the trajectories of BC progression, we performed exome sequencing (Illumina HiSeq2000, Truseq DNA sample preparation kit v2 and Exome Enrichment Kit v2; alignement done with BWA, substitutions and indels called with GATK and DINDEL respectively) coupled to validation by Sequenom and deep re-sequencing (ongoing), as well as copy number (CN) analyses (Oncoscan, Affymetrix) on DNA from matched primary (n=1-6/pt), axillary lymph node (ALN, n=1-6 for 4 pts), local recurrence (for 1 pt) and distant metastatic (n=1-5/pt) FFPE samples obtained from an autopsy series of 10 BC pts.

Study EGAS00001000760

Indonesian Genome Diversity Project 2

High-coverage whole genome sequences were collected to study patterns of genomic variation across the broad geography of Indonesia and New Guinea. This region has experienced an extremely complex demographic history, including repeated bouts of admixture with archaic and modern human groups. This dataset reports whole genome sequences for 82 individuals from different populations from Mentawai, New Guinea, Sumatra and Sumba islands. Particular attention has been paid in the original study to genomic signals that are informative for population history, including admixture with archaic hominins and the role of modern human admixture during the late Pleistocene and Holocene.

Study EGAS00001003654

NRG1 Fusions in KRAS Wild-type Pancreatic Cancer (H021)

We used whole-genome and transcriptome sequencing to identify clinically actionable gene fusions in young adults with KRAS wild-type (KRASwt) pancreatic ductal adenocarcinoma (PDAC). These alterations included recurrent NRG1 rearrangements predicted to drive PDAC development through aberrant ERBB receptor-mediated signaling, and pharmacologic ERBB inhibition resulted in clinical improvement and remission of liver metastases in two patients with NRG1-rearranged tumors that had proved resistant to standard treatment. Our findings demonstrate that systematic screening of KRASwt tumors for oncogenic fusion genes will substantially improve the therapeutic prospects for a sizeable fraction of PDAC patients.

Study EGAS00001002759

NanoSring of PBMC from bladder cancer and RCC patients

PBMCs from patients were isolated using 50 mL Leucosep™ tubes (Greiner Bio-One International, Germany) and Ficoll-Paque™ PLUS (GE Healthcare, Sweden). Whole blood drawn into sodium heparin blood collection tubes were diluted 3x with phosphate-buffered saline (PBS) without calcium or magnesium (Lonza, Walkersville, MD). Diluted cell suspensions were carefully layered on Leucosep tubes and centrifuged for 15 minutes at 800 x g at room temperature (RT). Interphase containing PBMCs were harvested and washed with PBS and subsequently centrifuged for 10 minutes at 250 x g at RT before further processing.

Study EGAS00001004229

Exploring_the_heterogeneity_of_sarcoma_using_single_cell_sequencing_

Multi region samples are collected from patients, with consent, immediately after resection of the tumour. Samples are digested and sorted using FACS as single cells into lysis buffer. Cells are then stored until further processing for G&T-seq. After sequencing, we will explore intra-tumour heterogeneity using computational approaches to integrate RNA and DNA data onto the tumour phylogeny 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 EGAS00001002866

Single cell exome sequencing of lung adenocarcinoma

Single cell exome sequencing was performed on 200 single cells isolated from a lung adenocarcinoma, female, non-smoker patient. The cells were derived from two distant tumour sectors and a far normal sector. The single cells were amplified on the Fluidigm C1 machine using the GE Healthcare illustra GenomiPhi V2 DNA Amplification Kit prior to sequencing library preparation. In addition, bulk exome sequencing was also conducted for each tumour sectors. For this study, somatic variations were detected from the single cells and phylogenetic clustering was conducted to elucidate the evolutionary trajectory of lung cancer.

Study EGAS00001002972

Single-cell DNA sequencing on Pediatric MDS

Single-cell DNA sequencing with antibody-oligonucleotide staining was performed using the Mission Bio Tapestri single-cell DNA sequencing platform, per the manufacturer’s instructions. All libraries were sized and quantified using an Agilent Bioanalyzer and pooled for sequencing on an Illumina NovaSeq6000 with 150 base-paired ending multiplexed runs. Fastq files generated by the sequencers were processed using the Tapestri Pipeline V2 and included adapter trimming, sequence alignment (BWA), barcode correction, cell finding, and variant calling (GATK v4/Haplotypecaller). FastQ files are provided for the 5 samples that were processed using the aforementioned pipeline.

Study EGAS00001005433

molecular profiles of serum-derived extracellular vesicles in high-grade serous ovarian cancer

Patients with high-grade serous ovarian cancer (HGSC) who have no visible residual disease (R0) after primary surgery have the best clinical outcomes, followed by patients who undergo neoadjuvant chemotherapy (NACT) and have response enabling interval cytoreductive surgery. Clinically useful predictive biomarkers for such distinctions are still lacking. Extracellular vesicles (EVs) have been recognized as liquid biopsy-based biomarkers for early cancer detection and disease surveillance in other disease settings. In this study, we performed extensive molecular characterization of serum-derived EVs and correlated them with therapeutic outcomes in patients with HGSC.

Study EGAS00001006350

Juntendo Muscle Study (JMS)

Skeletal muscle fiber type distribution has implications for human health, muscle function and performance. This knowledge has been gathered using labor intensive and costly methodology that limited these studies. This data was used to present a new method based on muscle tissue RNA sequencing data to estimate the distribution of skeletal muscle fiber types from frozen human samples, allowing for larger number of individuals to be tested in a cost and labor efficient way. Muscle biopsies were taken from the vastus lateralis muscle of 23 human subjects from Japan. Samples were sequenced on the NextSeq® 500/550 (Illumina).

Study EGAS00001006362

Muscle SATellite cell study (MSAT)

Skeletal muscle fiber type distribution has implications for human health, muscle function and performance. This knowledge has been gathered using labor-intensive and costly methodology that limited these studies. This data was used to present a new method based on muscle tissue RNA sequencing data to estimate the distribution of skeletal muscle fiber types from frozen human samples, allowing for larger number of individuals to be tested in a cost and labor efficient way. Muscle biopsies were taken from the vastus lateralis muscle of 39 Swedish male subjects. Samples were sequenced on the NextSeq® 500/550 (Illumina).

Study EGAS00001006363

scRNAseq and scATACseq of placebo controlled-trial on MMR non-specific effects

The project includes sequencing data of isolated PBMCs extracted from donors that received the MMR vaccine and donors that received placebo, both containg samples before and after the vaccine. In total, 6 MMR-vaccinated and 6 Placebo donors for both timepoints were included in the scRNAseq data. In total, 12 MMR-vaccinated and 12 Placebo donors for both timepoints were included in the scATACseq data. The main goal of the project is studying the non-specific effects of the vaccine and its possible use as an inducer of trained immunity in vivo.

Study EGAS00001006787

Selective advantage of mutant stem cells in human clonal hematopoiesis is associated with attenuated response to inflammation and aging

In this study we characterised the consequences of DNMT3A- and TET2-mutations in human clonal haematopoiesis. We screened 195 bone marrow samples for mutations by targeted DNA sequencing, and sequenced 99 paired peripheral blood samples to compare mutation detection between bone marrow and blood. We performed single-cell analysis with TARGET-seq+ on 13 samples (4 controls and 9 clonal haematopoiesis samples), which combines single-cell genotyping for targeted mutations with whole transcriptome sequencing on FACS sorted cells. Single-cell genotyping and transcriptomes were sequenced separately and linked by common single-cell identifiers.

Study EGAS00001007358

A genome-wide CRISPR screen identifies CALCOCO2 as a regulator of beta cell function influencing type 2 diabetes risk

Identification of the genes and processes mediating genetic association signals for complex diseases represents a major challenge. As many of the genetic signals for type 2 diabetes (T2D) exert their effects through pancreatic islet-cell dysfunction, we performed a genome-wide pooled CRISPR loss-of-function screen in a human pancreatic beta cell line. We assessed the regulation of insulin content as a disease-relevant readout of beta cell function and identified 580 genes influencing this phenotype. Integration with genetic and genomic data provided experimental support for 20 candidate T2D effector transcripts including the autophagy receptor CALCOCO2. Loss of CALCOCO2 was associated with distorted mitochondria, less proinsulin-containing immature granules and accumulation of autophagosomes upon inhibition of late-stage autophagy. Carriers of T2D-associated variants at the CALCOCO2 locus further displayed altered insulin secretion. Our study highlights how cellular screens can augment existing multi-omic efforts to support mechanistic understanding and provide evidence for causal effects at genome-wide association studies loci.

Dataset EGAD50000000518

An Integrated Approach to Patient Stratification and Therapy Selection in Acute Myeloid Leukemia

As more clinically-relevant genomic features of myeloid malignancies are revealed, it has become clear that targeted clinical genetic testing is inadequate for risk stratification. Here, we developed and validated a clinical transcriptome-based assay for stratification of acute myeloid leukemia (AML). Comparison of RNA-Seq to whole genome and exome sequencing revealed that as a standalone assay, RNA-Seq offered the greatest diagnostic return, enabling identification of expressed gene fusions, single nucleotide and short insertion/deletion variants, and whole-transcriptome expression information. Expression data were used to develop a novel risk score which, when combined with molecular risk guidelines, allowed for the re-stratification of 22.1 to 25.3% of AML patients from three independent cohorts into correct risk groups. Within the adverse-risk subgroup, we identified a subset of patients characterized by dysregulated integrin signaling and RUNX1 or TP53 mutation. We show that these patients may benefit from therapy with inhibitors of focal adhesion kinase (PTK2), demonstrating additional utility of transcriptome-based testing for therapy selection in myeloid malignancy.

Dataset EGAD00001006701

Glioblastoma stem cell lines RNA-seq (Guilhamon et al.)

RNA was extracted from GSCs using the Qiagen RNeasy Plus kit. RNA sample quality was measured by Qubit (Life Technologies) for concentration and by Agilent Bioanalyzer for RNA integrity. All samples had RIN above 9. Libraries were prepared using the TruSeq Stranded mRNA kit (Illumina). Two hundred nanograms from each sample were purified for polyA tail containing mRNA molecules using poly-T oligo attached magnetic beads, then fragmented post-purification. The cleaved RNA fragments were copied into first strand cDNA using reverse transcriptase and random primers. This is followed by second strand cDNA synthesis using RNase H and DNA Polymerase I. A single “A” base was added and adapter ligated followed by purification and enrichment with PCR to create cDNA libraries. Final cDNA libraries were verified by the Agilent Bioanalyzer for size and concentration quantified by qPCR. All libraries were pooled to a final concentration of 1.8nM, clustered and sequenced on the Illumina NextSeq500 as a pair-end 75 cycle sequencing run using v2 reagents to achieve a minimum of ~40 million reads per sample.

Dataset EGAD00001006813

GWAS for IgA Nephropathy

IgA nephropathy (IgAN) is a common form of immune-mediated glomerulonephritis characterized by glomerular deposition of IgA-containing immune complexes and manifesting with hematuria, proteinuria, and often kidney failure. This dataset is from a genome-wide association study (GWAS) designed to identify novel genetic risk loci for IgAN. In this dataset, seven cohorts were genotyped by Illumina Multi-Ethnic Global Array, consisting of 3,337 IgAN cases and 2,177 healthy controls of European and East Asian ancestries. Three of the provided cohorts were genotyped by Illumina HumanImmuno Chip, which are composed of 2007 cases of IgAN of European ancestry. These cohorts were used in the GWAS meta-analysis, as described in the manuscript entitled "GWAS defines pathogenic signaling pathways and prioritizes drug targets for IgA nephropathy" (Nature Genetics 2022, in press).

Study phs000431

Comprehensive Molecular Analysis of Colon Cancer for the Identification and Validation of New Biomarkers

This study aims to identify diagnostic and prognostic biomarkers in colon cancer and to study understand the complex genomic, genetic and epigenetic relationships with a multiomic approach. The study includes tumor and paired normal mucosa of 100 colon cancer patients diagnosed at stage II, untreated and with microsatellite stable tumours. Also 50 normal mucosa samples from healthy donors are included as reference. The samples have been analysed for gene expression with Affymetrix U219 array (available at GEO GSE44076), SNP/CNV with the Affymetrix 6.0 Human SNP array, for DNA methylation with the Illumina 450K array. Small RNAs were sequenced with the Solid technology and 42 of the tumours and paired normal mucosa DNA were whole exome sequenced with Illumina Genome Analyzer.

Study EGAS00001002453

Transcriptional and DNA binding profiling of CEBPE in REH acute lymphoblastic leukaemia cells using shRNA RNA-Seq and ChIP-Seq

ChIP-Seq - CEBPE - REH. The ETV6/RUNX1 translocated acute lymphoblastic leukaemia cell line. REH was used to perform ChIP-Seq using a CEBPE antibody. Cells were fixed in 1% formaldehyde for 10mins, prior to preparation of chromatin using Active Motif Express ChIP-IT. 2ug of antibody (anti CEBPE Atlas Antibodies HPA002928)was added to 25ug of chromatin O/N at 4C with rotation. Duplicate reactions were pooled and purified. 10ng of ChIP’d and input DNA used for Illumina NGS preparation (NEBNext ChIP-Seq Library kit; New England Biolabs), CEBPE and Input DNA ChIP samples were sequenced on a MiSeq using 150bp Kit v3 paired end and a HiSeq 2500 using 2x101 version 4 paired end (Illumina) respectively. Reactions performed in duplicate. shCEBPE RNA-Seq - REH. REH cells were lentivirally transduced with a pTRIPZ shRNA vector for transcriptional profiling of CEBPE. Two controls (empty and non-targeting) and two CEBPE shRNAs (V3THS_150517(A13), V3THS_404312(G3) Dharmacon, GE) were transduced into REH cells. Cells were treated with 1ug/ml doxycyclin for 144hrs and total RNA purified using Qiagen RNeasy. Knock down of CEBPE was validated by qRT. RNA integrity >7.7 for all samples. Libraries were prepared using NEBNext Ultra II Directional RNA Library Prep Kit and sequenced on an Illuimna HiSeq 2500 using 2x101 version 4 paired end chemistry. 3 biological replicates of each samples were prepared.

Dataset EGAD00001004084

Open Targets 020 Epigenomes of Cell Lines (2018-10-23)

There is currently a drive to establish cell based assay systems of greater human biological and disease relevance through the use of well characterised transformed cell lines, primary cells and complex cellular models (e.g. co-culture, 3D models). However, although the field is gaining valuable experience in running more non-standard & complex cell assays for target validation and compound pharmacology studies, there is the lack of a systematic approach to determine if this expansion in cell assay models is reflected in increased human biological and disease relevance. The increasing wealth of publically available transcriptomic, and epigenome (ENCODE and Epigenome Roadmap) data represents an ideal reference mechanism for determining the relationship between cell types used for target & compound studies to primary human cells and tissues from both healthy volunteers & patients. The CTTV020 epigenomes of cell line project aims to generate epigenetic and transcriptomic profiles of cell lines and compare these with existing and newly generated reference data sets from human tissue and cell types. The aim is to identify assay systems which will provide greater confidence in translating target biology and compound pharmacology to patients. Multiple cell types commonly used within research have been grouped according to biology. Examples include erythroid, lung epithelial, hepatocyte cell types and immortalised models of monocyte / macrophage biology. This data is part of a pre-publication release. For information on the proper use of pre-publication data shared by the Wellcome Trust Sanger Institute please see http://www.sanger.ac.uk/datasharing/ . This dataset contains all the data available for this study on 2018-10-23.

Dataset EGAD00001004415

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