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• To determine the genomic profiles of Ovarian carcinomas (UW cohort)

  Specific BRCA and immune configurations determine optimal response to platinum-based chemotherapy in triple negative breast and ovarian carcinomas (UW_OvCa_WGS)

  Study EGAS00001006048

• Epigenome mapping in purified cells of the digestive and urogenital organs: Participation in International Human Epigenome Consortium (IHEC) (RNA-Seq)

  The aim of this study is to participate in International Human Epigenome Consortium (IHEC, http://ihec-epigenomes.org/), through disclosure of quality epigenome profiles of various normal cell lineages and diseased cells purified from surgically resected materials of multiple Japanese people.

  Study JGAS000028

• Epigenome mapping in purified cells of the digestive and urogenital organs: Participation in International Human Epigenome Consortium (IHEC) (WGBS)

  The aim of this study is to participate in International Human Epigenome Consortium (IHEC, http://ihec-epigenomes.org/), through disclosure of quality epigenome profiles of various normal cell lineages and diseased cells purified from surgically resected materials of multiple Japanese people.

  Study JGAS000026

• Epigenome mapping in purified cells of the digestive and urogenital organs: Participation in International Human Epigenome Consortium (IHEC) (ChIP-seq)

  The aim of this study is to participate in International Human Epigenome Consortium (IHEC, http://ihec-epigenomes.org/), through disclosure of quality epigenome profiles of various normal cell lineages and diseased cells purified from surgically resected materials of multiple Japanese people.

  Study JGAS000027

• Exome sequencing of a novel cervical cancer cell line

  Human cancer cell lines are largely used in the searching for new antineoplastic agents. However, due to the artifacts of a long-term in culture, cell lines do not always represent the realistic tumor cell behavior. This has motivated the development of models that better mimetics the tumor tissue, among them, the establishment of primary cell cultures. In this work, we establish and characterized a low-passage cervix cancer cell line from a Brazilian patient with squamous cell carcinoma. The phenotype confirms the epithelial and tumor origin, through cytokeratins, EpCAM, and p16 staining. Whole exome sequencing showed relevant somatic mutations in several genes including BRCA2, TGFBR1, and IRX2 genes. CNV analysis by nanostring and WGS revealed amplification in genes related mainly with kinases proteins, involved in proliferation, migration and cell differentiation, such as EGFR, PIK3CA, and MAPK7. Overexpression of EGFR was confirmed by phospho RTK-array and western blot analysis. Furthermore, the cell was sensitivity to cisplatin, with IC50 13 times lower than SiHa cell line. In conclusion, this cervical cancer cell line presents molecular alterations that are an important tool for leading pre-clinical studies of new drugs that target one or more of the altered pathways.

  Study EGAS00001003343

• Genetics of Hypertension Associated Treatment (GenHAT) Study, Genomic Background of Blood Pressure Response to Treatment in African Americans

  The GenHAT study is an ancillary pharmacogenomics study to the Antihypertensive and Lipid Lowering Treatment to Prevent Heart Attack Trial (ALLHAT). ALLHAT was a randomized, double-blind, multi-center trial that enrolled 42,418 participants ≥55 years of age with hypertension and at least one additional risk factor for cardiovascular disease (CVD). Participants were randomized to treatment with one of four primary antihypertensive drugs: chlorthalidone (thiazide-like diuretic), amlodipine (CCB), lisinopril (ACE-i), or doxazosin (alpha-adrenergic blocker) in a ratio of 1.7:1:1:1, respectively. Participants were followed for treatment response and CVD outcomes. The original GenHAT study (N=39,114) evaluated the association of candidate hypertensive genetic variants with CVD outcomes for participants consenting to genetic research. The current GWAS study focuses on a subset of African American GenHAT participants (n=6908).  

  Study phs002716

• Population Genetic Testing and SERPINA1 Sequencing Identifies Unidentified Alpha-1 Antitrypsin Deficiency Alleles and Gene-Environment Interaction with Hepatitis C Infection

  Design and aims: Data were available through an institutional effort to combine de-identified electronic health records (EHR) data with genomic information from a DNA biobank. The design of this work was a retrospective cohort study in which we investigated the rates of liver transplantation and hepatitis C infection as it relates to genotype risk for alpha-1 antitrypsin deficiency (AATD). We used regression analysis to look for relationships between these risks and outcomes. This led to further investigation of environmental exposures and phenotypes as it related to the underlying genotypic risk. Population information: The cohort consisted of 72,027 individuals of European ancestry for whom genotype and EHR data were available. Molecular technologies employed: Genome-wide genotyping was previously performed using the Illumina Infinium Expanded Multi-Ethnic Genotyping Array plus custom content (referred to as the Vanderbilt University Medical Center BioVU MEGAEX). Individuals' AATD-related alleles were inferred from the genotyping data. Principal findings of the study: Liver transplantation was associated with presence of the Z AATD allele and with hepatitis C infection. There was a significant interaction between AATD genotype groups and hepatitis C infection as it related to liver transplantation. Data available through dbGaP: Inferred AATD genotype status, sex, age, length of EHR in years, presence/absence of AATD clinical diagnosis, liver transplant status, and hepatitis C infection status.

  Study phs003297

• Innate myeloid cell sbuset-specific gene expression patterns in the human colon are altered in Crohn's disease patients

  Background: There is a heterogeneous subset innate myeloid cells, such as macrophages and dendritic cells, in the human intestinal lamina propria. Several studies have demonstrated that these cells contribute to the maintenance of gut homeostasis through induction of inflammatory responses and tolerance via cell type-specific mechanisms; whereas, disrupted innate immune responses are implicated in the pathogenesis of Crohn?s disease. However, the detailed mechanisms by which each innate myeloid subset regulates gut homeostasis and inflammation largely remains unknown. Aim: We aimed to clarify the comprehensive gene expression profiles of innate myeloid cell subsets in the lamina propria from normal human colons (NC) and the inflamed colon sites from patients with Crohn?s disease (CDi). Methods: We performed RNA-sequencing analysis and precise bioinformatics analysis on three innate myeloid cell subsets, CD14-CD11c-, CD14-CD11c+, and CD14+CD11c+CD163low cells from NC and CDi. Results: Transcriptional analysis of the three subsets from the NC showed distinct gene expression patterns and gene ontology (GO) enrichment analysis revealed the associated innate myeloid subset-specific biological process (BP) terms. In addition, changes in gene expression patterns were observed in innate myeloid subsets from CDi. Furthermore, the core GO-BP terms for the genes upregulated in the innate myeloid cells from CDi were distinct from those found in NC. Conclusion: Our data identified the innate myeloid cell subset-specific transcriptomes and the associated enriched GO-BP terms in the NC and found these patterns were altered in CDi.

  Study JGAS000127

• Identification of point mutations, expression perturbations, and gene fusions in T-cell acute lymphoblastic leukemia by RNA-seq

  RNA-seq is a promising technology to re-sequence protein-coding genes for the identification of single nucleotide variants, while simultaneously obtaining information on structural variations and gene expression perturbations. T-cell acute lymphoblastic leukemia (T-ALL) is caused by a combination of gene fusions leading to the over-expression of transcription factors reinforced by point mutations in oncogenes and tumor suppressor genes. We asked whether RNA-seq is suitable for the detection of driver mutations in these leukemias. We analyzed 31 T-ALL patient samples and 18 T-ALL cell lines by high-coverage paired-end RNA-seq.

  Study EGAS00001000536

• RNA sequencing in primary inflammatory (TPP) macrophages treated with a MEK1/2 inhibitor

  By investigating an intergenic haplotype on chr21q22, independently linked to inflammatory bowel disease (IBD), ankylosing spondylitis, primary sclerosing cholangitis and Takayasu’s arteritis, we discover that the causal gene, ETS2, is a master regulator of inflammatory responses in human macrophages and delineate how the risk haplotype increases ETS2 expression. Using a database of cellular signatures, we identified drugs that could modulate this pathway. To validate the anti-inflammatory activity of one class of small molecules in vitro, we treated TPP macrophages with vehicle control or a selective, non-ATP competitive MEK inhibitor (PD-0325901) at 2 doses (100nM and 500nM) and performed RNA sequencing.

  Study EGAS00001007552

• Susceptibility loci for tanning ability in Japanese population identified by genome-wide association study

  We performed genome-wide association studies of Japanese Skin Type for Japanese individuals from a cohort study by the Tohoku Medical Megabank Project in order to identify variants related to tanning ability and skin pigmentation. We used imputed genotype data of 9,187 Japanese individuals, of which 4,475 individuals are from Miyagi prefecture and 4,712 individuals are from Iwate prefecture. Skin Type information was obtained from the questionnaire for cohort participants.

  Study JGAS000160

• Transcriptomic intra-tumor heterogeneity of colorectal cancer varies depending on tumor location within the colorectum

  Background; Intra-tumor heterogeneity (ITH) of colorectal cancer (CRC) complicates molecular tumor classification, such as transcriptional subtyping. Differences in cellular states, biopsy cell composition, and tumor microenvironment may all lead to ITH. Here we analyze ITH at the transcriptomic and proteomic levels to ascertain whether subtype discordance between multiregional biopsies reflects relevant biological ITH or lack of classifier robustness. Further, we study the impact of tumor location on ITH.MethodsMultiregional biopsies from stage II and III CRC tumors were analyzed by RNA sequencing (41 biopsies, 14 tumors) and multiplex immune protein analysis (89 biopsies, 29 tumors). CRC subtyping were performed using consensus molecular subtypes (CMS), CRC intrinsic subtypes (CRIS), and TUMOR types. ITH-scores and network maps were defined to determine the origin of heterogeneity. A validation cohort was used with one biopsy per tumor (162 tumors).ResultsOverall, inter-tumor transcriptional variation exceeded ITH, and subtyping calls were frequently concordant between multiregional biopsies. Still, some tumors had high ITH and were classified discordantly. Proximal tumors showed ITH related to differences in the microenvironment, while ITH within distal tumors were cancer-cell related. This subtype discordancy reflected actual molecular ITH within the tumors. The relevance of the subtypes was reflected at protein level where several inflammation markers were significantly increased in immune related transcriptional subtypes, which was verified in an independent cohort (Wilcoxon rank sum test; p<0.05).ConclusionOur transcriptomic and proteomic analyses show that the tumor location along the colorectum influence the ITH of CRC, which again influence the concordance of subtyping.

  Study EGAS00001004668

• Genomic and Phenotypic Profile of Sickle Cell Disease in Human Population in Cameroon

  Sickle cell disease (SCD) is caused by a biallelic single nucleotide substitution in the beta-globin gene (HBB), resulting in an amino acid substitution, Glu7Val, formerly known as Glu6Val. An estimated 300,000 babies with SCD are born worldwide each year, with nearly 75% of these births being in sub-Saharan Africa. In Africa, at least 30-50% of children with untreated SCD die before the age of 5 years. Therefore, accelerating the path for novel therapies for SCD through genomics research on fetal hemoglobin (HbF) is critical. Variants in the currently known HbF‐modulating genes/loci, i.e., BCL11A, HBS1L-MYB, and XmnI-HBG2, explain only 10-20% of the variation of HbF levels in African individuals with SCD, compared with nearly 50% of the variation in HbF levels among Europeans. Expanding genomic research in populations of African ancestry could uncover the missing heritability of HbF-promoting loci. This study used the Human Heredity and Health (H3Africa) consortium SNP genotyping array developed from whole genome enriched for common variants in sub-Saharan Africans to investigate genomic variations associated with HbF levels in individuals with sickle cell anemia of the HbSS genotype from Cameroon. Two of the previously described loci, BCL11A and HBS1L-MYB were replicated, while a novel locus on chromosome 13 mapping to FLT1 was uncovered. The study therefore opens up a novel avenue for exploring SCD gene therapy.

  Study phs003748

• Alzheimer's Disease Sequencing Project (ADSP)

  NOTICE OF CHANGE IN LOCATION FOR ALZHEIMER'S DISEASE SEQUENCING PROJECT (ADSP) GENETIC AND PHENOTYPIC DATA: ADSP whole exome and whole genome sequence data that are shared through dbGaP were mapped to the Genome Reference Consortium human genome GRCh37 (build 37). These data are from the Discovery Phase of the project (described below) and will continue to be available at this site. Please see the ADSP Design page for the complete study description. All data that are mapped to GRCh38 (hg38) are being shared through the NIA Genetics of Alzheimer's Disease Data Storage Site (NIAGADS) Data Sharing Service (DSS). For instructions on how access the ADSP Build 38 data that are shared through NIAGADS DSS, visit the Application Instructions page. STUDY DESCRIPTION FOR dbGaP BUILD 37 ADSP DATA: The overarching goals of the Alzheimer's Disease Sequencing Project (ADSP) are to: (1) identify new genomic variants contributing to increased risk of developing Alzheimer's Disease (AD), (2) identify new genomic variants contributing to protection against developing AD, and (3) provide insight as to why individuals with known risk factor variants escape from developing AD. These factors will be studied in multi-ethnic populations in order to identify new pathways for disease prevention. Such a study of human genomic variation and its relationship to health and disease requires examination of a large number of study participants and needs to capture information about common and rare variants (both single nucleotide and copy number) in well phenotyped individuals. Using existing samples from NIH funded and other studies, three NHGRI funded Large Scale Sequencing and Analysis Centers (LSAC) - Broad, Baylor, and Washington University - produced the DNA sequence data. Variant call data are being made available to the scientific community through NIH-approved data repositories. Statistical analysis of the sequence data is anticipated to identify new genetic risk and protective factors. The ADSP will conduct and facilitate analysis of sequence data to extend previous discoveries that may ultimately result in new directions for AD therapeutics. Analysis of ADSP data will be done in two phases. The Discovery Phase analysis (2014-2018) is funded under PAR-12-183. The entire Discovery dataset contains whole-genome sequencing data on 584 subjects from 113 families, and pedigree data for > 4000 subjects; whole exome sequencing data on 5096 cases 4965 controls; and whole exome sequence data on an additional 853 (682 Cases [510 Non-Hispanic, 172 Hispanic]), and 171 Hispanic Control subjects from families that are multiply affected with AD. The Replication Phase (2016-2021) analysis will be funded under RFA-AG-16-001 and RFA-AG-16-002 and is expected to include a combination of genotyping and sequencing approaches on at least 30,000 subjects. Targeted sequencing will be done by the LSACs. GRCh37 Data Releases The first ADSP data release occurred on November 25, 2013. It included the whole-genome sequencing data in BAM file format on 410 individuals. The second ADSP data release occurred on March 31, 2014, and included the whole-genome sequencing data in BAM file format for an additional 168 individuals. The third ADSP data release occurred on November 03, 2014 and included whole-exome sequencing data in BAM file format for 10,939 individuals. The fourth ADSP data release occurred on February 13, 2015 and included revised ethnic data for subjects with whole-exome sequencing data. The fifth ADSP data release occurred on July 13, 2015 and included whole-genome genotypes and updated phenotypes as well as changes to pedigree structures and sample IDs. The sixth ADSP data release occurred on December 8, 2015, and included whole-exome genotypes and updated phenotypes as well as changes to subject IDs. This seventh ADSP data release on April 12, 2016 includes: (1) WES and WGS SNV VCF files (2) WES and WGS Indel PLINK files ADSP Data Available through dbGaP: ADSP - Whole Genome Sequencing ADSP - Whole Exome Sequencing Comments DNA-Seq (BAM) n=578 n=10913 Sequence data available (plus n=38 replications w/out genotype data) Concordant SNV Genotypes (PLINK format) N/A n=10913 QC'ed genotypes that are concordant between the Atlas (Baylor's) and GATK (Broad's) calling pipelines (a subset of the consensus genotype set) Consensus Genotypes (PLINK and VCF format) n=578 n=10913 QC'ed genotypes that are concordant between Atlas and GATK pipelines as well as those that that were called uniquely by Atlas or GATK Concordant Indel Genotypes (PLINK format) n=578 n=10913 QC'ed genotypes that are concordant between the Atlas and GATK calling pipelines Phenotype Data n=4735 n=10913 Data of n=53 phenotype variables available (plus administrative data), including APOE genotype. WGS phenotypes include data of connecting family members. Please use the release notes provided by dbGaP to obtain detailed information about study release updates. The ADSP data portal provides a customized interface for users to quickly identify and retrieve files by covariates, phenotypes, and data properties such as sequencing facility or coverage. For more information about the ADSP study and the data portal, please visit https://www.niagads.org/adsp/.

  Study phs000572

• Urothelial Cancer - Genomic Analysis to Improve Patient Outcomes and Research (UC-GENOME): a Bladder Cancer Advocacy Network (BCAN)-Led Collaborative Research Pilot Study - for Samples Collected at Johns Hopkins

  UC-GENOME is a real-world cohort of patients with metastatic urothelial carcinoma (UC). The study consisted of two co-equal aims: 1) to provide targeted DNA sequencing for clinical decision making at no cost to patients; and 2) to create a resource for collaborative translational science including a clinically annotated biobank. The submitted data represents the foundational analysis of 218 patients accrued at 7 academic medical centers. The analysis includes 176 patients with both targeted DNA sequencing and RNA sequencing. The genomic data was combined with clinical variables to model response to chemotherapy and immune checkpoint inhibitors (ICI).The larger dataset for this study can be found in dbGaP study phs003066.

  Study phs003094

• A Single-Cell Atlas of the Multicellular Ecosystem of Primary and Metastatic Hepatocellular Carcinoma

  Hepatocellular carcinoma (HCC) represents a paradigm of the relation between tumor microenvironment (TME) and tumor development. Here, we generated a single-cell atlas of the multicellular ecosystem of HCCs from four relevant sites. Antitumor central memory T (TCM) cells were found in tertiary lymphoid structures (TLSs). Chronic HBV/HCV infection increases infiltration of CD8+ T cells in tumors but aggravates T cell exhaustion. We identified CD11b+ macrophages to be terminally differentiated tumor-associated macrophages (TAMs) and two distinct differentiation trajectories contribute to their accumulation. We further demonstrated CD11b+ TAMs promote HCC cells invasion and migration, and angiogenesis. We characterized the heterogeneous subpopulations of malignant hepatocytes and their multifaceted roles in shaping the immune microenvironment. Finally, we identified seven TME subtypes that can predict patient prognosis. Collectively, this large-scale atlas deepens our understanding of the ecosystem in primary and metastatic HCCs, might facilitating the development of new immune therapy strategies for this malignancy.

  Study EGAS00001004468

• Evolutionary analysis of primary tumors and metastatic lesions from 20 breast cancer patients (99 samples in total) using exome sequencing data.

  Metastatic breast cancers are still incurable. Characterizing their evolutionary landscape including the role of metastatic axillary lymph nodes to seed distant organ metastasis can provide rational basis for effective treatments. Here, we describe the genomic analyses of the primary tumors and metastatic lesions from 20 breast cancer patients (99 samples). Our evolutionary analyses revealed diverse spreading and seeding patterns governing tumor progression. Although linear evolution to successive metastatic sites was common, parallel evolution from the primary tumor to multiple distant sites was also evident. Metastatic spreading was frequently coupled with polyclonal seeding, where multiple metastatic subclones originated from primary tumor and/or other distant metastases. Synchronous axillary lymph node metastasis, a well-established prognosticator of breast cancer, was not involved in seeding distant metastasis, suggesting haematogenous route for cancer dissemination. Clonal evolution coincided frequently with emerging driver alterations and evolving mutational processes, notably a significant increase in apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like (APOBEC) associated mutagenesis. Our data provide the first genomic evidence regarding the role of axillary lymph node metastasis in seeding distant organ metastasis and elucidates the evolving mutational landscape during cancer progression.

  Study EGAS00001002737

• Endocrine therapy reprogramming of breast cancer facilitates metastatic escape via upregulation of P-Rex1/Rac1 signalling

  The estrogen receptor (ER) drives growth in ~70% of breast cancers, and endocrine therapies prevent recurrence. However, 30% of patients recur, with many fatal cases occurring late (after >5 years), presenting a challenge due to their often non-proliferative nature and limited options. To study this, we generated slow-growing resistant cells that form small primary tumours but readily metastasize. Single-cell RNA sequencing (scRNAseq) revealed that endocrine therapy reprograms these cells, notably upregulating the Rac1 signalling component P-Rex1. We found in clinical cohorts that P-Rex1 in high in ER+ breast cancer. Intravital imaging demonstrated that Rac1 signalling is active in ER+ cells following endocrine therapy. Targeting the Rac1 pathway with small molecule inhibitors (NSC23766, R-ketorolac) reduced survival and motility in resistant cells, inhibited in vivo Rac1 activity, and reduced tumour burden when combined with tamoxifen in a drug-refractory patient-derived xenograft. This work identifies the P-Rex1/Rac1 axis, as a potential therapeutic target for late recurring ER+ breast cancer.

  Study EGAS00001008353

• WNT-dependent interaction between inflammatory fibroblasts and FOLR2+ macrophages promotes fibrosis in chronic kidney disease

  Chronic kidney disease (CKD) is a public health problem driven by myofibroblast accumulation, leading to interstitial fibrosis. Heterogeneity is a recently recognised characteristics in kidney fibroblasts in CKD, but the role of different populations is still unclear. Here, we characterize a proinflammatory fibroblast population (named CXCL-iFibro), which corresponds to an early state of myofibroblast differentiation in CKD. By performing immunofluorescence studies and spatial transcriptomics analysis on 2 patients, we demonstrate that CXCL-iFibro co-localize with macrophages in the kidney and participate in their attraction, accumulation, and switch into FOLR2+ macrophages from early CKD stages on. In vitro, we cultured primary inflammatory fibroblasts that we characterized by using bulk RNAseq. We show that macrophages promote the switch of CXCL-iFibro into ECM-secreting myofibroblasts through a WNT/-catenin-dependent pathway, thereby suggesting a reciprocal crosstalk between these populations of fibroblasts and macrophages. Finally, the detection of CXCL-iFibro at early stages of CKD is predictive of poor patient prognosis, which shows that the CXCL-iFibro population is an early player in CKD progression and demonstrates the clinical relevance of our findings.

  Study EGAS50000000101

• Alterations in the gut microbiome in inflammatory arthritis implicate key taxa and metabolic pathways across arthritis phenotypes

  The gut microbiome has been implicated in several inflammatory conditions, but large-scale metagenomic evaluations have not yet traced the routes by which immunity in the gut affects systemic inflammatory diseases, such as inflammatory arthritis. To characterize the community structure and associated functional processes driving gut microbial involvement in arthritis, we investigated a total of 440 stool shotgun metagenomes comprising 275 adults diagnosed with rheumatoid arthritis, ankylosing spondylitis, or psoriatic arthritis, along with 165 healthy control individuals and individuals with joint pain without an underlying inflammatory cause.

  Study EGAS00001005525

• Pharmacogenomic profiling reveals molecular features of chemotherapy resistance in IDH wild type primary glioblastoma

  We stratified 69 primary IDH-wt GBM patients into TMZ-resistant (n = 29) and sensitive (n = 40) groups, using TMZ screening of the corresponding patient-derived glioma stem-like cells (GSCs). Genomic and transcriptomic features were then examined to identify TMZ-associated molecular alterations. Subsequently, we developed a machine learning (ML) model to predict TMZ response from combined signatures. Moreover, TMZ response in multisector samples (52 tumor sectors from 18 cases) was evaluated to validate findings and investigate the impact of intra-tumoral heterogeneity on TMZ efficacy.

  Study EGAS00001006989

• Germ Cell and Associated Heme Malignancies Evolve from a Common Shared Precursor

  Germ cell tumors (GCTs) are the most common cancer in men between the ages of 15-40. While most patients are cured, those with disease arising in the mediastinum have distinctly poor outcomes. One in every 17 patients with primary mediastinal non-seminomatous GCTs develop an incurable hematologic malignancy and prior data intriguingly suggests a clonal relationship exists between hematologic malignancies and GCTs in these cases. To date however, the precise clonal relationship between GCTs and the diverse additional somatic malignancies arising in such individuals has not been determined. Here, we traced the clonal evolution and characterized the genetic features of each neoplasm from a cohort of fifteen patients with GCTs and associated hematologic malignancies. We discovered that GCTs and hematologic malignancies developing in such individuals evolved from a common shared precursor, nearly all of which harbored allelically imbalanced TP53 and/or RAS pathway mutations. Hematologic malignancies arising in this setting genetically resembled mediastinal GCTs rather than de novo myeloid neoplasms. Our findings argue that this scenario represents a unique clinical syndrome, distinct from de novo GCTs or hematologic malignancies, initiated by an ancestral precursor which gives rise to the parallel evolution of GCTs and blood cancers in these patients. Reprinted from PMID: 32897884, with permission from JCI.

  Study phs002231

• HTAN Pilot Project: Single-Cell Transcriptomics Toolbox for Fresh and Frozen Human Tumors (Lung, Breast, Ovarian, Melanoma, Neuroblastoma, Sarcoma, Glioblastoma, Glioma, and Leukemia)

  Single cell transcriptomics is a powerful tool to map the complex tumor microenvironment, providing unprecedented resolution into cell types, cell states, cell interactions, and tumor heterogeneity. Depending on their tissue site and whether processing begins from a fresh or frozen sample, different tumors require different optimizations in order to obtain high quality single cell transcriptomic data. In this study, we profiled a range of tumor types, varying in cell-of-origin, solid and non-solid forms, patient ages, and transitions. The tumors also further varied in their tissue site and in their sample collection method. To enable profiling of such diverse samples, we developed a systematic toolbox for single cell RNA-Seq and single nucleus RNA-Seq of fresh and frozen clinical tumor samples. We validated this toolbox for 216,490 cells and nuclei extracted from 39 samples across 23 tumors (some tumors were split into multiple samples). The 23 tumors represented eight unique tumor types. This toolbox comprises both experimental and computational protocols as well as guidelines for comparing and selecting protocols for the biological problem of interest.

  Study phs001983

• Bioinformatic Methods and Bridging of Assay Results for Reliable Tumor Mutational Burden Assessment in Non-Small Cell Lung Cancer

  Tumor mutational burden (TMB) has emerged as a predictive biomarker of response to immune checkpoint inhibitors. Standardization of TMB measurement is essential for implementing diagnostic tools to guide treatment. Here we evaluate bioinformatic TMB analysis by whole exome sequencing (WES) in formalin-fixed, paraffin-embedded samples. In CheckMate 026, TMB was retrospectively assessed in 312 patients with non-small cell lung cancer (58% of the intent-to-treat population) who received first-line nivolumab treatment or chemotherapy. We examined the sensitivity of TMB assessment to bioinformatic filtering methods and assessed concordance between TMB data derived by WES and the FoundationOne CDx™ assay. TMB scores comprising synonymous, indel, frameshift, and nonsense mutations (all mutations) were 3.1-fold higher than data including missense mutations only, but values were highly correlated (Spearman’s r = 0.99). Scores from CheckMate 026 samples including missense mutations only were similar to those generated from data in The Cancer Genome Atlas, but those including all mutations were generally higher. Using databases for germline subtraction (instead of matched controls) showed a trend for race-dependent increases in TMB scores. Parameter variation can therefore impact TMB calculations, highlighting the need for standardization. Encouragingly, WES and FoundationOne CDx outputs were highly correlated (Spearman’s r = 0.90) and differences could be accounted for by empirical calibration, suggesting that reliable TMB assessment across assays, platforms and centers is achievable.

  Dataset EGAD00001005035

• Integrated Genomic Analysis of Periampullary Tumors at The Human Genome Sequencing Center, Baylor College of Medicine (HGSC-BCM)

  Adenocarcinomas arising in the complex environment of the ampulla of Vater constitute a histopathological heterogenous group, presumably originating from the different epithelial cellular constituents present at the site: pancreas, bile duct, and intestinal duodenum. These tumors have been described in many different ways: intra-ampullary, periampullary, intra-ampullary papillary-tubular neoplasm, ampullary-ductal, periampullary-duodenal, and ampullary-not otherwise specified. These varied classifications reflect the difficulty in classifying these tumors into specific groups. Only the tumors clearly localized in the bile duct or duodenum are identified as distal cholangiocarcinomas (CAC) or duodenal adenocarcinomas (DUOAC). The current classification is based on macroscopic features that may distinguish the epithelium of origin, microscopic features, clinicopathological criteria, histopathology and expression of differential markers. This classification is subjective and prone to inter-observer variability and significantly impacts on treatment selection and therapeutic development. In order to define subtypes of periampullary cancer with clinical relevance, we performed whole exome sequencing and copy number analysis of 160 cancers arising in the periampullary region, 62 of these clearly arising from either the bile duct (n = 44), or the duodenum (n = 18) and 98 periampullary cancers (AMPAC) where the epithelium of origin could not be clearly defined.

  Study phs000895

• NHLBI TOPMed: SubPopulations and InteRmediate Outcome Measures In COPD Study (SPIROMICS)

  Subpopulations and Intermediate Outcome Measures in COPD Study Description Subpopulations and intermediate outcome measures in COPD study (SPIROMICS) supports the prospective collection and analysis of phenotypic, biomarker, genetic, genomic, and clinical data from subjects with COPD for the purpose of identifying subpopulations and intermediate outcome measures. It is funded by the National Heart, Lung, and Blood Institute and is coordinated by the University of North Carolina at Chapel Hill. Molecular fingerprinting and extensive subject phenotyping will be performed to identify disease subpopulations and to identify and validate surrogate markers of disease severity, which will be useful as intermediate outcome measures for future clinical trials. Secondary aims are to clarify the natural history of COPD, to develop bioinformatic resources that will enable the utilization and sharing of data in studies of COPD and related diseases, and to create a collection of clinical, biomarker, radiographic, and genetic data that can be used by external investigators for other studies of COPD. Participating Institutions: Research participants for SPIROMICS will be enrolled, phenotyped, and followed at twelve SPIROMICS Clinical Centers: Columbia University, Temple University, Johns Hopkins University, Wake Forest University, University of Michigan, University of Illinois at Chicago, University of Iowa, University of Utah, National Jewish Health, University of California at San Francisco, and University of California at Los Angeles. The University of North Carolina at Chapel Hill serves as the Genomics and Informatics Center. The Radiology Reading Center is based at the University of Iowa. The PFT Reading Center is based at the University of California at Los Angeles. Study Design: SPIROMICS is a prospective cohort study that enrolled approximately 2,981 participants at twelve clinical centers over five years. Participants are distributed across four enrollment strata (i.e., Never-smokers, Smokers without COPD, Mild/Moderate COPD, and Severe COPD). Study Visits: Participants have up to four in-person visits (Baseline and Annual Clinic Visits at years 1, 2, 3 after Baseline). Study questionnaires and spirometry are completed at all main study visits. Blood and urine samples are collected at visits 1, 2, and 4. Sputum samples are collected at Visit 1. The CT scans are completed and Baseline and Visit 2. Participants also receive quarterly follow-up calls to assess health status and determine if an exacerbation has occurred. Substudies Bronchoscopy and Immunophenotyping Substudy The Bronchoscopy Substudy will enroll 50 subjects per site, for a total of 300 participants. These participants will be recruited across all four study strata. This substudy includes two study visits. During the first visit, sputum samples are collected for Immunophenotyping analysis at the Immunophenotyping Core Lab based at the University of Michigan. Participants then return for a second visit during which samples are collected via bronchoscopy, including bronchoalveolar lavage, epithelial brushings and bronchial biopsies. Immunophenotyping analysis is also conducted on BAL and blood collected during the bronchoscopy study visit. Repeatability Substudy The entire baseline clinic visit was repeated on 98 volunteers to determine reliability of measurement procedures. All baseline study-related procedures and questionnaires, including the CT scan, were re-administered and new samples of blood, urine, saliva, and sputum were collected. Field center staff processed these biospecimen samples according to the same protocol. Exacerbation Substudy The Exacerbation Substudy is a prospective, longitudinal observational study of up to 400 participants, which will allow the assessment of participant-driven health care utilization (HCU) events and symptom-defined exacerbation events over time. HCU-driven events are defined by change in medical treatment in response to a perceived COPD Exacerbation. Symptom-based events will be defined by using EXACT-PRO (EXacerbations of Chronic Pulmonary Disease Tool - Patient Reported Outcome), a daily symptom diary developed to measure the frequency, severity, and duration of exacerbations in clinical trials. All participants are provided with a PDA on which to complete the diary. Participants reporting a possible COPD exacerbation will be brought into the study clinic for a study visit to collect biological specimens and questionnaire data. The overall objectives of the Exacerbation Substudy are to: Obtain clinical data and specimens on SPIROMICS participants before and during an acute COPD exacerbation defined by Health care utilization triggered by the subject, or Symptomatic change (triggered by an EXACT defined threshold) Describe symptomatic changes occurring around HCU and symptom-defined (EXACT) events and their relationship to clinical and specimen data, Characterize the non-exacerbation "stable" state in COPD using the EXACT, including: Symptom variability (EXACT), Clinical data and specimen parameters during a stable state (baseline), The relationship between clinical and specimen data and symptom severity and variability. Explore the characteristics of stable patients, defined as those who do not experience HCU or symptom-defined (EXACT) events during the sub-study period, using baseline clinical data and specimens, and compare these characteristics with patients who experience HCU and/or symptom-defined events. Examine the relationship between HCU and symptom-defined exacerbation frequency during the first one-year period of follow-up for exacerbations and health outcomes, including 12-month change in lung function and COPD health status, and longer-term morbidity and mortality, with the latter derived from long-term data from the larger SPIROMICS study.

  Study phs001927

• Integrative analysis of exome-seq, RNA-seq, ATAC-seq (bulk and single-cell), and Hi-C data generated from 3-D spatially mapped samples acquired during surgical resection from 10 patients diagnosed with IDH-WT glioblastoma

  Glioblastoma (GBM), the most common primary brain cancer in adults, remains incurable with no targeted therapies approved despite decades of investigation into its molecular landscape. Treatment failure is attributed to intratumoral heterogeneity in the GBM genome and epigenome, which foster tumor evolution and selection of resistant clones. However, tumor evolution and intratumoral heterogeneity remain poorly understood on the level of the whole tumor as most studies are based on single samples per patient and lack spatial context. Here, we have used 3-D neuronavigation during surgical resection for 10 primary IDH-WT GBM patients to collect 102 samples representing maximal tumor diversity, each mapped by 3-D spatial coordinates. We have applied a strategic set of genomic and epigenomic assays spanning multiple levels of resolution to discover, orthogonally validate, and functionally assess drivers of tumor evolution and intratumoral heterogeneity. These include extrachromosomal DNA amplifications, chromothripsis events, inversions, and translocations that disrupt both the GBM genome and epigenome while generating fusion transcripts and opportunities for therapeutic intervention. We define epigenomic programs that contribute to GBM evolution and intratumoral heterogeneity, revealing their 3-D spatial patterning within whole tumors and their cell type(s) of origin in single-cell data from the same tumor samples. Notably, we distinguish neuronal, glial, and immune programs aberrantly active in tumor cells from their counterparts in normal cells and discover NEUROD1, JUN/FOS, and NF1 transcription factors as key drivers of GBM evolution and growth. Collectively, these data provide unprecedented insight into GBM evolution and intratumoral heterogeneity from single-cell to whole-tumor resolution, redefining current understanding and providing a rich resource of targets for therapeutic investigation.

  Study EGAS00001006785

• Transcriptomics-driven classification of ALAL

  Acute leukemia of ambiguous lineage (ALAL) is a rare, poor-prognosis subtype of acute leukemia that cannot be assigned to a single hematopoietic lineage. Although ALAL patients are typically treated with acute myeloid leukemia (AML) or acute lymphoblastic leukemia (ALL) regimens, optimal treatment choice is hindered by their lineage ambiguity. Therefore, we investigated the added value of transcriptomics for improving lineage assignment, currently based mainly on surface markers. First, we used an in-house pipeline to comprehensively detect genetic lesions in RNA-sequencing data (n=23) with a sensitivity >90%, as confirmed by targeted DNA sequencing. Second, we compared ALAL gene expression profiles (GEPs) with representative AML (n=145), B-ALL (n=223) and T-ALL (n=85) cases. In a principal component analysis, ALALs did not form a separate group, but clustered with AML, B-ALL or T-ALL. Accordingly, a machine learning classifier trained with GEPs of acute leukemias segregated 20/23 ALALs into myeloid-, B- or T-lymphoid leukemia. These 20 cases harbored genetic abnormalities consistent with the classifier-assigned leukemia. Furthermore, ALAL GEPs were deconvoluted with single-cell transcriptional profiles of normal hematopoietic cells using CIBERSORTx, revealing enrichment for signatures of lineages corresponding to the leukemic type predicted by our algorithm. The classifier was validated on an external ALAL cohort (n=24), assigning 75% of the patients to a lineage matching their immunophenotypic and methylation profiles. In conclusion, integrative analysis of RNA-sequencing data can accurately classify most ALAL cases. The pipeline and classifier developed in our study are valuable tools to improve ALAL diagnosis and guide therapeutic decisions.

  Study EGAS00001007967

• DDX50 Cooperates With STAU1 to Effect Stabilization of Pro-Differentiation RNAs

  This study was undertaken to determine the role of DDX50 in epidermal homeostasis. Primary neonatal foreskin keratinocytes were obtained from discarded surgical samples at a California hospital. Samples were anonymous and not selected. DDX50, which was associated with STAU1, was found to promote epidermal differentiation and transcript stability using the methods of RNA-seq, CLIP-seq and DMS-MaP-seq.

  Study phs003766

• NIDDM-Atherosclerosis Study (NIDDM-Athero)

  The NIDDM-Atherosclerosis Study, funded by NHLBI, was designed as a family study to examine the genetic basis of subclinical atherosclerosis and diabetes in Hispanic families. Family members of probands with T2D were recruited in the Los Angeles area. The baseline examination of the cohort included the euglycemic hyperinsulinemic clamp test from which the two key phenotypes were obtained: insulin sensitivity (M) and metabolic clearance rate of insulin (MCRI). Genome-wide genotyping was obtained under separate funding by NIDDK as a part of the GUARDIAN (Genetics Underlying Diabetes in Hispanics) Consortium.

  Study phs001130

• Comparative Sequence Analysis Between Primary and Metastatic Colorectal Cancer Lesions

  Molecular profiling for somatic mutations that predict response to anti-EGFR therapy in colorectal cancer (CRC) has become standard practice. However, abundant tissue from metastatic lesions is not always available from patients with metastatic CRC. Concerns involving genetic heterogeneity between primary and metastatic lesions have called into question the suitability of profiling primary tumors in patients with metastatic disease. Further, the identification of discordant mutations between matched primary and metastatic tumors would be of biological interest for the delineation of biomarkers of tumor progression and metastasis. To explore the degree of genetic heterogeneity between matched primary and metastatic tumors in CRC, we performed whole genome sequencing of four patient "trios" comprised of a primary colon tumor, a liver metastasis, and matched normal (non-cancerous) tissue. Somatic mutations and indels were called in each tumor and compared between primary and metastatic lesions.

  Study phs000790

• Diverse mutational landscapes in human lymphocytes

  A lymphocyte suffers many threats to its genome, including programmed mutation during differentiation, antigen-driven proliferation and residency in diverse microenvironments. After developing protocols for single-cell lymphocyte expansions, we sequenced whole genomes from 717 normal naive and memory B and T lymphocytes and hematopoietic stem cells. All lymphocyte subsets carried more point mutations and structural variants than haematopoietic stem cells – the extra mutations were mostly acquired during differentiation, with burdens higher in memory than naive lymphocytes, although T cells also had a higher rate of mutation accumulation throughout life. Off-target effects of immunological diversification accounted for most of the additional differentiation-associated mutations in lymphocytes. Memory B cells acquired, on average, 18 off-target mutations genome-wide for every one on-target IGV mutation during the germinal centre reaction. Structural variation was 16-fold higher in lymphocytes than stem cells, with ~15% of deletions being attributable to off-target RAG activity. Mutational processes associated with ultraviolet light exposure and other sporadic mutational processes generated hundreds to thousands of mutations in some memory lymphocytes. The mutation burden and signatures of normal B lymphocytes were broadly comparable to those seen in many B-cell cancers, suggesting that malignant transformation of lymphocytes arises from the same mutational processes active across normal ontogeny. The mutational landscape of normal lymphocytes chronicles the off-target effects of programmed genome engineering during immunological diversification and the consequences of differentiation, proliferation and residency in diverse microenvironments.

  Dataset EGAD00001008107

• Cohort-Based Genome-Wide Association Study of Glioma (GliomaScan)

  We conducted a new independent GWAS of adult glioma using 1,856 cases and 4,955 controls from 14 cohort studies belonging to the Cohort Consortium, 3 case-control studies, and 1 population-based case only study. Cases were newly diagnosed glioma [ICDO-3 codes 9380-9480 or equivalent], and controls were cancer-free at the time of glioma diagnosis. We found evidence of strong replication for three of the seven previously reported associations at 20q13.33 (RTEL), 5p15.33 (TERT), and 9p21.3 (CDKN2BAS), and consistent association signals for the remaining four at 7p11.2 (EGFR both loci), 8q24.21 (CCDC26) and 11q23.3 (PHLDB1). The direction and magnitude of the signal were consistent for samples from cohort and case-control studies, but the strength of the association was more pronounced for loci rs6010620 (20q,13.33; RTEL) and rs2736100 (5p15.33, TERT) in cohort studies despite the smaller number of cases in this group, likely due to higher grade tumors being captured in the cohort studies. Our findings suggest that larger studies focusing on novel approaches as well as specific tumor subtypes or subgroups will be required to identify additional common susceptibility loci for glioma risk.

  Study phs000652

• Suppressors and activators of JAK-STAT signaling at diagnosis and relapse of acute lymphoblastic leukemia in Down Syndrome

  Children with Down Syndrome (DS) are prone to development of high risk B-cell precursor ALL (DS-ALL) that differs genetically from most sporadic pediatric ALLs. Increased expression of CRLF2, the receptor to thymic stromal lymphopoietin (TSLP) characterizes about half of DS-ALLs and also a subgroup of sporadic “Philadelphia-like ALLs”. To understand the pathogenesis of relapsed DS-ALL we performed integrative genomic analysis of 25 matched diagnosis remission and relapse DS-ALLs. We found that the CRLF2 rearrangements are early events during DS-ALL evolution and generally stable between diagnoses and relapse. Secondary activating signaling events in JAK-STAT/RAS pathway were ubiquitous but highly redundant between diagnosis and relapse, suggesting that signaling is essential but that no specific mutations are “relapse driving”. We further found that activated JAK2 may be naturally suppressed in 25% of CRLF2 positive DS-ALL by loss-of-function aberrations in USP9X, a deubuitinase previously shown to stabilize the activated phosphorylated JAK2. Interrogation of large ALL genomic databases extended our findings up to 25% of CRLF2 positive, “Ph like” ALLs. Pharmacological or genetic inhibition of USP9X as well as treatment with low-dose Ruxolitinib enhanced the survival of pre-B ALL cells overexpressing mutated JAK2. Thus, somehow counterintuitive, we found that suppression of JAK-STAT “hyper-signaling” may be beneficial to leukemic B cell precursors. This and the reduction of JAK mutated clones at relapse suggest that the therapeutic effect of JAK specific inhibitors may be limited. Rather, combined signaling inhibitors or direct targeting of the TSLP receptor may be a useful therapeutic strategy for DS-ALL.

  Study EGAS00001002410

• Denisova admixture in Southeast Asia and Oceania

  Genome-wide data from populations from south Asia, Southeast Asia, and Oceania were analyzed for signatures of Denisova admixture to infer the history of human migration to this part of the world

  Study EGAS00001006132

• High number of somatic mutations found in the healthy blood compartment of a 115-year-old woman reveals oligoclonal hematopoiesis

  The somatic mutation burden in healthy white blood cells (WBCs) is not well known. Based on deep whole genome sequencing, we estimate that ~450 somatic mutations accumulated in the non-repetitive genome within the healthy blood compartment of a 115-year old woman. The detected mutations appear to have been harmless passenger mutations: they were enriched in non-coding, AT-rich regions that are not evolutionarily conserved, and they were depleted for genomic elements where mutations might have favorable or adverse effects on cellular fitness, such as regions with actively transcribed genes.The distribution of variant allele frequencies of these mutations suggests that the majority of the peripheral white blood cells were offspring of two related hematopoietic stem cell (HSC) clones. Moreover, telomere lengths of the WBCs were significantly shorter than telomere lengths from other tissues. Together, this suggests that the finite lifespan of HSCs, rather than somatic mutation effects, may lead to hematopoietic clonal evolution at extreme ages.

  Study EGAS00001000660

• Gut microbiome modulates response to anti PD1 immunotherapy in metastatic melanoma patients

  There is a growing appreciation of the role of the microbiome in cancer, and evidence in pre-clinical models that the gut microbiome may modulate responses to immune checkpoint blockade though this has not been well-characterized in patients. We analyzed the oral (n=86)and gut (n=43) 16S microbiome in melanoma patients on PD-1 blockade. Significant differences were noted in the diversity and composition of the gut microbiome between responders and non-responders in patients with a fecal microbiome sample, with significantly higher alpha diversity and relative abundance of Ruminococcaceae bacteria) in R. Metagenomic studies (n=25) revealed functional differences in gut bacteria in R including enrichment of anabolic pathways. Immune profiling demonstrated enhanced systemic and anti-tumor immunity in patients with a favorable gut microbiome as well as in germ-free mice receiving fecal transplant from responding patients. Together, these data have important implications for treatment with immune checkpoint blockade in cancer. The fastq files associated with this dataset are stored at ENA under the following accession numbers: PRJEB22894 (Fecal 16S), PRJEB22874 (Oral 16S), PRJEB22895 (Murine 16S), PRJEB22893 (Fecal WGS).

  Study EGAS00001002698

• HYPERMUTATION AND MALIGNANT PROGRESSION IN LOW-GRADE GLIOMA PATIENTS

  While patients with primary glioblastoma (GBM) have benefited from increased overall survival due to treatment with the alkylating chemotherapy temozolomide (TMZ), the benefit ofTMZ for patients with diffuse low-grade gliomas (LGG) remains unknown. To further explore the relationships among TMZ treatment, hypermutation and malignant progression, we studied tumor evolution in an expanded cohort of untreated and TMZ-treated patients by exome-sequencing of paired IDH1/2-mutant LGGs and their post-TMZ recurrences. Together, these findings further our understanding of the molecular features and clinical behavior of hypermutated clones.

  Study EGAS00001002368

• Genetic characterization patients affected by Cancer of Unknown Primary

  The genetic changes sustaining the development of human Cancers of Unknown Primary (CUP) remain elusive. Here we report the genomic profiling of 14 rigorously selected CUP samples. Although no specific recurring mutation in known driver genes was found, a comparison of the mutational landscape of CUPs with that of most other human tumor types revealed a consistent change in diverse genes belonging to the axon guidance KEGG pathway.

  Study EGAS00001006621

• FGFP and TR-MDD shotgun sequencing samples (N=157)

  Study exploring the neuroactive potential of the human gut microbiota in relation to quality of life and depression. This dataset includes shotgun sequenced samples from the FGFP (Flemish Gut Flora Project: N=150) and TR-MDD (Treatment-Resistant Major Depression Disorder: N=7). Samples were provided by volunteers from the Flanders region in Belgium, and the dataset is balanced for age, gender, and stool consistency.

  Study EGAS00001003298

• Leiden_melanomafamilies

  Around 10% of patients who present in melanoma clinics have a first degree relative with a previous diagnosis of melanoma. While around 3% have three or more relatives who have been diagnosed with the disease. In this project we will whole genome sequence patients from large Dutch familial melanoma pedigrees to identify mutations in genes that drive melanomagenesis. The identification of these genes will facilitate the management of familial melanoma patients and their families.

  Study EGAS00001000627

• Engineering large chromosomal deletions by CRISPR-Cas9

  Arm-level chromosomal deletions are a prevalent and defining feature of cancer. A high degree of tumor-type and subtype specific recurrencies suggest a selective oncogenic advantage. However, due to their large size it has been difficult to pinpoint the oncogenic drivers that confer this advantage. Suitable functional genomics approaches to study the oncogenic driving capacity of arm-level deletions are limited. Here we present an effective technique to engineer arm-level deletions by CRISPR-Cas9 and create isogenic cell line models. We simultaneously induce double-strand breaks (DSBs) at two ends of a chromosomal arm and select the cells that have lost the intermittent region. Using this technique, we induced arm-level deletions on chromosome 11q (65 MB) and chromosome 6q (53 MB) in neuroblastoma cell lines. Such isogenic models enable further research on the role of arm-level deletions in tumor development and growth, and their possible therapeutic potential.

  Study EGAS00001005134

• Genome analysis of oesophageal cancer and Barrett's oesophagus

  The median survival of oesophageal cancer this year is only 13 to 19 months after diagnosis and more than 90% will die from their disease. Therefore better treatment options are needed. The likelihood of cure for early screen-detected cancers is much higher. Barrett's oesophagus is a pre-cancerous lesion associated with a 30-40 fold increased risk of developing cancer. In an attempt to detect cancer early many patients with Barrett's are enrolled into surveillance programs involving regular endoscopies. A major problem with this approach is that the prevalence of BO in the population is estimated to be around 2%, but most patients with BO will never develop cancer. We are undertaking genomic and/or transcriptomic analysis of oesophageal tumours, Barrett's oesophagus and matched normal samples. The aim is to identify oesophageal-related genomic and transcriptomic alterations, which may reveal mutational process occurring, suggest biomarkers of tumour progression and treatment and identify novel treatment strategies.

  Study EGAS00001002864

• Autozygosity pilot - British-Pakistani from Birmingham 2

  The offspring of first cousin marriages have ~6% of their genome autozygous, i.e. homozygous identical by descent, or even more if there was further consanguinity in their ancestry. In the UK there are large populations with very high first cousin marriage rates of 20-50%. Sequencing the exomes of a sample of these individuals has the potential both to support genetic health programmes in these populations, and to provide genetic research information about rare loss of function mutations. This pilot study based on existing British-Pakistani cohort samples from Birmingham will identify homozygous individuals for almost all variants down to an allele frequency around 1%, plus individuals carrying hundreds of new homozygous rare loss-of-function variants, and will support development of community relations and ethics for a wider study currently being designed. The data deposited in the EGA consists of low coverage whole exome sequencing on these samples.

  Dataset EGAD00001001026

• Autozygosity pilot - British-Pakistani from Birmingham

  The offspring of first cousin marriages have ~6% of their genome autozygous, i.e. homozygous identical by descent, or even more if there was further consanguinity in their ancestry. In the UK there are large populations with very high first cousin marriage rates of 50-80%. Sequencing the exomes of a sample of these individuals has the potential both to support genetic health programmes in these populations, and to provide genetic research information about rare loss of function mutations. This pilot study based on existing British-Pakistani cohort samples from Birmingham will identify homozygous individuals for almost all variants down to an allele frequency around 1%, plus individuals carrying hundreds of new homozygous rare loss-of-function variants, and will support development of community relations and ethics for a wider study currently being designed. The data deposited in the EGA consist of low coverage whole exome sequencing on these samples.

  Dataset EGAD00001001025

• Single cell long read whole genome sequencing reveals somatic transposon activity in human brain

  The advent of single cell DNA sequencing revealed astonishing dynamics of genomic variability, but failed at characterizing smaller to mid size variants that on the germline level have a profound impact. In this work we discover novel dynamics in three brains utilizing single cell long-read sequencing. This provides key insights into the dynamic of the genomes of individual cells and further highlights brain specific activity of transposable elements.

  Study EGAS50000001156

• The Role of CTCF in the Organization of the Centromeric 11p15 Imprinted Domain Interactome

  Beckwith-Wiedemann Syndrome (BWS) is the most common human imprinting disorder and comprises a spectrum of overgrowth phenotypes. While approximately 35% of BWS cases are caused by isolated loss of methylation (LOM) at the human imprinting center 2 (IC2) on chromosome 11p15, only around 5% of patients develop BWS due to structural alterations to the IC2 domain. In this study, we identify a BWS-causing 7.6 kB familial deletion within the IC2 domain by performing Whole Genome Sequencing (WGS) on patient-derived human fibroblasts. Using a chromatin conformation capture technique, Capture C, on 3 control fibroblast samples, 3 BWS LOM fibroblast samples, and the single familial deletion sample, we investigate how the domain interactome changes in BWS patients. We find that high-strength interactions occur within the IC2 domain between the 5’ end of KCNQ1, KCNQ1 intron 2, and the 5’ end of CDKN1C in the control profile that are largely abrogated in the BWS samples, but some lower frequency contacts between the imprinting control region (ICR), KCNQ1 intron 2, and the 5’ end of CDKN1C are unaffected. We conclude that the strong KCNQ1-CDKN1C interactions are important in maintaining the domain imprint.

  Study phs002408

• Integrated genetic and epigenetic analysis defines novel molecular clusters in rhabdomyosarcoma

  Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in childhood. To unravel the genetic/epigenetic basis of RMS, we studied 60 RMS cases using whole exome/transcriptome sequencing, copy number (CN) profiling, and DNA methylome analysis. Based on DNA methylation patterns, RMS was clustered into 4 distinct subtypes, which exhibited remarkable correlation with mutation/CN profiles, histological phenotypes, and clinical behaviors. A1 and A2 subtypes, especially A1, largely corresponded to alveolar histology with frequent PAX3/7 fusions and alterations in cell cycle regulators. In contrast, mostly showing embryonal histology, both E1 and E2 subtypes were characterized by high frequency of CN alterations and/or allelic imbalances, FGFR4/RAS/AKT pathway mutations and PTEN mutations/methylation and in E2, also by p53 inactivation. Despite generally poor prognosis in RMS, patients in E1 cluster exceptionally showed excellent prognosis. Our results highlight the close relationship between DNA methylation status and gene mutations and biological behavior in RMS.

  Study EGAS00001000884

• Oncogenic cooperation in a human de novo T-ALL model

  We previously modelled T-cell acute lymphoblastic leukemia (T-ALL) by over-expressing four oncogenes (activated NOTCH1, LMO2, TAL1, and BMI1) in healthy human umbilical cord blood cells through lentiviral transduction (Kusakabe et al, Nat Commun 10, 2913 (2019). https://doi.org/10.1038/s41467-019-10510-8). We have now tested individual effects of the four oncogenes by transducing human cells with all possible combinations of one, two, three, and four oncogenes. We also tested the effects of additional known T-ALL oncogenes including HOXA9, LYL1, MEF2C, NXK2.1, TLX1, and TLX3. In addition to culture in vitro on OP9-DL1 feeders, we also injected transduced cells into immunodeficient NSG mice. We observed differing abilities of the various oncogene combinations to support growth in vitro and frank leukemia in vivo. This study includes RNA-seq data from samples of transduced cells cultured in vitro as well as cells harvested from clinically moribund mice after development of leukemia in vivo.

  Study EGAS00001006055

• RNA-seq of cultured human hematopoietic stem and progenitor cells from umblical cord blood in cytokine-rich ex vivo culture conditions following sphingolipid modulation

  To elucidate the biological pathways altered by sphingolipid modulation with N-(4-hydroxyphenyl) retinamide (4HPR) treatment in human HSPC that may contribute to the restraint in proliferation while promoting persistence of HSC self-renewal as well as determine the mechanism of synergy in enhancement of HSC self-renewal with CB CD34+ agonists UM171 and StemRegenin 1 (SR1), we performed RNA-sequencing (RNA-Seq) of 3 pools of lin-CB cells following 2 or 4 days with DMSO, 4HPR, UM171+SR1 or 3-Factor (4HPR+UM171+SR1). We identified modulation of sphingolipid metabolism regulates self-renewal through activating coordinated stress pathways that coalesce on endoplasmic reticulum stress and autophagy programs.

  Study EGAS00001003756