Search Results - EGA European Genome-Phenome Archive

Characterization of Human Transcriptome by Computational and HTS Approaches

We have deeply sequenced total RNA and whole exome of six different post-mortem human snap-frozen tissues (brain, liver, lung, striated muscle, kidney, heart) from three unrelated healthy Caucasian individuals (males, aged 47-54) with the aim to characterize post-transcriptional events due to alternative splicing and RNA editing. In addition, to facilitate the detection of RNA editing sites we have also resequenced the entire genome of each individual. Data are also used to investigate tissue-specific abundance of mitochondrial DNA from exomes and its correlation with mitochondrial transcription, mass and respiratory activity. Tissues were obtained from Cureline (South San Francisco, CA, USA). DNA and RNA were purified using the DNeasy Blood and Tissue Kit (Qiagen, Hilden, Germany) and the RNeasy Plus Mini Kit (Qiagen, Hilden, Germany), respectively. Exome capture was performed using the TruSeq Exome Enrichment Kit (Illumina, San Diego, CA) and the sequencing was carried out at IGA Technology Services in Udine (Italy) on Illumina HiSeq 2000 (generating for each tissue approximately 40 millions of 100bp paired-end reads). Strand-oriented RNA reads (2x100bp) were generated using the TruSeq Stranded Total RNA Sample Prep Kit (Illumina, San Diego, CA, USA) and sequenced at IGA Technology Services in Udine (Italy) on Illumina HiSeq 2000 platform. Whole genome resequencing was performed at Personal Genomics in Italy on an Illumina HiSeq 2000 platform (2x100bp) with a mean coverage of 30X. MiRNA-Seq was instead performed at IBBE Institute (Italy) using the TruSeq Small RNA Sample Prep Kit on Illumina MiSeq platform. Preliminary bioinformatics analyses have been performed to check quality using FASTQC software and inconsistent read regions have been trimmed using trim_galore. All reads were mapped onto the human reference genome (version hg19) using GSNAP.

Study phs000870

Chromothripsis in Patient WHIM-09

We are studying the natural history, pathogenesis and treatment of patients with WHIM syndrome, an immunodeficiency disorder characterized by warts, hypogammaglobulinemia, recurrent infections and neutropenia usually due to autosomal dominant gain-of-function mutations in chemokine receptor CXCR4. We have identified a patient born with WHIM syndrome and the WHIM mutation CXCR4R334X who has been disease-free for 20 years and who lacks CXCR4R334X in myeloid cells, the cells that drive disease manifestations. She is a genetic and hematopoietic mosaic, since she still has the mutation in lymphoid cells and non-hematopoietic cells. Cytogenetics and microarray analysis revealed that the mechanism of loss of the mutation was deletion of the mutant allele from one copy of chromosome 2. Whole genome sequencing of patient neutrophil and skin fibroblast genomic DNA revealed that the mechanism of deletion was chromothripsis, a process of chromosome shattering resulting in deletions and rearrangements of the non-deleted chromosomal segments. In the patient, this process evidently occurred in a single hematopoietic stem cell (HSC), resulting in deletion of the disease allele CXCR4R334X and one copy of 163 other genes on chromosome 2. This HSC evidently acquired a growth advantage and repopulated the HSC population and the myeloid lineage. Consistent with this, studies using gene targeted mice in competitive bone marrow transplantation experiments revealed that selective Cxcr4 haploinsufficiency (inactivation of one copy of Cxcr4 and not of any other genes) was sufficient to confer a strong engraftment advantage over bone marrow cells from wild type mice as well as bone marrow cells from a mouse model of WHIM syndrome. These results suggest that CXCR4 knockdown may be a useful strategy to enhance bone marrow engraftment in the absence of toxic bone marrow conditioning regimens.

Study phs000856

CIP: Differential Response to Hydroxyurea and Incidence of Stroke in Sickle Cell Disease

Sickle cell disease (SCD) is a severe debilitating hematological disorder associated with a high degree of morbidity and mortality. There are approximately 200,000 babies born with sickle cell disease each year, with the disease predominately affecting individuals in Africa. The overall global burden of the disease is tremendous, with more than 100,000 patients currently in the US and further millions worldwide. The governing bodies of the World Health Organization have recently adopted a resolution to strengthen the response to sickle disease in all affected countries and there is a definite need for high quality sickle cell disease research that has the potential to improve the treatment and prognosis of patients with this devastating disease. The clinical manifestations of SCD arise from a complex pathophysiology that includes hemolysis, acute vaso-occlusion, endothelial dysfunction, inflammation, and chronic organ damage. While the individual clinical course of this disease is highly variable, many of the associated complications demonstrate some degree of heritability. Intensive research into identifying genetic modifiers that can affect the pathophysiology of SCD has been limited to date and there is an urgent need to improve of our knowledge the molecular mechanisms underlying the clinical complications of SCD. The Sickle cell CIP project is investigating complication of stroke and pharmacogenomics of hydroxyurea response in patients with sickle cell anemia. The major benefit of hydroxyurea comes from its ability to induce fetal hemoglobin (HbF) and higher HbF levels are associated with reduced morbidity and mortality in SCA patients. We will perform whole exome and whole genome sequencing of SCA patients in order to identify genome variants associated with incidences of stroke and HbF response to hydroxyurea.

Study phs000691

National Cancer Institute (NCI) Non-Hodgkin Lymphoma Genome-wide Association Study (GWAS)

Within the framework of the International Lymphoma Epidemiology Consortium (InterLymph) Consortium and NCI-sponsored Cohort Consortium, investigators from 22 studies came together to conduct a genome-wide association study (GWAS) of non-Hodgkin lymphoma (NHL). The goal of the study was to identify common genetic variants associated with the risk of NHL. As described previously (Berndt SI et al., Nature Genetics, 2013, PMID: 23770605 ), cases and controls from 22 studies, including nine prospective cohort studies, eight population-based case-control studies, and five clinic or hospital-based case-control studies were included in this effort. Cases of NHL were ascertained from cancer registries, clinics or hospitals, or through self-report verified by medical and pathology reports. The phenotype information for all NHL cases was reviewed centrally at the InterLymph Data Coordinating Center and harmonized according to the hierarchical classification proposed by the InterLymph Pathology Working Group based on the World Health Organization (WHO) classification (2008). Controls were frequency matched to cases on age and gender by study, and the study was limited to subjects of European ancestry. Participants were genotyped using the Illumina Omni Express. The NCI Non-Hodgkin Lymphoma GWAS is utilized in the following dbGaP sub-studies. To view genotypes and derived variables collected in these sub-studies, please click on the following sub-studies below or in the "Sub-studies" box located on the right hand side of this top-level study page phs000801 NCI Non-Hodgkin Lymphoma GWAS. phs000802 NCI Non-Hodgkin Lymphoma GWAS: CLL phs000818 NCI Non-Hodgkin Lymphoma GWAS: Controls phs000889 NCI Non-Hodgkin Lymphoma GWAS: DLBCL phs000890 NCI Non-Hodgkin Lymphoma GWAS: FL phs000891 NCI Non-Hodgkin Lymphoma GWAS: MZL

Study phs000801

Intraductal transplantation models of human pancreatic ductal adenocarcinoma reveal progressive transition of molecular subtypes

Pancreatic ductal adenocarcinoma (PDAC) is the most lethal common malignancy, with little improvement in patient outcomes over the past few decades. Recently, subtypes of pancreatic cancer with different prognoses have been elaborated. However, the inability to model these subtypes has precluded mechanistic investigation of their origins. Here, we present an orthotopic xenotransplantation model of human patient-derived PDAC organoids in which organoids are grafted via direct injection into the pancreatic duct of immunocompromised (NSG) mice. Tumors that arise from these intraductally grafted organoids (IGOs) enable the distinction of the two main PDAC subtypes: intraepithelial neoplasms from this model progress in an indolent or invasive manner representing the classical or basal-like subtypes of PDAC, respectively, depending on which organoid they originate from. RNA-sequencing was used to characterize the tumors that resulted from IGO xenografts, and to compare the fast- and slow-progressing IGO tumors. Tumors generated using a traditional orthotopic xenograft model, in which organoids are injected into the parenchyma of the pancreas (i.e. orthotopically grafted organoids, or OGOs), were also sequenced for comparison with the IGO tumors. To further characterize the model, laser-capture microdissection followed by RNA-sequencing was used to compare intraductal to invasive lesions resulting from IGOs. To better understand the connection between KRAS signaling and the fast-progressing phenotype, RNA-sequencing was used to compare cultured organoids in which oncogenic KRAS (GeneID: 3845) was hyperactivated to control organoids, and to compare laser-capture microdissected lesions derived from IGOs of KRAS-hyperactivated organoids to lesions derived from control organoids. To ascertain copy number, low-coverage, whole genome DNA-sequencing was performed on organoids cultured in vitro, and on laser-capture microdissected lesions derived from IGO tumors.

Study phs002045

Clinical Sequencing Exploratory Research Consortium: Incorporation of Genomic Sequencing into Pediatric Cancer Care

This project includes data derived from subjects enrolled in the BASIC3 (Baylor College of Medicine Advancing Sequencing in Childhood Cancer Care) study. BASIC3 is a National Genome Human Research Institute (NHGRI) and National Cancer Institute-funded Clinical Sequencing Exploratory Research (CSER) consortium project that focused on prospective implementation of clinical whole exome sequencing in the pediatric oncology clinic. The primary study objective were to integrate information from CLIA-certified germline and tumor exome sequencing into the care of newly diagnosed solid tumor patients at Texas Children's Cancer Center, and to perform parallel evaluation of the impact of tumor and germline exomes on families and physicians. Blood and frozen tumor (if available) samples were collected from children undergoing surgery or biopsy of newly diagnosed solid tumors and subjected to exome sequencing in a CLIA-certified laboratory. Germline and tumor (if applicable) exome sequencing reports were generated and submitted into the electronic health record and returned to each patient/family by their primary oncologist. In addition to the clinical exome sequencing, specific (optional) consent was requested for research sequencing studies. If this consent was obtained then research studies of some of the children and parents participating in the BASIC3 study (including tumor transcriptome and whole genome sequencing of blood and/or tumor) are performed. The Clinical Sequencing Exploratory Research Consortium Cohort is utilized in the following dbGaP sub-study. To view molecular data, and derived variables collected in this sub-study, please click on the following sub-studies below or in the "Sub-study" section of this top-level study page phs001383 Clinical Sequencing Exploratory Research Consortium Cohort. phs001026 BASIC3 phs001878 GMKF BASIC3

Study phs001683

Resistance to Checkpoint Blockade Therapy Through Inactivation of Antigen Presentation

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

Study phs001427

Systems Analysis of the PfSPZ Vaccine in Kenyan Infants

The PfSPZ vaccine is a candidate vaccine targeting the Plasmodium falciparum parasite, which is the cause of the most lethal type of human malaria. This study used de-identified blood samples collected from 316 infants aged 5 to 12 months (inclusive) enrolled in a Phase 2, randomized, placebo-controlled trial (NCT02687373) of the PfSPZ malaria vaccine, conducted in a region of western Kenya where malaria transmission is high and perennial. The study began in July, 2016, after peak malaria transmission, and was completed in August, 2018. In this study, three doses of the PfSPZ Vaccine (4.5x105 PfSPZ, 9.0x105 PfSPZ, and 1.8x106 PfSPZ) were tested against a normal saline placebo comparator. In all four groups, doses were administered in 3 doses at 0, 8, and 16 weeks by direct venous inoculation. Vaccine efficacy was determined at 3, 6, and 12 months of follow-up after the third immunization. RNA was extracted from whole blood collected in PAXgene tubes before PfSPZ immunization, and two weeks after the third (final) dose, from 262 children for which sufficient material was available at both time points (82% of 316 enrolled children; 525 samples). RNA was depleted of globin and ribosome transcripts before library generation and sequencing on the Illumina NovaSeq platform. RNA-seq expression data was analyzed to determine blood transcriptional signatures, before and after PfSPZ vaccine immunization, that predicts sterile protection from naturally occurring P. falciparum infection or delayed parasitemia at 3 months post-immunization. Gene expression data will also be correlated to P. falciparum-specific IgG antibody and cellular responses to identify possible molecular correlates of vaccine-induced immunogenicity

Study phs002196

National Cancer Institute Clinical and Laboratory Analysis of Familial Cancer

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

Study phs001935

The Human Pancreas Analysis Program (HPAP)

The past decade has seen a dramatic improvement in our ability to phenotype and molecularly profile human tissues with unprecedented resolution at the genomic, epigenomic, protein, and functional levels. The Human Pancreas Analysis Program (HPAP), part of the Human Islet Research Network and supported by the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) through multiple NIH grants, is performing deep phenotyping of the human endocrine pancreas to better understand the cellular and molecular events that precede and lead to beta-cell loss and/or dysfunction in type 1 diabetes (T1D) and type 2 diabetes (T2D) as well as to accumulate, analyze, and distribute high-value data sets to the diabetes research community through the HPAP PANC-DB database (and additional details provided at that site). To this end, HPAP employs state-of-the-art technologies to perform comprehensive analyses of pancreas biology as it pertains to organ donors with T1D, autoantibody-positive donors without diabetes, donors with T2D, and control donors. Pancreas procurement and analyses take advantage of the expertise and extensive network of Organ Procurement Organizations (OPOs) and autoantibody screening centers established through the JDRF/The Lenona M. and Harry B. Helmsley Charitable Trust (HCT) – funded nPOD program. In contrast to nPOD, the major product provided by HPAP is not archived biomaterial subject to broad distribution, but rather, the delivery of extensive and high-quality molecular data sets to the diabetes research community in order to facilitate further discovery. Together, nPOD and HPAP are complementary programs that assist the diabetes community and afford the maximal opportunity for advancing knowledge about the pathogenesis of T1D and T2D.

Study phs002465

Kids First Pediatric Research Study in Familial Predisposition to Hematopoietic Malignancies (SJFAMILY-HM)

The Gabriella Miller Kids First Pediatric Research Program (Kids First) is a trans-NIH effort initiated in response to the 2014 Gabriella Miller Kids First Research Act and supported by the NIH Common Fund. This program focuses on gene discovery in pediatric cancers and structural birth defects and the development of the Gabriella Miller Kids First Pediatric Data Resource (Kids First Data Resource). All of the WGS and phenotypic data from this study are accessible through dbGaP and kidsfirstdrc.org, where other Kids First datasets can also be accessed. Acute lymphoblastic leukemia (ALL) is the commonest childhood tumor and a leading cause of cancer death in children, adolescents and young adults. Hodgkin and non-Hodgkin lymphoma are also important hematologic malignancies (HM) that occur in children. Each are genetic diseases with growing evidence for a germline predisposition of both familial and sporadic cases, however the inherited genetic basis of ALL/lymphoma are poorly understood. Such knowledge is essential to gain mechanistic insight into the basis of tumor formation, and to guide genetic counseling and genetic management. Here we have assembled a large collection of familial HM kindreds, and extended recurrence cohorts of ALL and HL which will be used to identify the genetic basis of familial HM, examine the frequency of germline variants in sporadic ALL and HL, and to integrate inherited and somatic genomic data. These studies have high potential to provide fundamental new insights into the inherited genetic basis of HM, to provide important information to guide clinical management, and to provide an invaluable public resource of genomic data.

Study phs001738

Columbia University Study of Caribbean Hispanics with Familial and Sporadic Late Onset Alzheimer's disease

Compared to Caucasians residing in the same community the incidence rate of Alzheimer's disease is approximately twice as high in Caribbean Hispanics. Moreover, Caribbean Hispanics represent a homogenous population with only a few founders. Replication of genetic associations in other ethnic groups provides supporting evidence that a putative gene is involved in the disease pathogenesis, and when different allelic variants within the same genes are associated with disease it can help to localize the pathogenic variant. SORL1 is an example of a candidate gene that was first identified in Hispanics and then confirmed in multiple ethnic and racial groups in a meta-analysis. For this project, we are genotyping 704 individuals in families multiply affected by Alzheimer's disease (166 families). These families were recruited in the United States, Puerto Rico and the Dominican Republic. In addition, we are genotyping 2491 individuals (960 patients with sporadic Alzheimer's disease and 1531 unrelated controls) phenotyped in a similar fashion totaling 3195 individuals. Both cohorts are followed at regular intervals of 18 to 24 months, and potential phenotypes available other than those related to Alzheimer's disease include body mass index, measured blood pressure, neurological history and examinations. More importantly, both studies are funded through 2014 and additional phenotypes could be added in successive waves. We propose a genome wide association (GWA) study of Alzheimer's disease and longitudinal changes in cognition and other age-related neurological and medical phenotypes. The goal will be to identify the chromosomal locations of genes underlying this disease and its related endophenotypes.

Study phs000496

Genomic Studies in Charcot-Marie-Tooth Disease

The various forms of Charcot-Marie-Tooth disease (CMT) comprise a genetically heterogeneous set of peripheral neuropathies. Currently, >90 different CMT genes have been identified; yet, for the axonal CMT2 subtype these genes explain only 30-40% of the genetic effect. In particular, the application of exome sequencing has led to an unprecedented pace in identifying about half of those genes in the past five years. We have been at the forefront of this development with 22+ published new gene identifications in the past seven years. This success was only possible through broad national and international collaborations, data sharing, and the development of advanced analysis tools and methods. We also learned that despite an accumulation of exome sequencing data, over 50% of axonal CMT patients remain undiagnosed, suggesting even more genes are involved; additionally, our preliminary data also support the existence of non-coding variations as Mendelian allele contributors. In this study, we will continue our highly impactful work in CMT family recruitment, exome sequencing and traditional gene identification. In addition, we will expand the genetic studies to whole genome sequencing with a focus on families already explored unsuccessfully in whole exome studies. We further plan to perform the first large rare variant burden analysis in the rare disorder CMT2. This is only possible through the exceptional clinical resources we have built over the past decade. Finally, with many CMT genes available, we will perform statistical network analyses on a CMT-ome to identify gene modules and pathways that will be the starting point of polypharmacology, and provide insight into pathophysiology of peripheral nerve degeneration. All data will be available in real-time to an existing network of CMT investigators worldwide, and deposited in dbGaP annually.

Study phs003389

A Phase I Study With a Personalized Neoantigen Cancer Vaccine in Glioblastoma Multiforme

Neoantigens, which are derived from tumor-specific protein-coding gene mutations, are exempt from central tolerance, can generate robust immune responses and function as bona fide antigens that facilitate tumor rejection. A strategy of using multi-epitope and personalized neoantigen vaccination has previously been tested in patients with high-risk melanoma. We demonstrated that this strategy is feasible for glioblastoma (GBM) which typically has a relatively low mutation load and an immunologically 'cold' tumor microenvironment. We conducted whole exome sequencing (WES) of tumor and normal cells from individual GBM patients to identify tumor-specific mutations. We assessed the expression of mutated alleles by RNA-sequencing of tumor, and used personalized neoantigen-targeting vaccines to immunize newly diagnosed GBM patients following surgical resection and conventional radiotherapy in a phase I/Ib study. Patients who did not receive dexamethasone, a highly potent corticosteroid that is frequently prescribed to treat cerebral oedema in GBM patients, generated circulating polyfunctional neoantigen-specific CD4+ and CD8+ T cell responses that were enriched in a memory phenotype and showed an increase in the number of tumor-infiltrating T cells. Using single-cell T cell receptor analysis, we provided an evidence that neoantigen-specific T cells from the peripheral blood can migrate into an intracranial GBM tumor. Neoantigen-targeting vaccines thus have the potential to favorably alter the immune environment of GBM.In order to discover cancer antigens derived from annotated and unannotated protein-coding regions of the genome, we carried out matched ribosome profiling (Ribo-Seq) on two GBM samples. We discovered novel or unannotated open reading frames (nuORFs) and their expression levels as well as somatic mutations within the nuORFs, which could be used to predict potential neoantigens.

Study phs001519

Stockholm-Tartu Atherosclerosis Reverse Network Engineering Task (STARNET)

STARNET is a genetics of RNA expression study of multiple disease-relevant tissues obtained from living patients with cardiovascular disease. Tissue samples are obtained from blood, atherosclerotic-lesion-free internal mammary artery (MAM) and atherosclerotic aortic root (AOR), subcutaneous fat (SF), visceral abdominal fat (VAF), skeletal muscle (SKLM), and liver (LIV) during open thorax surgery of 600 coronary artery disease (CAD) patients. All patients gave written informed consent. The inclusion criterion was eligibility for coronary artery by-pass graft (CABG) surgery. Patients with other severe systemic diseases, such as active systemic inflammatory disease or cancer, were excluded. The primary clinical end points were the SYNTAX score based on the extent of coronary atherosclerosis assessed from preoperative angiograms. The STARNET patients are Caucasians (31% females); 32% had diabetes, 75% had hypertension, and 67% had hyperlipidemia; and 33% had an MI before age 60. By New York Heart Association criteria, 45% were class I, 42% class II, 9% class III, and 1% class IV. TYPES AND RNA SEQUENCING: 566 DNA genotype and 3577 RNA-seq profiles from seven tissues from 600 STARNET CABG patients passed quality control (on average 511 RNA-seq profiles/tissue). DNA was genotyped with the OmniExpress Exome array (Illumina, ~900k SNPs) and imputed to a total of 14,098,063 DNA variant calls (6,245,505 with minor allele frequency >5%). The STARNET subjects mainly overlap with Caucasian of Northern European (Finnish) descent. RNA sequencing was performed using the HighSeq2000 platform, poly-A (LIV, SKLM, VAF, SF and blood) and ribo-zero (AOR, MAM) protocols with 50-100 bp read lengths, single end to 15-30 million read depth.

Study phs001203

Genomic Landscape of Multiple Myeloma and of its Precursor Conditions, and its Clinical Implications

Reliable strategies to capture patients at risk of progression from precursor stages of multiple myeloma (MM) to overt disease are still missing. In this study, we generated whole-genome sequencing (WGS) data from matched tumor and normal samples from 98 individuals from the PCROWD study (Precursor Crowd, NCT02269592). These individuals have either a precursor condition for MM, which can be a monoclonal gammopathy of undetermined significance (MGUS) or smoldering multiple myeloma (SMM), or an active multiple myeloma (MM). To do so, we enriched tumor plasma cells from the bone marrow by magnetic bead selection (CD138+), sequenced the DNA of these samples to a target depth of 60X (tumor), and analyzed them against normal white blood cells (peripheral blood mononuclear cells) from the same individuals sequenced to a target depth of 30X (normal). In our first publication [Alberge, Dutta, Poletti, et al., 2025], we integrated and homogeneously analyzed these data together with data from mostly publicly available datasets (phs003084 and phs000748) to reach 1,030 total samples. We used these data to identify recurrent coding and non-coding candidate drivers, as well as significant hotspots of structural variation, and used those drivers to define and validate a simple “MM-like” score which we could use to place patients' tumors on a gradual axis of progression toward active disease. Our MM precursor genomic map provides new insights on the time of initiation and cell of origin of the disease, on the order of acquisition of genomic alterations, and on mutational processes found across stages of the transformation. Taken together, we highlight here the potential of genome sequencing to better inform risk assessment and monitoring of MM precursor conditions.

Study phs003846

Cellular Profiling Identifies Targetable T Cell Phenotypes in Lymphocytic Variant Hypereosinophilic Syndrome

Lymphocytic variant hypereosinophilic syndrome (LHES) is driven by aberrant Th2‑skewed lymphocyte clones that promote tissue eosinophilia. We performed multimodal single‑cell RNA‑sequencing (scRNA‑seq), cell‑surface protein (ADT) profiling, and paired αβ‑TCR sequencing on peripheral‑blood mononuclear cells (PBMCs) from two clinically characterized LHES patients and one idiopathic HES control. One LHES patient was sequenced at two timepoints, pre-JAK inhibitor treatment and post-JAK inhibitor treatment. Single‑cell regulatory‑network inference (SCENIC), CellChat ligand–receptor modelling, and scRNA variant calling were integrated with flow‑cytometric validation and 5,000× amplicon sequencing. Two clonally expanded CD3‑CD4⁺ central‑/effector‑memory T‑cell clusters were observed exclusively in LHES. These cells showed Th2‑associated regulons (GATA3, BATF, MAF), high expression of CCR4, CD52, PTGDR2 and IL13, and down‑regulation of CD247 (CD3ζ), a mechanism experimentally confirmed to abolish surface CD3/TCR. Autocrine IL‑7 signaling, predicted by CellChat, provides an antigen‑independent proliferative cue. Therapeutically actionable targets (CCR4, CD52, IL‑5, IL‑7, TNF) were prioritized; one refractory patient achieved complete clinical and cytologic remission after mogamulizumab (anti‑CCR4) therapy. Longitudinal sequencing revealed clonal evolution with acquisition of a STAT3 p.G618R mutation under JAK‑inhibition, highlighting a resistance mechanism. Raw scRNA‑seq (5′ cDNA, VDJ, ADT) data, and sample‑level metadata will be distributed through dbGaP. These resources provide the first high‑resolution immune atlas of LHES and a framework for rational, precision immunotherapy in eosinophilic disorders.

Study phs004041

Convergent evolution towards CD38 biallelic loss is a recurrent mechanism of resistance to anti-CD38 antibodies in multiple myeloma.

Monoclonal antibodies directed against CD38 are a therapeutic mainstay in multiple myeloma (MM). While they have contributed to improved outcomes, most patients will still experience disease relapse. Despite their ubiquity, little is known about tumor-intrinsic mechanisms of resistance to these drugs. Antigen escape, whereby tumor cells alter expression of cell surface immunotherapy targets, has been proposed as a potential avenue, and has been seen with other immunotherapies in MM. Yet, it is unknown whether MM cells can develop permanent resistance to anti-CD38 antibodies by acquiring genomic events leading to biallelic disruption of the CD38 gene locus. Here, by using whole genome and whole exome sequencing data from 701 newly diagnosed patients, 72 patients at relapse with naivety to anti-CD38 antibodies, and 52 patients collected at relapse following anti-CD38 antibodies, we report biallelic inactivation of CD38 as a recurrent mechanism of resistance in 6% of patients, often via convergent evolution where distinct subclones are advantaged towards the same selective endpoint within the same patient. Monoallelic loss of the CD38 locus was seen in another 14% post-CD38-directed therapy. Three of the cases with biallelic events on CD38 presented with missense mutations and retained CD38 expression. Functional work demonstrated that two of these mutations decreased binding affinity and antibody-dependent cellular cytotoxicity of commercial antibodies Daratumumab and Isatuximab. However, the third mutation (R140G) provided relative resistance only towards one commercial product. Given that patients are often rechallenged with CD38 antibodies following disease progression, these data support the need for next generation sequencing to guide treatment selection.

Study EGAS50000000709

CCR8-targeted specific depletion of clonally expanded Treg cells in tumor tissues evokes potent tumor immunity with long-lasting memory

Foxp3-expressing CD25+CD4+ regulatory T cells (Tregs) are abundantly infiltrating into tumor tissues, hindering effective anti-tumor immunity. Here, assuming that such tumor Tregs were clonally expanding by recognizing tumor-associated antigens, we attempted to characterize them by their gene expression profiling and T-cell receptor (TCR) clonotyping at the single-cell level. We found that they predominantly expressed the chemokine receptor CCR8: ~40% of tumor Tregs, less than 10% of Tregs in other tissues, and few tumor-infiltrating conventional T cells expressed CCR8 in mice and humans. CCR8+ tumor Tregs were phenotypically highly differentiated and functionally stable. One injection of anti-CCR8 mAb with high antibody-dependent cellular cytotoxic activity was indeed able to specifically deplete clonally expanded tumor Tregs and thereby eradicate established tumors in mice. The depletion predominantly expanded tumor-infiltrating CD8+ effector T cells, which were different in activated or exhausted patterns from those seen after immune checkpoint blockade by anti-PD-1 mAb. The treatment also evoked potent secondary immune responses against the same tumor cell line inoculated several months after tumor eradication, indicating that the anti-CCR8-induced memory-type tumor-reactive effector T cells were, once induced, resistant to recovered Treg suppression. In contrast to such evocation of potent tumor immunity, anti-CCR8 treatment scarcely elicited discernable autoimmunity in mice, contrasting with a similar treatment for systemic whole Treg depletion, which induced histologically and serologically evident autoimmunity. Thus, temporary and specific depletion of clonally expanded Tregs in tumor tissues by cell-depleting anti-CCR8 mAb is sufficient to evoke potent tumor immunity with long-lasting memory and without serious autoimmunity.

Study JGAS000312

Influence of age on molecular changes and treatment stratification in multimodal glioblastoma therapy

Background: The increasing life expectancy has led to a rise in high-grade gliomas, particularly in patients of older age. The impact of chronological age, associated molecular changes and contributing factors on therapy response, progression-free survival (PFS) and overall survival (OS) remain uncertain. Methods: Between January 1st, 2016, and December 31th, 2021, patients with glioblastoma IDH-wt (CNS WHO grade 4) were included. RNA- and T-cell receptor sequencing was performed in a subset of patients. Primary outcome parameters were extent of resection (EOR), progression free- and overall survival. Multivariate analysis was performed to identify independent predictors of survival. Results: A total of 286 patients were included. The mean age was 66.4 years. Chronological age did not correlate with overall survival (r=0.12). With age, tumor immune composition changed towards macrophages with M2 phenotype and decreased numbers of CD8+ and neutrophiles. Aging related genes were not enriched. Beyond 69.5 years of age, despite similar PFS (p=0.0783), OS (p<0.0001) was significantly shorter, compared to younger patients. Extent of resection and adjuvant radiochemotherapy were significant predictors of OS (p<0.0001). No difference in PFS and OS was found between younger and older patients treated with maximal safe resection, followed by concomitant RT and chemotherapy and adjuvant chemotherapy. Conclusions: This study suggests that age alone should not be the sole determinant in therapeutic decision-making for glioblastoma patients. Gross total resection and adjuvant therapy, tailored to individual clinical status, significantly impact survival outcomes. Patient-specific treatment strategies are important to minimize undertreatment and optimize therapeutic outcomes in older patients.

Study EGAS00001008246

RNAseq: Acquired non-permissive bone marrow microenvironment impairs hematopoietic stem cell proliferation and maintenance and B-cell development post-HSCT

Defects in B-cell reconstitution upon hematopoietic stem-cell (HSC) transplantation (HSCT) are a common observation, yet the mechanism remains unexplained. The bone marrow (BM) stroma, including mesenchymal stromal cells (MSCs), guides HSC maintenance and B-lymphopoiesis by secreting crucial cytokines. We report acquired, permanent, selective and complete B-cell deficiency in the context of full donor-chimerism in a patient with X-linked lymphoproliferative disease and aimed to identify the contribution of the BM-microenvironment in disrupted B-cell reconstitution post-HSCT. We studied longitudinal BM samples from the patient and his identical twin, both of whom underwent HSCT with the same donor with opposite outcomes in B-cell reconstitution. In the index patient BM, we observed progressive loss of proliferation of HSCs and a selective block at the pre-BI cell stage. In vitro modeling studies showed limited survival of patient-HSCs and a relative accumulation of pre-B cells. Patient-derived MSCs failed to support survival and proliferation of HSCs and B-cell development of healthy-HSCs which was correlated with reduced CXCL12 levels. Using bulk RNA-sequencing of MSCs and in vitro functional studies, we showed global changes in the patients’ MSCs and a progressive loss of CXCL12 expression. Indeed, survival of patients HSCs improved supplementing in vitro development culture with CXCL12, suggesting a contribution of defective CXCL12 signaling to the phenotype. In summary, our data show that an acquired defect in the BM-stromal microenvironment and exhaustion of HSCs and committed progenitors may cause a permanent non-permissive state for normal B-cell development.

Study EGAS50000001437

ATAC-seq: Acquired non-permissive bone marrow microenvironment impairs hematopoietic stem cell proliferation and maintenance and B-cell development post-HSCT

Defects in B-cell reconstitution upon hematopoietic stem-cell (HSC) transplantation (HSCT) are a common observation, yet the mechanism remains unexplained. The bone marrow (BM) stroma, including mesenchymal stromal cells (MSCs), guides HSC maintenance and B-lymphopoiesis by secreting crucial cytokines. We report acquired, permanent, selective and complete B-cell deficiency in the context of full donor-chimerism in a patient with X-linked lymphoproliferative disease and aimed to identify the contribution of the BM-microenvironment in disrupted B-cell reconstitution post-HSCT. We studied longitudinal BM samples from the patient and his identical twin, both of whom underwent HSCT with the same donor with opposite outcomes in B-cell reconstitution. In the index patient BM, we observed progressive loss of proliferation of HSCs and a selective block at the pre-BI cell stage. In vitro modeling studies showed limited survival of patient-HSCs and a relative accumulation of pre-B cells. Patient-derived MSCs failed to support survival and proliferation of HSCs and B-cell development of healthy-HSCs which was correlated with reduced CXCL12 levels. Using bulk RNA-sequencing of MSCs and in vitro functional studies, we showed global changes in the patients’ MSCs and a progressive loss of CXCL12 expression. Indeed, survival of patients HSCs improved supplementing in vitro development culture with CXCL12, suggesting a contribution of defective CXCL12 signaling to the phenotype. In summary, our data show that an acquired defect in the BM-stromal microenvironment and exhaustion of HSCs and committed progenitors may cause a permanent non-permissive state for normal B-cell development.

Study EGAS50000001438

Indonesian Genome Diversity Project

Indonesia, the world’s fourth largest country by population, comprises an archipelago of about 900 permanently inhabited islands in tropical Asia, and hosts an astonishing array of human diversity that remains largely underrepresented in modern biological surveys. The region is unique for its key role in both the early and recent evolution of Homo outside of Africa. It has one of the first traces of anatomically modern humans in Eurasia, and was the focus of extensive interaction between archaic hominins and humans – both in the likely co-existence of H. floresiensis with modern humans, and in the introgression of Denisovans into the ancestors of its present inhabitants. More recently, Indonesia was a center of the spread of Neolithic culture by Austronesian speakers. Advancing maritime technologies allowed farming populations to treat this region as a springboard to reach oceanic islands as remote as Madagascar, Hawaii, Easter Island and New Zealand. Here we present complete human genomes from healthy individuals from multiple key islands in Indonesia and Papua. The genomes were sequenced to high depth (30x). The purpose of this dataset is to address different questions of human evolution in Island Southeast Asia and beyond, including early settlement of the area by anatomically modern humans, population structure, genetic admixture and adaptation, and contacts with archaic hominins. We employed a sampling strategy to capture a broad range of diversity across the region incorporating an average of 12 samples from each of 14 islands spanning Indonesia and Papua, from Sumatra in the west to New Britain in the east. Genomes from this dataset form part of an ongoing project to describe and understand human evolutionary history in Island Southeast Asia and Eurasia in general.

Study EGAS00001003054

Molecular subtypes of pancreatic neuroendocrine tumors mutated in MEN1/DAXX/ ATRX explain biological variability

Pancreatic Neuroendocrine Tumors (PanNETs) most probably arise from the islets of Langerhans, although direct evidence is still lacking. About 40% of non-functioning (NF) PanNETs harbour mutations in MEN1, often in conjunction with alterations in DAXX/ATRX. Despite the homogeneous genetic background, the ADM (mutation in DAXX/ATRX and MEN1) group is heterogeneous regarding molecular features and response to treatment. To explore the molecular and clinical heterogeneity within this group, we integrated transcriptomic and DNA methylation data from 36 and 93 samples, respectively. Using a multi-omic approach we described a two-level hierarchy separating PanNET subtypes mutated in MEN1, DAXX and ATRX. First, DNA methylation discriminates ADM-PanNET from PanNET mutated in MEN1 only (α-like). Compared to α-like, all ADM tumors exhibit DNA methylation changes at enhancer and lowly methylated regions as well as at peri-centromeric and telomeric regions, associated with alternative lengthening of telomeres, increased chromosomal instability, and enhanced proliferative capacity. At a second level, transcriptomic analysis revealed three distinct ADM subtypes: ADM hypoxic, ADM NST (No Special Type), and ADM immunosuppressive. The ADM hypoxic subtype is characterized by strong hypoxia signature, likely regulated epigenetically. The ADM NST subtype appears to be primarily driven by epigenetic changes that promote proliferation. The ADM immunosuppressive is highly enriched in immune component and strong metastasis-like signature. While all the three subtypes are highly associated with liver metastasis formation and invasion, the ADM immunosuppressive are significantly smaller (<2.5 cm p.value=0.023) despite displaying aggressive molecular features. By defining these three novel ADM subtypes, our study provides a refined framework for understanding PanNET heterogeneity. This classification underscores potential diagnostic markers and highlights distinct biological vulnerabilities that may inform the development of subtype-tailored therapeutic strategies.

Study EGAS00001008272

Multi-site tumor sampling highlights molecular intra-tumor heterogeneity in malignant pleural mesothelioma

Background:Malignant pleural mesothelioma (MPM) is a heterogeneous cancer. Better knowledge of molecular and cellular intra-tumor heterogeneity throughout the thoracic cavity is required to develop efficient therapies. This study focuses on molecular intra-tumor heterogeneity using the largest series to date in MPM and is the first to report on the multi-omics profiling of a substantial series of multi-site tumor samples. Methods:Intra-tumor heterogeneity was investigated in 16 patients from whom biopsies were taken at distinct anatomical sites. The paired biopsies collected from apex, side wall, costo-diaphragmatic or highest metabolic sites as well as 5 derived cell lines were screened using targeted sequencing. Whole exome sequencing, RNA sequencing, and DNA methylation were performed on a subset of the cohort for deep characterization. Molecular classification, recently defined histo-molecular gradients, and cell populations of the tumor microenvironment were assessed. Results:Sequencing analysis identified heterogeneous variants notably in NF2, a key tumor suppressor gene of mesothelial carcinogenesis. Subclonal tumor populations were shared among paired biopsies, suggesting a polyclonal dissemination of the tumor. Transcriptome analysis highlighted dysregulation of cell adhesion and extracellular matrix pathways, linked to changes in histo-molecular gradient proportions between anatomic sites. Methylome analysis revealed contribution of epigenetic mechanisms in two patients. Finally, significant changes in the expression of immune mediators and genes related to immunological synapse, as well as differential infiltration of immune populations in the tumor environment were observed and led to a switch from a hot to a cold immune profile in three patients. Conclusions:This comprehensive analysis reveals patient-dependent spatial intra-tumor heterogeneity at the genetic, transcriptomic and epigenetic levels and in the immune landscape of the tumor microenvironment. Results support

Study EGAS00001005328

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