European Genome-phenome Archive Research Articles

Successful Retreatment With Venetoclax in a Patient With Chronic Lymphocytic Leukemia.

Successful Retreatment With Venetoclax in a Patient With Chronic Lymphocytic Leukemia.

Claudin 18.2 is a potential therapeutic target for zolbetuximab in pancreatic ductal adenocarcinoma.

BACKGROUNDPancreatic ductal adenocarcinoma (PDAC) is frequently diagnosed and treated in advanced tumor stages with poor prognosis. More effective screening programs and novel therapeutic means are urgently needed. Recent studies have regarded tight junction protein claudin 18.2 (CLDN18.2) as a candidate target for cancer treatment, and zolbetuximab (formerly known as IMAB362) has been developed against CLDN18.2. However, there are few data reported thus far related to the clinicopathological characteristics of CLDN18.2 expression for PDAC.AIMTo investigate the expression of CLDN18.2 in PDAC patients and subsequently propose a new target for the treatment of PDAC.METHODSThe Cancer Genome Atlas, Genotype-Tissue Expression, Gene Expression Omnibus, and European Genome-phenome Archive databases were first employed to analyze the CLDN18 gene expression in normal pancreatic tissue compared to that in pancreatic cancer tissue. Second, we analyzed the expression of CLDN18.2 in 93 primary PDACs, 86 para-cancer tissues, and 13 normal pancreatic tissues by immunohistochemistry. Immunostained tissues were assessed applying the histoscore. subsequently, they fell into two groups according to the expression state of CLDN18.2. Furthermore, the correlations between CLDN18.2 expression and diverse clinicopathological characteristics, including survival, were investigated.RESULTSThe gene expression of CLDN18 was statistically higher (P < 0.01) in pancreatic tumors than in normal tissues. However, there was no significant correlation between CLDN18 expression and survival in pancreatic cancer patients. CLDN18.2 was expressed in 88 (94.6%) of the reported PDACs. Among these tumors, 50 (56.8%) cases showed strong immunostaining. The para-cancer tissues were positive in 81 (94.2%) cases, among which 32 (39.5%) of cases were characterized for strong staining intensities. Normal pancreatic tissue was identified solely via weak immunostaining. Finally, CLDN18.2 expression significantly correlated with lymph node metastasis, distant metastasis, nerve invasion, stage, and survival of PDAC patients, while there was no correlation between CLDN18.2 expression and localization, tumor size, patient age and sex, nor any other clinicopathological characteristic. CONCLUSIONCLDN18.2 expression is frequently increased in PDAC patients. Thus, it may act as a potential therapeutic target for zolbetuximab in PDAC.

A quality control portal for sequencing data deposited at the European genome-phenome archive.

Since its launch in 2008, the European Genome–Phenome Archive (EGA) has been leading the archiving and distribution of human identifiable genomic data. In this regard, one of the community concerns is the potential usability of the stored data, as of now, data submitters are not mandated to perform any quality control (QC) before uploading their data and associated metadata information. Here, we present a new File QC Portal developed at EGA, along with QC reports performed and created for 1 694 442 files [Fastq, sequence alignment map (SAM)/binary alignment map (BAM)/CRAM and variant call format (VCF)] submitted at EGA. QC reports allow anonymous EGA users to view summary-level information regarding the files within a specific dataset, such as quality of reads, alignment quality, number and type of variants and other features. Researchers benefit from being able to assess the quality of data prior to the data access decision and thereby, increasing the reusability of data (https://ega-archive.org/blog/data-upcycling-powered-by-ega/).

SmMIP-tools: a computational toolset for processing and analysis of single-molecule molecular inversion probes-derived data.

MotivationSingle-molecule molecular inversion probes (smMIPs) provide an exceptionally cost-effective and modular approach for routine or large-cohort next-generation sequencing. However, processing the derived raw data to generate highly accurate variants calls remains challenging.ResultsWe introduce SmMIP-tools, a comprehensive computational method that promotes the detection of single nucleotide variants and short insertions and deletions from smMIP-based sequencing. Our approach delivered near-perfect performance when benchmarked against a set of known mutations in controlled experiments involving DNA dilutions and outperformed other commonly used computational methods for mutation detection. Comparison against clinically approved diagnostic testing of leukaemia patients demonstrated the ability to detect both previously reported variants and a set of pathogenic mutations that did not pass detection by clinical testing. Collectively, our results indicate that increased performance can be achieved when tailoring data processing and analysis to its related technology. The feasibility of using our method in research and clinical settings to benefit from low-cost smMIP technology is demonstrated.Availability and implementationThe source code for SmMIP-tools, its manual and additional scripts aimed to foster large-scale data processing and analysis are all available on github (https://github.com/abelson-lab/smMIP-tools). Raw sequencing data generated in this study have been submitted to the European Genome-Phenome Archive (EGA; https://ega-archive.org) and can be accessed under accession number EGAS00001005359.Supplementary information Supplementary data are available at Bioinformatics online.

The European Genome-phenome Archive in 2021.

The European Genome-phenome Archive (EGA - https://ega-archive.org/) is a resource for long term secure archiving of all types of potentially identifiable genetic, phenotypic, and clinical data resulting from biomedical research projects. Its mission is to foster hosted data reuse, enable reproducibility, and accelerate biomedical and translational research in line with the FAIR principles. Launched in 2008, the EGA has grown quickly, currently archiving over 4,500 studies from nearly one thousand institutions. The EGA operates a distributed data access model in which requests are made to the data controller, not to the EGA, therefore, the submitter keeps control on who has access to the data and under which conditions. Given the size and value of data hosted, the EGA is constantly improving its value chain, that is, how the EGA can contribute to enhancing the value of human health data by facilitating its submission, discovery, access, and distribution, as well as leading the design and implementation of standards and methods necessary to deliver the value chain. The EGA has become a key GA4GH Driver Project, leading multiple development efforts and implementing new standards and tools, and has been appointed as an ELIXIR Core Data Resource.

A Systematic Review of Transcriptional Dysregulation in Huntington's Disease Studied by RNA Sequencing.

Huntington’s disease (HD) is a chronic neurodegenerative disorder caused by an expansion of polyglutamine repeats in exon 1 of the Huntingtin gene. Transcriptional dysregulation accompanied by epigenetic alterations is an early and central disease mechanism in HD yet, the exact mechanisms and regulators, and their associated gene expression programs remain incompletely understood. This systematic review investigates genome-wide transcriptional studies that were conducted using RNA sequencing (RNA-seq) technology in HD patients and models. The review protocol was registered at the Open Science Framework (OSF). The biomedical literature and gene expression databases, PubMed and NCBI BioProject, Array Express, European Nucleotide Archive (ENA), European Genome-Phenome Archive (EGA), respectively, were searched using the defined terms specified in the protocol following the PRISMA guidelines. We conducted a complete literature and database search to retrieve all RNA-seq-based gene expression studies in HD published until August 2020, retrieving 288 articles and 237 datasets from PubMed and the databases, respectively. A total of 27 studies meeting the eligibility criteria were included in this review. Collectively, comparative analysis of the datasets revealed frequent genes that are consistently dysregulated in HD. In postmortem brains from HD patients, DNAJB1, HSPA1B and HSPB1 genes were commonly upregulated across all brain regions and cell types except for medium spiny neurons (MSNs) at symptomatic disease stage, and HSPH1 and SAT1 genes were altered in expression in all symptomatic brain datasets, indicating early and sustained changes in the expression of genes related to heat shock response as well as response to misfolded proteins. Specifically in indirect pathway medium spiny neurons (iMSNs), mitochondria related genes were among the top uniquely dysregulated genes. Interestingly, blood from HD patients showed commonly differentially expressed genes with a number of brain regions and cells, with the highest number of overlapping genes with MSNs and BA9 region at symptomatic stage. We also found the differential expression and predicted altered activity of a set of transcription factors and epigenetic regulators, including BCL6, EGR1, FOSL2 and CREBBP, HDAC1, KDM4C, respectively, which may underlie the observed transcriptional changes in HD. Altogether, our work provides a complete overview of the transcriptional studies in HD, and by data synthesis, reveals a number of common and unique gene expression and regulatory changes across different cell and tissue types in HD. These changes could elucidate new insights into molecular mechanisms of differential vulnerability in HD. Systematic Review Registration: https://osf.io/pm3wq

Physical activity attenuates postprandial hyperglycaemia in homozygous TBC1D4 loss-of-function mutation carriers

Aims/hypothesisThe common muscle-specific TBC1D4 p.Arg684Ter loss-of-function variant defines a subtype of non-autoimmune diabetes in Arctic populations. Homozygous carriers are characterised by elevated postprandial glucose and insulin levels. Because 3.8% of the Greenlandic population are homozygous carriers, it is important to explore possibilities for precision medicine. We aimed to investigate whether physical activity attenuates the effect of this variant on 2 h plasma glucose levels after an oral glucose load.MethodsIn a Greenlandic population cohort (n = 2655), 2 h plasma glucose levels were obtained after an OGTT, physical activity was estimated as physical activity energy expenditure and TBC1D4 genotype was determined. We performed TBC1D4–physical activity interaction analysis, applying a linear mixed model to correct for genetic admixture and relatedness.ResultsPhysical activity was inversely associated with 2 h plasma glucose levels (β[main effect of physical activity] −0.0033 [mmol/l] / [kJ kg−1 day−1], p = 6.5 × 10−5), and significantly more so among homozygous carriers of the TBC1D4 risk variant compared with heterozygous carriers and non-carriers (β[interaction] −0.015 [mmol/l] / [kJ kg−1 day−1], p = 0.0085). The estimated effect size suggests that 1 h of vigorous physical activity per day (compared with resting) reduces 2 h plasma glucose levels by an additional ~0.7 mmol/l in homozygous carriers of the risk variant.Conclusions/interpretationPhysical activity improves glucose homeostasis particularly in homozygous TBC1D4 risk variant carriers via a skeletal muscle TBC1 domain family member 4-independent pathway. This provides a rationale to implement physical activity as lifestyle precision medicine in Arctic populations.Data repositoryThe Greenlandic Cardio-Metabochip data for the Inuit Health in Transition study has been deposited at the European Genome-phenome Archive (https://www.ebi.ac.uk/ega/dacs/EGAC00001000736) under accession EGAD00010001428.Graphical abstract

Identification of a 15-pseudogene based prognostic signature for predicting survival and antitumor immune response in breast cancer.

Pseudogenes are noncoding RNAs that have been revealed to play critical roles in oncogenesis and tumor progression. However, their functional roles have not been comprehensively clarified in breast cancer. Here, we systematically analyzed the RNA sequencing data of 13931 pseudogenes in 775 breast cancer patients from The Cancer Genome Atlas dataset, and ultimately identified 15 prognostic pseudogenes by univariate Cox proportional hazard regression. A risk score model was constructed based on the prognostic pseudogenes via LASSO analysis and dichotomized patients into low- and high-risk subgroups. Patients in the high-risk group had a significantly shorter overall survival than those in the low-risk group. The prognostic value of these 15 pseudogenes and the risk score model were further validated in the European Genome-Phenome Archive dataset. Furthermore, we performed consensus clustering of the 15 prognostic pseudogenes and found that their expression pattern was significantly associated with tumor malignancy and host antitumor immune response, in terms of infiltrating immune cell compositions, antigen presenting genes expression, cytolytic activity and T-cell exhausted markers. This study indicated that these 15 prognostic pseudogenes were significantly correlated with tumor malignancy and host antitumor immune response in breast cancer, and might serve as potential targets for immunotherapy.

A molecular map of lung neuroendocrine neoplasms.

BackgroundLung neuroendocrine neoplasms (LNENs) are rare solid cancers, with most genomic studies including a limited number of samples. Recently, generating the first multi-omic dataset for atypical pulmonary carcinoids and the first methylation dataset for large-cell neuroendocrine carcinomas led us to the discovery of clinically relevant molecular groups, as well as a new entity of pulmonary carcinoids (supra-carcinoids).ResultsTo promote the integration of LNENs molecular data, we provide here detailed information on data generation and quality control for whole-genome/exome sequencing, RNA sequencing, and EPIC 850K methylation arrays for a total of 84 patients with LNENs. We integrate the transcriptomic data with other previously published data and generate the first comprehensive molecular map of LNENs using the Uniform Manifold Approximation and Projection (UMAP) dimension reduction technique. We show that this map captures the main biological findings of previous studies and can be used as reference to integrate datasets for which RNA sequencing is available. The generated map can be interactively explored and interrogated on the UCSC TumorMap portal (https://tumormap.ucsc.edu/?p=RCG_lungNENomics/LNEN). The data, source code, and compute environments used to generate and evaluate the map as well as the raw data are available, respectively, in a Nextjournal interactive notebook (https://nextjournal.com/rarecancersgenomics/a-molecular-map-of-lung-neuroendocrine-neoplasms/) and at the EMBL-EBI European Genome-phenome Archive and Gene Expression Omnibus data repositories.ConclusionsWe provide data and all resources needed to integrate them with future LNENs transcriptomic studies, allowing meaningful conclusions to be drawn that will eventually lead to a better understanding of this rare understudied disease.

Abstract 225: Frequent abnormalities in TP53 and increased genetic instability in myxofibrosarcoma

Abstract Myxofibrosarcoma (MFS) is a rare subtype of soft tissue sarcomas (STSs) preferentially affecting the elderly. Histologically, MFS is distinct from other STSs, in that it is characterized by the proliferation of pleomorphic spindle cells with varying degrees of myxoid components. However, the molecular pathogenesis of MSF is poorly understood. We conducted an integrated molecular study involving 70 samples from primary MFS patients, in which samples were analyzed by whole-genome sequencing (WGS, n=5), whole-exome sequencing (WES, n=44), targeted-capture sequencing (TS, n=65), RNA sequencing (n=3), and immunohistochemistry (IHC, n=50). Copy number (CN) alterations were detected by sequencing-based CN analysis. We combined our STS data with those from from the Cancer Genome Atlas and European Genome-phenome Archive cohort, including 116 MSF samples, and compared to the data from other STS samples (n=189). To further investigate the genetic basis of mixed histological components and chronological changes in MFS, we performed WES of multi-regional and/or multi-time-point samples from 8 MSF cases. A total of 9,246 mutations were identified by WES in 130 primary MFS samples with a median of 55.5 mutations/sample. Mutations were dominated by age-related C to T transitions at CpG sites. When combining the results from TS, among 188 primary MFS samples, most frequently mutated and/or CN-altered genes included TP53 (n=82, 43.6%), RB1 (n=31, 16.5%), ATRX (n=29, 15.4%), and CDKN2A (n=19, 10.2%). A fusion gene involving TRIO was newly identified by RNA sequencing (n=1). A similar mutational profile was observed in undifferentiated pleomorphic sarcoma (UPS), in which TP53 (59.1%), ATRX (34.0%), RB1 (22.7%), and CDKN2A (20.5%) were the major mutational targets, suggesting the common molecular pathogenesis between these two subtypes. Combined with frequent positive staining in IHC (n=22, 44.0%), TP53 was affected in 55.3% of the MFS cases (n=104). Five MFS cases evaluated by WGS showed complex structural abnormalities suggestive of increased genetic instability, where a median of 179 structural variations were detected per sample. WES with multi-regional sampling (n=5) disclosed a high level of intratumor heterogeneity, in which less than 29.0% of mutations were shared by different samples taken from the same tumor. An analysis of longitudinal samples (n=6) revealed significantly higher numbers of mutations in relapse samples (1.6 times on average, p = 0.03). In all cases, TP53 lesions were present from at the time of the initial diagnosis, while most of other lesions were subclonal and acquired during the clinical course. In summary, the genetic profile of MFS is characterized by frequent abnormalities in TP53, RB1, CDKN2A and ATRX, and closely related to other STSs, particularly to UPS. Clonal TP53 abnormalities resulted in complex chromosomal structure and a high degree of intratumor heterogeneity. Citation Format: Yasuhide Takeuchi, Annegret Kunitz, Adriane Halik, Hiromichi Suzuki, Kenichi Yoshida, Yuichi Shiraishi, Kenichi Chiba, Hiroko Tanaka, Nobuyuki Kakiuchi, Yusuke Shiozawa, Akira Yokoyama, Yoshikage Inoue, Tetsuichi Yoshizato, Kosuke Aoki, Yoichi Fujii, Yasuhito Nannya, Hideki Makishima, Satoru Miyano, Hironori Haga, Frederik Damm, Seishi Ogawa. Frequent abnormalities in TP53 and increased genetic instability in myxofibrosarcoma [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 225.

Molecular and Functional Characterization of the Somatic PIWIL1/piRNA Pathway in Colorectal Cancer Cells.

PIWI-like (PIWIL) proteins and small non-coding piRNAs, involved in genome regulation in germline cells, are found aberrantly expressed in human tumors. Gene expression data from The Cancer Genome Atlas (TCGA), the Genotype-Tissue Expression (GTEx) project, and the European Genome-Phenome Archive (EGA) indicate that the PIWIL1 gene is ectopically activated in a significant fraction of colorectal cancers (CRCs), where this is accompanied by promoter demethylation, together with germline factors required for piRNA production. Starting from this observation, the PIWIL/piRNA pathway was studied in detail in COLO 205 CRC cells, which express significant levels of this protein, to investigate role and significance of ectopic PIWIL1 expression in human tumors. RNA sequencing and cell and computational biology led to the demonstration that PIWIL1 localizes in a nuage-like structure located in the perinuclear region of the cell and that a significant fraction of the piRNAs expressed in these cells are methylated, and, therefore, present in an active form. This was further supported by RNA immunoprecipitation, which revealed how several piRNAs can be found loaded into PIWIL1 to form complexes also comprising their target mRNAs. The mature transcripts associated with the PIWIL–piRNA complex encode key regulatory proteins involved in the molecular mechanisms sustaining colorectal carcinogenesis, suggesting that the PIWI/piRNA pathway may actively contribute to the establishment and/or maintenance of clinico-pathological features of CRCs.

Building Infrastructure for African Human Genomic Data Management

Human genomic data are large and complex, and require adequate infrastructure for secure storage and transfer. The NIH and The Wellcome Trust have funded multiple projects on genomic research, including the Human Heredity and Health in Africa (H3Africa) initiative, and data are required to be deposited into the public domain. The European Genome-phenome Archive (EGA) is a repository for sequence and genotype data where the data access is controlled by access committees. Access is determined by a formal application procedure for the purpose of secure storage and distribution, and must be in line with the informed consent of the study participants. H3Africa researchers based in Africa and generating their own data can benefit tremendously from the data sharing capabilities of the internet by using the appropriate technologies. The H3Africa Data Archive is an effort between the H3Africa data generating projects, H3ABioNet and the EGA to store and submit genomic data to public repositories. H3ABioNet maintains the security of the H3Africa Data Archive, ensures ethical security compliance, supports users with data submission and facilitates the data transfer. The goal is to ensure efficient data flow between researchers, the archive and the EGA or other public repositories. To comply with the H3Africa data sharing and release policy, nine months after the data is in secure storage, H3ABioNet converts the data into an XML format ready for submission to EGA. This article describes the infrastructure that has been developed for African human genomic data management.

Neoepiscope improves neoepitope prediction with multivariant phasing.

The vast majority of tools for neoepitope prediction from DNA sequencing of complementary tumor and normal patient samples do not consider germline context or the potential for the co-occurrence of two or more somatic variants on the same mRNA transcript. Without consideration of these phenomena, existing approaches are likely to produce both false-positive and false-negative results, resulting in an inaccurate and incomplete picture of the cancer neoepitope landscape. We developed neoepiscope chiefly to address this issue for single nucleotide variants (SNVs) and insertions/deletions (indels). Herein, we illustrate how germline and somatic variant phasing affects neoepitope prediction across multiple datasets. We estimate that up to ∼5% of neoepitopes arising from SNVs and indels may require variant phasing for their accurate assessment. neoepiscope is performant, flexible and supports several major histocompatibility complex binding affinity prediction tools. neoepiscope is available on GitHub at https://github.com/pdxgx/neoepiscope under the MIT license. Scripts for reproducing results described in the text are available at https://github.com/pdxgx/neoepiscope-paper under the MIT license. Additional data from this study, including summaries of variant phasing incidence and benchmarking wallclock times, are available in Supplementary Files 1, 2 and 3. Supplementary File 1 contains Supplementary Table 1, Supplementary Figures 1 and 2, and descriptions of Supplementary Tables 2-8. Supplementary File 2 contains Supplementary Tables 2-6 and 8. Supplementary File 3 contains Supplementary Table 7. Raw sequencing data used for the analyses in this manuscript are available from the Sequence Read Archive under accessions PRJNA278450, PRJNA312948, PRJNA307199, PRJNA343789, PRJNA357321, PRJNA293912, PRJNA369259, PRJNA305077, PRJNA306070, PRJNA82745 and PRJNA324705; from the European Genome-phenome Archive under accessions EGAD00001004352 and EGAD00001002731; and by direct request to the authors. Supplementary data are available at Bioinformatics online.

Abstract 3173: Clinical significance of Fanconi anemia complementation group E(FANCE)DNA repair-related gene expression in hepatocellular carcinoma

Abstract Background : Hepatocellular carcinoma (HCC) is one of the most threatening malignancies because of the limited availability of radical therapeutic options. Thus, identification of prognostic biomarkers as well as molecular therapeutic targets of HCC should be very important for HCC patients. Fanconi anemia complementation group E (FANCE) is a DNA repair-related gene and it’s deletion is one of causes of Fanconi anemia. Recent studies reported that FANCD2 which is activated by FA complex involving FANCE shows high expression in HCC and expression of FANCD2 is in direct proportion to the grade of malignancy of HCC (Anticancer Res, 2017). And other studies reported that FANCE shows high expression in breast cancer (J Mol Biol, 2015). However clinical significance of FANCE expression in HCC is unknown. Objective : To clarify the clinical significance of FANCE expression in HCC. Material and method: Firstly, we assessed the relation between mRNA expression of FANCE and prognosis using large scale HCC gene data sets (The Cancer Genome Atlas; TCGA, Gene Expression Omnibus; GEO, and European Genome-phenome Archive; EGA). Secondly, the mRNA expression of FANCE was measured in 72 surgically resected HCC in our hospital during the period from 2000 to 2004 by RT-qPCR (normalized by internal control GAPDH), and we compared the expression of FANCE in between tumors and normal tissues. Thirdly, we assessed the associations between expression of FANCE and clinicopathological factors. Finally, we investigated the localization of FANCE by immunohistochemical staining data of THE HUMAN PROTEIN ATLAS. Result: In large scale HCC gene data sets and our samples, tumor tissues have higher mRNA expression of FANCE than normal tissues(t test. p&amp;lt;0.001). The high expression of FANCE was significantly associated with poor prognosis (Kaplan-Meier method, Log rank test. p&amp;lt;0.05). In the clinicopathological analysis, FANCE expression was not associated with any clinicopathological factors except age (p&amp;lt;0.05). THE HUMAN PROTEIN ATLAS showed that FANCE is expressed in the nuclei of HCC cells. Discussion: FANCE was overexpressed in HCC cells, and the high expression of FANCE was associated with poor prognosis. So high expression of FANCE could be a prognostic biomarker in HCC. Now we are performing knockdown experiment for FANCE to clarify the biological significance of FANCE expression in HCC.Conclusion: FANCE could be a prognostic biomarker in HCC. Citation Format: Junichi Takahashi, Takaaki Masuda, Yosuke Kuroda, Akihiro Kitagawa, Yushi Motomura, Kensuke Koike, Dai Shimizu, Shotaro Kuramitsu, Atsushi Fujii, Miwa Noda, Kuniaki Sato, Yusuke Tsuruda, Hajime Otsu, Hidetoshi Eguchi, Keishi Sugimachi, Masaki Mori, Koshi Mimori. Clinical significance of Fanconi anemia complementation group E(FANCE)DNA repair-related gene expression in hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3173.

Cellular circadian period length inversely correlates with HbA1c levels in individuals with type 2 diabetes.

The circadian system plays an essential role in regulating the timing of human metabolism. Indeed, circadian misalignment is strongly associated with high rates of metabolic disorders. The properties of the circadian oscillator can be measured in cells cultured in vitro and these cellular rhythms are highly informative of the physiological circadian rhythm in vivo. We aimed to discover whether molecular properties of the circadian oscillator are altered as a result of type 2 diabetes. We assessed molecular clock properties in dermal fibroblasts established from skin biopsies taken from nine obese and eight non-obese individuals with type 2 diabetes and 11 non-diabetic control individuals. Following in vitro synchronisation, primary fibroblast cultures were subjected to continuous assessment of circadian bioluminescence profiles based on lentiviral luciferase reporters. We observed a significant inverse correlation (ρ = -0.592; p < 0.05) between HbA1c values and circadian period length within cells from the type 2 diabetes group. RNA sequencing analysis conducted on samples from this group revealed that ICAM1, encoding the endothelial adhesion protein, was differentially expressed in fibroblasts from individuals with poorly controlled vs well-controlled type 2 diabetes and its levels correlated with cellular period length. Consistent with this circadian link, the ICAM1 gene also displayed rhythmic binding of the circadian locomotor output cycles kaput (CLOCK) protein that correlated with gene expression. We provide for the first time a potential molecular link between glycaemic control in individuals with type 2 diabetes and circadian clock machinery. This paves the way for further mechanistic understanding of circadian oscillator changes upon type 2 diabetes development in humans. RNA sequencing data and clinical phenotypic data have been deposited at the European Genome-phenome Archive (EGA), which is hosted by the European Bioinformatics Institute (EBI) and the Centre for Genomic Regulation (CRG), ega-box-1210, under accession no. EGAS00001003622.

Circulating Tumor DNA Abundance and Potential Utility in De Novo Metastatic Prostate Cancer

BackgroundSeveral systemic therapeutic options exist for metastatic castrate-sensitive prostate cancer (mCSPC). Circulating tumor DNA (ctDNA) can molecularly profile metastatic castration-resistant prostate cancer and can influence decision-making, but remains untested in mCSPC. ObjectiveTo determine ctDNA abundance at de novo mCSPC diagnosis and whether ctDNA provides complementary clinically relevant information to a prostate biopsy. Design, setting, and participantsWe collected plasma cell-free DNA (cfDNA) from 53 patients newly diagnosed with mCSPC and, where possible, during treatment. Targeted sequencing was performed on cfDNA and DNA from diagnostic prostate tissue. Results and limitationsThe median ctDNA fraction was 11% (range 0–84%) among untreated patients but was lower (1.0%, range 0-51%) among patients after short-term (median 22d) androgen deprivation therapy (ADT). TP53 mutations and DNA repair defects were identified in 47% and 21% of the cohort, respectively. The concordance for mutation detection in matched samples was 80%. Combined ctDNA and tissue analysis identified potential driver alterations in 94% of patients, whereas ctDNA or prostate biopsy alone was insufficient in 19 cases (36%). Limitations include the use of a narrow gene panel and undersampling of primary disease by prostate biopsy. ConclusionsctDNA provides additional information to a prostate biopsy in men with de novo mCSPC, but ADT rapidly reduces ctDNA availability. Primary tissue and ctDNA share relevant somatic alterations, suggesting that either is suitable for molecular subtyping in de novo mCSPC. The optimal approach for biomarker development should utilize both a tissue and liquid biopsy at diagnosis, as neither captures clinically relevant somatic alterations in all patients. Patient summaryIn men with advanced prostate cancer, tumor DNA shed into the bloodstream can be measured via a blood test. The information from this test provides complementary information to a prostate needle biopsy and could be used to guide management strategies.Sequencing data were deposited in the European Genome-phenome Archive (EGA) under study identifier EGAS00001003351.

The 1000IBD project: multi-omics data of 1000 inflammatory bowel disease patients; data release 1

BackgroundInflammatory bowel disease (IBD) is a chronic complex disease of the gastrointestinal tract. Patients with IBD can experience a wide range of symptoms, but the pathophysiological mechanisms that cause these individual differences in clinical presentation remain largely unknown. In consequence, IBD is currently classified into subtypes using clinical characteristics. If we are to develop a more targeted treatment approach, molecular subtypes of IBD need to be discovered that can be used as new drug targets. To achieve this, we need multiple layers of molecular data generated from the same IBD patients.Construction and contentWe initiated the 1000IBD project (https://1000ibd.org) to prospectively follow more than 1000 IBD patients from the Northern provinces of the Netherlands. For these patients, we have collected a uniquely large number of phenotypes and generated multi-omics profiles. To date, 1215 participants have been enrolled in the project and enrolment is on-going. Phenotype data collected for these participants includes information on dietary and environmental factors, drug responses and adverse drug events. Genome information has been generated using genotyping (ImmunoChip, Global Screening Array and HumanExomeChip) and sequencing (whole exome sequencing and targeted resequencing of IBD susceptibility loci), transcriptome information generated using RNA-sequencing of intestinal biopsies and microbiome information generated using both sequencing of the 16S rRNA gene and whole genome shotgun metagenomic sequencing.Utility and discussionAll molecular data generated within the 1000IBD project will be shared on the European Genome-Phenome Archive (https://ega-archive.org, accession no: EGAS00001002702). The first data release, detailed in this announcement and released simultaneously with this publication, will contain basic phenotypes for 1215 participants, genotypes of 314 participants and gut microbiome data from stool samples (315 participants) and biopsies (107 participants) generated by tag sequencing the 16S gene. Future releases will comprise many more additional phenotypes and -omics data layers. 1000IBD data can be used by other researchers as a replication cohort, a dataset to test new software tools, or a dataset for applying new statistical models.ConclusionsWe report on the establishment and future development of the 1000IBD project: the first comprehensive multi-omics dataset aimed at discovering IBD biomarker profiles and treatment targets.

Very low-depth whole-genome sequencing in complex trait association studies.

MotivationVery low-depth sequencing has been proposed as a cost-effective approach to capture low-frequency and rare variation in complex trait association studies. However, a full characterization of the genotype quality and association power for very low-depth sequencing designs is still lacking.ResultsWe perform cohort-wide whole-genome sequencing (WGS) at low depth in 1239 individuals (990 at 1× depth and 249 at 4× depth) from an isolated population, and establish a robust pipeline for calling and imputing very low-depth WGS genotypes from standard bioinformatics tools. Using genotyping chip, whole-exome sequencing (75× depth) and high-depth (22×) WGS data in the same samples, we examine in detail the sensitivity of this approach, and show that imputed 1× WGS recapitulates 95.2% of variants found by imputed GWAS with an average minor allele concordance of 97% for common and low-frequency variants. In our study, 1× further allowed the discovery of 140 844 true low-frequency variants with 73% genotype concordance when compared to high-depth WGS data. Finally, using association results for 57 quantitative traits, we show that very low-depth WGS is an efficient alternative to imputed GWAS chip designs, allowing the discovery of up to twice as many true association signals than the classical imputed GWAS design.Availability and implementationThe HELIC genotype and WGS datasets have been deposited to the European Genome-phenome Archive (https://www.ebi.ac.uk/ega/home): EGAD00010000518; EGAD00010000522; EGAD00010000610; EGAD00001001636, EGAD00001001637. The peakplotter software is available at https://github.com/wtsi-team144/peakplotter, the transformPhenotype app can be downloaded at https://github.com/wtsi-team144/transformPhenotype.Supplementary information Supplementary data are available at Bioinformatics online.

The ICR142 NGS validation series: a resource for orthogonal assessment of NGS analysis.

To provide a useful community resource for orthogonal assessment of NGS analysis software, we present the ICR142 NGS validation series. The dataset includes high-quality exome sequence data from 142 samples together with Sanger sequence data at 704 sites; 416 sites with variants and 288 sites at which variants were called by an NGS analysis tool, but no variant is present in the corresponding Sanger sequence. The dataset includes 293 indel variants and 247 negative indel sites, and thus the ICR142 validation dataset is of particular utility in evaluating indel calling performance. The FASTQ files and Sanger sequence results can be accessed in the European Genome-phenome Archive under the accession number EGAS00001001332.

Identification of novel high-impact recessively inherited type 2 diabetes risk variants in the Greenlandic population

Aims/hypothesisIn a recent study using a standard additive genetic model, we identified a TBC1D4 loss-of-function variant with a large recessive impact on risk of type 2 diabetes in Greenlanders. The aim of the current study was to identify additional genetic variation underlying type 2 diabetes using a recessive genetic model, thereby increasing the power to detect variants with recessive effects.MethodsWe investigated three cohorts of Greenlanders (B99, n = 1401; IHIT, n = 3115; and BBH, n = 547), which were genotyped using Illumina MetaboChip. Of the 4674 genotyped individuals passing quality control, 4648 had phenotype data available, and type 2 diabetes association analyses were performed for 317 individuals with type 2 diabetes and 2631 participants with normal glucose tolerance. Statistical association analyses were performed using a linear mixed model.ResultsUsing a recessive genetic model, we identified two novel loci associated with type 2 diabetes in Greenlanders, namely rs870992 in ITGA1 on chromosome 5 (OR 2.79, p = 1.8 × 10−8), and rs16993330 upstream of LARGE1 on chromosome 22 (OR 3.52, p = 1.3 × 10−7). The LARGE1 variant did not reach the conventional threshold for genome-wide significance (p < 5 × 10−8) but did withstand a study-wide Bonferroni-corrected significance threshold. Both variants were common in Greenlanders, with minor allele frequencies of 23% and 16%, respectively, and were estimated to have large recessive effects on risk of type 2 diabetes in Greenlanders, compared with additively inherited variants previously observed in European populations.Conclusions/interpretationWe demonstrate the value of using a recessive genetic model in a historically small and isolated population to identify genetic risk variants. Our findings give new insights into the genetic architecture of type 2 diabetes, and further support the existence of high-effect genetic risk factors of potential clinical relevance, particularly in isolated populations.Data availabilityThe Greenlandic MetaboChip-genotype data are available at European Genome-Phenome Archive (EGA; https://ega-archive.org/) under the accession EGAS00001002641.