11917 results for "division 2 coins fc 26 Visit Buyfc26coins.com Kupiłem(a) duży pakiet monet FC 26 i dostałem(a) dodatkowy rabat. Świetnie!.fXzt"

in 66.11 milliseconds.

• Total RNAseq data from 8 patients with muscle invasive bladder cancer

  For this dataset we performed total RNAseq on blood samples from 8 patients with muscle invasive bladder cancer. The data consists of 8 sets of paired fastq files with accompanying UMI fastq files. Sequenced on the Illumina NoveSeq6000 platform using S2 flow cells (v1.5,2x150 cycles), with a mean yield of 313 million reads.

  Dataset EGAD50000001380

• Smart-seq2 analysis of 11 ETMR patient samples (4,031 high-quality single cells/nuclei).

  We analyzed 11 ETMR patient samples using single-cell transcriptomics (6 from fresh and 5 from frozen material). Samples were profilied by paired-end scRNA-sequencing using the Smart-Seq2 protocol on a NextSeq 500 sequencer (Illumina). The dataset contains 8,062 fastq files of 4,031 high-quality single cells/nuclei. Raw fastq files of high-quality cells are shared.

  Dataset EGAD50000001379

• M116 DNA Methylation Array

  DNA samples were quantified with Qubit BR kit on a Qubit2.0 fluorimeter (ThermoFisher Scientific), and integrity was checked using agarose gel electrophoresis. DNA from samples passing the quality control was bisulfite-converted using the EZ DNA Methylation-Gold™ Kit (Zymo Research, Cat. D5005) following the manufacturer's instructions. Then, bisulfite-converted DNA was hybridized into the Methylation EPICv1.0 BeadChip (Illumina) array interrogating > 850,000 CpG sites according to manufacturer's instructions

  Dataset EGAD50000001678

• Neuroblastoma cell-lines and healthy cfDNA used for the benchmarking study

  This dataset comprises nine neuroblastoma cell lines used to construct the reference matrix for the RRBS-CL dataset in the corresponding benchmarking study. In addition, it includes one extra neuroblastoma cell line and a pooled cfDNA sample from multiple donors, which were used to generate the in silico mixtures. All the samples were profiled using DNA methylation sequencing.

  Dataset EGAD50000002199

• scRNAseq of fresh CRC punch biopsies

  The dataset includes single-cell RNAseq data generated by Plate-Seq technology from scDiscoveries. The dataset contains samples from a total of 5 different early CRC biobank patients (pt5, pt11, pt13, pt14, pt16) from 4 different sequencing runs. Punch biopsies of different regions of the tumor (Core, Invasive front and, if available, Adenoma) were FACS sorted onto plates prior to sequencing.

  Dataset EGAD50000002203

• Multi-omic single-nucleus ATAC-seq + RNA-seq of nuclei from the motor cortex of ALS/ALS-FTD patients and unaffected controls

  A multi-omic dataset of single-nucleus paired ATAC-seq + RNA-seq data of nuclei from the post-mortem human primary motor cortex from patients with ALS/ALS-FTD and unaffected controls.

  Dataset EGAD50000002240

• ALS-FTD brain motor cortex single-nucleus RNA-seq of NeuN+ nuclei sorted for TDP-43 low vs. high

  A dataset of single-nucleus RNA-seq data of nuclei from the post-mortem human primary motor cortex from patients with ALS-FTD. The nuclei were sorted by flow cytomotetry into NeuN+ (neuronal) TDP-43 low vs. TDP-43 high fractions.

  Dataset EGAD50000002243

• Multimodal epigenetic sequencing analysis of cell-free DNA identifies biomarkers for ALS diagnosis and progression

  The role of the epigenome in age-related neurodegenerative disorders remains understudied. Here, we analyzed circulating cell-free DNA (cfDNA) from blood to detect methylation changes as a liquid-biopsy for Amyotrophic Lateral Sclerosis (ALS). Our study included 20 patients with sporadic ALS, 10 patients with C9orf72-associated ALS, 10 asymptomatic carriers of the C9orf72 repeat expansion mutation, and 21 non-disease controls.

  Dac EGAC50000000739

• Metadata Submission

  Metadata Submission Welcome to the EGA Metadata Submission landing page. Here, we provide you with essential resources to facilitate a seamless submission process. Before starting your submission, please make sure you check the following pages: Submission FAQ Submission Quickguide Submission account terms EGA metadata schema If you want to submit sequencing and/or phenotypic data check these pages: Submitter Portal Submitter Portal API If you want to submit array data check these pages: Array documentation Webin Webin API

  Documentation submission/metadata/submission

• Single-cell DNA sequencing dataset of aplastic anemia and RCC (31 samples)

  This dataset contains raw single-cell DNA sequencing data generated from 31 patients with aplastic anemia using the Mission Bio Tapestri platform. Libraries were prepared with a targeted DNA amplicon panel and sequenced on an Illumina platform. These raw data capture single-cell variant profiles and clonal architecture and serve as the basis for downstream analysis of somatic mutations and clonal hematopoiesis.

  Dataset EGAD50000002189

• Hepatitis B Viral sequence reads

  Deep sequencing of viral samples (average ~9,000x coverage) from patients chronically infected with Hepatitis B (HBV). Whole HBV genome sequencing of 1467 patients (1102 in discovery and 365 in validation cohort) chronically infected with HBV at baseline. The patient population contained HBV genotypes A (98), B (285), C (716), D (356), E (7), and F (5) with 977 HBeAg-positive and 490 HBeAg-negative patients.

  Dataset EGAD00001005075

• Whole Exome Sequencing of Spanish Patients diagnosed with rare ophtalmogenetic disorders.

  Whole Exome sequencing of a set of Spanish patients suffering rare genetic diseases. The set consists of 4 patients, one was diagnosed with Retinitis Pigmentosa (RP-1629), another one was diagnosed with Macular Dystrophy (MD-0235) and two were diagnosed with Leber's Congenital Amaurosis (LCA-0081 and LCA-0103).

  Dataset EGAD00001005746

• Genomic analysis of pancreatic neuroendocrine tumour with MEN1, ATRX, or DAXX mutations

  This dataset includes 52 samples from 19 individuals with pancreatic neuroendocrine tumours. These samples were analyzed using RNA-sequencing, whole-exome sequencing, shallow (~0.3x) whole-genome sequencing, and a 21-gene panel targeted capture sequencing. Everything was sequenced using the Illumina HiSeq and is provided in BAM or FASTQ format.

  Dataset EGAD00001006001

• Genetic screening of GPI-anchor protein synthesis

  This study will analyse the guide sequence which were used for making mutations in the Cas9-expressing cells. We used GeCKO v2 library which were released by Feng Zhang, 2014. This data is part of a pre-publication release. For information on the proper use of pre-publication data shared by the Wellcome Trust Sanger Institute (including details of any publication moratoria), please see http://www.sanger.ac.uk/datasharing/

  Dataset EGAD00001001928

• Whole genome sequencing data of lung adenocarcinomas

  This dataset includes the whole-genome sequencing data from a study entitled "Tracing Oncogene Rearrangements in the Mutational History of Lung Adenocarcinoma". Whole-genome sequencing libraries were generated by PCR-free methods, and sequencing run was made in HiSeq X Ten machines. PCR duplicates-marked, indel-realigned, and base-recalibrarted BAM files are provided in our dataset.

  Dataset EGAD00001004793

• Whole Genome Sequencing of INTERVAL (2019-06-12)

  15x Whole Genome Sequencing of 15,000 individuals from the INTERVAL study cohort, phase III. This data is part of a pre-publication release. For information on the proper use of pre-publication data shared by the Wellcome Trust Sanger Institute (including details of any publication moratoria), please see http://www.sanger.ac.uk/datasharing/ . This dataset contains all the data available for this study on 2019-06-12.

  Dataset EGAD00001005086

• Sequence Data for Paper: Epigenetic reprogramming during differentiation of human CD4+ T lymphocytes into memory stages

  Sequence Data of total RNA, miRNA, WGB, mRNA, NOMe, Chip (H3K27ac,H3K27me, H3K36me3, H3K4me1, H3K4me3, H3K9me3, Input) Short Desrciption: Epigenetic profiling of human CD4+ memory T cells reveals their proliferative history and argues in favor of a progressive differentiation model driven by epigenetically controlled master regulators.

  Dataset EGAD00001001865

• RNA sequencing of high-risk paediatric cancers for identifying T-cell infiltration signatures

  Transcriptome profiling of 121 high-risk paediatric cancer samples for identifying T-cell infiltration signatures using poly-A capture by Truseq and sequenced on NextSeq 500

  Dataset EGAD00001010920

• PIAMA nasal RNAseq dataset

  This study included nasal gene expression data collected from nasal brushes of adolescents in PIAMA birth cohort, which is used in the project Nasal DNA methylation at three CpG sites predicts childhood allergic disease. 186 samples were included in this analysis, and phenotypes (age and sex) were provided together with a gene expression count table. Gene expression was measured by the Illumina HiSeq2500 sequencer.

  Dataset EGAD00001008767

• Enhanced reduced representation bisulfite sequencing (eRRBS) on 45 multiple myeloma samples and 3 normal plasma cell

  Enhanced reduced representation bisulfite sequencing (eRRBS) on 45 multiple myeloma samples and 3 normal plasma cell. Enhanced reduced representation bisulfite sequencing (eRRBS) on 45 multiple myeloma samples and 3 normal plasma cell. Libraries were sequenced on a HiSeqTM4000 Illumina machine using 75bp paired-end reads

  Dataset EGAD00001007822

• Sequence-based gene expression in uterine and ovarian carcinosarcomas

  Gynecologic carcinosarcomas (CS), including more generally uterine (endometrial) and less frequently ovarian localization, are histologically defined as biphasic neoplasms composed of carcinomatous (C) epithelial and sarcomatous (S) malignant components. We report a comprehensive analysis of 20 patients of macro-dissected samples of C and S components through RNA sequencing.

  Dataset EGAD00001009099

• Paired-end Whole Exome-seq analysis of multi-centric lower grade glioma

  This dataset includes Nimblegen SeqCap EZ Exome v3 data for each lesion of three patients with multicentric glioma. For two patients, each lesion was sequenced along with whole blood. For a third patient, 3 pieces from the right lesion and 4 pieces from the left were sequenced along with whole blood. In each case BAM files that have been aligned with BWA mem alignment are available.

  Dataset EGAD00001003795

• Exome sequencing of 102 gastric cancer and matched normal from Japanese cohorts (YCU)

  As a contribution to the International Cancer Genome Consortium, exome sequencing of 102 Japanese gastric cancer with various histological subtypes have been conducted. This study aims to identify unique and common driver genes and molecular subtypes in Japanese gastric cancer. Please refer ICGC website for detail: http://icgc.org/icgc/cgp/69/420/1012357

  Dataset EGAD00001004042

• Paired-WGS-two-families-with-GBA-variants-and-Parkinsons-disease

  We performed whole- genome sequencing, rare variant filtering, segregation analysis and functional validation of PD cosegregating rare genetic variation in two families (6 samples) segregating PD associated GBA variants c.115+1G>A (ClinVar ID: 93445, ) and p.L444P (ClinVar ID: 4288) respectively. The paired WGS sequencing was run on HiSeq X Ten and the library preparation kit was Illumina TruSeq DNA nano.

  Dataset EGAD00001006561

• 48 EXOMES FAMILIAL MYELOID LEUKEMIA (QMUL)

  Whole exome sequencing (WES) libraries were prepared from 200ng of genomic DNA using the Agilent SureSelect XT Target Enrichment System for Illumina Paired-End Multiplexed Sequencing Library coupled with the Agilent SureSelect XT Human all exon v6 capture reagent. Libraries were sequenced on a NextSeq 550 sequencer using the High output 300 cycles kit generating 150bp paired end single-indexed reads. Alignment against b37 using Novoalign (version 3.02.08).

  Dataset EGAD00001004539

• ChIP sequencing data to study therapeutic targeting of ependymoma

  This dataset contains ChIP sequencing data from 24 patients. ChIP of 5–10 mg flash-frozen primary ependymoma tumour was performed using 5 mg H3K27ac antibody per ChIP experiment. The enriched DNA has been sequenced on a Illumina HiSeq2000 instrument in paired-end mode. Up to two lanes per sample have been sequenced resulting in 70 Fastq files.

  Dataset EGAD00001003979

• RNAseq Atlas in HCC tumors

  The atlas provides a comprehensive exploration of genetic and transcriptomic landscapes within HCC, offering insights into key genomic alterations and gene expression patterns. The integration of DNA sequencing and RNAseq data enhances our understanding of the molecular complexity underlying HCC, potentially paving the way for targeted therapeutic strategies and biomarker discovery in the context of hepatocellular carcinoma.

  Dataset EGAD00001015341

• CRISPR iPSC methods paper

  This submission is of the sequencing data used in the CRISPR iPSC methods paper. Specifically it is 3 fastq files that each represent a replicate of an experiment to transduce the Toronto KnockOut CRISPR Library - Version 3 (TKOv3) into inferred pluripotent stem cell (iPSC) derived macrophages. The sequencing is of the guide RNAs from the TKOv3 having been extracted from the transduced iPSC derived macrophages.

  Dataset EGAD00001007020

• Australia and New Guinea haplotype phasing (2017-06-27)

  Whole-genome sequencing and phasing of admixed Aboriginal Australian genomes and Papua New Guinean genomes using 10x Genomics Chromium technology. This data is part of a pre-publication release. For information on the proper use of pre-publication data shared by the Wellcome Trust Sanger Institute please see http://www.sanger.ac.uk/datasharing/ This dataset contains all the data available for this study on 2017-06-27.

  Dataset EGAD00001003407

• Isotype-resolved sequencing of B-cell receptor in health and disease (2017-09-13)

  Sequencing of B-cell receptor repertoires in healthy individuals and patients with chronic lymphocytic leukemia. 1) This data is part of a pre-publication release. For information on the proper use of pre-publication data shared by the Wellcome Trust Sanger Institute please see http://www.sanger.ac.uk/datasharing/ This dataset contains all the data available for this study on 2017-09-13.

  Dataset EGAD00001003748

• BulkTCR-sequencing data from different human intestinal CD8 T cell subsets.

  5000 cells of each subset of CD8 T cells (CD103-KLRG1+, CD103-KLRG1- and CD103+ from LP and CD103+ IELs) were sorted into tubes. A modified SMART protocol was used in first-strand cDNA synthesis, and TCRalpha / TCRbeta genes were amplified in two rounds of semi-nested PCR reaction, following the method described in detail in Risnes et al., 2018.

  Dataset EGAD00001005049

• Human and rat skeletal muscle multiomic profiling sequencing data

  Illumina sequencing data (fastq files) representing single-nucleus (sn) ATAC-seq, snRNA-seq, bulk ATAC-seq, and snATACseq+snRNAseq multiomics data from human and rat skeletal muscle samples (19 libraries total). Includes a README file that describes the relationship between libraries, samples, and files.

  Dataset EGAD00001008323

• ctDNA whole exome genome sequencing dataset

  Dataset contains all available exome sequencing paired-end fastq files from our study "A generalizable machine learning framework for classifying DNA repair defects using ctDNA exomes"

  Dataset EGAD00001010069

• Targeted DNA sequencing on bulk bone marrow and peripheral blood

  Targeted DNA sequencing was performed on 195 bone marrow samples to identify cases of clonal haematopoiesis, and on 99 paired peripheral blood samples. The SeqCap EZ HyperCap protocol was followed, and targeted capture performed against a panel of 97 genes recurrently mutated in myeloid malignancies and clonal hematopoiesis. One BAM file (mapped to the hg38 reference genome) is provided per sample.

  Dataset EGAD00001011083

• Exome sequencing of 142 gastric cancer and matched normal from Japanese cohorts (UT)

  As a contribution to the International Cancer Genome Consortium, exome sequencing of 142 Japanese gastric cancer with various histological subtypes have been conducted. This study aims to identify unique and common driver genes and molecular subtypes in Japanese gastric cancer. Please refer ICGC website for detail: http://icgc.org/icgc/cgp/69/420/1012357

  Dataset EGAD00001004041

• RNA Sequencing of Colorectal Liver Metastases

  Reverse-stranded paired-end 75 base-pair RNA sequencing libraries of 93 metastatic FFPE samples were constructed using Illumina Total RNA Stranded Kits. Ribosomal RNAs (rRNAs) were depleted by using the Ribo-Zero rRNA Removal Kit (Illumina). Libraries were sequenced on a HiSEQ2500 machine. Five samples were re-sequenced using paired-end 50 base-pair libraries due to the smaller insert sizes.

  Dataset EGAD00001004111

• RNA-seq transcriptome of bronchial brush samples from COPD and control

  Human subjects (COPD patients or apparently healthy controls) where investigated by bronchoscopy and a 5 mm brush was used to sample the subsegment airways of the right lung. The material obtained mainly consist of bronchial epithelial cells plus some contamination with leukocytes. For further details see Ziegler-Heitbrock et al, European Respiratory Journal, 40:823-829, 2012.

  Dataset EGAD00001002003

• The Chinese University of Hong Kong Hereditary Spastic Paraplegia Data

  The dataset contains the whole genome sequencing data of a family with two unaffected parents and two probands that showed Hereditary spastic paraplegias symptoms. Sequencing reads were aligned to human genome (GRCh38) using BWA-MEM, followed by indel-realignment and PCR-duplicates marking. Alignment results are available for download in BAM format.

  Dataset EGAD00001002146

• Cell type-specific transcriptomics of esophageal adenocarcinoma

  3' mRNA-Seq obtained from distinct isolated cell types (epithelia cells,immune cells, fibroblasts) of endoscopically obtained esophageal adenocarcinoma tissue as well as normal esophageal mucosa. Libraries for RNA-sequencing were prepared using the QuantSeq 3' mRNA-Seq Library Prep Kit FWD for Illumina according to the low input protocol. Libraries were sequenced on a HiSeq 4000 (Illumina) by 1x 50 bases.

  Dataset EGAD00001005508

• Whole Genome Sequencing of Human Organoid Lines (2020-02-20)

  The study will use WGS to aid in benchmarking different culture conditions in a set of genetically annotated human organoid lines. The data will be used to assess whether there is any clonal differences introduced when culturing these lines in different conditions. . This dataset contains all the data available for this study on 2020-02-20.

  Dataset EGAD00001005995

• RNA-seq data of Metabolic profiling of patient-derived organoids reveals nucleotide synthesis as a metabolic vulnerability in malignant rhabdoid tumors

  Patient-matched normal kidney organoid (103H) and MRT tumoroid (103T2) models were treated for 24h with either DMSO (ctrl), 400nM MTX, or 50nM BAY to investigate the direct effects of drug treatment on the expression of key metabolic enzymes in the nucleotide biosynthesis pathways.

  Dataset EGAD00001015391

• 47 urothelial cancer patients WES and 38 RNAseq

  We performed a global mutational landscape analysis using tumor samples from the 47 urothelial cancer patients included in MATCH-R or MOSCATO studies, with advanced metastatic disease and WES available (RNAseq is included for 38 patient samples).

  Dataset EGAD00001011063

• Spatially resolved cellular and molecular drivers of cardiac remodelling in healthy and failing human hearts: Adult RNA (2025-07-31)

  Cell Atlas of the diseased human heart. This data is part of a pre-publication release. For information on the proper use of pre-publication data shared by the Wellcome Trust Sanger Institute (including details of any publication moratoria), please see http://www.sanger.ac.uk/datasharing/ . This dataset contains all the data available for this study on 2025-07-31.

  Dataset EGAD00001015668

• Long-Term Outcome in Offspring and Mothers of Dexamethasone-Treated Pregnancies at Risk for Classical Congenital Adrenal Hyperplasia Owing to 21-Hydroxylase Deficiency

  Summary The purpose of this study is to perform the first long-term follow up study both of adolescents and young adults with a history of prenatal treatment with dexamethasone and of their mothers and to test for adverse medical or behavioral side effects. The emphasis will be on the outcome of this prenatal treatment in those fetuses who are not affected with steroid 21-hydroxylase deficiency (21OHD) form of congenital adrenal hyperplasia (CAH) and are either heterozygotes or homozygote-unaffected. Prenatal treatment of 46,XX fetuses with 21OHD (via administration of dexamethasone to the pregnant mother) has been shown to reduce the masculinization of the genitalia and, thereby, the later need for 'corrective' feminizing genital surgery and potential impairment of sexual functioning. Along with the suppression of excess prenatal androgen production and reduction of prenatal masculinization of the genitalia in 46,XX fetuses with 21OHD, prenatal dexamethasone treatment may reduce the behavioral masculinization that is well documented in untreated 46, XX patients with 21OHD. Those fetuses who are partially treated until diagnosis of an unaffected status (heterozygous or homozygous-unaffected) will be studied as well. This latter group is of importance because these fetuses are being unnecessarily treated, but we have no way of diagnosing the unaffected status before the 8th week of gestation when treatment must begin for the female fetus who is affected. As treatment is necessary only until term in the female fetus affected with classical CAH, male fetuses and unaffected or heterozygous female fetuses do not require treatment. Thus, only one out of eight fetuses will require prenatal treatment until term. Findings of adverse effects of glucocorticoid treatment in non-human mammals [1, 2] have raised concerns among other clinicians and investigators about potential adverse side effects of such treatment on the developing human. Thus, this study will address this concern and investigate the potential adverse side effects of prenatal treatment. Detailed Description Females with classical CAH owing to 21-hydroxylase deficiency are born with ambiguous genitalia due to the production of excess androgens in utero. Prenatal treatment with dexamethasone was inaugurated in 1978 by Maguelone Forest and has been the standard of practice in the United States since 1986. Dexamethasone, which crosses the placental barrier, suppresses the fetal adrenal gland production of androgens, thus preventing ambiguous genitalia in the affected female. Children of both sexes are prenatally treated as soon as pregnancies at risk are confirmed. Treatment in females with 21OHD continues to term, but is discontinued in males and unaffected females. To date, 685 pregnancies have been diagnosed, of which 366 fetuses were treated. These investigators will study prenatally treated adolescents and adults 12 years and older with respect to medical and behavioral outcomes (see Table 5). In addition, mothers of children prenatally treated for varying periods of time for suspected 21OHD will be studied for long-term side effects of dexamethasone treatment administered during pregnancy. The long-term outcome in these children and their mothers has never before been studied.

  Study phs001317

• Primary Prostate Tumor Tissue DNA Methylation Profiles

  The study cohort includes 525 Caucasian and African American prostate cancer (PCa) patients who underwent radical prostatectomy (RP) as primary therapy for clinically localized adenocarcinoma of the prostate. These patients were previously enrolled in one of two population-based studies: 1) men 40-64 years of age, diagnosed between January 1993 and December 1996; and, 2) men 35-74 years of age, diagnosed between January 2002 and December 2005. Gleason grade and sum, diagnostic PSA, and pathological tumor stage were obtained from the SEER cancer registry. Formalin-fixed paraffin-embedded prostate tumor tissue blocks were obtained from RP specimens and used to make hematoxalin and eosin stained slides, which were reviewed by pathologists to confirm the presence/location of adenocarcinoma. For each patient two 1-mm tumor tissue cores from the dominant lesion that were enriched with >=75% tumor cells were taken for DNA purification. The RecoverAll Total Nucleic Acid Isolation Kit (Ambion/Applied Biosciences, Austin, TX) was used to extract DNA, which was then quantified (PicoGreen), aliquoted onto 96-well plates and shipped to Illumina for DNA methylation profiling using the Infinium® HumanMethylation450 BeadChip.

  Study phs001921

• Systems Analysis of Single-Cell Heterogeneity Underlying Glioma Drug Resistance

  This study characterizes the transcriptional regulatory mechanisms that drive responses of two patient-derived glioblastoma multiforme (GBM) stem-like cells (PD-GSCs) that have distinct phenotypes - one sensitive and another resistant to the drug pitavastatin. This study investigates the differing mechanisms driving drug response in the two PD-GSCs at the single-cell level, and provides an approach that can be used to infer transcriptional regulatory network models, which can then be used to identify mechanisms driving cell-state transitions, e.g., proneural-to-mesenchymal transitions (PMT), which has been observed experimentally and clinically in GBM. The use of these network models enabled the identification of transcription factor and gene targets that perturbed drug-induced PMT through 1) simulations of network dynamics network, and 2) characterization of gene program activities over the time-course response to drug treatment. These results enabled the identification of multiple siRNAs and the drug vinflunine as secondary components that potentiate the efficacy of pitavastatin. This work demonstrates an approach to uncover the transcriptional network topology (TRN) topology of PD-GSCs, and use it to rationally predict combinatorial treatments that block treatment escape and acquired resistance to drugs in GBM.

  Study phs003501

• Transcriptomes of human CD4+ T lymphocytes - Metabolic project

  Human CD4+ T cells are essential mediators of immune responses. By altering the mitochondrial and metabolic states, we defined metabolic requirements of human CD4+ T cells for in vitro activation, expansion, and effector function. T cell activation and proliferation were reduced by inhibiting oxidative phosphorylation, while early cytokine production was maintained by either OXPHOS or glycolytic activity. Glucose deprivation in the presence of mild mitochondrial stress markedly reduced all three T cell functions, contrasting the exposure to resveratrol, an antioxidant and sirtuin-1 activator, which specifically inhibited cytokine production and T cell proliferation, but not T cell activation. Conditions that inhibited T cell activation were associated with the downregulation of 2′,5′-oligoadenylate synthetase genes via interferon response pathways. Our findings indicate that T cell function is grossly impaired by stressors combined with nutrient deprivation, suggesting that correcting nutrient availability, metabolic stress and/or the function of T cells in these conditions will improve the efficacy of T cell-based therapies.

  Study EGAS00001005565

• Effect of ETS2 modulation on chromatin accessibility and enhancer activity in inflammatory (TPP) macrophages

  This dataset consists of H3K27ac ChIP-seq and ATAC-seq in primary human macrophages following modulation of ETS2 expression. There are three datasets: H3K27ac ChIP-seq in ETS2-edited and unedited inflammatory (TPP) macrophages, H3K27ac ChIP-seq in ETS2-overexpressing and control M0 macrophages, and ATAC-seq in ETS2-edited and unedited inflammatory macrophages. For editing experiments, monocytes were transfected with CRISPR-Cas9 ribonucleoproteins containing a gRNA that was validated to edit ETS2, or a non-targeting control gRNA. Inflammatory macrophage differentiation was performed using conditions that model chronic inflammation (TPP): 3 days GM-CSF (50ng/mL) followed by 3 days GM-CSF, TNFa (50ng/mL), PGE2 (1mg/mL), and Pam3CSK4 (1mg/mL). For overexpression experiments, controlled overexpression of ETS2 mRNA or control mRNA (an equivalent amount of mRNA encoding the reverse complement of ETS2 – thereby controlling for the quantity, length and purine/pyrimidine composition of the transfected RNA but with a transcript that would not be translated) was induced in resting, non-activated (M0) macrophages by transfecting 500ng mRNA. To minimise non-specific activation due to the transfected RNA, in vitro transcription was performed using co-transcriptional capping (to minimise uncapped products), and incorporating modified, minimally immunogenic nucleotides (replacing uridine with N1-methyl-pseudouridine and cytidine with methylcytidine). After 18 hours, transfected cells were were activated with low dose LPS and harvested 6 hours later. For H3K27ac ChIP-seq, cells were harvested, cross-linked, quenched, lysed, and sheared. Immunoprecipitation of histone-DNA complexes was performed overnight at 4C with rotation using an anti-H3K27ac antibody and the SimpleChIP Plus Sonication ChIP kit (Cell Signaling Technology). Following reverse cross-linking, 50ng of immunoprecipitated DNA or input DNA were used to prepare sequencing libraries using the iDeal Library Preparation kit (Diagenode), according to manufacturer instructions. 10 PCR cycles were used for the amplification step and size selection was not performed. The quality and molarity of all libraries was assessed using a BioAnalyzer 2100 (Agilent) and the libraries were sequenced on an Illumina NovaSeq 6000. Raw data are provided as raw and aligned sequencing reads from H3K27ac-bound DNA and the input chromatin. ATAC-seq in TPP macrophages was performed using the Omni-ATAC protocol (Corces et al 2017) with the following modifications: cell number was increased to 75,000 cells, cell lysis time was increased to 5 minutes; volume of Tn5 transposase in the transposition mixture was doubled; duration of the transposition step was extended to 40 minutes. Sequencing libraries were prepared by PCR with the NEBNext Ultra II Q5Master Mix (NEB) as described previously (Corces et al 2017). Amplified libraries were cleaned using AMPure XP beads (Beckman Coulter). Sequencing reads were trimmed using TrimGalore! (Phred score 24), filtered to remove reads < 20 bp and aligned to the human genome (hg19) using Burrows-Wheeler Aligner (BWA). Aligned reads were converted to BAM files, sorted and indexed using SAMtools (Li et al, 2009). PCR duplicates and unmapped reads were removed, and the resulting BAM files were re-sorted and indexed. Raw data are provided as 100 bp paired-end Illumina reads from n = 3 donors for each experiment. BAM files are also provided.

  Study EGAS50000000109

• Small molecule inhibitors of LOXL synergize with 5-AZA to restore erythropoiesis in myeloid neoplasms

  Objectives: Bone marrow (BM) stroma and microenvironmental factors in the BM milieu of myeloid neoplasms are increasingly being recognized as novel perspective therapeutic targets. Lysyl oxidases (LOX/LOXL) are crucial enzymes that cross-link collagen and contribute to the deposition of aberrant stiff extracellular matrix (ECM) in the BM of patients with myelodysplastic syndromes (MDS) and myeloproliferative neoplasms (MPN). Here, we studied whether inhibition of LOX/LOXL enzymes produced by BM-derived MSCs favors the differentiation of MDS/MPN hematopoietic stem cell and progenitor cells (HPSCs) and enhances the effects of 5-azacytidine (5-AZA). Methods: Primary co-cultures of MSCs and HPSCs were established from n=31 MDS/MPN patients and treated with the pan-LOX/LOXL inhibitor PXS-5505, 5-AZA or the combination of both for 4 days. At the end of co-culture, HSPC differentiation was assessed by CFU assays and flow cytometry. In some of the experiments, MSCs monolayers were pre-treated or not with PXS-5505 for 6 days and decellularized using 20 mM ammonia. Subsequently, autologous HSPCs were cultured on ECM matrices in the presence of 5-AZA or PXS-5505 + 5-AZA combination with or without inhibitors of integrin signaling such as blebbistatin, Y-27632 and αvβ3-integrin blocking antibodies. Patient-derived xenografts (PDX) were established in NSG mice based on intrafemoral co-injection of HSPCs and autologous MSCs. PDX were treated with 3 cycles of 5-AZA (75 mg/m2), PXS-5505 (30 mg/kg on alternate days for the duration of 5-AZA treatment) or combination of both. Results: MSCs of MDS and MPN patients displayed significant overexpression of LOX/LOXL transcripts as compared to healthy age-matched controls. Remarkably, LOX/LOXL inhibition in MSCs/HSPCs co-cultures increased erythroid differentiation of patient HSPCs that had suboptimal or no response to 5-AZA alone. From a total of n=31 advanced stage MDS/MPN patients, n=11 (35.5%) responded to the combination treatment, which was evidenced by a significant increase of CD235a+CD45- erythroid cell production compared to 5-AZA/PXS/untreated arms (p&lt;0.05). Of note, combination treatment with 5-AZA + PXS-5505 facilitated differentiation of CD235+ erythroid progenitors from HSPCs sub-clones with low mutational VAFs. The presence of MSCs was necessary for the effects of the combination treatment and required direct cell contacts between MSCs and HSPCs. Moreover, PXS-5505-mediated alterations of the ECM composition alone were sufficient to induce erythroid differentiation. Inhibition of integrin-mediated signaling using blebbistatin, Y-27632 and αvβ3-integrin antibodies completely abrogated the effects of PXS-5505 induced erythroid differentiation. In vivo, the combination of PXS-5505 + 5-AZA favored erythroid differentiation of HSPCs and confirmed the in-vitro results. The combination treatment of 5-AZA + PXS-5505 induced the greatest reduction of disease burden as assessed by decrease of spleen indexes, total engraftment rates, BM fibrosis and mutational VAF profiles. Conclusions: The combination of 5-AZA + LOX/LOXL inhibitor PXS-5505 in MDS and MPN synergistically restores erythropoietic differentiation of primary HSPCs. The effect is dependent of contact interaction between HSPCs and niche components, such as MSCs and MSCs-derived ECM. Our data propose a strong rationale for a BM niche targeted combination treatment with 5-AZA + PXS-5505 in transfusion dependent MDS and MPN, which will be assessed in forthcoming clinical trials.

  Study EGAS00001006174

• Innate Immune Anti-Viral Deaminase Deregulation Fuels Pre-Leukemia Stem Cell Evolution

  This study evaluates the effect of Rebecsinib treatment on gene expression, isoform splicing, and ADAR1 mediated RNA editing in cells from an in vivo humanized mouse model which have been engrafted with human secondary AML cells. For analysis of cells from Rebesinib-treated animals, CD34+ cells were obtained from bone marrow and spleens of sAML-engrafted mice following once weekly treatment with 0 (Vehicle) or 5 or 10 mg/kg Rebecsinib. For analysis of cells from serial transplant recipients of cells harvested from Rebecsinib-treated mice, CD34+ cells were obtained from bone marrow and spleens of sAML-engrafted mice that were transplanted with cells from animals that received twice weekly treatment with 0 (Vehicle) or 10 mg/kg Rebecsinib. No further treatment was given to serial transplant recipients, thus allowing the analysis of molecular changes that are sustained following serial transplantation. Whole transcriptome sequencing was performed at The Scripps Research Institute Next Generation Sequencing Core on Illumina NextSeq 500 sequencers with 150bp paired-end reads.These data revealed a uniquely expressed ADAR1 splice isoform (with a retrained intronic region), global downregulation of normalized RNA editing events, and isoform switching from MCL1-L to MCL1-S in Rebecsinib-treated sAML-engrafted mice. These results demonstrate the anti-survival and anti-self-renewal effects of Rebecsinib treatment on the leukemia stem cell (LSC) population in sAML-engrafted mice as well as the reduction in the inflammation-induced, hyper-editing p150 isoform of ADAR1. Thus, Rebecsinib treatment has promise for reducing the LSC population in myeloproliferative disorders and associated cancers.

  Study phs002228