Digital gene expression profiling of primary acute lymphoblastic leukemia cells
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE26530 | GEO | 2011/12/20
SECONDARY ACCESSION(S): PRJNA142169
REPOSITORIES: GEO
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Project description:We determined the genome-wide digital gene expression (DGE) profiles of primary acute lymphoblastic leukemia (ALL) cells from 28 patients and fractionated blood cells from healthy blood donors taking advantage of “second generation” sequencing technology. The patients included in the study represent distinct subtypes of B-cell precursor (BCP) ALL and T-cell lineage ALL (T-ALL) and the controls are fractionated CD19+ and CD3+ cells. Gene expression analysis of 28 ALL patient samples with different immunophenotypic backgrounds including T-ALL (n=4) and patients with BCP ALL with diverse cytogenetic backgrounds: High Hyperploidy (HeH) (n=10), t(9;22) BCR-ABL1 (n=3), t(12;21) ETV6-RUNX1 (n=4), dic(9;20) (n=3), t(1;19)TCF3-PBX1, MLL/11q23 (n=1) and undefined/non-recurrent aberrations (n=1). Fractionated b-cells (CD19+) and t-cells (CD3+) isolated from peripheral blood of healthy donors were used as controls. Sequencing libraries were prepared from 1 µg of total RNA using reagents from the NlaIII Digital Gene Expression Tag Profiling kit (Illumina Inc, San Diego, CA, USA). mRNA was captured on magnetic oligo(dT) beads and reverse transcribed into double-stranded cDNA. The cDNA was cleaved using the restriction enzyme NlaIII. An adapter sequence containing the recognition sequence for the restriction enzyme MmeI was ligated to the NlaIII cleavage sites. The adapter-ligated cDNA was digested with MmeI to release the cDNA from the magnetic bead, while leaving 17 base-pairs of sequence in the fragment. The fragments were dephosphorylated and purified by phenol-chloroform. A second adapter was ligated at the MmeI cleavage sites. The adapter-ligated cDNA fragments were amplified by PCR, and the PCR products were purified on a 6% polyacrylamide gel. The ~96 base pair PCR products were excised from the gel and eluted overnight, followed by ethanol precipitation and re-suspension. Purified libraries were quality controlled and quantified on a Bioanalyzer using DNA 1000 series or High Sensitivity chips. The DGE libraries were diluted to a 10 nM concentration and sequenced on one lane of an Illumina GAII or GAIIx for 18 cycles.
2011-12-20 | E-GEOD-26530 | biostudies-arrayexpress
Project description:Analysis of RNA samples by massive parallel sequencing holds the promise to assay gene expression in both a quantitative and qualitative fashion and therefore allows for digital gene expression (DGE) profiling. We assessed the effect of different experimental approaches by generating small RNA libraries from a biological sample as well as an equimolar pool of synthetic miRNAs and analyzed the results using capillary dideoxy sequencing and next-generation sequencing platforms (Roche/454, AB/SOLiD and Illumina/Solexa). Whereas different sequencing platforms provided highly similar results, large differences in DGE profiles were observed depending on the library preparation method used. Nevertheless, our results indicate that the preferential nature of the library preparation methods is systematic and highly reproducible and we show that DGE is well suited for the quantification of relative expression differences between samples. This SuperSeries is composed of the following subset Series: GSE16369: Limitations and possibilities of small RNA digital gene expression profiling: library preparation comparison (454) GSE16370: Limitations and possibilities of small RNA digital gene expression profiling: library preparation comparison (SOLiD) GSE16371: Limitations and possibilities of small RNA digital gene expression profiling: spleen and liver comparison (SOLiD) GSE16372: Limitations and possibilities of small RNA digital gene expression profiling: synthetic miRNAs replicates (SOLiD) GSE16373: Limitations and possibilities of small RNA digital gene expression profiling: synthetic miRNA replicates (Illumina) Refer to individual Series
2009-07-28 | E-GEOD-16374 | biostudies-arrayexpress
Project description:The aim of this study was to benchmark digital gene expression (DGE) profiling by massively parallel sequencing against the most commonly used method for gene expression analysis. We compared the DGE levels to expression levels from Affymetrix arrays. Data from Affymetrix Human Genome U133 plus 2.0 GeneChips was available for 12 of the 21 RNA samples from ALL patient cells analyzed by DGE.
2011-12-20 | GSE26865 | GEO
Project description:In present study, digital gene expression (DGE) profiling was performed to obtain a general picture of the transcriptomic implicated in the early development of R. venosa. Eighteen DGE libraries at six developmental stages of R. venosa were constructed, sequenced by IIIumina HiSeq 2500 platform.
2016-07-22 | GSE70548 | GEO
Project description:Here we performed a transcriptomic study on the effect of sodium nitroprusside (SNP) on cucumber leaves under alkali stress using Solexa/Illumina's high-throughput digital gene expression (DGE) system. Two DGE libraries (from one NaHCO3-treated sample and one NaHCO3 + SNP-treated sample) were constructed, and the gene expression variations between the two samples were compared. Hundreds of differentially expressed genes were obtained by the comparison, and GO analysis of these genes suggested that many biological processes, molecular function, cellular components were related to SNP’s mitigated effect on alkaline stress. The authenticity of the DGE data was further confirmed by analyzing real-time RT-PCR using several random-selected genes.
2013-11-26 | GSE42439 | GEO
Project description:Here we performed a transcriptomic study on PaWB phytoplasma-infected Paulownia sp. using Solexa/Illumina’s high-throughput digital gene expression (DGE) system. 4 DGE libraries (from 2 virus-infected samples and 2 healthy samples) were constructed, and the gene expression variations between the PaWB phytoplasma-infected (diseased) sample and the corresponding healthy sample were compared. Thousands of differentially expressed genes were obtained by the comparison, and KEGG pathway analysis of these genes suggested that many biological processes were responded to PaWB infection.
2014-10-01 | GSE42087 | GEO
Project description:Here we performed a transcriptomic study on PaWB phytoplasma-infected Paulownia sp. using Solexa/IlluminaM-bM-^@M-^Ys high-throughput digital gene expression (DGE) system. 4 DGE libraries (from 2 virus-infected samples and 2 healthy samples) were constructed, and the gene expression variations between the PaWB phytoplasma-infected (diseased) sample and the corresponding healthy sample were compared. Thousands of differentially expressed genes were obtained by the comparison, and KEGG pathway analysis of these genes suggested that many biological processes were responded to PaWB infection. To investigate the response of Paulownia sp. to PaWB infection, we collected four samples in two groups, namely the tissue cultured group (containing healthy sample TH and diseased sample TD) and field-grown group (containing healthy sample FH and diseased sample FD). Four individual tag libraries from these samples were constructed in parallel. For the gene expression analysis, the digital gene expression (DGE) data of diseased sample were compared to that of healthy sample in each group to obtain the gene expression variations.
2014-10-01 | E-GEOD-42087 | biostudies-arrayexpress
Project description:Recent improvements in relapse/refractory B-cell progenitor acute lymphoblastic leukemia (BCP-ALL) outcome can be attributed, in part, to the emergence of CD19-directed immune-based therapies including chimeric antigen receptor (CAR)-T cell therapy and bi-specific T cell engager (BiTE) therapy. Longitudinal BCP-ALL studies have highlighted individual cases of progressive surface CD19 antigen reduction associated with disease persistence following CAR-T infusion. CD19 genomic deletions and mRNA alternative splicing have been previously shown to confer CAR-T and BiTE resistance, however gene regulatory programs that modulate BCP-ALL CD19 surface antigen abundance remains poorly understood. To address this, we performed genome-wide CRISPR/Cas9 screening to identify positive and negative regulators of surface CD19 abundance in human BCP-ALL as compared to mature B cell neoplasms, chronic lymphocytic leukemia and B cell lymphoma. Here, we show DNA binding protein, ZNF143, occupies the CD19 promoter and transcriptionally regulates CD19 mRNA expression in human B-cell malignancies. Conversely, we show that RNA-binding protein, NUDT21, limits surface CD19 abundance on BCP-ALL. NUDT21 acts to regulate CD19 3’ UTR length and CD19 mRNA stability. NUDT21 mRNA displays high concordance with CD19 mRNA expression in primary healthy and transformed human B cell progenitors. Using primary BCP-ALL patient single cell data, we show that reduction of CD19 mRNA expression following CAR-T therapy coincides with increased NUDT21 mRNA expression. Importantly, NUDT21 ablation sensitizes BCP-ALL to BiTE and CD19 CAR-T killing ex vivo. These findings reveal previously unknown modulators of CD19 antigen abundance that can be applied to therapeutically enhance or determine resistance to CD19-based therapies in B-cell leukemias.
2022-07-27 | GSE190841 | GEO
Project description:Recent improvements in relapse/refractory B-cell progenitor acute lymphoblastic leukemia (BCP-ALL) outcome can be attributed, in part, to the emergence of CD19-directed immune-based therapies including chimeric antigen receptor (CAR)-T cell therapy and bi-specific T cell engager (BiTE) therapy. Longitudinal BCP-ALL studies have highlighted individual cases of progressive surface CD19 antigen reduction associated with disease persistence following CAR-T infusion. CD19 genomic deletions and mRNA alternative splicing have been previously shown to confer CAR-T and BiTE resistance, however gene regulatory programs that modulate BCP-ALL CD19 surface antigen abundance remains poorly understood. To address this, we performed genome-wide CRISPR/Cas9 screening to identify positive and negative regulators of surface CD19 abundance in human BCP-ALL as compared to mature B cell neoplasms, chronic lymphocytic leukemia and B cell lymphoma. Here, we show DNA binding protein, ZNF143, occupies the CD19 promoter and transcriptionally regulates CD19 mRNA expression in human B-cell malignancies. Conversely, we show that RNA-binding protein, NUDT21, limits surface CD19 abundance on BCP-ALL. NUDT21 acts to regulate CD19 3’ UTR length and CD19 mRNA stability. NUDT21 mRNA displays high concordance with CD19 mRNA expression in primary healthy and transformed human B cell progenitors. Using primary BCP-ALL patient single cell data, we show that reduction of CD19 mRNA expression following CAR-T therapy coincides with increased NUDT21 mRNA expression. Importantly, NUDT21 ablation sensitizes BCP-ALL to BiTE and CD19 CAR-T killing ex vivo. These findings reveal previously unknown modulators of CD19 antigen abundance that can be applied to therapeutically enhance or determine resistance to CD19-based therapies in B-cell leukemias.
2022-07-27 | GSE191007 | GEO
Project description:Recent improvements in relapse/refractory B-cell progenitor acute lymphoblastic leukemia (BCP-ALL) outcome can be attributed, in part, to the emergence of CD19-directed immune-based therapies including chimeric antigen receptor (CAR)-T cell therapy and bi-specific T cell engager (BiTE) therapy. Longitudinal BCP-ALL studies have highlighted individual cases of progressive surface CD19 antigen reduction associated with disease persistence following CAR-T infusion. CD19 genomic deletions and mRNA alternative splicing have been previously shown to confer CAR-T and BiTE resistance, however gene regulatory programs that modulate BCP-ALL CD19 surface antigen abundance remains poorly understood. To address this, we performed genome-wide CRISPR/Cas9 screening to identify positive and negative regulators of surface CD19 abundance in human BCP-ALL as compared to mature B cell neoplasms, chronic lymphocytic leukemia and B cell lymphoma. Here, we show DNA binding protein, ZNF143, occupies the CD19 promoter and transcriptionally regulates CD19 mRNA expression in human B-cell malignancies. Conversely, we show that RNA-binding protein, NUDT21, limits surface CD19 abundance on BCP-ALL. NUDT21 acts to regulate CD19 3’ UTR length and CD19 mRNA stability. NUDT21 mRNA displays high concordance with CD19 mRNA expression in primary healthy and transformed human B cell progenitors. Using primary BCP-ALL patient single cell data, we show that reduction of CD19 mRNA expression following CAR-T therapy coincides with increased NUDT21 mRNA expression. Importantly, NUDT21 ablation sensitizes BCP-ALL to BiTE and CD19 CAR-T killing ex vivo. These findings reveal previously unknown modulators of CD19 antigen abundance that can be applied to therapeutically enhance or determine resistance to CD19-based therapies in B-cell leukemias.
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