Ontology highlight
ABSTRACT:
PROVIDER: PRJNA1237884 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR32762567_1.fastq.gz | Fastqsanger.gz | |||
| SRR32762567_2.fastq.gz | Fastqsanger.gz | |||
| SRR32762568_1.fastq.gz | Fastqsanger.gz | |||
| SRR32762568_2.fastq.gz | Fastqsanger.gz | |||
| SRR32762569_1.fastq.gz | Fastqsanger.gz |
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Project description:The prognosis of infant B-cell acute lymphoblastic leukemia (iB-ALL) remains dismal, especially in patients harboring the MLL-AF4 (KTM2A-AFF1) rearrangement, which arises prenatally in early hematopoietic stem/progenitor cells (HSPCs). MLL-AF4+ B-ALL shows a bimodal localization of the MLL gene breakpoint within the MLL break cluster region, and two subgroups of patients based on the gene expression pattern of the HOXA/MEIS cluster have been identified. The pathogenic mechanisms in MLL-AF4+ B-ALL are challenging to study functionally due to the absence of faithful human cellular models recapitulating the disease phenotype and latency. Here, we assess the molecular contribution and leukemogenic capacity of MLL breakpoints occurring in either intron 10 (MLLi10, centromeric) or intron 12 (MLLi12, telomeric) in ontogenically-different human HSPCs sourced prenatally (fetal liver) and neonatally (cord blood). CRISPR-Cas9-induced MLL-AF4 (MA) targeting either MLLi10 (Mi10A) or MLLi12 (Mi12A) causes MA-driven in vitro myeloid immortalization in both fetal liver- and cord blood-CD34+ HSPCs. The cellular ontogeny and the location of the MLL breakpoint influenced the capacity of MLL-edited CD34+ HSPCs to initiate pro-B-ALL in vivo, which faithfully recapitulated the molecular, transcriptomic and methylome profiles of patients with primary MA+ iB-ALL. This dataset contains the DNA methylation information (Illumina MethylationEPIC Beadchip platform) of MLL-edited CD34+ cells (i10 or i12). BCPs, used as controls, were obtained from E-MTAB-8505
2023-10-29 | E-MTAB-12806 | biostudies-arrayexpress
Project description:Site-specific correction of a point mutation causing a monogenic disease in autologous hematopoietic stem and progenitor cells (HSPCs) can be used as a treatment of inherited disorders of the blood cells. Sickle Cell Disease (SCD) is an ideal model to investigate the potential use of gene editing to transvert a single point mutation at the β-globin locus (HBB). We compared the activity of ZFNs and CRISPR/Cas9 for editing and homologous donor templates delivered as single-stranded oligodeoxynucleotides (ssODN), adeno-associated virus serotype 6 (AAV6), integrase-deficient lentiviral vectors (IDLV) and adenovirus 5/35 serotype (Ad5/35) to transvert the base-pair responsible for SCD in HBB in primary human CD34+ HSPCs. We found that the ZFNs and Cas9 directed similar frequencies of nuclease activity. In vitro, AAV6 led to the highest frequencies of homology-directed repair (HDR), but levels of base-pair transversions were significantly reduced when analyzing cells in vivo in immune-deficient mouse xenografts, with similar frequencies achieved with either AAV6 or ssODN. AAV6 also caused significant impairment of colony-forming progenitors and human cell engraftment. Gene correction in engrafting hematopoietic stem cells may be limited by the capacity of the cells to mediate HDR, suggesting additional manipulations may be needed for high efficiency gene correction in HSPCs
2019-05-18 | GSE131387 | GEO
Project description:CD3δ SCID is a devastating inborn error of immunity caused by mutations in CD3D, encoding the invariant CD3δ chain of the CD3/TCR complex necessary for normal thymopoiesis. We demonstrate an adenine base editing (ABE) strategy to restore CD3δ in autologous hematopoietic stem and progenitor cells (HSPC). Delivery of mRNA encoding a laboratory-evolved ABE and guide RNA into CD3δ SCID patient’s HSPCs resulted in 71.2±7.85% (n=3) correction of the pathogenic mutation. Edited HSPCs differentiated in artificial thymic organoids produced mature T cells exhibiting diverse TCR repertoires and TCR-dependent functions. Edited human HSPCs transplanted into immunodeficient mice showed 88% reversion of the CD3D defect in human CD34+ cells isolated from mouse bone marrow after 16 weeks, indicating correction of long-term repopulating HSCs. These findings demonstrate preclinical efficacy of ABE in HSPC for the treatment of CD3δ SCID, providing a foundation for the development of a one-time treatment for CD3δ SCID patients.
2023-03-20 | GSE220611 | GEO
Project description:Chronic granulomatous disease (CGD) is a severe inborn error of immunity (IEI) caused by defects in the NADPH oxidase. The best current treatment option for patients with CGD, allogeneic hematopoietic stem cell (HSC) transplantation, is associated with severe adverse effects such as graft-versus-host disease, highlighting a need for improved treatment options based on transplantation of autologous ex vivo gene-edited HSCs. Here, we generate CRISPR/Cas9-based gene editing strategies for the correction of two CGD-causing variants; CYBA c.287+1G>T (p22phox) causing autosomal CGD and CYBB c.252G>A (NOX2) causing X-linked CGD. We find that rAAV6 outperforms ssODNs and IDLVs as HDR repair templates in CD34+ hematopoietic stem and progenitor cells (HSPCs), and we optimize gene editing strategies further by including mRNA-encoded inhibitors/effectors. In addition, we develop a near-universal gene editing strategy for X-CGD by targeted integration of a truncated CYBB cDNA, covering 86% of X-CGD patients, and show functional ROS production in edited cells. We find prevalent off-target editing and chromosomal translocations that severely decreased the ability of gene edited HSPCs to engraft in immunodeficient mice, however, by limiting off-target editing through the use of a high-fidelity Cas9, we can show that we can substantially rescue the multilineage engraftment potential of the gene edited HSPCs. To further improve safety, we finally we develop a novel paired D10A Cas9n gene editing approach targeting CYBB. We demonstrate that using this D10A Cas9 approach, we can retain high on-target efficacy while we do not detect any off-target editing or translocations between on- or off-target cleavage sites. Collectively, we bring new insights into to safety and off-target effects of gene editing approaches targeting CYBA and CYBB, highlighting key challenges of these approaches while offering a potentially curative D10A Cas9n-based treatment option for patients with CGD with improved safety.
2025-02-24 | GSE287370 | GEO
Project description:Ex-vivo gene editing in T cells and hematopoietic stem/progenitor cells (HSPCs) holds promise for treating diseases by non-homologous end joining (NHEJ) gene disruption or homology-driven repair (HDR) gene correction. Gene editing encompasses delivery of nucleases by electroporation and, when aiming to HDR, of a DNA template often provided by viral vectors. Whereas HSPCs activate robust p53-dependent DNA damage response (DDR) upon editing, the responses triggered in T cells remain poorly characterized. Here, we performed comprehensive multi-omics analyses and found that electroporation is the culprit of cytotoxicity in T cells, causing death and cell cycle delay, perturbing metabolism and inducing inflammatory response. Nuclease delivery by lipid nanoparticles (LNPs) nearly abolished cell death and ameliorated cell growth, improving tolerance to the procedure and yielding higher number of edited cells compared to electroporation. Transient transcriptomic changes upon LNP treatment were mostly caused by cellular loading with exogenous cholesterol, whose potentially detrimental impact could be overcome by limiting exposure. Notably, LNP-based HSPC editing dampened p53 pathway induction and supported higher reconstitution by long-term repopulating HSPCs compared to electroporation, reaching similar editing efficiencies. Overall, LNPs may allow efficient and stealthier ex-vivo gene editing in hematopoietic cells for treatment of human diseases.
2024-01-26 | PXD037529 | Pride
Project description:We have developed a therapeutic strategy for beta-hemoglobinopathies aimed at reactivating fetal hemoglobin expression in red blood cells derived from human hematopoietic stem/progenitor cells edited with CRISPR/Cas9 nucleases, cytidine or adenine base editors targeting the fetal gamma-globin promoters. Here, we report the transcriptomic changes occurring in human hematopoietic stem/progenitor cells (obtained from healthy donors) 48 h after transfection with CRISPR/Cas9 nucleases, cytidine or adenine base editors.
2022-09-06 | GSE191135 | GEO
Project description:Purpose: Illumina next-generation sequencing (NGS) has been used to interrogate the transcriptome profiling (bulk RNA-seq) of primary human HSPCs in the presence and absence of RIOK2. Primary human hematopoietic stem and progenitor cells (HSPCs) isolated from 3 different donors were genome edited to obtain knockdown (KD) and knockout (KO) of RIOK2. The genome edited HSPCs were then differentiated for 48 hours and their total RNA was isolated to perform cDNA synthesis and bulk RNA sequencing. The overall goal of this study was to investigate the global alterations in gene expressions with dose-dependent loss of RIOK2 in primary human HSPCs, that would expand our understanding of RIOK2-dependent transcriptomic changes involved in hematopoietic differentiation.
2021-12-02 | GSE185922 | GEO
Project description:Transforming growth factor-β (TGFβ) is a potent inhibitor of hematopoietic stem cell (HSC) proliferation. However, the precise mechanism for this effect is unknown. Here, we have identified the transcription factor Gata2, previously described as an important regulator of HSC function, as an early and direct target gene for TGFβ-induced Smad signaling in hematopoietic stem and progenitor cells (HSPCs). Interestingly, TGFβ-induced Gata2 upregulation is critical for subsequent transcriptional activation of the TGFβ signaling effector molecule p57 and resulting growth arrest of HSPCs. Importantly, both Gata2 and p57 are abundantly expressed in freshly isolated highly purified HSCs, demonstrating the relevance of this circuit in HSC regulation within the HSC niche. Our results connect key molecules involved in HSC self-renewal and reveal a functionally relevant network regulating proliferation of primitive hematopoietic cells. To identify early gene targets of TGFβ signaling in hematopoietic progenitor cells, we performed high-throughput gene expression profiling of a primitive murine hematopoietic cell line. One of the revealed target genes was the transcription factor Gata2, which became the base for the rest of the study. Three independent RNA harvests were separately analyzed. Untreated cells were used as controls to the 10ng/ml TGFb-treated cells.
2016-09-01 | E-GEOD-73567 | biostudies-arrayexpress
Project description:β-thalassemia is a highly prevalent monogenic recessive disease caused by mutations affecting the synthesis of the adult hemoglobin β-chains. Transplantation of autologous, genetically modified hematopoietic stem/progenitor cells (HSPCs) is an attractive therapeutic option. However, current gene therapy strategies based on the use of lentiviral vectors or CRISPR/Cas9 nuclease are not equally effective in all the patients and/or raise safety concerns. Base editing (BE), a new CRISPR/Cas9 derived genome editing tool, allows the effective introduction of point mutations at precise locations within the genome without generating double strand breaks. The two β0 mutations CD39 (CAG>TAG) and IVS2-1 (G>A) are among the most common and severe β-thalassemia mutations in the Mediterranean area and Middle East. We exploited the capacity of BE to correct these mutations in HSPCs from β-thalassemia patients. We demonstrated that red blood cells in vitro derived from edited HSPCs exhibited high β-globin levels and that the delayed erythroid differentiation typically observed in β-thalassemic cell cultures was corrected by our treatment. Finally, xenotransplantation experiments showed base editing in HSCs and correction of the β-thalassemic phenotype in vivo. Overall, our study provides in vitro and in vivo proof of efficacy of a BE approach to treat patients with prevalent and severe β-thalassemia mutations.
2025-05-05 | GSE273814 | GEO
Project description:β-thalassemia is a highly prevalent monogenic recessive disease caused by mutations affecting the synthesis of the adult hemoglobin β-chains. Transplantation of autologous, genetically modified hematopoietic stem/progenitor cells (HSPCs) is an attractive therapeutic option. However, current gene therapy strategies based on the use of lentiviral vectors or CRISPR/Cas9 nuclease are not equally effective in all the patients and/or raise safety concerns. Base editing (BE), a new CRISPR/Cas9 derived genome editing tool, allows the effective introduction of point mutations at precise locations within the genome without generating double strand breaks. The two β0 mutations CD39 (CAG>TAG) and IVS2-1 (G>A) are among the most common and severe β-thalassemia mutations in the Mediterranean area and Middle East. We exploited the capacity of BE to correct these mutations in HSPCs from β-thalassemia patients. We demonstrated that red blood cells in vitro derived from edited HSPCs exhibited high β-globin levels and that the delayed erythroid differentiation typically observed in β-thalassemic cell cultures was corrected by our treatment. Finally, xenotransplantation experiments showed base editing in HSCs and correction of the β-thalassemic phenotype in vivo. Overall, our study provides in vitro and in vivo proof of efficacy of a BE approach to treat patients with prevalent and severe β-thalassemia mutations.
2025-05-05 | GSE292238 | GEO