Epigenetic Suppression of Transgenic T-cell Receptor Expression via Gamma-Retroviral Vector Methylation in Adoptive Cell Transfer Therapy
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA644286 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR12150000_1.fastq.gz | Fastqsanger.gz | |||
| SRR12150000_2.fastq.gz | Fastqsanger.gz | |||
| SRR12150001_1.fastq.gz | Fastqsanger.gz | |||
| SRR12150001_2.fastq.gz | Fastqsanger.gz | |||
| SRR12150002_1.fastq.gz | Fastqsanger.gz |
Items per page: 5 1 - 5 of 96 |
Similar Datasets
Project description:Transgenic T-cell receptor (TCR) adoptive cell therapies recognizing tumor antigens are associated with robust initial response rates, but frequent disease relapse. This usually occurs in the setting of poor long-term persistence of cells expressing the transgenic TCR, generated using murine stem cell virus (MSCV) -retroviral vectors. Analysis of clinical transgenic adoptive cell therapy products in vivo revealed that despite strong persistence of the transgenic TCR DNA sequence over time, its expression was profoundly decreased over time at the RNA and protein levels. Patients with the greatest degrees of expression suppression displayed significant increases in DNA methylation over time within the MSCV promoter region, as well as progressive increases in DNA methylation within the entire MSCV vector over time. These increases in vector methylation occurred independently of its integration site within the host genomes. These results have significant implications for the design of future viral-vector gene engineered adoptive cell transfer therapies.
2020-07-06 | GSE153820 | GEO
Project description:Transgenic T-cell receptor (TCR) adoptive cell therapies recognizing tumor antigens are associated with robust initial response rates, but frequent disease relapse. This usually occurs in the setting of poor long-term persistence of cells expressing the transgenic TCR, generated using murine stem cell virus (MSCV) γ-retroviral vectors. Analysis of clinical transgenic adoptive cell therapy products in vivo revealed that despite strong persistence of the transgenic TCR DNA sequence over time, its expression was profoundly decreased over time at the RNA and protein levels. Patients with the greatest degrees of expression suppression displayed significant increases in DNA methylation over time within the MSCV promoter region, as well as progressive increases in DNA methylation within the entire MSCV vector over time. These increases in vector methylation occurred independently of its integration site within the host genomes. These results have significant implications for the design of future viral vector gene-engineered adoptive cell transfer therapies. SIGNIFICANCE: Cellular immunotherapies' reliance on retroviral vectors encoding foreign genetic material can be vulnerable to progressive acquisition of DNA methylation and subsequent epigenetic suppression of the transgenic product in TCR adoptive cell therapy. This must be considered in the design of future generations of cellular immunotherapies for cancer.This article is highlighted in the In This Issue feature, p. 1611.
| S-EPMC7641915 | biostudies-literature
Project description:Gene transfer of distinct cocktails of transcription factors enables the targeted modification of cell fate. Reprogramming somatic cells to induced pluripotent stem cells (iPSCs) via ectopic expression of the transcription factors Oct4, Sox2, Klf4 and c-Myc allows creation of pluripotent cells with the potential to differentiate into any cell type of the body. To achieve a high degree of biosafety for future application of converted cells, technologies are required to express the required transcription factors without affecting the target cell genome. Retroviral vectors are widely employed gene transfer vehicles and can be converted into transient gene delivery vehicles by mutation of the viral integrase, thus avoiding risks associated with retroviral integration. We improved the retroviral episome transfer system by identifying the most suitable vector architecture, by enhancing episomal expression and by tuning the activity of the transcription factor Oct4. Repeated episomal transductions allowed establishment of stable, fully reversible expression levels over time. Providing proof-of-principle in the reprogramming context, we were able to generate human iPSCs following retroviral episomal transfer of Oct4 in fibroblasts stably expressing Klf4, Sox2 and c-Myc, with a high percentage of clones not carrying residually integrated vector copies.
2015-05-06 | GSE57378 | GEO
Project description:Adoptive T-Cell therapy is being considered as a promising method for cancer treatment. In this approach, patient's T cells are isolated, modified, expanded, and administered back to the patient. Modifications may include adding specific T cell receptors (TCR) or chimeric antigen receptors (CAR) to the isolated cells by using retroviral vectors. PG13 cells, derivatives of NIH3T3 mouse fibroblasts, are being used to stably produce retroviral vectors that transduce the T cells. PG13 cells are anchorage-dependent cells that grow in roller bottles or cell factories and lately also in fixed bed bioreactors to produce the needed viral vector. To scale up viral vector production, PG13 cells were propagated on microcarriers in a stirred tank bioreactor utilizing an alternating tangential flow perfusion system. Microcarriers are 10 µm - 0.5 mm beads that support the attachment of cells and are suspended in the bioreactor that provides controlled growth conditions. As a result, growth parameters, such as dissolved oxygen concentration, pH, and nutrients are monitored and continuously controlled. There were no detrimental effects on the specific viral vector titer or on the efficacy of the vector in transducing the T cells of several patients. Viral vector titer increased throughout the 11 days perfusion period, a total of 4.8 × 1011 transducing units (TU) were obtained with an average titer of 4.4 × 107 TU/mL and average specific productivity of 10.3 (TU) per cell, suggesting that this method can be an efficient way to produce large quantities of active vector suitable for clinical use.
| S-EPMC6028049 | biostudies-literature
Project description:Gene transfer of distinct cocktails of transcription factors enables the targeted modification of cell fate. Reprogramming somatic cells to induced pluripotent stem cells (iPSCs) via ectopic expression of the transcription factors Oct4, Sox2, Klf4 and c-Myc allows creation of pluripotent cells with the potential to differentiate into any cell type of the body. To achieve a high degree of biosafety for future application of converted cells, technologies are required to express the required transcription factors without affecting the target cell genome. Retroviral vectors are widely employed gene transfer vehicles and can be converted into transient gene delivery vehicles by mutation of the viral integrase, thus avoiding risks associated with retroviral integration. We improved the retroviral episome transfer system by identifying the most suitable vector architecture, by enhancing episomal expression and by tuning the activity of the transcription factor Oct4. Repeated episomal transductions allowed establishment of stable, fully reversible expression levels over time. Providing proof-of-principle in the reprogramming context, we were able to generate human iPSCs following retroviral episomal transfer of Oct4 in fibroblasts stably expressing Klf4, Sox2 and c-Myc, with a high percentage of clones not carrying residually integrated vector copies. RNA obtained from human embryonic stem cells, fibroblasts and induced pluripotent stem cell lines reprogrammed with the mentioned reprogramming protocol
2015-05-06 | E-GEOD-57378 | biostudies-arrayexpress
Project description:Smad7 is a member of the TGF-B superfamily that plays a signigicant role in fate determination of hematopoietic stem cells. This experiment compares expression between megakaryotic and fibroblast cell lines that have been transduced with either Smad7 recombinant retroviral vector or retroviral vector alone. Experiment Overall Design: this experiment include 4 samples and 30 replicates
2008-06-12 | E-GEOD-3771 | biostudies-arrayexpress
Project description:Safety assessment in retroviral vector-mediated gene therapy remains challenging. In clinical trials for different blood and immune disorders, insertional mutagenesis led to myeloid and lymphoid leukemia. We previously developed the In Vitro Immortalization Assay (IVIM) and Surrogate Assay for Genotoxicity Assessment (SAGA) for pre-clinical genotoxicity prediction of integrating vectors. Murine hematopoietic stem and progenitor cells (mHSPC) transduced with mutagenic vectors acquire a proliferation advantage under limiting dilution (IVIM) and activate stem cell- and cancer-related transcriptional programs (SAGA). However, both assays present an intrinsic myeloid bias due to culture conditions. To detect lymphoid mutants, we differentiated mHSPC to mature T cells and analyzed their phenotype, insertion site pattern, and gene expression changes after transduction with retroviral vectors. Mutagenic vectors induced a block in differentiation at an early progenitor stage (double-negative 2) compared to fully differentiated untransduced mock cultures. Arrested samples harbored high-risk insertions close to Lmo2, frequently observed in clinical trials with severe adverse events. Lymphoid insertional mutants displayed a unique gene expression signature identified by the machine learning algorithm of SAGA. The gene expression-based highly sensitive molecular readout will broaden our understanding of vector-induced oncogenicity and help in pre-clinical prediction of retroviral genotoxicity.
2023-09-21 | GSE241155 | GEO
Project description:Safety assessment in retroviral vector-mediated gene therapy remains challenging. In clinical trials for different blood and immune disorders, insertional mutagenesis led to myeloid and lymphoid leukemia. We previously developed the In Vitro Immortalization Assay (IVIM) and Surrogate Assay for Genotoxicity Assessment (SAGA) for pre-clinical genotoxicity prediction of integrating vectors. Murine hematopoietic stem and progenitor cells (mHSPC) transduced with mutagenic vectors acquire a proliferation advantage under limiting dilution (IVIM) and activate stem cell- and cancer-related transcriptional programs (SAGA). However, both assays present an intrinsic myeloid bias due to culture conditions. To detect lymphoid mutants, we differentiated mHSPC to mature T cells and analyzed their phenotype, insertion site pattern, and gene expression changes after transduction with retroviral vectors. Mutagenic vectors induced a block in differentiation at an early progenitor stage (double-negative 2) compared to fully differentiated untransduced mock cultures. Arrested samples harbored high-risk insertions close to Lmo2, frequently observed in clinical trials with severe adverse events. Lymphoid insertional mutants displayed a unique gene expression signature identified by the machine learning algorithm of SAGA. The gene expression-based highly sensitive molecular readout will broaden our understanding of vector-induced oncogenicity and help in pre-clinical prediction of retroviral genotoxicity.
2023-09-21 | GSE241146 | GEO
Project description:Cultured cancer cells exhibit substantial phenotypic heterogeneity when measured in a variety of ways such as sensitivity to drugs or the capacity to grow under various conditions. Among these, the ability to exhibit anchorage-independent cell growth (colony forming capacity in semisolid media) has been considered to be fundamental in cancer biology because it has been connected with tumor cell aggressiveness in vivo such as tumorigenic and metastatic potentials, and also utilized as a marker for in vitro transformation. Although multiple genetic factors for anchorage-independence have been identified, the molecular basis for this capacity is still largely unknown. To investigate the molecular mechanisms underlying anchorage-independent cell growth, we have used genome-wide DNA microarray studies to develop an expression signature associated with this phenotype. Using this signature, we identify a program of activated mitochondrial biogenesis associated with the phenotype of anchorage-independent growth and importantly, we demonstrate that this phenotype predicts potential for metastasis in primary breast and lung tumors. Keywords: c-Myc or v-Src retroviral vector-infected immortalized mouse embryonic fibroblasts.
2009-04-10 | GSE15161 | GEO