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RNA sequencing revealed the different transcriptional profiles of long-term cultured and short-term cultured CAR-T cells

ABSTRACT: RNA sequencing revealed the different transcriptional profiles of long-term cultured and short-term cultured CAR-T cells

PROVIDER: PRJNA739669 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR14871064_1.fastq.gz	Fastqsanger.gz
	SRR14871064_2.fastq.gz	Fastqsanger.gz
	SRR14871065_1.fastq.gz	Fastqsanger.gz
	SRR14871065_2.fastq.gz	Fastqsanger.gz
	SRR14871066_1.fastq.gz	Fastqsanger.gz

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RNA sequencing revealed the different transcriptional profiles of long-term cultured and short-term cultured CAR-T cells

Project description:Chimeric antigen receptor (CAR) T cells are known to be potent in recognizing and eliminating tumor cells. They induce marked responses in patients with refractory leukemia and lymphoma, but exhaustion is an inevitable obstacle important barrier for to persistant curative effect. CD19-targeted CAR incorporating the 4-1BB costimulation domain showed a lower tendency toward exhaustion during ex vivo expansion, and downstream differentiation was still observed with a prolonged culture time. We explored the transcriptional changes during the culture process using 4-1BB CD19-targeted CAR-T cells . In contrast to T cells (cultured 5 days in vitro) and short-term cultured (5 days) CAR-T cells, the pathway for regulation of the cytosolic calcium ion concentration was significantly upregulated in the long-term cultured (11 days) group.

2022-04-05 | GSE178570 | GEO

Analysis of the transcriptomic profiles for HER2 CD8+ CAR T cells cultured under short-term resting conditions.

Project description:T cell exhaustion and metabolic dysfunction induced by chronic antigen stimulation within the tumor microenvironment represent formidable obstacles in CAR T cell therapy for solid tumors. Previously, we developed a method to convert effector/exhausted CAR T cells that emerge from repeated antigen stimulation into stem cell memory-like CAR T (termed CAR-iTSCM) cells. These CAR-iTSCM cells exhibit expression of early memory markers and enhanced mitochondrial metabolism after quiescence under culture conditions containing IL-7, CXCL12, and the NOTCH-ligand. However, we noticed that this strategy was effective for CD19 CAR used for blood cancers, but not for HER2 CAR commonly used to treat solid tumors, due to tonic CAR signals. In this study, we developed a new culture condition for CAR iTSCM cells suitable for HER2 CAR and elucidated the mechanism of induction of stemness and mitochondrial fitness. We found that short-term resting condition with IL-7, CXCL12, and pan-tyrosine kinase inhibitor, Dasatinib can induce stem-like CAR T cells at the epigenetic level and enhance mitochondrial metabolism. We identified FOXO1 as a critical regulator in the process of epigenetic and metabolic reprogramming, and anti-tumor activity. We found that overexpression of FOXO1 leads to enhanced mitochondrial metabolism, efficient CAR T cell expansion, and potent anti-tumor effects through epigenetic modifications. These findings suggest that regulating the transcriptional activity of FOXO1 is a promising strategy for engineering effective CAR T cells for treating solid tumors. In this analysis, we aimed to investigate the effects of the resting conditions (IL-7+CXCL12+IGF-I +Dasatinib) on the transcriptomic profiles in 4-1BB-based HER2 CD8+ CAR T cells which exhibit tonic signals-induced exhaustion hallmarks.

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Analysis of the transcriptomic profiles for HER2 CD8+ CAR T cells cultured under short-term resting conditions.

Project description:Analysis of the transcriptomic profiles for HER2 CD8+ CAR T cells cultured under short-term resting conditions.

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Analysis of the epigenetic profiles for HER2 CD8+ CAR T cells cultured under short-term resting conditions and overexpressing FOXO1WT

Project description:T cell exhaustion and metabolic dysfunction induced by chronic antigen stimulation within the tumor microenvironment represent formidable obstacles in CAR T cell therapy for solid tumors. Previously, we developed a method to convert effector/exhausted CAR T cells that emerge from repeated antigen stimulation into stem cell memory-like CAR T (termed CAR-iTSCM) cells. These CAR-iTSCM cells exhibit expression of early memory markers and enhanced mitochondrial metabolism after quiescence under culture conditions containing IL-7, CXCL12, and the NOTCH-ligand. However, we noticed that this strategy was effective for CD19-CAR used for blood cancers, but not for HER2 CAR commonly used to treat solid tumors, due to tonic CAR signals. In this study, we developed a new culture condition for CAR iTSCM cells suitable for HER2 CAR and elucidated the mechanism of induction of stemness and mitochondrial fitness. We found that short-term resting condition with IL-7, CXCL12, and pan-tyrosine kinase inhibitor, Dasatinib can induce stem-like CAR T cells at the epigenetic level and enhance mitochondrial metabolism. We identified FOXO1 as a critical regulator in the process of epigenetic and metabolic reprogramming, and anti-tumor activity. We found that overexpression of FOXO1 leads to enhanced mitochondrial metabolism, efficient CAR T cell expansion, and potent anti-tumor effects through epigenetic modifications. These findings suggest that regulating the transcriptional activity of FOXO1 is a promising strategy for engineering effective CAR T cells for treating solid tumors. In this analysis, we aimed to investigate the effects of the resting conditions (IL-7+CXCL12+IGF-I +Dasatinib) and FOXO1 overexpression on the epigenetic profiles in 4-1BB-based HER2 CAR T cells which exhibit tonic signals-induced exhaustion hallmarks.

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Analysis of the epigenetic profiles for HER2 CD8+ CAR T cells cultured under short-term resting conditions and overexpressing FOXO1WT

Project description:Analysis of the epigenetic profiles for HER2 CD8+ CAR T cells cultured under short-term resting conditions and overexpressing FOXO1WT

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Short term perturbation

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5`aza-dC demethylation of three short term cultured glioblastomas

Project description:Glioblastoma, the most aggressive and least treatable form of malignant glioma, is the most common human brain tumor. Although many regions of allelic loss occur in glioblastomas, relatively few tumor suppressor genes have been found mutated at such loci. To address the possibility that epigenetic alterations are an alternative means of glioblastoma gene inactivation, we coupled pharmacological manipulation of methylation with gene profiling to identify potential methylation-regulated, tumor-related genes. Triplicates of three short-term cultured glioblastomas were exposed to 5μM 5-aza-dC for 96 hours followed by cRNA hybridization to an oligonucleotide microarray (Affymetrix U133A). We based candidate gene selection on bioinformatics, RT-PCR, bisulfite sequencing, methylation-specific PCR and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Two genes identified in this manner, RUNX3 and Testin (TES), were subsequently shown to harbor frequent tumor-specific epigenetic alterations in primary glioblastomas. This overall approach therefore provides a powerful means to identify candidate tumor suppressor genes for subsequent evaluation and may lead to the identification of genes whose epigenetic dysregulation is integral to glioblastoma tumorigenesis. Keywords: expression profile following global demethylation

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