RNA seq analysis of T cells，EpCAM CAR-T cells and rapamycin-pretreated EpCAM CAR-T cells
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ABSTRACT:
PROVIDER: PRJNA683033 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR13208500_1.fastq.gz | Fastqsanger.gz | |||
| SRR13208500_2.fastq.gz | Fastqsanger.gz | |||
| SRR13208501_1.fastq.gz | Fastqsanger.gz | |||
| SRR13208501_2.fastq.gz | Fastqsanger.gz | |||
| SRR13208502_1.fastq.gz | Fastqsanger.gz |
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Project description:Purpose: Compared the differences in the transcriptome of T cells versus EpCAM CAR-T cells and EpCAM CAR-T cells versus rapamycin-pretreated EpCAM CAR-T cells.Methods: Next-generation sequencing (NGS) has revolutionized the systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived transcriptome profiling (RNA-seq) between T cells and EpCAM CAR-T cells，and between EpCAM CAR-T cells and rapamycin-pretreated EpCAM CAR-T cells. Results: RNA-seq showed that CAR-T cells significantly upregulated the expression of exhaustion markers compared to T cells. Gene Set Enrichment Analysis (GSEA) analysis showed that CAR-T cells cells were enriched for signatures of apoptosis. Compared with T cells, Kyoto Encyclopedia of Genes and Genomessignaling(KEGG) pathway analysis showed that the PI3K/AKT/mTOR signaling pathway was significantly enriched. Likewise, GSEA showed that the mTOR signaling pathway was significantly upregulated in CAR-T cells. Next, we analyzed the RNA-seq data between CAR-T cells and rapamycin-pretreated CAR-T cells. RNA-seq showed that rapamycin-treated CAR T down-regulated the expression of exhaustion markers compared with untreated CAR-T cells. GSEA analysis showed that the apoptosis-related gene set was enriched in untreated CAR-T cells.Conclusions:In vitro culture promotes the terminal differentiation of CAR-T cells.CAR-T cells present a highly activated mTOR phenotype. Rapamycin pretreatment prevents CAR-T cells terminal differentiation.
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Project description:Chimeric Antigen Receptor (CAR) T cell therapy shows promise for various diseases. Our studies in humanized mice and non-human primates (NHPs) demonstrate that hematopoietic stem cell (HSCs) modified with anti-HIV CAR achieve lifelong engraftment, providing functional anti-viral CAR-T cells that reduce viral rebound after ART withdrawal. However, T cell exhaustion due to chronic immune activation remains a key obstacle for sustained CAR-T efficacy, necessitating additional measures to achieve functional cure. We recently showed that low dose rapamycin treatment reduced inflammation and improved anti-HIV T cell function in HIV-infected humanized mice. Here, we report that rapamycin improved CAR-T cell function both in vitro and in vivo. In vitro treatment with rapamycin enhanced CAR-T cell mitochondria respiration and cytotoxicity. In vivo treatment with low-dose rapamycin in HIV-infected, CAR-HSC mice decreased chronic inflammation, prevented exhaustion of CAR-T cells and improved CAR-T control of viral replication. RNAseq analysis of CAR-T cells from humanized mice showed that rapamycin downregulated multiple checkpoint inhibitors and the upregulated key survival genes. Mice treated with CAR-HSCs and rapamycin had delayed viral rebound post-ART and reduced HIV reservoir compared to CAR-HSCs alone. These findings suggest that HSCs-based anti-HIV CAR-T combined with rapamycin treatment is a promising approach for treating persistent inflammation and improving immune control of HIV replication.
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