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Pooled screening identifies combinatorial CAR signaling domains for next-generation CAR-M immunotherapies [scRNA-seq]

ABSTRACT: Pooled screening identifies combinatorial CAR signaling domains for next-generation CAR-M immunotherapies [scRNA-seq]

PROVIDER: PRJNA1221813 | ENA |

REPOSITORIES: ENA

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Pooled screening identifies combinatorial CAR signaling domains for next-generation CAR-M immunotherapies

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Pooled screening identifies combinatorial CAR signaling domains for next-generation CAR-M immunotherapies [RNA-Seq]

Project description:Pooled screening identifies combinatorial CAR signaling domains for next-generation CAR-M immunotherapies [RNA-Seq]

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Pooled screening identifies combinatorial CAR signaling domains for next-generation CAR-M immunotherapies [scCAR-seq]

Project description:Chimeric antigen receptor-engineered macrophages (CAR-Ms) hold great promise for solid tumor immunotherapy. The intracellular domains ICDs of CARs determine the phenotypic output of therapeutic macrophages but remain largely unexplored. Here, we constructed a library of CARs containing 131 unique signaling domains derived from native immune receptors and revealed 17 ICDs that enhanced macrophage phagocytosis, the inflammatory response and tumor infiltration in vitro and in vivo. We further developed a scalable 3’ barcode technology, CARode, to uniquely label and trace ICD variants in a large-scale combinatorial signaling domain CAR library and applied it to perform single-cell RNA sequencing and single-cell CAR analysis to measure the synergetic effects of ICD combinations in promoting macrophage activation. This platform includes a novel CD40-LY9-FCRL1 chimeric receptor that modulates the tumor microenvironment and improves solid tumor clearance. In conclusion, pooled screening facilitates the discovery of complex ICD constructs to program macrophage functions for therapeutic applications.

2025-02-17 | GSE289201 | GEO

Pooled screening identifies combinatorial CAR signaling domains for next-generation CAR-M immunotherapies [scRNA-seq]

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Pooled screening identifies combinatorial CAR signaling domains for next-generation CAR-M immunotherapies [RNA-Seq]

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Pooled screening identifies combinatorial CAR signaling domains for next-generation CAR-M immunotherapies

Project description:This SuperSeries is composed of the SubSeries listed below.

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Dissecting the role of CAR signaling architectures on T cell activation and persistence using pooled screening and single-cell sequencing

Project description:Chimeric antigen receptor (CAR) T cells represent a promising approach for cancer treatment, yet challenges remain such as limited efficacy due to a lack of T cell persistence. Given its critical role in promoting and modulating T cell responses, it is crucial to understand how alterations in the CAR signaling architecture influence T cell function. Here, we designed a combinatorial CAR signaling domain library and performed repeated antigen stimulation assays, pooled screening and single-cell sequencing to investigate T-cell responses triggered by different CAR architectures. Parallel comparisons of CAR variants, at early, middle and late timepoints during chronic antigen stimulation systematically assessed the impact of modifying signaling domains on T cell activation and persistence. Our data reveal the predominant influence of membrane-proximal domains in driving T cell phenotype. Additionally, we highlight the critical role of CD40 costimulation in promoting potent and persistent T cell responses, followed by CTLA4, which induces a long-term cytotoxic phenotype. This work deepens the understanding of CAR T cell biology and may be used to guide the future engineering of CAR T cell therapies.

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Pooled screening for CAR function in glioblastoma

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Pooled screening for CAR function in glioblastoma

Project description:Human primary CD3+ T cells expressing IL13Ra2-targeted chimeric antigen receptors (CARs) that had been identified through high throughput pooled screening were rechallenged with an IL13Ra2+ human glioblastoma cell line , then stained with CITESEQ antibodies indicative of T cell memory and exhaustion state and prepared for single cell sequencing in order to determine their phenotypic responses to chronic tumor challenge relative to that of a clinical standard CAR.

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