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Trogocytosis and fratricide killing impede MSLN-directed CAR T cell functionality.

ABSTRACT: Successful translation of chimeric antigen receptor (CAR) T cell therapy for the treatment of solid tumors has proved to be troublesome, mainly due to the complex tumor microenvironment promoting T cell dysfunction and antigen heterogeneity. Mesothelin (MSLN) has emerged as an attractive target for CAR T cell therapy of several solid malignancies, including ovarian cancer. To improve clinical response rates with MSLN-CAR T cells, a better understanding of the mechanisms impacting CAR T cell functionality in vitro is crucial. Here, we demonstrated superior cytolytic capacity of CD28-costimulated MSLN-CAR T cells (M28z) relative to 4-1BB-costimulated MSLN-CAR T cells (MBBz). Furthermore, CD28-costimulated MSLN CAR T cells displayed enhanced cytolytic capacity against tumor spheroids with heterogeneous MSLN expression compared to MBBz CAR T cells. In this study, we identified CAR-mediated trogocytosis as a potential impeding factor for successful MSLN-CAR T cell therapy due to fratricide killing and contributing to tumor antigen heterogeneity. Moreover, we link antigen-dependent upregulation of LAG-3 with reduced CAR T cell functionality. Taken together, our study highlights the therapeutic potential and bottlenecks of MSLN-CAR T cells, providing a rationale for combinatorial treatment strategies.

SUBMITTER: Schoutrop E

PROVIDER: S-EPMC9313125 | biostudies-literature |

REPOSITORIES: biostudies-literature

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CAR T cell trogocytosis and cooperative killing regulate tumour antigen escape.

Project description:Chimeric antigen receptors (CARs) are synthetic antigen receptors that reprogram T cell specificity, function and persistence1. Patient-derived CAR T cells have demonstrated remarkable efficacy against a range of B-cell malignancies1-3, and the results of early clinical trials suggest activity in multiple myeloma4. Despite high complete response rates, relapses occur in a large fraction of patients; some of these are antigen-negative and others are antigen-low1,2,4-9. Unlike the mechanisms that result in complete and permanent antigen loss6,8,9, those that lead to escape of antigen-low tumours remain unclear. Here, using mouse models of leukaemia, we show that CARs provoke reversible antigen loss through trogocytosis, an active process in which the target antigen is transferred to T cells, thereby decreasing target density on tumour cells and abating T cell activity by promoting fratricide T cell killing and T cell exhaustion. These mechanisms affect both CD28- and 4-1BB-based CARs, albeit differentially, depending on antigen density. These dynamic features can be offset by cooperative killing and combinatorial targeting to augment tumour responses to immunotherapy.

| S-EPMC6707377 | biostudies-literature

CAR T cell trogocytosis and cooperative killing regulate tumour antigen escape

Project description:Chimeric antigen receptors (CARs) are synthetic receptors for antigen that reprogram T cell specificity, function and persistence. Patient-derived CAR T cells have demonstrated remarkable efficacy against a range of B-cell malignancies, and early trial results suggest activity in multiple myeloma. Despite high complete response rates, relapses occur in a large fraction of patients, some of which are antigen-negative and others antigen-low. Unlike mechanisms resulting in complete and permanent antigen loss, those leading to antigen-low tumour escape remain obscure. In murine leukaemia models, we show that CARs provoke reversible antigen loss through trogocytosis, an active process whereby target antigen is transferred to T cells, thereby decreasing target density on tumour cells and abating T cell activity by promoting fratricide killing and exhaustion. These mechanisms affect both CD28 and 4-1BB-based CARs, albeit differentially, depending on antigen density. They can be offset by cooperative killing and combinatorial targeting.

2019-03-27 | GSE126753 | GEO

Reversible Transgene Expression Reduces Fratricide and Permits 4-1BB Costimulation of CAR T Cells Directed to T-cell Malignancies.

Project description:T cells expressing second-generation chimeric antigen receptors (CARs) specific for CD5, a T-cell surface marker present on normal and malignant T cells, can selectively kill tumor cells. We aimed to improve this killing by substituting the CD28 costimulatory endodomain (28.z) with 4-1BB (BB.z), as 28.z CD5 CAR T cells rapidly differentiated into short-lived effector cells. In contrast, 4-1BB costimulation is known to promote development of the central memory subpopulation. Here, we found BB.z CD5 CAR T cells had impaired growth compared with 28.z CD5.CAR T cells, due to increased T-cell-T-cell fratricide. We demonstrate that TRAF signaling from the 4-1BB endodomain upregulated the intercellular adhesion molecule 1, which stabilized the fratricidal immunologic synapse between CD5 CAR T cells. As the surviving BB.z CD5 CAR T cells retained the desired central memory phenotype, we aimed to circumvent the 4-1BB-mediated toxicity using a regulated expression system that reversibly inhibits CAR expression. This system minimized CAR signaling and T-cell fratricide during in vitro expansion in the presence of a small-molecule inhibitor, and restored CAR expression and antitumor function of transduced T cells in vivo These studies reveal a mechanism by which 4-1BB costimulation impairs expansion of CD5 CAR T cells and offer a solution to mitigate this toxicity. Cancer Immunol Res; 6(1); 47-58. ©2017 AACR.

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Trogocytosis by Entamoeba histolytica contributes to cell killing and tissue invasion.

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CAR T cell trogocytosis and cooperative killing regulate tumour antigen escape

Project description:CAR T cell trogocytosis and cooperative killing regulate tumour antigen escape

| PRJNA523212 | ENA

Low-affinity CAR T cells exhibit reduced trogocytosis, preventing rapid antigen loss, and increasing CAR T cell expansion.

Project description: Not available

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Direct and high-throughput assays for human cell killing through trogocytosis by Entamoeba histolytica.

Project description:Entamoeba histolytica is the causative agent of amoebiasis. Pathogenesis is associated with profound damage to human tissues. We previously showed that amoebae kill human cells through trogocytosis. Trogocytosis is likely to underlie tissue damage during infection, although the mechanism is still unknown. Trogocytosis is difficult to assay quantitatively, which makes it difficult to study. Here, we developed two new, complementary assays to measure trogocytosis by quantifying human cell death. One assay uses CellTiterGlo, a luminescent readout for ATP, as a proxy for cell death. We found that the CellTiterGlo could be used to detect death of human cells after co-incubation with amoebae, and that it was sensitive to inhibition of actin or the amoeba surface Gal/GalNAc lectin, two conditions that are known to inhibit amoebic trogocytosis. The other assay uses two fluorescent nuclear stains to directly differentiate live and dead human cells by microscopy, and is also sensitive to inhibition of amoebic trogocytosis through interference with actin. Both assays are simple and inexpensive, can be used with suspension and adherent human cell types, and are amenable to high-throughput approaches. These new assays are tools to improve understanding of trogocytosis and amoebiasis pathogenesis.

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An "off-the-shelf" fratricide-resistant CAR-T for the treatment of T cell hematologic malignancies.

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| S-EPMC6102094 | biostudies-literature

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