A Versatile Safeguard for Chimeric Antigen Receptor T-Cell Immunotherapies

Valton, Julien; Guyot, Valérie; Boldajipour, Bijan; Sommer, Cesar; Pertel, Thomas; Juillerat, Alexandre; Duclert, Aymeric; Sasu, Barbra J.; Duchâteau, Philippe; Poirot, Laurent

doi:10.1038/s41598-018-27264-w

Cited by 29 publications

(35 citation statements)

References 30 publications

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“…Furthermore, it did not affect our ability to purify and efficiently deplete TRAC CAR T cells with the FDA-approved drug Rituximab (RTX), as demonstrated previously using a lentiviral-based vectorization approach ( Supplementary Figs. 1 and 2 36 ).…”

Section: Resultsmentioning

confidence: 97%

“…To do so, we generated three different AAV6 promoter-less repair matrices. The first matrix, TRAC CARm , was designed to integrate an anti-CD22 CAR expression cassette at the TRAC locus 26,36 . This CAR architecture contains integrated purification and safeguard domains that allow purification and Rituximab-dependent depletion of TRAC CAR T cells ( Supplementary Fig.…”

Section: Resultsmentioning

confidence: 99%

“…This CAR architecture contains integrated purification and safeguard domains that allow purification and Rituximab-dependent depletion of TRAC CAR T cells ( Supplementary Fig. 1a) 36 . The second and third matrices, IL2rα IL12m and PDCD1 IL12m , were designed to integrate the IL-12P70 heterodimer (formed by IL-12A p35 and IL-12B p40 subunits) along with a truncated lowaffinity nerve growth factor receptor (ΔLNGFR) surface marker 37 at the IL2rα and PDCD1 loci, respectively.…”

Section: Resultsmentioning

confidence: 99%

“…Thus, we hypothesized that the IL-12P70mediated increase in tumor cell clearance rate might translate into long-term tumor cell control by TRAC CAR T cells, even after repeated tumor cell challenges. We performed an in vitro serial killing assay 36 that allowed us to repeatedly challenge TRAC CAR T cells with daily doses of tumor cells for 8 consecutive days (Fig. 6b).…”

Section: Secretion Of Il-12p70 Improves Trac Car T Cells Activitymentioning

confidence: 99%

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Repurposing endogenous immune pathways to tailor and control chimeric antigen receptor T cell functionality

et al. 2019

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Endowing chimeric antigen receptor (CAR) T cells with additional potent functionalities holds strong potential for improving their antitumor activity. However, because potency could be deleterious without control, these additional features need to be tightly regulated. Immune pathways offer a wide array of tightly regulated genes that can be repurposed to express potent functionalities in a highly controlled manner. Here, we explore this concept by repurposing TCR, CD25 and PD1, three major players of the T cell activation pathway. We insert the CAR into the TCRα gene (TRAC CAR), and IL-12P70 into either IL2Rα or PDCD1 genes. This process results in transient, antigen concentration-dependent IL-12P70 secretion, increases TRAC CAR T cell cytotoxicity and extends survival of tumor-bearing mice. This gene network repurposing strategy can be extended to other cellular pathways, thus paving the way for generating smart CAR T cells able to integrate biological inputs and to translate them into therapeutic outputs in a highly regulated manner.

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Section: Resultsmentioning

confidence: 97%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Secretion Of Il-12p70 Improves Trac Car T Cells Activitymentioning

confidence: 99%

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Repurposing endogenous immune pathways to tailor and control chimeric antigen receptor T cell functionality

et al. 2019

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“…In addition, the use of corticosteroids has been shown to alleviate the neurotoxic symptoms of CRS in some patients (13,15). Although these treatments are promising, they failed to prevent CRS-associated death in some cases (16,17), and treatment with an antibody can cause further toxicities such as infection and hepatic dysfunction (18). Thus, alternative strategies are needed to control CRS symptoms better or even to prevent CRS in patients receiving CAR T-cell therapies.…”

mentioning

confidence: 99%

Granulocyte–macrophage colony-stimulating factor inactivation in CAR T-cells prevents monocyte-dependent release of key cytokine release syndrome mediators

Sachdeva

Duchâteau²,

Depil³

et al. 2019

Journal of Biological Chemistry

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126

132

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Edited by Peter Cresswell Chimeric antigen receptor T-cell (CAR T-cell) therapy has been shown to be clinically effective for managing a variety of hematological cancers. However, CAR T-cell therapy is associated with multiple adverse effects, including neurotoxicity and cytokine release syndrome (CRS). CRS arises from massive cytokine secretion and can be life-threatening, but it is typically managed with an anti-IL-6Ra mAb or glucocorticoid administration. However, these treatments add to a patient's medication burden and address only the CRS symptoms. Therefore, alternative strategies that can prevent CRS and neurotoxicity associated with CAR T-cell treatment are urgently needed. Here, we explored a therapeutic route aimed at preventing CRS rather than limiting its consequences. Using a cytokine-profiling assay, we show that granulocyte-macrophage colony-stimulating factor (GMCSF) is a key CRS-promoting protein. Through a combination of in vitro experiments and gene-editing technology, we further demonstrate that antibody-mediated neutralization or TALEN-mediated genetic inactivation of GMCSF in CAR T-cells drastically decreases available GMCSF and abolishes macrophage-dependent secretion of CRS biomarkers, including monocyte chemoattractant protein 1 (MCP-1), interleukin (IL) 6, and IL-8. Of note, we also found that the genetic inactivation of GMCSF does not impair the antitumor function or proliferative capacity of CAR T-cells in vitro. We conclude that it is possible to prevent CRS by using "all-in-one" GMCSF-knockout CAR T-cells. This approach may eliminate the need for anti-CRS treatment and may improve the overall safety of CAR T-cell therapies for cancer patients.

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High‐Throughput Image Cytometry Detection Method for CAR‐T Transduction, Cell Proliferation, and Cytotoxicity Assays

Wang

Chan²,

Grimaud

et al. 2020

Cytometry Pt A

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Chimeric antigen receptor (CAR)‐T cell therapy has drawn much attention due to its recent clinical success in B‐cell malignancies. In general, the CAR‐T cell discovery process consists of CAR identification, T‐cell activation, transduction, and expansion, as well as assessment of CAR‐T cytotoxicity. The current evaluation methods for the CAR‐T discovery process can be time‐consuming, low‐throughput and requires the preparation of multiple sacrificial samples in order to produce kinetic data. In this study, we employed the use of a plate‐based image cytometer to monitor anti‐CAIX (carbonic anhydrase IX) G36 CAR‐T generation and assess its cytotoxic potency of direct and selective killing against CAIX+ SKRC‐59 human renal cell carcinoma cells. The transduction efficiency and cytotoxicity results were analyzed using image cytometry and compared directly to flow cytometry and Chromium 51 (51Cr) release assays, showing that image cytometry was comparable against these conventional methods. Image cytometry method streamlines the assays required during the CAR‐T cell discovery process by analyzing a plate of T cells from CAR‐T generation to in vitro functional assays with minimum disruption. The proposed method can reduce assay time and uses less cell samples by imaging and analyze the same plate over time without the need to sacrifice any cells. The ability to monitor kinetic data can allow additional insights into the behavior and interaction between CAR‐T and target tumor cells. © 2020 International Society for Advancement of Cytometry

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A Versatile Safeguard for Chimeric Antigen Receptor T-Cell Immunotherapies

Cited by 29 publications

References 30 publications

Repurposing endogenous immune pathways to tailor and control chimeric antigen receptor T cell functionality

Repurposing endogenous immune pathways to tailor and control chimeric antigen receptor T cell functionality

Granulocyte–macrophage colony-stimulating factor inactivation in CAR T-cells prevents monocyte-dependent release of key cytokine release syndrome mediators

High‐Throughput Image Cytometry Detection Method for CAR‐T Transduction, Cell Proliferation, and Cytotoxicity Assays

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