Biology subjects

Dutoit, V.

Publications and source records attributed to Dutoit, V..

3 recordsLinked to original sources

bioRxiv2026

Safety profiling of CAR-T cells using an organotypic human tissue platform

CAR-T-cell-associated on-target off-tumor (OTOT) toxicity represents a major safety concern, as recognition of target antigens on healthy tissues can trigger severe and potentially life-threatening complications. Predicting OTOT toxicity remains a challenge because current preclinical models fail to capture the complexity of native human tissues. Here, we developed a human organotypic tissue platform that enables functional assessment of CAR-T-cell activity in intact human tissues across organ-specific and inflammatory contexts. Using a panel of clinically relevant CAR-T-cell products with known OTOT toxicities, we demonstrate that the platform faithfully recapitulates clinically observed tissue-specific toxicity profiles. CAR-T cells targeting EGFR, HER2, and mesothelin induced inflammatory and cytotoxic responses in healthy human lung tissue, whereas CD19 CAR-T cells remained inactive. We further show that OTOT toxicity cannot be reliably predicted from antigen abundance alone but instead results from the integration of multiple target-dependent determinants, including CAR affinity, inflammatory context, antigen accessibility, and effector-cell dose. The platform also enables quantitative assessment of inflammatory and cytotoxic responses and supports evaluation of pharmacological and CAR design-based strategies to mitigate toxicity. Together, this work establishes the first human organotypic platform for functional modeling of CAR-T-cell-associated OTOT toxicity, providing a clinically relevant framework for preclinical safety evaluation and the rational development of safer engineered cell therapies. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=167 SRC="FIGDIR/small/740527v1_ufig1.gif" ALT="Figure 1"> View larger version (38K): org.highwire.dtl.DTLVardef@12c8addorg.highwire.dtl.DTLVardef@150d392org.highwire.dtl.DTLVardef@17218f7org.highwire.dtl.DTLVardef@1c54078_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2025

Unveiling the Molecular Architecture of T Cells and Immune Synapses with Cryo-Expansion Microscopy

Cellular communication is critical for anti-cancer immunity, with tumor cell killing occurring at immunological synapses (IS) formed between effector immune cells and target tumor cells. While optical super-resolution microscopy (SRM) has enlightened the spatial organization of the IS mostly in regular immune cells, visualizing the nanoscale architectural features of IS in its native state, including 3D receptor distribution and the ultrastructural details of the lytic granule release remains challenging. Using cryo-expansion microscopy (cryo-ExM), we unravel the cellular architecture of activated T cells and T cell-target cell pairs. Our approach visualizes actin and microtubule networks during synapse formation, membrane topography, and the distribution of signaling molecules and lytic granules of different types, offering novel insights into IS organization. Finally, we apply U-ExM to glioblastoma tissue, visualizing T cells and their lytic content in situ, highlighting its potential for pre-clinical immunotherapy studies.

immunology↗

bioRxiv2023

PTPRZ1-targeting RNA CAR-T cells exert antigen-specific and bystander antitumor activity in glioblastoma

The great success of chimeric antigen receptor (CAR)-T cell therapy in B-cell malignancies has prompted its translation to solid tumors. In the case of glioblastoma (GBM), clinical trials have shown modest efficacy, but anti-GBM CAR-T cells are being intensely developed. In this study, we selected PTPRZ1 as an attractive new target for GBM treatment. We isolated six anti-human PTPRZ1 scFv from a human phage display library and produced 2nd generation CAR-T cells in an RNA format. Patient-derived GBM PTPRZ1-knock-in cell lines were used to select the CAR construct (471_28z), which showed high cytotoxicity while consistently displaying high CAR expression. CAR-T cells incorporating 471_28z were able to release IFN-{gamma}, IL-2, TNF-, Granzyme B, IL-17A, IL-6, and soluble FasL, and displayed low tonic signaling. Additionally, they maintained an effector memory phenotype after in vitro killing. Importantly, 471_28z CAR-T cells displayed strong bystander killing against PTPRZ1-negative cell lines after pre-activation by PTPRZ1-positive tumor cells, but did not kill antigen-negative non-tumor cells. In an orthotopic xenograft tumor model using NSG mice, a single dose of anti-PTPRZ1 CAR-T cells significantly delayed tumor growth. Taken together, these results validate the use of PTPRZ1 as a new GBM target and prompt the use of anti-PTPRZ1 CAR-T cells for clinical translation.

immunology↗