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    Cellular and T cell engager Immunotherapy

    Category: Cellular and T cell engager Immunotherapy

    BCMA CAR-T cells capable of IL-6-neutralization bifunctionally reduce cytokine release syndrome and IL-6-driven myeloma growth in preclinical models

    (PA-052) BCMA CAR-T Cells Capable of il-6-neutralization Bifunctionally Reduce Cytokine Release Syndrome and IL-6-driven Myeloma Growth in Preclinical Models

    Wednesday, September 17, 2025

    Introduction:      The widespread adoption of BCMA-directed CAR-T cells for relapsed multiple myeloma (MM), especially in outpatient and community settings, is limited due to inflammatory toxicity concerns, including cytokine release syndrome (CRS). We therefore seek to develop CAR-T cells that can modulate the signalling involved in CRS to improve the therapeutic window of CAR-T cell therapy.

    Methods:   We developed single domain antibody (sdAb)-based BCMA CAR constructs (sdBCMA.BBz) and generated CAR-T cells using lentiviral vectors and healthy donor T-cells. We performed in vitro testing of cytotoxicity, cytokine release, proliferation, and repetitive stimulation against human cell lines, and in vivo with OPM2 xenografts in NSG mice. We then developed membrane-bound versus secreted versions of a sdAb that neutralizes IL-6 and incorporated this into the lead sdBCMA.BBz CAR construct. These underwent the same in vitro and in vivo assessment as above. We also developed an in vitro tri-culture system with CAR-T cells, tumour cells, and monocyte-derived macrophages, to model CRS cytokine production. Finally, we tested our sdIL-6 neutralizing domains against IL6-dependent MM cell lines.

    Results:   CAR-T cells generated from healthy donor T-cells carrying anti-BCMA sdAb’s with varying affinities, or control binders derived from ide-cel and cilta-cel, were tested using co-culture experiments with MM target cell lines. These in vitro studies confirmed that all sdBCMA.BBz CAR-T cells effectively lysed MM targets, released inflammatory cytokines, and proliferated. Effective disease control was demonstrated in NSG mice bearing OPM2 tumours with our sdAb-based CARs, comparable to our mimics of clinically approved CAR-T cells. We synthesized an anti-human IL-6 sdAb and show effective binding with an EC50 of 0.84nM in a direct binding assay, and an IC50 of 2.8nM in a competitive binding assay with the native IL-6 receptor. We show effective clearance of exogenous IL-6 in culture supernatants, as well as clearance of macrophage-derived IL-6 in tri-cultures incorporating CAR-T cells, OPM2 or MM1S tumour cells, and monocyte-derived macrophages. We also show that this IL-6 binder can slow the growth of IL-6-dependent cell lines, ANBL-6 and INA-6. In NSG mice with OPM2 tumours, we demonstrate equivalent survival with CAR-T cells incorporating the IL-6-neutralizing domains.

    Conclusions:   We show that our sdAb-based CAR T-cells are functional in vitro and in vivo. We further show that our IL-6-neutralizing CAR-T cells can clear macrophage-derived IL-6 derived in vitro, the major source of IL-6 in CAR-T cell-related CRS. We show that this IL-6 binder can impede the growth of IL-6 dependent cell lines, highlighting microenvironment-derived IL-6 as a potential therapeutic enhancement. The assessment of these CAR-T cells in humanized mouse models of MM is on-going. Taken together, we show a CAR-T cell product that can neutralize IL-6, which underlies the pathophysiology of CRS and is a key myeloma growth-promoting cytokine.