Systemic Lupus Erythematosus (SLE) is a progressive disease leading to immune-mediated tissue damage, associated with an alteration of lymphoid organs. Therapeutic strategies involving regulatory T (Treg) lymphocytes, which physiologically quench autoimmunity and support long-term immune tolerance, are considered, as conventional treatment often fails. We describe here a therapeutic strategy based on Tregs overexpressing FoxP3 and harboring anti-CD19 CAR (Fox19CAR-Tregs). Fox19CAR-Tregs efficiently suppress proliferation and activity of B cells in vitro, which are relevant for SLE pathogenesis. In an humanized mouse model of SLE, a single infusion of Fox19CAR-Tregs restricts autoantibody generation, delay lymphopenia (a key feature of SLE) and restore the human immune system composition in lymphoid organs, without detectable toxicity. Although a short survival, SLE target organs appear to be protected. In summary, Fox19CAR-Tregs can break the vicious cycle leading to autoimmunity and persistent tissue damage, representing an efficacious and safe strategy allowing restoration of homeostasis in SLE.
© 2024. The Author(s).

As checkpoint inhibitor immunotherapies gain traction among cancer researchers and clinicians, the need grows for assays that can definitively phenotype patient immune cells. Herein, we present an 8-color flow cytometry panel for lineage and immune checkpoint markers and validate it using healthy human donor peripheral blood mononuclear cells (PBMCs). Flow cytometry data was generated on a BD LSR Fortessa and supported by Luminex multiplex soluble immunoassay. Our data showed significant variation between donors at both baseline and different stages of activation, as well as a trend in increasing expression of checkpoint markers on stimulated CD4+ and CD8+ T-cells with time. Soluble immune checkpoint quantification assays revealed that LAG-3, TIM-3, CTLA-4, and PD-1 soluble isoforms are upregulated after stimulation. This 8-color flow cytometry panel, supported here by soluble immunoassay, can be used to identify and evaluate immune checkpoints on T-lymphocytes in cryopreserved human PBMC samples. This panel is ideal for characterizing checkpoint expression in clinical samples for which cryopreservation is necessary.