Background/Purpose: Chimeric antigen receptor (CAR)-T cell therapies have revolutionized the treatment of cancer and can be curative. CD19-targeted CAR-T cells hold promise for the treatment of refractory autoimmune diseases, but therapies that indiscriminately deplete all B cells are limited by infection risk. In contrast, precision immunotherapies that selectively eliminate autoreactive B cells, while maintaining the normal B cell pool, have the potential to treat autoimmunity without impairing protective immunity. Antiphospholipid syndrome (APS) is characterized by autoantibodies against beta-2-glycoprotein I (B2GPI) domain I (DI) which are pathogenic and promote thrombosis and fetal loss. We therefore aimed to develop an autoantigen-specific T-cell therapy targeting anti-B2GPI B cells for the treatment of APS.

Methods: We employed a CRISPR-Cas9/12a-based genome-editing platform to incorporate APS-relevant autoantigen into the T cell receptor (TCR)-CD3 complex of primary human T cells. Homology-directed repair template encoding partial or full nucleotide sequences of B2GPI and Cas-gRNA ribonucleoprotein complexes targeting CD3G, CD3E, or TRAC were transfected into activated T cells. CATCR expression was quantified by flow cytometry. Ramos B cells were CRISPR-Cas9-edited to replace their native BCR with monoclonal anti-B2GPI DI BCRs cloned from patients with APS. The potency and specificity of 36 B2GPI-CATCR-T cells against different anti-B2GPI or wild-type Ramos B cells was interrogated in co-culture, and cytotoxicity was quantified by flow cytometry, live-cell imaging, and cytokine assays.

Results: We hypothesized that T cells can be redirected to selectively kill anti-B2GPI B cells in APS. To this end, T cells were engineered to express chimeric autoantigen-TCRs (CATCRs) which confer the ability to bind autoantigen-specific B-cell receptors (BCRs), thereby inducing immune synapse formation and killing of autoreactive B cells via the perforin–granzyme effector pathway (Fig.1A). B2GPI-CATCR-T cells, expressing the APS autoantigen B2GPI as part of CD3-gamma, CD3-epsilon, or the TCR-beta/alpha chains, were successfully generated from primary T cells, as confirmed by flow cytometry (Fig.1B). In co-culture of T cells with autoreactive Ramos B cells (expressing pathogenic anti-B2GPI BCRs/autoantibody) or unedited Ramos cells (expressing irrelevant BCR), B2GPI-CATCR-T cells -but not control T cells- efficiently and selectively eliminated anti-B2GPI DI B cells. In contrast, B2GPI-CATCR-T cells did not deplete B cells expressing irrelevant BCR (Fig.1C). Target cell killing was observed at physiologically relevant effector:target cell ratios (Fig.1D).

Conclusion: We describe a precision cellular immunotherapy approach for the autoantigen-specific depletion of autoreactive B cells in autoimmune diseases, which we here first applied to APS. B2GPI-CATCR-T cells selectively eliminated anti-B2GPI DI B cells—pathogenic drivers of thrombosis and fetal loss in APS—in a dose-dependent manner, providing an opportunity to treat and prevent APS. Beyond APS, these autoantigen-specific immunotherapies promise a future of treating autoimmunity without increasing the risk of infection.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/chimeric-autoantigen-t-cell-receptor-catcr-t-cell-therapies-to-selectively-target-autoreactive-b-cells/