CAR- and TRuC-redirected regulatory T cells differ in capacity to control adaptive immunity to FVIII

TLDR

Regulatory T cells suppress immune responses in autoimmune disease, transplantation, and protein‑replacement therapies by using antigen‑specific TCR signals that direct tissue‑specific suppression. The study aimed to engineer antigen‑specific Tregs that redirect polyclonal Tregs to suppress anti‑factor VIII antibody formation in hemophilia A, independent of MHC. They engineered polyclonal Tregs with antigen‑specific receptors (CAR or TRuC) to redirect them toward factor VIII, enabling MHC‑independent suppression. CAR‑Tregs triggered a strong effector phenotype that abrogated suppression, whereas TRuC‑Tregs engaged the full TCR complex to deliver controlled signaling and effectively suppressed anti‑FVIII antibody production, indicating that activation thresholds must be regulated for engineered Treg therapies.

Abstract

Regulatory T cells (Tregs) control immune responses in autoimmune disease, transplantation, and enable antigen-specific tolerance induction in protein-replacement therapies. Tregs can exert a broad array of suppressive functions through their T cell receptor (TCR) in a tissue-directed and antigen-specific manner. This capacity can now be harnessed for tolerance induction by "redirecting" polyclonal Tregs to overcome low inherent precursor frequencies and simultaneously augment suppressive functions. With the use of hemophilia A as a model, we sought to engineer antigen-specific Tregs to suppress antibody formation against the soluble therapeutic protein factor (F)VIII in a major histocompatibility complex (MHC)-independent fashion. Surprisingly, high-affinity chimeric antigen receptor (CAR)-Treg engagement induced a robust effector phenotype that was distinct from the activation signature observed for endogenous thymic Tregs, which resulted in the loss of suppressive activity. Targeted mutations in the CD3ζ or CD28 signaling motifs or interleukin (IL)-10 overexpression were not sufficient to restore tolerance. In contrast, complexing TCR-based signaling with single-chain variable fragment (scFv) recognition to generate TCR fusion construct (TRuC)-Tregs delivered controlled antigen-specific signaling via engagement of the entire TCR complex, thereby directing functional suppression of the FVIII-specific antibody response. These data suggest that cellular therapies employing engineered receptor Tregs will require regulation of activation thresholds to maintain optimal suppressive function.