Enhancing stability and function of induced regulatory T cells through membrane-anchored transforming growth factor-β1 and interleukin-10 secretion to protect against graft-versus-host disease.

Graft-versus-host disease (GvHD) remains a key challenge in allogeneic transplantation, underscoring the need for effective therapies to establish immune tolerance. Adoptive regulatory T cell therapy shows promise but is restricted by the limited expansion of natural Tregs (nTregs) and the instability of induced Tregs (iTregs). The latter are prone to losing Foxp3 expression and suppressive function in inflammatory microenvironments. Here, we developed membrane-anchored TGF-β1 and IL-10-secreting iTregs (MAT-iTregs), a novel engineered iTreg platform designed to enhance lineage stability and suppressive potency. MAT-iTregs express membrane-anchored TGF-β1 and produce high levels of IL-10 to amplify immunosuppression and maintain functional stability. We optimized construct design and manufacture process to yield highly enriched MAT-iTregs. In preclinical studies, MAT-iTregs outperformed conventional iTregs in safety profiles and therapeutic efficacies. MAT-iTregs significantly prolonged survival, reduced tissue injury, and attenuated pathogenic effector T cell responses in xenogeneic GvHD mice. Notably, MAT-iTregs maintained suppressive function, durable persistence, and a favorable safety profile in mice models, with no evidence of converting to effector T cells and minimal tissue toxicity. Together, these findings support MAT-iTregs as a clinically translatable cell therapy to promote transplant tolerance and improve outcomes in GvHD.