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bioRxiv · 10.1101/2025.07.19.665688

Receptor-tethered orthogonal IL-2 enhances regulatory T cell therapy

Indart, A.·Lyu, H.·Nguyen, V.·Rosenthal, W.·Jang, S.·Chen, Y.·Jude, K.·Su, L.·Garcia, K. C.·Bluestone, J. A.·Tang, Q.
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Abstract

Regulatory T cell (Treg) therapy is an emerging platform for controlling immune overactivation. Efficacy of Treg therapy is limited by the poor persistence of infused Tregs due to insufficient IL-2, which is essential for Treg survival and function. IL-2 activates many immune cells besides Tregs, imposing a challenge for the selective provision of IL-2 to infused Tregs. In this study, we found infusions of orthogonal (ortho) IL-2 failed to enhance Tregs engineered with a corresponding orthoIL-2 receptor (IL-2R) in a mouse model of autoimmune diabetes. We then developed a receptor-tethered orthoIL-2 by optimizing the combination of IL-2, IL-2R, and the linker connecting them to achieve autocrine signaling selectively in engineered Tregs. Tregs expressing the tethered orthoIL-2 showed autocrine IL-2 signaling in vitro, enhanced CD25, CTLA-4, and Foxp3 expression, persisted without exogenous IL-2 in vivo, and prevented autoimmune diabetes using as few as 2,000 Tregs. Knocking the tethered orthoIL-2 construct into the Foxp3 locus enabled Treg-specific expression, with the additional benefit of positively reinforcing tethered orthoIL-2 expression through the activation of the Foxp3 gene by enhanced IL-2 signaling. Together, these results illustrate a safe and effective cell-engineering solution for overcoming Tregs dependency on exogenous IL-2, thereby achieving superior therapeutic efficacy.

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Indart, A., Lyu, H., Nguyen, V., Rosenthal, W., Jang, S., Chen, Y., Jude, K., Su, L., Garcia, K. C., Bluestone, J. A., Tang, Q.. 2025-07-23. Receptor-tethered orthogonal IL-2 enhances regulatory T cell therapy. https://doi.org/10.1101/2025.07.19.665688

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