Abstract Text: Regulatory T cells (Tregs) are essential for keeping the immune system in check, thus dysregulation of Tregs can lead to autoimmune disorders. Treg cell therapy has the potential to address this problem, however limitations of sorted and cultured Treg (cTregs) cell therapies include inherent plasticity and instability. Indeed, cTregs can lose FOXP3 expression and acquire the ability to express T effector (Teff) cytokines in non-favorable environments. To address these limitations, we have engineered human CD4 T cells into Tregs (EngTregs) endowed with stable FOXP3 expression and a rapamycin-activated, chemically induced IL-2 signaling complex (CISC). To address the stability and functionality of EngTregs, we compared them to cTregs at both steady state and in inflammatory conditions. At steady state, EngTregs demonstrate enrichment of core T regulatory cell gene signatures (CTLA-4, IL2RB, TNFRSF1B, TNFRSF18) compared to cTregs. EngTregs but not cTregs, maintain stability as measured by expression of FOXP3, CD25 and other Treg stability markers (e.g. EOS, CD27+CD70-). Stable FOXP3 expression in EngTregs is reflected in their suppressive activity against Teff cells. Importantly, EngTregs maintain FOXP3 expression in inflammatory environments and secrete IL-10 similarly to cTreg. However, unlike cTregs, EngTregs express little or no key Th2 cytokines (e.g. IL4 and IL13) in such environments. Engineering human Tregs has shown the potential to overcome key limitations of sorted Treg therapies by enhancing their stability and function and thus providing a novel therapeutic modality to restore immune tolerance.
