Intranuclear interactomic inhibition of FoxP3 suppresses functions of Treg cells

•This study presents a novel strategy to modulate FoxP3 function directly.•The nt-FoxP3-subdomains are effectively delivered into the nucleus without toxicity.•The nt-FoxP3-FKH is intranuclear interactomic inhibitor of FoxP3. Regulatory T cells (Treg cells) are crucial for the maintenance of immunological tolerance, and it has been reported that Treg cells are enriched within the tumor micro-environment for immune evasion due to their immunosuppressive functions. To inhibit Treg cells functions, FoxP3, a lineage-specific transcription factor responsible for the differentiation and functions of Treg cells, was functionally targeted by a nucleus-transducible (nt) form of various FoxP3 functional subdomains. These nt modified domains can be delivered into the nucleus effectively and work as interactomic inhibitors via disruption of the endogenous FoxP3-mediated transcription complex. Among these domains, nt-FoxP3-FKH (Forkhead DNA binding domain) is most effective at restoring NFAT activity suppressed by FoxP3, and inhibiting the binding of endogenous FKH-containing proteins to FKH DNA binding sequences without influencing the viability and activation of T cells. The suppressive functions of TGF-β-induced iTreg cells and thymus-derived tTreg cells were substantially blocked by nt-FoxP3-FKH, accompanied with down-regulation of CTLA-4 surface expression and IL-10 secretion of Treg cells. In addition, nt-FoxP3-FKH upregulated the expression of IL-2 and IFN-γ in Treg cells. Therefore, nt-FoxP3-FKH has the potential to be a novel therapeutic agent to modulate the immune-evasive tumor environment created by Treg cells without the need for genetic modifications.