Regulatory T Cells Restrain Interleukin-2- and Blimp-1-Dependent Acquisition of Cytotoxic Function by CD4⁺ T Cells

In addition to helper and regulatory potential, CD4⁺ T cells also acquire cytotoxic activity marked by granzyme B (GzmB) expression and the ability to promote rejection of established tumors. Here, we examined the molecular and cellular mechanisms underpinning the differentiation of cytotoxic CD4⁺ T cells following immunotherapy. CD4⁺ transfer into lymphodepleted animals or regulatory T (Treg) cell depletion promoted GzmB expression by tumor-infiltrating CD4⁺, and this was prevented by interleukin-2 (IL-2) neutralization. Transcriptional analysis revealed a polyfunctional helper and cytotoxic phenotype characterized by the expression of the transcription factors T-bet and Blimp-1. While T-bet ablation restricted interferon-γ (IFN-γ) production, loss of Blimp-1 prevented GzmB expression in response to IL-2, suggesting two independent programs required for polyfunctionality of tumor-reactive CD4⁺ T cells. Our findings underscore the role of Treg cells, IL-2, and Blimp-1 in controlling the differentiation of cytotoxic CD4⁺ T cells and offer a pathway to enhancement of anti-tumor activity through their manipulation. Śledzińska et al. show that upon depletion of regulatory T cells, a surplus of interleukin-2 in the tumor microenvironment supports acquisition of cytotoxic activity by T helper cells orchestrated by the transcription factor Blimp-1. These polyfunctional CD4⁺ T cells exhibit potent anti-tumor activity in an adoptive transfer setting with therapeutic implications.