Reprogramming of Regulatory T cells in the Tumour Microenvironment

Abstract

Foxp3+ (CD4+) regulatory T (Treg) cells are a stable lymphocyte lineage central to the maintenance of immune homeostasis. They are required to regulate anti-pathogen responses and prevent autoimmune disease, but detrimental in cancer patients through their suppression of anti-tumour immune responses. Recent studies have however challenged the notion of a stable, immune-suppressive Treg cell lineage and suggested that Treg cells can be “converted” to various effector T helper (Th)-like subsets in response to environmental cues. Additionally, it has been suggested that the expression of IFN-γ by “converted” intratumoural Treg cell is required for tumour rejection in response to anti-PD-1 checkpoint therapy. However, it is not known through which mechanisms these Treg cells are converted; and how their generation is limited. Furthermore, controversy exists on the retention of the suppressive capabilities of these Treg cells upon expressing Th-lineage transcription factors. We have previously shown that the disruption of the CARMA1-BCL10-MALT1 (CBM) signalosome complex induces significant IFN-γ expression by tumour infiltrating Treg cells (Di Pilato et al., 2019, Nature). In line with this, our bulk assay for transposase-accessible chromatin sequencing (ATAC-seq) analysis of intratumoural Foxp3+ Treg cells has shown a gain of chromatin accessibility across several pro-inflammatory genes, specifically Ifng, suggesting the coexistence of suppressive Treg and inflammatory Th programmes. At a single cell level, we have identified distinct programmes that contribute to the preservation of Treg cell stability (maintenance of Foxp3 expression), as well as resistance to pro-inflammatory cytokine expression (such as IFN-γ) that has been associated with the proinflammatory conversion of Foxp3 positive Treg beyond the classical definition of ‘Treg plasticity’ (upregulation of Teff cell-associated chemokine receptors and transcription factors) upon Treg cell activation and infiltration to the tumour. And lastly, we have identified a DNA accessible site at position -28.5kb in the Ifng locus as a novel cis-regulatory sequence that is crucial for IFN-γ expression. Based on these results we propose that Treg cell instability and Treg cell plasticity are independent, such that Foxp3+ cells can secrete IFN- while Foxp3– ex-Treg cells can, but do not need to commit to doing so.