The cancer immunoediting hypothesis postulates a dual role of the immune system: protecting the host by eliminating tumor cells, and shaping the tumor by editing its genome. Here, we elucidate the impact of evolutionary and immune-related forces on editing the tumor in a mouse model for hypermutated and microsatellite-instable colorectal cancer. Analyses of wild-type and immunodeficient RAG1 knockout mice transplanted with MC38 cells reveal that upregulation of checkpoint molecules and infiltration by Tregs are the major tumor escape mechanisms. Our results show that the effects of immunoediting are weak and that neutral accumulation of mutations dominates. Targeting the PD-1/PD-L1 pathway using immune checkpoint blocker effectively potentiates immunoediting. The immunoediting effects are less pronounced in the CT26 cell line, a non-hypermutated/microsatellite-instable model. Our study demonstrates that neutral evolution is another force that contributes to sculpting the tumor and that checkpoint blockade effectively enforces T-cell-dependent immunoselective pressure.

Objective: Nicotinamide phosphoribosyltransferase (NAMPT, also referred to as pre-B cell colony-enhancing factor or visfatin) is critically required for the maintenance of cellular nicotinamide adenine dinucleotide (NAD) supply catalysing the rate-limiting step of the NAD salvage pathway. NAMPT is strongly upregulated in inflammation including IBD and counteracts an increased cellular NAD turnover mediated by NAD-depleting enzymes. These constitute an important mechanistic link between inflammatory, metabolic and transcriptional pathways and NAD metabolism. Design: We investigated the impact of NAMPT inhibition by the small-molecule inhibitor FK866 in the dextran sulfate sodium (DSS) model of colitis and the azoxymethane/DSS model of colitis-associated cancer. The impact of NAD depletion on differentiation of mouse and human primary monocytes/macrophages was studied in vitro. Finally, we tested the efficacy of FK866 compared with dexamethasone and infliximab in lamina propria mononuclear cells (LPMNC) isolated from patients with IBD. Results: FK866 ameliorated DSS-induced colitis and suppressed inflammation-associated tumorigenesis in mice. FK866 potently inhibited NAMPT activity as demonstrated by reduced mucosal NAD, resulting in reduced abundances and activities of NAD-dependent enzymes including PARP1, Sirt6 and CD38, reduced nuclear factor kappa B activation, and decreased cellular infiltration by inflammatory monocytes, macrophages and activated T cells. Remarkably, FK866 effectively supressed cytokine release from LPMNCs of patients with IBD. As FK866 was also effective in Rag1^−⁄− mice, we mechanistically linked FK866 treatment with altered monocyte/macrophage biology and skewed macrophage polarisation by reducing CD86, CD38, MHC-II and interleukin (IL)-6 and promoting CD206, Egr2 and IL-10. Conclusion: Our data emphasise the importance of NAD immunometabolism for mucosal immunity and highlight FK866-mediated NAMPT blockade as a promising therapeutic approach in acute intestinal inflammation.

Chronic hepatitis B virus (HBV) infection affects 300 million patients worldwide^1,2, in whom virus-specific CD8 T cells by still ill-defined mechanisms lose their function and cannot eliminate HBV-infected hepatocytes^3–7. Here we demonstrate that a liver immune rheostat renders virus-specific CD8 T cells refractory to activation and leads to their loss of effector functions. In preclinical models of persistent infection with hepatotropic viruses such as HBV, dysfunctional virus-specific CXCR6⁺ CD8 T cells accumulated in the liver and, as a characteristic hallmark, showed enhanced transcriptional activity of cAMP-responsive element modulator (CREM) distinct from T cell exhaustion. In patients with chronic hepatitis B, circulating and intrahepatic HBV-specific CXCR6⁺ CD8 T cells with enhanced CREM expression and transcriptional activity were detected at a frequency of 12–22% of HBV-specific CD8 T cells. Knocking out the inhibitory CREM/ICER isoform in T cells, however, failed to rescue T cell immunity. This indicates that CREM activity was a consequence, rather than the cause, of loss in T cell function, further supported by the observation of enhanced phosphorylation of protein kinase A (PKA) which is upstream of CREM. Indeed, we found that enhanced cAMP–PKA-signalling from increased T cell adenylyl cyclase activity augmented CREM activity and curbed T cell activation and effector function in persistent hepatic infection. Mechanistically, CD8 T cells recognizing their antigen on hepatocytes established close and extensive contact with liver sinusoidal endothelial cells, thereby enhancing adenylyl cyclase–cAMP–PKA signalling in T cells. In these hepatic CD8 T cells, which recognize their antigen on hepatocytes, phosphorylation of key signalling kinases of the T cell receptor signalling pathway was impaired, which rendered them refractory to activation. Thus, close contact with liver sinusoidal endothelial cells curbs the activation and effector function of HBV-specific CD8 T cells that target hepatocytes expressing viral antigens by means of the adenylyl cyclase–cAMP–PKA axis in an immune rheostat-like fashion.

Analyzing mouse tumor models in vivo, human T cells ex vivo, and human lung cancer samples, we provide direct evidence that NR2F6 acts as an immune checkpoint. Genetic ablation of Nr2f6, particularly in combination with established cancer immune checkpoint blockade, efficiently delays tumor progression and improves survival in experimental mouse models. The target genes deregulated in intratumoral T lymphocytes upon genetic ablation of Nr2f6 alone or together with PD-L1 blockade reveal multiple advantageous transcriptional alterations. Acute Nr2f6 silencing in both mouse and human T cells induces hyper-responsiveness that establishes a non-redundant T-cell-inhibitory function of NR2F6. NR2F6 protein expression in T-cell-infiltrating human NSCLC is upregulated in 54% of the cases (n = 303) and significantly correlates with PD-1 and CTLA-4 expression. Our data define NR2F6 as an intracellular immune checkpoint that suppresses adaptive anti-cancer immune responses and set the stage for clinical validation of targeting NR2F6 for next-generation immuno-oncological regimens.

Gastrointestinal (GI) homeostasis is strongly dependent on nuclear receptor (NR) functions. They play a variety of roles ranging from nutrient uptake, sensing of microbial metabolites, regulation of epithelial intestinal cell integrity to shaping of the intestinal immune cell repertoire. Several NRs are associated with GI pathologies; therefore, systematic analysis of NR biology, the underlying molecular mechanisms, and regulation of target genes can be expected to help greatly in uncovering the course of GI diseases. Recently, an increasing number of NRs has been validated as potential drug targets for therapeutic intervention in patients with inflammatory bowel disease (IBD). Besides the classical glucocorticoids, especially PPARγ, VDR, or PXR-selective ligands are currently being tested with promising results in clinical IBD trials. Also, several pre-clinical animal studies are being performed with NRs. This review focuses on the complex biology of NRs and their context-dependent anti- or pro-inflammatory activities in the regulation of gastrointestinal barrier with special attention to NRs already pharmacologically targeted in clinic and pre-clinical IBD treatment regimens.