Using whole-genome CRISPR/Cas9 screening on MC38 colon adenocarcinoma and B16 melanoma cell lines, Kearney et al. demonstrated that tumors evade CD8+ T cell killing by disrupting antigen presentation, IFNγ, and TNF signaling, and evade NK cells by disrupting TNF signaling. Perforin-related pathways were not involved in evasion mechanisms. Recombinant TNF (but not IFNγ) was able to directly kill tumor cells and, upon antigen recognition, TNF mediated bystander killing of antigen-negative tumor cells in vitro. Analysis of patient data in TCGA confirmed these observations.
Immunotherapy has revolutionized outcomes for cancer patients, but the mechanisms of resistance remain poorly defined. We used a series of whole-genome clustered regularly interspaced short palindromic repeat (CRISPR)-based screens performed in vitro and in vivo to identify mechanisms of tumor immune evasion from cytotoxic lymphocytes [CD8(+) T cells and natural killer (NK) cells]. Deletion of key genes within the tumor necrosis factor (TNF) signaling, interferon-gamma (IFN-gamma) signaling, and antigen presentation pathways provided protection of tumor cells from CD8(+) T cell-mediated killing and blunted antitumor immune responses in vivo. Deletion of a number of genes in the TNF pathway also emerged as the key mechanism of immune evasion from primary NK cells. Our screens also identified that the metabolic protein 2-aminoethanethiol dioxygenase (Ado) modulates sensitivity to TNF-mediated killing by cytotoxic lymphocytes and is required for optimal control of tumors in vivo. Remarkably, we found that tumors delete the same genes when exposed to perforin-deficient CD8(+) T cells, demonstrating that the dominant immune evasion strategy used by tumor cells is acquired resistance to T cell-derived cytokine-mediated antitumor effects. We demonstrate that TNF-mediated bystander killing is a potent T cell effector mechanism capable of killing antigen-negative tumor cells. In addition to highlighting the importance of TNF in CD8(+) T cell- and NK cell-mediated killing of tumor cells, our study also provides a comprehensive picture of the roles of the TNF, IFN, and antigen presentation pathways in immune-mediated tumor surveillance.