Immune-intrinsic MLKL regulates dendritic cell maturation and CD8⁺ T cell effector function in anti-tumor immunity

Anti-tumor immunity relies on coordinated interactions between dendritic cells (DCs) and cytotoxic CD8⁺ T cells to eliminate malignant cells. Mixed lineage kinase domain-like protein (MLKL), a key executioner of necroptosis, has recently been implicated in cancer through necroptosis-independent functions and is being explored as a therapeutic target. However, most studies have focused on tumor cells, whereas the immune-intrinsic functions of MLKL remain poorly understood. In particular, MLKL expression across immune cell subsets and its role in anti-tumor immunity have not been systematically investigated.

Transcriptomic analyses revealed broad MLKL expression across multiple immune cell subsets, including neutrophils, monocytes, dendritic cells, and activated/memory Tcell subsets. Consistent with these findings, MLKL was upregulated in primary mouse immune cells following activation, including Tcell stimulation, bone marrow-derived dendritic cell (BMDC) maturation, and M1 macrophages. Pharmacological inhibition of MLKL impaired BMDC maturation, reduced DC-mediated CD8⁺ T cell priming, and suppressed CD8⁺ T cell activation, proliferation, and cytotoxic killing of B16-OVA tumor cells. Integrated single-cell RNA sequencing analyses from cancer patients receiving immune checkpoint blockade (ICB) further showed that MLKL expression was enriched in activated and effector CD8⁺ T cell subsets. Donor-level analyses showed higher MLKL expression in T cells from ICB responders than in those from non-responders, particularly in post-treatment samples. These findings associate MLKL expression with activated and effector anti-tumor Tcell states and suggest its potential relevance to ICB response.

Collectively, these findings show that MLKL is upregulated during immune activation and contributes to DC maturation and CD8⁺ T cell effector responses. Our findings highlight the context-dependent roles of MLKL across tumor and immune compartments and emphasize the importance of considering immune-intrinsic MLKL functions in the development of MLKL-targeted therapies for precision cancer immunotherapy.