Recently, a research team led by Jean-Christophe Marine and Joanna Pozniak of the Flemish Institute of Biotechnology (VIB) in Belgium, and Oliver Bechter of the University Hospital of Leuven, published an important research paper in the famous journal Cancer Discovery. When analyzing tumor samples from melanoma patients who did not respond to immunotherapy, they found that the number of natural killer (NK) cells increased significantly, and both NK cells and killer T cells gathered around the tumor. The amazing thing is that if the NK cells around the tumor are eliminated and then supplemented with immune checkpoint inhibitors, the killer T cells will infiltrate and destroy the tumor.
Even more unexpectedly, the researchers also found that after receiving immune checkpoint inhibitor treatment, the expression of the chemokine receptor CX3CR1 of NK cells increased, and they were recruited to the periphery of the tumor, thereby inhibiting the infiltration and anti-tumor ability of killer T cells to the tumor.
As we all know, immunotherapy has rewritten the paradigm of cancer treatment. However, only a small number of cancer patients have truly benefited from immunotherapy, and most patients do not respond to immunotherapy. There are many reasons behind this, and the "blackening" of immune cells is one of the more important ones. Needless to say, there are too many studies on macrophages and neutrophils being "blackened" by tumors. However, in the past one or two years, studies have successively found that NK cells, the key effector cells of anti-tumor immune response, also seem to do something beneficial to tumors. However, it is still unknown whether NK cells directly mediate immunotherapy resistance.
In order to answer the above questions, the Marine team collected tumor tissue samples from more than 20 newly diagnosed melanoma patients before and after treatment with immune checkpoint inhibitors, and conducted in-depth analysis. What puzzled them was that for patients who did not respond to immunotherapy, NK cells were significantly enriched in their tumor samples after treatment, while other immune cells did not show a correlation with treatment response. NK cells are enriched in samples that do not respond to immunotherapy. Aren't NK cells the main anti-cancer force? Is it wrong? So they found an external melanoma dataset and came to the same conclusion; they even found a similar phenomenon in a breast cancer dataset.
Although previous studies have found that some NK cell subpopulations are associated with immunotherapy resistance, the Marine team noticed that the NK cells they found were significantly different from the previous NK cells, or they were not a subtype. Because the NK cell subtypes discovered by the Marine team mostly express the cytotoxic genes GNLY, GZMB and PRF1, indicating that they are cytotoxic.
As for the source of these effector/cytotoxic NK cells around the tumor, the researchers analyzed the changes in peripheral blood NK cells before and after immunotherapy. It was found that after immunotherapy, the percentage of GZMB-positive and PRF1-positive NK cells in the peripheral blood of patients who did not respond to treatment decreased. This shows that immunotherapy promotes the aggregation of NK cells around the tumors of patients who do not respond to immunotherapy.
It is worth noting that these effector/cytotoxic NK cells co-localize with killer T cells around the tumors of patients who do not respond to immunotherapy. Simply put, in patients who do not respond to immunotherapy, effector/cytotoxic NK cells and killer T cells do not enter the tumor, but gather around the tumor.
In order to explore the role of these NK cells, the Marine team conducted research using an immune rejection mouse model. They found that neither PD-1 inhibitors alone nor anti-NK1.1 monoclonal antibodies that eliminate NK cells have an effect on tumors (anti-NK1.1 monoclonal antibodies can promote the increase of cytotoxic T cells). However, the combination of the two can further promote the proliferation of cytotoxic T cells and increase the infiltration of immune cells into tumors, ultimately inhibiting tumor growth. If cytotoxic T cells are eliminated, the anti-tumor effect of PD-1 inhibitors combined with anti-NK1.1 monoclonal antibodies disappears.
It is not difficult to see that the interaction between NK cells and cytotoxic T cells plays an important role in tumors that do not respond to immunotherapy.
Figure 1. NK cells as modulators of ICB sensitivity in mice. (Pozniak, Joanna, et al. 2025)
As for why PD-1 inhibitors promote the aggregation of effector/cytotoxic NK cells around tumors, the Marine team found that PD-1 inhibitors lead to increased expression of chemokine receptor CX3CR1 of NK cells, which promotes the aggregation of NK cells around tumors. In addition, upregulation of CD355 (Nkp46) and CXCR1 was observed, indicating that NK cells become mature after treatment with immune checkpoint inhibitors. Combining the CX3CR1 inhibitor JMS-17-2 with PD-1 inhibitors can significantly inhibit tumor growth.
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At the end of the study, the Marine team tested the effect of daratumumab (anti-CD38 monoclonal antibody), which can clear peripheral blood NK cells, in combination with PD-1 inhibitors and found that it can significantly inhibit tumor growth. At the same time, the number of NK cells around the tumor decreased and the infiltration of cytotoxic T cells in the tumor increased. Obviously, this combined treatment may help some cancer patients overcome immunotherapy resistance.
In general, the research results of the Belgian VIB team confirmed that NK cells play a direct role in immunotherapy resistance and preliminarily revealed the mechanism behind it. More importantly, they also found potential treatment options for this resistance mechanism.
Reference
- Pozniak, Joanna, et al. Cytotoxic NK Cells Impede Response to Checkpoint Immunotherapy in Melanoma with an Immune-Excluded Phenotype. Cancer Discovery (2025): OF1-OF16.