CD73 immune checkpoint defines regulatory NK cells within the tumor microenvironment
Shi Yong Neo, Ying Yang, Julien Record, Ran Ma, Xinsong Chen, Ziqing Chen, Nicholas P. Tobin, Emily Blake, Christina Seitz, Ron Thomas, Arnika Kathleen Wagner, John Andersson, Jana de Boniface, Jonas Bergh, Shannon Murray, Evren Alici, Richard Childs, Martin Johansson, Lisa S. Westerberg, Felix Haglund, Johan Hartman, Andreas Lundqvist
Shi Yong Neo, Ying Yang, Julien Record, Ran Ma, Xinsong Chen, Ziqing Chen, Nicholas P. Tobin, Emily Blake, Christina Seitz, Ron Thomas, Arnika Kathleen Wagner, John Andersson, Jana de Boniface, Jonas Bergh, Shannon Murray, Evren Alici, Richard Childs, Martin Johansson, Lisa S. Westerberg, Felix Haglund, Johan Hartman, Andreas Lundqvist
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Research Article Immunology

CD73 immune checkpoint defines regulatory NK cells within the tumor microenvironment

Abstract

High levels of ecto-5′-nucleotidase (CD73) have been implicated in immune suppression and tumor progression, and have also been observed in cancer patients who progress on anti–PD-1 immunotherapy. Although regulatory T cells can express CD73 and inhibit T cell responses via the production of adenosine, less is known about CD73 expression in other immune cell populations. We found that tumor-infiltrating NK cells upregulate CD73 expression and the frequency of these CD73-positive NK cells correlated with larger tumor size in breast cancer patients. In addition, the expression of multiple alternative immune checkpoint receptors including LAG-3, VISTA, PD-1, and PD-L1 was significantly higher in CD73-positive NK cells than in CD73-negative NK cells. Mechanistically, NK cells transport CD73 in intracellular vesicles to the cell surface and the extracellular space via actin polymerization–dependent exocytosis upon engagement of 4-1BBL on tumor cells. These CD73-positive NK cells undergo transcriptional reprogramming and upregulate IL-10 production via STAT3 transcriptional activity, suppressing CD4-positive T cell proliferation and IFN-γ production. Taken together, our results support the notion that tumors can hijack NK cells as a means to escape immunity and that CD73 expression defines an inducible population of NK cells with immunoregulatory properties within the tumor microenvironment.

Authors

Shi Yong Neo, Ying Yang, Julien Record, Ran Ma, Xinsong Chen, Ziqing Chen, Nicholas P. Tobin, Emily Blake, Christina Seitz, Ron Thomas, Arnika Kathleen Wagner, John Andersson, Jana de Boniface, Jonas Bergh, Shannon Murray, Evren Alici, Richard Childs, Martin Johansson, Lisa S. Westerberg, Felix Haglund, Johan Hartman, Andreas Lundqvist

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Figure 5

Characterization of CD73+ NK cells based on differential gene expression.

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Characterization of CD73+ NK cells based on differential gene expression...
(A) CD73 enzymatic activity normalized to CD73+ T cells from PBMCs from healthy donors (n = 3). Student’s t test was used to assess significance. (B) Left: Representative histogram showing proliferation of CD4+ responder T cells after 2 days of coculture with different sorted populations on NK cells at a 10:1 ratio (n = 4). Right: Percentage of dividing CD4+ T cells in a 48-hour suppression assay. Significance was tested with Wilcoxon’s signed-rank test (n = 8). (C) Volcano plot generated to visualize the significance and magnitude of changes in gene expression comparing CD73+ NK cells versus CD73– NK cells. The x axis represents the fold change between the 2 groups and is on a log2 scale, and the y axis shows the negative log10 of the P values from mixed-model ANOVA. Genes with significant fold change less than 2 are represented in red (n = 5). (D) Gene expression heatmap generated to visualize immune-related genes that are differentially expressed in CD73+ NK cells based on Supplemental Table 1 (n = 5). (E) Gene ontology enrichment analysis showing functional pathways for the top 100 genes upregulated by CD73+ NK cells from the gene list in Supplemental Table 2. Each node has a minimum of 3 genes and the size of the nodes is proportional to the number of genes. Color of the nodes is determined by the significance of the enriched term assessed by 2-sided hypergeometric test with Bonferroni’s step-down method for P value correction (n = 5).