Fig. 6

Targeting CD39 on macrophages is a potential therapeutic strategy to overcome immunotherapy resistance in HCC. a Tumor size was measured over time after H22^Ctrl (H22 overexpressing Control) or H22^OE (H22 overexpressing circTMEM181) were used to construct tumor xenografts in the C57^wt (C57BL/6 wild-type) or C57^{Entpd1−/−} (C57BL/6 Entpd1-knocked-out) mouse models (n = 6 for each group; one-way ANOVA: ***: p < 0.001, **: p < 0.01, *: p < 0.05, ns: not significant). b Flow cytometry analysis shows the proportion of CD8⁺ T cells and GZMB⁺ CD8⁺ T cells in the tumor microenvironment of HCC mouse models (one-way ANOVA: ***: p < 0.001, **: p < 0.01, *: p < 0.05, ns: not significant). c Tumor size was measured over time following PD1 antibody (αPD1), POM1 or combination therapy (αPD1 + POM1) in C57 mouse models (n = 6 for each group; one-way ANOVA: ***: p < 0.001, ns: not significant). d Flow cytometry analysis shows proportion of CD8⁺ T cells and GZMB⁺ CD8⁺ T cells in the tumor microenvironment of HCC mouse models (one-way ANOVA: ***: p < 0.001, **: p < 0.01, *: p < 0.05, ns: not significant). e Tumor size was measured over time following treatment with PD1 antibody + infusion of exosome enriched from H22^OE medium (αPD1 + i.v. Exo^OE) or H22 Ctrl medium (αPD1 + i.v. Exo^Ctrl) in C57^wt mouse models (n = 5 for each group; t test). f Flow cytometry analysis of the tumor microenvironment indicates a significant increase in CD39 expression on TAM (CD11b⁺ F4/80⁺) after infusion of exosomes enriched from H22^OE medium and the CD39^high TAM tended to be co-expressed with CD163^high (Top). Statistical results show the proportion of CD8⁺ T cells and GZMB⁺ CD8⁺ T cells in the tumor microenvironment of HCC mouse models (t test). g Representative fluorescence photography of the two indicated groups at different time points after anti-PD1 therapy in the orthotopic xenograft model of liver cancer (t test, **: p < 0.01). h tSNE plot of different major clusters in CD45⁺ tumor-infiltrating immune cells from the two indicated groups. i tSNE plot of CD39 fluorescence intensity in different clusters between the two indicated groups. j Flow cytometry analysis shows proportion of CD8⁺ T cells and GZMB⁺ CD8⁺ T cells in the tumor microenvironment (t test). k Flow cytometry analysis shows CD73 expression on CD45⁻ H22 tumor cell in the HCC mouse model (t test). l Measurement of serum ATP and ADO concentrations 20 days after anti-PD1 therapy in the orthotopic xenograft model of liver cancer (t test)