Fig. 4: Genomic knockout of GPR171 exhibits stronger antitumor immunity.

a, b GPR171^LacZ/lacZ mice were inoculated with MC38 tumors for 14 days. The expression of GPR171 (β-gal) on T cells and NK cells in dLN and tumor were quantified by flow cytometry (a). In GPR171^lacZ/lacZ tumors, TILs were examined for the expressions of GPR171 together with immune checkpoints (ICs), including PD-1, LAG3, TIGIT, and TIM3 (b). c–h GPR171^lacZ/lacZ GPR171^+/lacZ, and WT littermates were inoculated subcutaneously with MC38 tumor cells. Tumor growth curve (c) and tumor weight (d) at day 16 after tumor inoculation were determined. Single-cell suspensions were prepared from MC38 tumors to determine the densities of immune cells (e). The frequency of CD4+ Treg cells (f), the ratio of CD8/Treg (g), as well as the percentages of CD8+ T cells expressing Granzyme B or CD137 (h), within tumors were quantified. (i) MC38 tumor weights after 19 days of tumor inoculation were determined. In some mice, NK cells, CD4 + T cells, or CD8+ T cells were eliminated by corresponding depleting antibody. j–n GPR171^lacZ/lacZ and WT littermates were injected intravenously with B16F10 tumor cells. Overall survival of mice was followed till 60 days after tumor inoculation (j). 4 weeks later, gross images of lung metastasis were taken (k); lung weight (l) and tumor nodules in the lung (m) were quantified. The incidences of tumor metastasis in organs other than lung were recorded (n). a n = 8 biologically independent samples. b n = 4 biologically independent samples. c, d n = 8 (WT littermates and GPR171^lacZ/lacZ) and n = 6 (GPR171^+/lacZ) biologically independent samples. e, i n = 8 (WT littermates) and n = 10 (GPR171^lacZ/lacZ) biologically independent samples. f–h n = 8 (WT littermates) and n = 8 (GPR171^lacZ/lacZ) biologically independent samples. j n = 7 biologically independent samples. k–n n = 7 biologically independent samples. Statistical significance was determined by one-way ANOVA for d and i, two-tailed Student’s t-test for a, b, d, e, f, g, h, l and m, two-way ANOVA for c, log-rank test for j or chi-square test for n. Unless otherwise denoted, values are mean ± SEM. Source data was provided as a Source Data file. Data are representative of two (a, b, i, j, k, l, m, and n) or three independent experiments (c, d, e, f, g, and h).