Extended Data Fig. 8: Cancer cell STING increases NK and T cell levels in metastases.

(a) Schematic of flow cytometry or immunofluorescence analysis of immune cells in bone metastasis. (b) Flow cytometry analysis and quantification of CD45⁺ leukocytes, CD45⁺CD19⁻TCRβ⁻XCR1⁺CD11c⁺ cDC1 cells, CD45⁺NK1.1⁺TCRβ⁻CD49b⁺NKp46⁺ NK cells, CD45⁺TCRβ⁺NK1.1⁻CD8⁺ T cells, and CD45⁺TCRβ⁺NK1.1⁻CD4⁺ T cells in KP-482T1 metastasis-bearing femurs after Dox-induced expression of STING for 2 days or 7 days. Cell numbers per gram of femur tissue were counted and normalized to control (-Dox). Metastasis-bearing femurs were collected 2 weeks after intracardiac inoculation of 2 x 10⁴ KP-482T1 cells in B6-albino mice. n = 7 mice per group. Mean ± s.e.m., two-sided unpaired t-test. p values comparing each Dox treatment group with control (−Dox). (c–f) Percentage of NK cells degranulating (CD107a⁺) (c) or producing IFNγ (d), and percentage of CD44⁺CD8⁺ T cells producing IFNγ (e) or TNF (f), isolated from KP-482T1 metastasis-bearing femurs after Dox-induced expression of STING for 2 days or 7 days, and then cultured ex vivo with phorbol 12-myristate 13-acetate (PMA) and ionomycin for 4h. n = 5 (+Dox 7 days) or 6 (other groups) mice per group. Mean ± s.e.m. (g) Schematic of the NK cell-mediated killing assay, and the percentage of WT or STING KO H2087-LCC cells killed by incubation with naïve NK cells for 4h at the indicated effector:target ratios. n = 3 per group. Mean ± s.e.m. (h) Schematic of the trans-well migration assay, and the number of NK cells migrated into cell culture media conditioned by WT or STING knockout H2087-LCC cells. n = 3 per group. Mean ± s.e.m., two-sided unpaired t-test.

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