Bortezomib sensitizes RENCA and LLC1 to syngeneic NK-cell lysis in vivo. (A left panel) Tumor size by palpation in BALB/c mice injected with RENCA tumor cells (100 000 cells subcutaneously) and treated with bortezomib (5 μg/mouse intravenously) on days 3, 10, and 17, followed by injection of IL-2–activated syngeneic NK cells (10⁶ cells intravenously) on days 4, 11, and 18. Each symbol represents an individual mouse on day 29 after tumor injection. (Right panel) Tumor size by palpation in C57BL/6 mice injected with LLC1 tumor cells (500 000 cells subcutaneously) that were treated with bortezomib (15 μg/mouse intraperitoneally) on day 14, followed by injection of IL-2–activated syngeneic NK cells (1 × 10⁶ cells intravenously) on day 15. Each symbol represents an individual mouse on day 28 after tumor injection (horizontal bars show mean). (B) Quantification of pulmonary tumor nodules in RENCA tumor–bearing BALB/c mice after treatment with bortezomib (days 5, 12, and 19) and wild-type (wt) or perforin-deficient (pfp−/−) NK cells (2 × 10⁶ cells; days 6, 13, and 20). All animals received IL-2. Animals were killed on day 28 (left panel) and on day 30 (right panel) and evaluated for number of tumor nodules in the lung by manual counting (horizontal bars show mean). (C) In vivo bioluminescence assay and survival of animals treated with bortezomib and/or NK-cell infusions. Luciferase-transduced RENCA cells were injected, and mice were treated with 9 weekly injections of bortezomib and 2 to 5 × 10⁶ syngeneic NK cells starting 4 days after tumor injection. Values in bioluminescence graph (left panel) represent tumor doubling time. Values in survival graph (right panel) represent median survival days. Images show animals on day 39 after tumor injection. Values below each image represent the average tumor flux (photons/second [p/s]; × 10³) ± SD. All experiments were performed with 5 or more mice per group and repeated at least once. P values were calculated by 2-tailed unpaired t test or 2-tailed log-rank test.