Mi-2 induces the immunogenic cell death via ER stress. (A-D) MM cells were treated with indicated concentrations of Mi-2 for 24 hours. The CRT expression was measured by flow-cytometric analysis, the bar graph shows the FC of CRT’s mean fluorescence intensity (MFI) compared with that of DMSO (A). The ATP released was assessed by ATP assay kit, the bar graph shows the FC of ATP compared with that of DMSO (B). The HSP70 and HSP90 protein exposure were measured by flow-cytometric analysis (left), the bar graph shows the FC of HSP proteins’ MFI compared with that of DMSO (right) (C). The HMGB1 secretion was assessed by enzyme-linked immunosorbent assay analysis, the bar graph shows the relative HMGB1 concentration in groups normalized to OD value at 450 nm (D). (E) DCs were cocultured with MM cells pretreated with DMSO or Mi-2 for 24 hours, and the expression of CD80 and CD86 were detected by flow cytometry (left). The bar graph shows the MFI of CD80 or CD86 (right). (F) The ration of dead cells of the following groups was determined by live/dead staining: H929, H929/DCs, H929/CD8⁺ T cells, 1 μM Mi-2–treated H929, H929/DCs/CD8⁺ T cells, and 1 μM Mi-2–treated H929/DCs/CD8⁺ T cells. The representative scatter plots are shown. (G) The bar graph shows that positive enriched pathways after Mi-2 treatment. FDR < 0.25. (H) Whole-cell lysates from OPM2 and H929 treated with the indicated concentration of Mi-2 for 24 hours were subjected to WB analysis and probed with indicated antibodies, with GAPDH as a loading control. ns, P > .05; ∗P < .05; ∗∗P < .005; ∗∗∗P < .001; and ∗∗∗∗P < .0001.