Geldanamycin abrogates increased immunogenicity of bortezomib-killed myeloma cells. (A) DCs were pulsed with U266 myeloma cells killed by γ irradiation, bortezomib alone, or with myeloma cells preincubated with geldamamycin for 1 hour before treatment by bortezomib. Tumor-loaded DCs with or without additional maturation stimulus with LPS were then used as stimulators of autologous T cells for 2 weeks. On day 7, T cells were restimulated with fresh tumor cell–pulsed DCs. On day 14, IFNγ production by tumor-reactive T cells in response to U266 myeloma cells was quantified by an overnight ELISPOT. Response to an HLA-A2–negative cell line cag or unpulsed autologous DCs was used as a control. Data shown are mean/SD of 3 independent experiments on 3 blood donors. (B) DCs were fed with apoptotic U266 tumor cells killed by γ irradiation or bortezomib, with or without 1 hour preincubation of U266 cells with geldanamycin. Pulsed DCs then were activated with LPS or left untreated. DCs then were used to stimulate T cells as in panel 2A. On day 14 of culture, IFNγ production in response to U266 tumor cells was monitored by intracellular cytokine flow cytometry. Data are representative of similar experiments on 3 donors. Response to HLA-A2 negative cag cells was used as a negative control and did not exceed 0.1% (not shown).