Myeloma killing by CD4⁺T cells is dependent on IFN-γ and depends on asialo-GM1–expressing effector cells. (A) ELISA quantitation of IFN-γ levels in the cell-free fraction of the tumor-infiltrated femoral bone marrow of TCR-Tg or wild-type BALB/c mice on day 12 after IV challenge with 1 × 10⁶ MOPC315.BM-luc cells (means ± SD; n = 6 mice per treatment group). (B) Survival of wild-type (WT) and TCR-Tg BALB/c mice challenged IV with 1 × 10⁶ MOPC315.BM-Luc cells. Mice were treated every 3 days with IP injection of 200 μg of a blocking mAb against IFN-γ (aIFNg) or isotype control mAb (Iso ctr.) as indicated, starting 1 day before tumor cell injection (n = 8 mice per treatment group). Representative dorsal bioluminescence imaging data for TCR-Tg mice treated with aIFNg or isotype mAb is shown on the right. (C) Serum M315 Id levels from experiment described in panel B. (D) Survival of GATA1^−/− BALB/c (GATA1^−/−) and wild-type BALB/c (WT) mice challenged IV with 1 × 10⁶ MOPC315.BM cells and treated by adoptive T-cell therapy. Mice were irradiated (500 cGy) on day 18 after tumor challenge and injected IV with 2 × 10⁶ Id-specific CD4⁺TCR-Tg cells (ACT) or irrelevant, polyclonal CD4⁺ T cells (Ctr.) the next day (n = 6 mice per treatment group). (E) Survival of TCR-Tg and wild-type BALB/c (WT) IV challenged with 1 × 10⁶ MOPC315.BM cells. Mice received IP injections of 200 μg anti-Ly6C mAb 1A8 (aLy6G) or isotype control mAb (Iso ctr.) every other day, starting 5 days before tumor challenge (n = 6 mice per treatment group). (F) Survival curves and representative bioluminescence imaging data for TCR-Tg BALB/c mice challenged IV with 1 × 10⁶ MOPC315.BM-Luc cells. Mice were treated every 3 days with IP injection of a blocking mAb against asialoGM1 (a-AGM1) or isotype control mAb (Iso ctr.) as indicated, starting 1 day before tumor cell injection (n = 8 mice per treatment group). (G) Survival of IV MOPC315.BM tumor-challenged wild-type BALB/c and BALB/c Rag^−/− IL2rgc^−/− (Rag/yC^−/−) mice treated with 2 × 10⁶ Id-specific CD4⁺TCR-Tg cells (ACT) or irrelevant, polyclonal CD4⁺ T cells (Ctr.) on day 18 after tumor cell injection (n = 6 mice per treatment group). (H) Representative flow cytometry staining of asialo-GM1⁺ cells isolated from femoral bone marrow cells harvested from TCR-Tg or nontransgenic BALB/c (WT) mice on day 5 after IV challenge with 1 × 10⁶ MOPC315.BM cells. Cells were isolated by incubation with anti-asialo GM1 antibody, followed by positive selection by anti-rabbit IgG MicroBeads.