Myeloma cells recruit Tregs and express B7H1. (A) Cytokines secreted by tumor-specific T cells in tumor-free and tumor-bearing mice. Balb/c mice were subcutaneously vaccinated twice with either normal g96N, autologous gp96A, or pooled heterologous gp96BC or gp96BCD, followed by challenge with tumor-A cells. Three weeks later, splenocytes were isolated from tumor-bearing and tumor-free mice, and restimulated for 5 days with irradiated tumor-A cells. Culture supernatants were collected and assayed by enzyme-linked immunosorbent assay to quantify IFN-γ, IL-4, and IL-10. (B) Percentages of CD4⁺Foxp3⁺ Tregs in the spleens of tumor-free and tumor-bearing mice and in subcutaneous tumors. (C) B7H1 expression on murine tumors A, B, C, D, E, and F. (D) Blocking B7H1 enhanced the killing activity of tumor-specific CTLs. Shown is the cytolytic activity of splenocytes from mice vaccinated with gp96BCD and ex vivo restimulated with irradiated tumor-A cells for 5 days, in cultures with or without the addition of anti-B7H1 mAb (M5H1; 20 μg/mL) or an isotype control rat IgG. Target cells were tumors A, B, C, or D cells. Representative results of 3 independent experiments are shown. *P < .05; **P < .01 (compared with controls). The error bars in panels A and D represent SD of 3 independent experiments.