Figure 7. Chaperone vaccines targeting dual antigens induce potent therapeutic antitumor immunity.

A. Superior therapeutic efficacy generated by the combined chaperone vaccines. Mice (n=5) were established with B16 tumors 3 days prior to vaccine treatment. Grp170-TRP2 complex, tumor-derived grp170 or grp170-gp100/TRP2 complexes were administrated every three days for a total 4 treatments (*, p< 0.01). B. Priming of T-cells reactive with gp100_25–33 following TRP2-chaperone vaccine treatment. Splenocytes from treated mice were stimulated with gp100_25–33 or TRP2_180–188 and subjected to an ELISPOT assay (*, p< 0.001; **, p< 0.01; ***, p < 0.001). Data shown are from two independent experiments. C. Potent antitumor immunity augmented by endogenous grp170 complexed with gp100/TRP2 proteins. B16 tumor-bearing mice were treated with tumor-derived grp170, antigen ‘stripped’ grp170, or ‘stripped’ grp170 complexed with gp100/TRP2 protein antigens. Tumor growth was followed as described (* and **, p< 0.01). D. High frequency of gp100/TRP2-specific TILs elicited by the chaperone vaccines. TILs were isolated from pooled tumor tissues after treatment and stimulated with gp100_25–33/TRP2_180–188 peptides, followed by intracellular staining for IFN-γ. Cells were analyzed using FACS by gating on CD8⁺ cells. E. Robust tumor-killing activity of TDLN cells from chaperone vaccine-treated mice. Single cell suspensions were prepared using TDLNs pooled from treated mice. Cells were stimulated with mitomycin and IFN-γ-treated B16 cells for 5 days. T-cells were used as effector cells in cytotoxic assays against B16 cells as targets (*, p< 0.005). Data shown are from two independent experiments.