FIGURE 2.

Lm-LLO-Flk-1 vaccines can induce regression of established Her-2/neu⁺ tumors in vivo. A, NT-2 tumor volume (mm³) from mice treated with each construct. Graph shows mean ± SEM; *, p < 0.05; Mann-Whitney statistical test, n = 8 mice per group, experiment repeated twice. B, IFN-γ ELISPOTs showing epitope spreading to various Her-2/neu regions. Splenocytes from the 64-day time point were restimulated ex vivo with Her-2/neu peptide epitopes. Graph shows mean ± SEM; *, p < 0.05; Mann-Whitney statistical test, n = 5 mice per group, experiment repeated once. C, Mice that had fully regressed tumors were rechallenged with NT-2 in the contralateral flank on day 100. A saline-treated group was used as our negative control for tumor growth. D, Tumor volume for mice that grew tumors after rechallenge on day 100 of tumor-free mice. Both graphs refer to a single experiment. Number of tumor-free mice was two of eight for Flk-E1 and Flk-I1 groups; the saline group had five mice. E, Mice were immunized thrice over the course of 3 wk after the initial establishment of NT-2 tumors. In this figure (right-hand panel), we show staining for the pan-endothelial marker CD31-PE and nucleus using DAPI. Isotype controls were used on sequential sections, as shown to the right. Quantitation of vessel density, shown in the left-hand panel, was performed using Image Pro software. Graph shows mean ± SEM; *, p < 0.05; Mann-Whitney U test.