Fig. 3

TEDbody-mediated in vivo CMV-pMHCI presentation suppresses tumor growth by redirecting adoptively transferred CMVp-CTLs to tumor cells in mice. A IHC detection of CMV-pMHCI (red) on MDA-MB-231 tumor tissues excised from NSG mice bearing a preestablished MDA-MB-231 cell-derived tumor xenograft (100–120 mm³), 24 h after a single i.p. injection of the indicated TEDbody or control Ab (20 mpk). Images are representative of three independent experiments; additional images are shown in Fig. S7A. Nuclei were stained with Hoechst 33342 (blue). Scale bar: 20 μm. B Functional phenotype analysis of CMVp-CTLs from MDA-MB-231 tumor tissues, excised from NSG mice bearing a preestablished MDA-MB-231 tumor (100–120 mm³), 24 h after a single i.p. injection of the indicated TEDbody, control Ab (20 mpk), or peptide (equivalent molar amount of 20 mpk TEDbody), followed 6 h later by peritumoral injection of ex vivo-expanded CMVp-CTLs (5 × 10⁶ cells), as described in the upper panel. Each symbol represents a value obtained from an individual mouse. The data from inCT and inCTp_480–503 were pooled from two independent experiments with at least three mice per group. The bar graphs present the mean ± SEM (n ≥ 3). C The treatment scheme for assessing in vivo antitumor efficacy of the TEDbody or a control Ab in conjunction with adoptive transfer of ex vivo-expanded CMVp-CTLs in NSG mice carrying a preestablished HCT116 cell-derived subcutaneous tumor xenograft or MDA-MB-231 cell-derived orthotopic tumor xenograft (100–120 mm³). The arrows indicate each time point for the treatment or assay. D Tumor growth, measured as the average tumor volume, in response to the indicated treatment, as described in (C). Error bars: ±SEM (n = 9 to 14 per group for HCT116 tumors, n = 8 to 13 per group for MDA-MB-231 tumors). Data were pooled from two independent experiments with at least four mice per group. E and F IHC detection of CMV-pMHCI (red) on tumor tissues (E) and the number of tumor-infiltrating CMVp-CTLs per gram of a tumor (F) excised from mice on day 3 after the last treatment, as described in (C). In (E), nuclei were stained with Hoechst 33342 (blue), and images are representative of three independent experiments; additional images are shown in Fig. S7B. Scale bar: 20 μm. The right panel shows the quantification of red fluorescence intensity, obtained by ImageJ software. Error bars, ±SD of 2 fields per tumor (n = 3 per group). In (F), bar graphs present the mean ± SEM (n ≥ 3 different tumors). In (B), (D), and (F), **P < 0.01 and ***P < 0.001 denote a significant difference between the indicated groups (B and F) or a significant difference from the inCT group (B, D, and F), as determined by one-way ANOVA with the Newman–Keuls post hoc test; ns: not significant