Fig. 1
Position and epitope context of heterologous antigens in MCMV-vectored vaccines impact vaccine-elicited T cell responses and associated tumor protection. a Schematic of MCMV-IE2-E6/E7 full length replace/fusion and MCV-IE2-E7. The MCMV genome region at ∼kb 186–187 corresponds to the MCMV gene ie2 (enlarged). Constructs encoding the E6/E7 fusion protein or the E7 epitope were generated by BAC recombineering. DNA sequences encoding the full-length E6/E7 fusion protein were inserted either to replace the ie2 gene (full length replace) or in frame at the very end of the ie2 gene (full length fusion). The E749–57 epitope RAHYNIVTF was fused to the end of ie2. b, c C57BL/6 mice were vaccinated IP with 1 × 105 PFU MCMV-IE2-E6/7 full length fusion, MCMV-IE2-E6/7 full length replace or MCMV-IE2-E7. Splenocytes were harvested 10 (b) or 21 (c) weeks after vaccination and stimulated with the immunodominant E749–57 peptide or with a pool of 15-mer peptides covering the entire E7 protein for 36 h. IFN-γ production was measured by ELISpot assay. Shown is the number of spots per 5 × 105 cells ± SEM. c Frequency of E749–57-specific T cells identified by MHC class I multimers within the total CD8+ cell population at 21 weeks post vaccination. Data represents mean values ± SEM (n = 4–5 mice per group). *, P < 0.05; **, P < 0.01. d Tumor outgrowth of TC-1 tumor cells in mice challenged with TC-1 tumor cells 21 weeks after vaccination with 1 × 105 PFU MCMV-IE2-E6/7 full length replace, MCMV-IE2-E6/7 full length fusion or MCMV-IE2-E7 (5–10 mice per group). The number of tumor-free/total mice is indicated in each tumor out growth graph