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. 2020 Jun 22;9(1):1771925. doi: 10.1080/2162402X.2020.1771925

Figure 3.

Figure 3.

Adding the CD4 neoantigen vaccine to LRT results in a potent polyantigenic CD8+ T cell response and T cell memory. (a) IFNγ ELISpot using splenocytes isolated from CT26 tumor-bearing BALB/c mice, vaccinated with CT26 PME1 or control RNA-LPX and locally irradiated at a mean tumor volume of 45 mm3 (n = 6 mice/group, 2 mice pooled each), against CT26 cells, gp70-AH1 peptide-pulsed BALB/c BMDC, CT26-gp70KO, and 20 Gy irradiated CT26-gp70KO cells. In vitro irradiation was performed to enhance tumor cell MHC class I presentation (Supplementary Figure 3(a)). (b-d) CT26-gp70KO tumor growth (b) and survival (c) of mice immunized with CT26 PME1 or control RNA-LPX and irradiated at a mean volume of 45 mm3 (n = 7–10/group). (d) IFNγ ELISpot using splenocytes isolated from CT26 tumor-bearing mice (n = 4 mice/group, 2 mice pooled each) against CT26 cells, 20 Gy irradiated CT26-gp70KO cells and gp70-AH1 peptide-pulsed BALB/c BMDC. As in (a), in vitro irradiation was performed to enhance tumor cell MHC class I presentation (Supplementary Figure 3(a)). (e) Survival of 12 Gy and CT26 PME1/ME1 RNA-LPX treated CT26 tumor-free mice, challenged with a tumorigenic dose of CT26-gp70KO cells 40 days after initial tumor rejection (n = 10 each). Naïve BALB/c mice served as control group (n = 10). Significance was determined using (c, e) log-rank test and (a, d) one-way ANOVA, Tukey’s multiple comparison test. (b) Tumor growth is displayed on a log2-scale. Ratios depict frequency of mice with complete tumor responses (CR). Mean±SEM.