FIGURE 5.
Squeezed cells prime antitumor activity in a mouse model of HPV16+ tumors. (A) Mice were left untreated (Untr.) or immunized with T cells squeezed with the E7 SLP containing the H2-Db-restricted epitope RAHYNIVTF and coinjected with CpG. Seven days later, E7-specific responses were measured in the spleen by intracellular cytokine staining (ICS). (B) Mice were prophylactically immunized with 1 × 106 T cells squeezed with E7 SLP and coinjected with CpG on day −7. On day 0, mice were challenged s.c. with TC-1. n = 10 mice per group. (C) Ten days after TC-1 inoculation, mice were immunized with 1 × 106 T cells squeezed with E7 SLP with or without coinjection of CpG. Mean tumor volume + SEM is shown. n = 10 mice per group. (D) Mice were left untreated or immunized with 5 × 106 B cells squeezed with E7 SLP and matured with CpG. Seven days later, E7-specific responses were measured by ICS. (E) Mice were immunized with 5 × 106 B cells squeezed with E7 SLP and matured with CpG overnight or left untreated. Seven days later, mice were challenged s.c. with TC-1. Survival is shown. n = 10 mice per group. (F) Mice were immunized 14 d after TC-1 implantation using 5 × 106 B cells pulsed with minimal epitope (Min. Epi.), RAHYNIVTF, or squeezed with E7 SLP and matured with CpG overnight. An additional group of mice was immunized and boosted on days 14 and 28 s.c. with 150 µg E7 SLP and 50 µg CpG. Mean tumor volume + SEM is shown. n = 10 mice per group. (G) Mice were immunized as indicated 14 d after TC-1 implantation. Seven days later, tumors were harvested for analysis by flow cytometry. Mean ± SD is shown. (H) Mice were left untreated or immunized with 1 × 106 splenocytes squeezed with E7 SLP and matured overnight with CpG. Seven days later, E7-specific responses were measured in the spleen by ICS. (I) Mice were immunized on days −14 and −7 with 1 × 106 M-SQZ-PBMC-HPV or left untreated. On day 0, TC-1 tumors were implanted, and tumor growth was monitored. Sixty days after inoculation, immunized mice were rechallenged with TC-1 on the opposite flank, and a new cohort of untreated mice was introduced. (J) Ten days after TC-1 implantation, mice were immunized with 1 × 106 splenocytes squeezed with E7 SLP that had been matured with CpG for 4 h or incubated without CpG. Tumor volume + SEM is shown. n = 10 mice per group. (K) Mice were primed with M-SQZ-PBMC-HPV at a dose of 1 × 106 or 1 × 105 cells on day 10. An additional group of mice was primed on day 10 and boosted two more times on days 17 and 24 (P/B/B). (L) Mice were immunized with 1 × 106 M-SQZ-PBMC-HPV or s.c. 150 μg E7 SLP with 50 μg CpG 16 d after TC-1 implantation. Twelve days later, tumor-infiltrating lymphocytes (TILs) were analyzed by flow cytometry. Mean percentage ± SD of indicated populations is shown. *p < 0.05, **p < 0.01, ***p < 0.001.