Figure 7.

Inhibition of tumor recurrence in vivo. A, Five-day established subcutaneous B16tk tumors were treated with ganciclovir intraperitoneally on days 6 to 10 and 13 to 17. On day 27, mice with no palpable tumors were treated with control IgG, anti-asialo GM-1 (NK depleting), anti-TNFα, or anti–PD-1 every other day for 3 weeks and survival was assessed. Survival analysis was conducted using log-rank tests. The threshold for significance was determined by using the Bonferroni correction for multiple comparisons. Mice that developed a recurrent tumor were euthanized when the tumor reached a diameter of 1.0 cm. Eight mice per group, except for the ganciclovir/anti-asialo GM-1 group n = 9. *, P < 0.01 Representative of two separate experiments. B, Triplicate cultures of 10⁶ splenocytes and lymph node cells from C57BL/6 mice were incubated with PBS, LPS (25 ng/mL), or CpG for 48 hours, and supernatants were assayed for TNFα by ELISA. Mean and SD of triplicates are shown; ***, P < 0.0001 PBS versus LPS (t test). C, Cumulative results from skin explants at the sites of tumor from tumor-regressed mice treated with ganciclovir (B16tk tumors), reovirus therapy (B16tk cells), or OT-I adoptive T-cell therapy (B16ova cells). Explants were cocultured with 10⁶ splenocytes and lymph node cells from C57BL/6 mice in the presence of PBS, LPS, CpG, or LPS plus anti-TNFα (0.4 μg/mL). Seven days later, adherent B16 tumor cells were counted and wells containing >10⁴ cells were scored for active growth of MRD cells. P < 0.001 LPS versus all other groups (ANOVA). D, Five-day established subcutaneous B16tk were treated with ganciclovir intraperitoneally on days 6 to 10 and 13 to 17. On days 27 and 29, mice with no palpable tumors were treated with LPS (25 μg/injection). Mice were treated in-parallel with control IgG, anti-asialo GM-1, anti-TNFα, or anti-PD-1 every other day for 3 weeks. Mice with recurrent tumors were euthanized when the tumors reached a diameter of 1.0 cm. Survival of mice with time is shown. **, P < 0.01; ***, P < 0.001. Survival analysis was conducted using log-rank tests. The threshold for significance was determined by using the Bonferroni correction for multiple comparisons. Representative of two experiments.