Fig. 4. Efficacy of intratumoral delivery of immune oncology agents.

a Left panel, C57Bl/6 mice were implanted with 1 × 10⁶ of 4MOSC1 cells into the tongue. After tumors reached ~30 mm³, mice were either treated IP or by intratumoral (IT) delivery of PBS, IP with 10 mg/kg or IT with 5 mg/kg anti-PD-1. Shown is the average volume of each tumor (n = 4 mice per group; two-sided Student’s t test; data are represented as mean ± SEM). Right panel, representative pictures of tongues from mice in a with tumors depicted with a dotted line. b Distribution of anti-PD-1 antibody in mice with 4MOSC1 tumors using IP or IT delivery of the treatment. Staining for anti-hamster IgG showed the localization of anti-PD-1 antibody in the tongue, lymph nodes, and spleen of treated mice (n = 4 mice per group). c RNA from each tumor was isolated and comprehensive immune profiling was analyzed using the NanoString nCounter PanCancer Mouse Immune Profiling gene expression platform. The advanced analysis module of the nSolver software was used to analyze genes associated with listed immune cells and given a score. Shown is the Z-score of each cell profile score (n = 3 mice per group). d Absolute number of live CD45⁺CD3⁺CD8⁺ T cells infiltrating 4MOSC1 tumors with or without anti-PD-1 or anti-CTLA-4 treatment. Shown is the average of the number of live CD8 T cells infiltrating per mm³ of tumor (n = 3 mice per group; two-sided Student’s t test; data are represented as mean ± SEM). e Frequency of live CD45⁺CD3⁺CD4⁺ FoxP3⁺ Tregs infiltrating 4MOSC1 tumors with or without anti-PD-1 or anti-CTLA-4 treatment. Left panel, a representative flow cytometry plot from one mouse showing the frequency of Tregs (CD4⁺FoxP3⁺) out of CD4⁺ cells is shown. Right panel, the frequency of Tregs out of CD4⁺ cells was quantified following treatment with anti-PD-1 or anti-CTLA-4 (n = 5 mice per group; two-sided Student’s t test; data are represented as mean ± SEM).