Figure 2. The impact of TAMs depletion on M1 virus viral load and on immune-activating transcriptional signals in tumor. Mice administration pattern. After MC38-derived tumor volumes reached 50–100 mm³, mice were randomly grouped. On Day 0 and Day 3, clodronate liposomes were administered via tail vein injection. On Days 1–3, M1 virus was administered via tail vein injection (I.V.). (B) Intratumoral infiltration of TAMs following clodronate liposome treatment. MC38-derived tumors were harvested 4 days after the first M1 virus dose (Day 10) and processed for flow cytometric analysis. Vehicle, control liposome. Clodronate, clodronate liposome. *p<0.05. (C) Viral copies of M1 virus in tumor. For virus copies testing, tumors were harvested at 2, 50, 96, and 144 hours after the first M1 dose, and were subject to qPCR. ns, no statistical difference. *p<0.05. **p<0.01. (D–F) Volcano plot of differentially expressed genes (D) and KEGG pathway enrichment signaling pathway map for differentially expressed genes (E) or for anti-pathogen immune pathways (F). MC38-derived tumors were harvested 4 days after the first M1 virus dose (Day 10) and processed for RNA sequencing. (G–K) GSEA analysis for T-cell receptor immune pathways (G), Th1 and Th2 cell immune pathways (H), PD-1 and PD-L1 immune checkpoint pathways (I), NK cell cytotoxic immune pathways (J) and apoptosis signaling pathways (K). ns, no statistical difference. *p<0.05. **p<0.01. GSEA, Gene Set Enrichment Analysis; I.V., intravenously; KEGG, Kyoto Encyclopedia of Genes and Genomes; NK, natural killer; PD-1, programmed cell death protein-1; PD-L1, programmed death-ligand 1; qPCR, quantitative PCR; TAM, tumor-associated macrophage.