Fig. 5: Tumor-directed immunoradiotherapy enhances dendritic cell antitumor surveillance across the tumor and sentinel lymph node.

A Cartoon schema of the experimental approach. WT animals injected with 4MOSC1 tumors were treated with tdRT→αPD-1 and then subjected to sentinel lymph node (SLN) mapping. RNA sequencing from the sentinel lymph node showing normalized enrichment scores for various immune response pathways post-treatment. X axis represents gene sets ranked by normalized enrichment score (NES); Y-axis represents the −log₁₀(FDR q value). B ELISA quantification of CCL19 levels in the TME post-treatment (n = 5 mice per group, p = 0.0008). Data are presented as mean values ± SEM (n = 5 biologically independent samples/group); p values calculated by two-sided unpaired Student’s t test. C Flow cytometric analysis of activated dendritic cells (MHCII+ CD11c + CCR7 + ) in the SLN post-treatment (n = 5 mice per group, p = 0.0432). Data are presented as mean values ± SEM; p values calculated by two-sided unpaired Student’s t test. D Cartoon schema of the experimental approach. ROSA-26 x Ai9 animals were injected with 4MOSC1 tumors were treated with tdRT→αPD-1, labeled with tamoxifen in the tumor and then subjected to sentinel lymph node (SLN) mapping. Sorted live tdTomato+ cells from the SLN were then sent for CITE-sequencing. CITE-sequencing was performed on sorted tdTomato+ cells isolated from the SLNs, as described in the methods. n = 2 biologically independent samples/group. E (left) Subsampled dendritic cell populations after CITE-sequencing, showing UMAP plots with clusters of dendritic cells (right) Seurat objects featuring the expression of CCR7, CD40, CD86, and MHCII across DC populations. UMAP clustering and Seurat object analysis were conducted on the CITE-sequencing data to identify dendritic cell subpopulations. F Multiplex immunofluorescence from sentinel lymph nodes showing CD11c, CCR7, CD11b, CD40, and DAPI staining in control and tdRT-treated groups. Representative of n = 3 biologically independent samples; experiment was independently repeated at least twice with similar results. G Analysis of average expression from the DC-3 population showing activation in programs of DC migration, phagocytosis, antigen processing and presentation, and interferon signaling pathways. Representative of n = 2 biologically independent samples. H Flow cytometric analysis of conventional dendritic cells (cDC1, MHCII + XCR1 + ) in the SLN post-treatment (n = 5 mice per group, p = 0.0398). Data are presented as mean values ± SEM; p values calculated by two-sided unpaired Student’s t test. I (left) Cartoon schema of the experimental approach for Batf3−/− animals, which lack conventional type I dendritic cells. (right) Tumor growth curves for control and tdRT→αPD-1 treatment in Batf3−/− animals (n = 5 mice per group, p = ns). Data are presented as mean values ± SEM (n = 5–6 biologically independent samples/group). Best-fit lines and p values calculated by simple linear regression (two-sided). Created in BioRender. Saddawi-Konefka, R. (2025) https://BioRender.com/m675tdd. Source data are provided as a Source Data file.