Fig. 2: Flt3L enhances cross-priming of anti-tumor CD8⁺ T cells upon NDV-cytolysis.

a Uninfected/NDV-preinfected GFP⁺ A20 cells were co-cultured with splenocytes from untreated/Flt3L-treated mice (contour plots show % cDCs of all splenocytes) and analyzed after 24 h. Representative viSNE plots showing myeloid cell populations (left) and relative expression (color code = mean MFI) of different markers (right). Bar graphs show mean MFI of cell-surface markers on cDCs (CD11c⁺I-Ad⁺). Repeated measures two-way ANOVA with Dunnett’s multiple comparisons test. n = 3 b Uninfected/NDV-preinfected GFP⁺ SUDHL4 cells were co-cultured with patient-derived PBMCs, pre- and post-Flt3L treatment, and analyzed after 24 h. Representative (n = 5) contour plots showing percent HLA-DR⁺CD11c⁺ cDCs (of all non-tumor cells) pre- and post-Flt3L treatment. Stacked bar graphs showing fold expression (vs ‘No NDV’) of activation markers in cDC1s (CD141⁺) and cDC2s (CD1c⁺). One-way ANOVA with Dunnett’s multiple comparisons test (n = 5). c Cells were cultured as in (a) with IFNAR-blocking or isotype-control antibodies and cDCs were analyzed. d Expression of Axl and Clec9A in cDCs cultured as in (a). c, d Graphs show No NDV vs 10 MOI data analyzed in triplicates, representative from at least 2 independent experiments. One-way ANOVA with Dunnett’s multiple comparisons test. e Tumor Ag (GFP)-uptake and MHC I expression in splenocytes from untreated/Flt3L-treated mice, after co-culture with uninfected/NDV-preinfected GFP⁺ A20 cells (n = 4); repeated measures One-way ANOVA with Sidak’s multiple comparisons test. f Tumor Ag (GFP)-uptake and MHC I expression in human cDC1 (CD141⁺) and cDC2 (CD1c⁺) DCs in PBMCs derived from Flt3L-treated patients, after co-culture with uninfected/NDV-preinfected GFP⁺ SUDHL4 cells. Repeated measures One-way ANOVA with Dunnett’s multiple comparisons test (n = 5). g NDV-preinfected MHC I-deficient GFP⁺ A20 cells were co-cultured with splenocytes (from untreated or Flt3L-treated mice) for 48 h and CellTrace violet-stained JEDI T cells were added to co-cultures and analyzed after 3–4 days. Representative flow cytometry data and quantification of proliferation (n = 7) and cytokine production (n = 4) in CD8⁺ T cells. h NDV-preinfected SUDHL4 cells were co-cultured with CellTrace violet-stained PBMCs +/− Staphylococcal enterotoxin B (SEB). CD8⁺ T cells (n = 5) were analyzed after 3 days. g, h Two-way ANOVA with Sidak’s multiple comparisons test. Data show mean ± SD.