Figure. 6. DCs regulate CAR T-cell-induced antigen spreading through enhanced tumor antigen acquisition and IFN-γ-IL-12 crosstalk.

(A) EGFRvIII⁺ CT-2A cell killing by WT, IFN-γ^−/−, or IFNGR1^−/− CAR T-cells in vitro (n=3).

(B) Enumeration of tumor-infiltrating immune cells in mice (n = 4–5) receiving WT CAR T ± vax. See supplemental methods for phenotyping details.

(C) IFN-γ ELISPOT. Tumor-bearing WT or Batf3^−/− mice (n=5) treated with WT CAR T ± vax.

(D) Tumor-bearing mice were left untreated (n=4) or treated with WT or IFN-γ^−/− CAR T-vax (n=5). Shown are chemokine expression in tumors 7 days post treatment.

(E-F) Ki67 (E) and CCR7(F) expression in intratumoral CD103⁺ DCs and CD11b⁺ DCs from mice (n=5) treated with WT CAR T ± vax.

(G-H) Mice bearing ZsGreen⁺EGFRvIII⁺CT-2A tumors were treated with WT CAR T, WT CAR T-vax or IFN-γ^−/− CAR T-vax (n=5), shown are tumor antigen (ZsGreen) uptake by intratumoral CD103⁺ DCs (G) and CD11b⁺ DCs (H).

(I-J) IFN-γ ELISPOT (I) and tumor growth (J) in mice (n = 5) treated with WT or IFNGR1^−/− CAR T ± vax.

(K-N) IFN-γ ELISPOT.

(K) WT vs. CD11c-specific IFNGR1 KO tumor-bearing mice (n=5) following WT CAR T ± vax.

(L) Tumor-bearing mice (n=5) following WT CAR T or WT CAR T-vax + anti-IFN-γ or anti-IL12(p70).

(M) Tumor-bearing WT mice (n=5) following WT or IL12rb2^−/− CAR T-vax therapy or in IL12p40^−/− mice following WT CAR T-vax.

(N) Tumor-bearing WT mice (n=7) following WT CAR T-vax or IFNGR1^−/− CAR T ± vax.

(O) Tumor growth in mice (n=5–7) left untreated or treated with WT or IFNGR1^−/− CAR T ± vax.

All mice except those in G-H bear EGFRvIII⁺CT-2A tumors. Error bars are mean ± 95% CI, ***, p<0.001; **, p<0.01; *, p<0.05; ns, not significant by Student’s t-test for B and E-F, by one-way ANOVA with Tukey’s post-test for A, C-D, G-I and K-N, by two-way ANOVA with Tukey’s post-test for J and O.