Fig. 2. Autocrine synthetic IL-2 circuits strongly improve T cell cytotoxicity against multiple models of immune-excluded syngeneic tumors.

(A) The synthetic IL-2 circuit was recapitulated in mouse T cells producing mouse IL-2 (mIL-2) to test circuits in presence of an intact immune system, suppressive TME and native IL-2 consumer cells.

(B) KPC CD19⁺ pancreatic tumors were engrafted subcutaneously into immunocompetent C57/B16 mice and treated 9 days later with synthetic IL-2 circuit T cells and anti-Mesothelin CAR T cells as a two-cell paracrine system. No tumor control was observed in this paracrine configuration, even though KPC tumors express mesothelin.

(C) KPC CD19⁺ pancreatic tumors were engrafted as in B and treated 9 days later with T cells engineered with both a synthetic IL-2 circuit and an anti-Mesothelin CAR (autocrine configuration). Significant improvement in tumor control was observed (red lines) compared to anti-Mesothelin CAR T cells combined with dummy synthetic cytokine circuit (synNotch only produces BFP, black lines).

(D) KPC CD19⁺ pancreatic tumors were engrafted orthotopically in the pancreas tail and treated 9 days later with engineered T cells. 100% survival was observed only with the addition of the IL-2 circuit out to 120 days (duration of study).

(E) B16F10 OVA CD19⁺ melanoma tumors were engrafted orthotopically into immunocompetent C57/B16 mice and treated 8 days later with 2e6 engineered mouse CD8⁺ OT-1 (anti-OVA) T cells. Tumor control was only observed in mice treated with T cells expressing the IL-2 circuit. For B-E All plots show tumor burden measured by average +/− S.E.M. of caliper or bioluminescence measurements and overall survival (n=4–5 per group, * = significant difference in survival with addition of IL-2 circuit using log-rank test, p < 0.05). See Fig. S7 and S8 for individual growth curves data.