Fig. 3

The synNotch system increases T cell specificity. A: Top, customized recognition domain of synNotch receptors (e.g., αCD19 scFV) detects a signal molecule (e.g., CD19) on target tumor cells. Middle, the core regulatory region of the Notch receptor that governs proteolysis or cleavage is activated by the interaction between the receptor and the tumor target signal (top), and a cytoplasmic orthogonal transcription factor is released; Bottom, the orthogonal transcription factor enters the nucleus of engineered T cells and controls the function-related transcriptional programs. B. I: αCD19-synNotch T cells recognize CD19⁺ tumor cells and release customized cytokines (e.g., IL-2, IL-12, and IL-10) that destroy CD19⁺ tumor cells; II: αCD19-synNotch T cells recognize CD19⁺ tumor cells, promote the transcription of differentiation-related genes (e.g., T-bet), and then promote T cells differentiation into Th1 cells; III: αGFP-synNotch T cells recognize GFP⁺ tumor cells, promote the transcription of TRAIL, and accelerate the lysis of tumor cells; IV: αGFP-synNotch T cells recognize GFP⁺ tumor cells, promote the production of antibodies (e.g., αPD-1, αCTLA-4 and αCD19/αCD3 BiTE), and accelerate tumor lysis. C. I: EpCAM/B7-H3-synNotch-αROR1-CAR-T cells recognize EpCAM⁺/B7-H3⁺ tumor cells, promote the expression of αROR1-CAR on T cells, and enable T cells to recognize and lyse EpCAM⁺/B7-H3⁺ROR1⁺ tumor cells; II: MET-synNotch-αMART1-T cells recognize MET⁺ tumor cells, and then enable T cells to lyse MET⁺MART1⁺ melanocytes; III: GFP-synNotch1-HER2-synNotch2-αCD19-CAR-T cells recognize GFP⁺ tumor cells, promote the expression of αCD19-CAR on T cells, and recognize and lyse GFP⁺CD19⁺ tumor cells. However, if GFP-synNotch1-HER2-synNotch2-αCD19-CAR-T cells recognize HER2⁺ normal cells, the proapoptotic factor tBID (truncated BH3-interacting domain death agonist) will be expressed by T cells to casue T cell apoptosis; IV: EGFR-synNotch1-MET-synNotch2-αHER2-CAR-T cells recognize two antigens from tumor cells (first one is EGFR and second one is MET). This promotes the expression of αHER2-CAR on T cells, and enable T cells to lyse EGFR⁺MET⁺HER2⁺ tumor cells; V: Both MET-synNotch-αHER2/αEGFR-CAR-T cells and EGFR/HER2-synNotch-αMET-CAR-T cells recognize and lyse EGFR⁺MET⁺/HER2⁺MET⁺ tumor cells; VI: GD2-synNotch-αB7-H3-CAR-T cells, APPL2-synNotch-αMCAM/αMSLN/αHER2-CAR-T cells, or EGFRvIII-synNotch-αEphA2/αIL13Rα2-CAR-T cells recognize the first antigen (GD2, APPL2, or EGFRvIII) on tumor cells, promote the expression of αB7-H3-CAR, αMCAM/αMSLN/αHER2-CAR, or αEphA2/αIL13Rα2-CAR on T cells, and enable T cells to lyse specific tumor cells (GD2⁺B7-H3⁺ tumor cells, APPL2⁺MCAM⁺/MSLN⁺/HER2⁺ tumor cells, or EGFRvIII⁺EphA2⁺/IL13Rα2⁺ tumor cells). Figure was created with BioRender.com