Supplementary Materials

The PDF file includes:

  • Materials and Methods
  • Fig. S1. Construction and testing of α-EGFRvIII synNotch–α-EphA2/IL13Rα2 CAR T cells against U87 GBM.
  • Fig. S2. Prime-and-kill circuit in T cells can overcome heterogeneity using a model antigen system in vitro.
  • Fig. S3. α-EGFRvIII synNotch–α-EphA2/IL13Rα2 CAR T cells mediate effective and localized antitumor response against U87 GBM that heterogeneously expresses EGFRvIII in the brain.
  • Fig. S4. α-EGFRvIII synNotch–α-EphA2/IL13Rα2 CAR T cells are effective against GBM6 tumor cells in vitro and in vivo.
  • Fig. S5. Fluorescence imaging of brain slices from NCG mice with implanted GBM6 tumors treated with α-EGFRvIII synNotch–α-EphA2/IL13Rα2 CAR T cells.
  • Fig. S6. SynNotch-CAR T cells show more naïve-like phenotype compared to comparable constitutively expressed CAR T cells.
  • Fig. S7. Mass cytometry shows that synNotch-induced circuits yield T cells with increased expression of stemness marker TCF1 and reduced expression of exhaustion marker CD39.
  • Fig. S8. Construction and testing of α-CDH10 synNotch–α-EphA2/IL13Rα2 CAR T cells.
  • Fig. S9. Brain-specific synNotch-CAR T cells mediate effective anti-GBM responses.
  • Fig. S10. Strategies for design of synNotch-CAR T cells to treat GBM.
  • Legends for movies S1 and S2

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Other Supplementary Material for this manuscript includes the following:

  • Movie S1 (.mp4 format). Real-time killing assays using different heterogeneous mixtures of EGFRvIII+ and EGFRvIII target cells show efficient trans-killing.
  • Movie S2 (.mp4 format). Intravital imaging of circuit CAR T cells shows dynamic priming within the GBM6 xenograft tumor.
  • Data file S1 (Microsoft Excel format). Raw data.