Fig. 1

Construction of CAR-TIM3 and its introduction into iPSCs. A (Upper) Diagram of the third-generation CAR-TIM3 construct harboring signal peptide, anti-TIM3 scFv fragment, CD8 hinge, CD28, 4-1BB, and CD3ζ, under the control of EF-1α promoter. (lower) Two sets of primers used for DNA sequencing and its predicted target regions are shown. B, C DNA sequencing results of CAR-TIM3 iPSCs (clone 5E1) spanning over scFv fragment (B) and CD3ζ (C). D CAR expression on the surface of CAR-TIM3 iPSCs as evaluated by flow cytometry using anti-F(ab′)2 antibody. E Western blot analysis of CD3ζ, which is a CAR signaling molecule