Figure 4.

Expression and function of hinge/transmembrane domains (HD/TMD)-modified CARs in human T cells introduced CAR mRNA by electroporation. (A) CAR mRNAs were analyzed by reverse transcription and quantitative polymerase chain reaction, and their amount were calculated relative to GAPDH mRNA. (B) CAR expression on T cells were analyzed by flow cytometry using anti-HA-tag mAbs (solid color histograms) or isotype control antibody (dashed white histograms). Each CAR expression level was calculated from the ratio of geometric mean fluorescence intensity (GMFI) when stained with the anti-HA-tag mAb to GMFI when stained with the isotype antibody. (C) Top panel, cytotoxic activity of human CAR-T cells cultured for 24 h after electroporation against hVEGFR2⁺ L1.2 cells at E/T ratio = 10; lower panel, cytotoxicity per CAR expression level. The data were obtained from three independent tests. Statistical analysis was performed using Student’s t-test; ^†† p < 0.01.