Fig. 6: In vivo efficacy of CUL5 knockout (KO)- and knockdown (KD)-CD19 CAR T cells.
a A schematic diagram of in vivo experiments. Nonobese diabetic/severe combined immunodeficiency (NOD/SCID) common-gamma chain KO (NOG) mice were injected with 5 × 105 Raji/ffluc-green fluorescent protein (GFP) cells via the tail vein on Day 0, followed by 1  × 106 truncated epidermal growth factor receptor (tEGFR)-, control (Ctrl)-, or CUL5 KO-CD19 CAR T cells on Day 7. Tumor burden was assessed using BLI at the indicated time points and weekly thereafter. b Representative BLI of Raji-inoculated mice treated with Ctrl- or CUL5 KO-CD19 CAR T cells over time. c Tumor burden in mice treated with Ctrl- or CUL5 KO-CD19 CAR T cells at the indicated time points. Data are presented as the mean and SEM, compared using the two-way ANOVA, and acquired from three independent experiments on three different donors (tEGFR-T cells: n = 4; Ctrl- and CUL5 KO-CD19 CAR T cells: n = 9 per group). d Kaplan-Meier survival analysis of Raji/ffluc-bearing NOG mice (log-rank test). e The number of CAR T cells derived from 1 × 106 CD8 + T cells on Day 7. Data are derived from three independent experiments on three different donors. Data were expressed as mean ± SEM. f The effect of CUL5 KD on activated CD19 CAR T cell proliferation. These cells were stimulated with γ-irradiated Raji cells in a 1:5 E: T ratio every 10 days. Data were expressed as mean ± SEM. (n = 3, different donors) Note that the paired t-test was used for e, whereas the one-way ANOVA was used for (f). g, h In vivo efficacy assessment using a subcutaneous tumor model. On Day 0, Raji/ffluc-GFP cells were subcutaneously inoculated. On Day 10, 1 × 106 tEGFR-, shGFP-, or shCUL5-CD19 CAR T cells were injected. g Subcutaneous tumor volume. The curve was censored when any of the mice in the group were euthanized. Data are presented as the mean and SEM, compared using the two-way ANOVA, and acquired from three independent experiments on three different donors (n = 9 per group). h Kaplan-Meier analysis of NOG mice subcutaneously injected with Raji/ffluc cells (log-rank test). (i) Kaplan-Meier analysis of NOG mice intravenously injected with Raji/ffluc cells as in (a). 1 × 106 tEGFR-T, CUL5 KO-, or shCUL5-CD19 CAR T cells were injected. Data are acquired from three independent experiments on four different donors (tEGFR-T cells: n = 8; CUL5 KO- and shCUL5-CD19 CAR T cells: n = 12 per group) (log-rank test). Source data are provided as a Source Data file.
