T cells with CD3/TCRαβ knock-down by PEBL and CAR expression kill leukemia cells in mice. (A) NSG mice were IV injected with 5 × 10⁵ Nalm6-luciferase cells. Three days later, mice received 2 × 10⁷ T-lymphocytes transduced with anti-CD19-41BB-CD3ζ CAR plus either PEBL or mCherry alone; other mice received tissue culture medium instead (“no T cells”). Bioluminescence images on day 3 are shown with enhanced sensitivity to illustrate Nalm6 engraftment. (B) Symbols correspond to the average bioluminescence signal in ventral and dorsal imaging. (C) Kaplan-Meier curves and log-rank test for overall survival. Mice were euthanized when the ventral and dorsal bioluminescence average signal reached 1 × 10¹⁰ photons per second. ****P < .0001. (D) NSG mice were IV injected with 5 × 10⁵ Nalm6-luciferase cells and with 2 × 10⁷ T lymphocytes on day 3 as described in panel A. Before T lymphocytes injection, mice received 2.5 Gy total body irradiation. Bioluminescence images on day 3 are shown with enhanced sensitivity to illustrate Nalm6 engraftment. (E) Symbols correspond to bioluminescence average by ventral and dorsal imaging. (F) Kaplan-Meier curves and log-rank test for overall survival. Mice were euthanized when the ventral and dorsal bioluminescence average signal reached 1 × 10¹⁰ photons per second, or when signs of GVHD (>20% weight reduction exceeded in 2 consecutive measurements, with reduced mobility and/or fur loss) were evident. GVHD occurred in 3 of the 5 CAR⁺mCherry mice and 0 of the 6 CAR⁺PEBL mice; relapse (“Rel.”) rates were 0 of 5 vs 2 of 6, respectively. **P = .0014; ***P = .0006.