Optimized low affinity Dual CAR CIK cells control AML growth while reducing off-targeted toxicities. (A) Long-term (E:T ratio of 1:10, n = 2 for NT and DC mut) cytotoxicity against CD123⁺/CD33⁺ KG-1 cell line, primary AML cells and TIME cell line. Scatted plot (mean ± SD) from two independent CAR-CIK donors is shown. (B) Cytokine release against CD123⁺/CD33⁺ primary AML cells and TIME cell line (n = 2 for NT and DC mut). (C) Schematic of the AML patient xenograft mouse model. NSG mice were injected via tail vein on day 0 with 1 × 10⁶ tertiary PDX AML cells. Mice were randomized to 2 treatment groups each receiving one injection of vehicle or gene-modified DC mut CIK cells at day 5 (n = 4 per group). (D) Analysis of hCD45⁺/CD33⁺/CD123⁺ cells in the PB, BM and spleen of untreated and treated mice. (E) Analysis of hCD45⁺/CD3⁺ cells in the PB, BM and spleen of Dual CAR-CIK treated mice. (F) Representative dot plot of PB analysis. P-values from the Wilcoxon test are referred to the comparison of DC mut-CIK-treated mice compared with untreated mice within PB, BM and spleen. ∗, p-value < 0.05.