In vivo antitumor effects by CD44v6-targeted T cells. T cells were RV-transduced with the CD44v6.CAR28z after activation with CD3/CD28 beads (28z beads) or with anti-CD3 and anti-CD28 mAbs (28z mAbs). Bead-activated T cells were also transduced with a CD44v6.CARz (z beads). CAR-redirected T cells were compared in vitro in terms of (A) expansion, measured as fold increase at different time points (mean ± SD from n = 5 donors); (B) release of IL-2 and interferon-γ upon recognition of CD44v6⁺ (n = 4) or CD44v6^– (n = 3) primary leukemic blasts (concentration, mean ± SD); (C) antileukemia and antimyeloma activity, measured as the elimination index (see “Material and methods”) of CD44v6⁺ AML (n = 3) and MM (n = 2) cell lines or CD44v6^– AML (n = 2) and MM (n = 2) cell lines. Results from a 2-way ANOVA are shown when statistically significant (*P < .05; ***P < .001). (D) NSG mice were infused with MM1.S myeloma cells and, after 3 days, treated with CD44v6.CAR28z⁺ T cells activated with beads (n = 14 mice) or mAbs (n = 6). The percentages of circulating T cells were analyzed at different time points by FACS (mean ± SD). Results from an unpaired Student t test are shown for each time point (***P < .001). (E) Left: percentages of MM1.S myeloma cells (CD38⁺/CD45^–) and T cells (CD38dim/CD45⁺) in the BM of representative mice at 5 weeks. Right: percentages of MM1.S myeloma cells in the BM of each mouse. Results from a 1-way ANOVA are shown when statistically significant (*P < .05; ***P < .001). (F) NSG mice were infused with THP-1 leukemia cells and, after 3 days, treated with bead-activated CD44v6.CAR28z⁺ T cells (n = 10 mice) or with CTR.CAR28z⁺ T cells (n = 7). A group of control mice received saline only (n = 4). Left: percentages of THP-1 leukemia cells (CD33⁺/CD45⁺) and T cells (CD33^–/CD45⁺) in the liver of representative mice at 4 weeks. Right: weights of THP1-infiltrated livers from each mouse. Results from a 1-way ANOVA are shown when statistically significant (*P < .05; **P < .01).