CD3-targeted LVs mediate functional CAR T-cell generation in vivo. For in vivo CAR delivery, huNSG mice were subcutaneously (SC) injected with human IL-7 or phosphate-buffered saline 4 days and 1 day before IV injection of TR66.opt-LV harboring the CD19-CAR or vehicle as control. (A) Experimental outline. (B) Representative plots show CAR⁺ cells in CD8⁺ and CD8⁻ T cells at day 26 postinjection of vector. (C) Line diagrams show CAR⁺ cells in CD3⁺ cells in the peripheral blood of individual mice in group 1 (IL-7), group 2 (TR66.opt-LV), and group 3 (TR66.opt-LV + IL-7). *^,#P < .05, **^,##P < .01, ***^,###P < .001 comparing group 3 to group 1 (*) and group 2 (^#) by 2-way ANOVA with Tukey's correction. Arrows point to tendencies for CAR T-cell detection in group 2, which were not statistically significant. (D) Scatter dot plots compare the percentages of CAR⁺ cells within the CD3⁺CD45⁺ T-cell compartment, CD45⁻ mouse cells and CD3⁻CD45⁺ non-T cells. (E-F) B-cell levels were determined in peripheral blood of the animals to assess CD19-CAR T-cell functionality. (E) Representative dot plots show CD19⁺CD20⁺ B cells at day 26 postinjection. (F) Line diagrams summarize relative levels of CD19⁺ cells compared with day 0 in the peripheral blood of individual mice in group 1 (IL-7), group 2 (TR66.opt-LV), and group 3 (TR66.opt-LV + IL-7). All data from n = 5-6 mice per group. *P < .05, **P < .01, ***P < .001 comparing groups 2 and 3 to group 1 by 2-way ANOVA with Turkey's correction.