Fig. 3
The combination of FLT3scFv/NKG2D-CAR T cells and gilteritinib act synergistically on mediating the regression of AML in a xenograft mouse model. A Schema of establishing the AML xenograft mouse model. NSG mice were injected with MOLM-13 luciferase-expressing cells (2 × 106 cells) via the tail vein. On day 7, gilteritinib was dissolved in 4% DMSO and administered via intraperitoneal (i.p.) injection at a dose of 15 mg/kg, which was continued 5 days/week for 3 weeks in Groups 2 and 4. On day 8, a single dose of FLT3scFv/NKG2D-CAR T cells (2.5 × 106 cells) was injected via the tail vein in Groups 3 and 4 (n = 4 mice/group). B Leukaemia progression was monitored by serial bioluminescent (BL) imaging using an IVIS Lumina imaging system following D-luciferin substrate administration (0.3 mg/g body weight, IP). The scale (right) shows the upper and lower BL imaging thresholds at each analysis time point (days 7, 14, 21, and 28). C AML burden was assessed by quantification of BL radiance obtained as photon/sec/cm2/sr in the target zone encompassing the entire body of each mouse, and the AML burden was significantly reduced in mice treated with FLT3scFv/NKG2D-CAR T cells alone and in combination with gilteritinib. The waterfall plot shows the ∆BL value (upper-increase/below-decrease) as absolute BL values obtained from each mouse between day 7 and day 14 after tumour inoculation. D Kaplan–Meier survival curves showed the survival of AML mice was significantly improved with CAR T cell monotherapy (p < 0.05 compared with gilteritinib monotherapy) and with combination therapy (p < 0.05 compared with CAR T cell monotherapy). The in vivo data shown are derived from two independent experiments with T cells provided from 2 different donors. E Immunofluorescence staining showed that the distribution of CAR T cells in the bone marrow of AML mice was significantly enhanced following combination with gilteritinib treatment. The diagram shows the MFI of GFP + CAR T cells from n = 3 mice in each group as assessed with ImageJ. MFI, mean fluorescence intensity. GFP, green fluorescent protein. * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001