Fig. 3: Efficacy of Fox19CAR-Tregs in an in vivo humanized mouse model of SLE.

A Generation of the humanized mouse model of SLE. 1 day-old pups of NSG mice were irradiated and transplanted with 0.8-1 × 10⁵ human cord-blood stem cells/mouse injected intra-liver. The engraftment was monitored assessing the presence and the composition of human leukocytes on peripheral blood weekly by flow cytometry. After the establishment of a human immune system, mice were injected i.p. with pristane to induce a chronic inflammation. After 3 weeks, UT-, Fox19CAR-Tregs or PBS were injected and their kinetic and the levels of human leukocytes in the peripheral blood were monitored weekly by flow cytometry. Mice were sacrificed 15 weeks after humanization. B, C Longitudinal assessment of human B and T lymphocytes in peripheral blood. Human B and T cells were identified as huCD45⁺CD19⁺ and huCD45⁺CD3⁺ lymphocytes, respectively. Absolute cell counts were assessed by flow cytometry. Pristane injection is indicated with an arrow. N = 53 mice. One-way ANOVA with Tukey correction for multiple comparison. *p-value 0.021, ****p-value < 0.0001. D Longitudinal assessment of circulating Fox19CAR-Tregs in mouse peripheral blood after their injection. CAR⁺ cells were identified with FITC-conjugated recombinant CD19 (rCD19). CAR-Treg absolute counts were assessed by flow cytometry. N = 16 mice. E–G Absolute numbers of circulating human cells in humanized mice in the different groups of treatment after the injection of Fox19CAR-Tregs, UT-Tregs or PBS. Human cells were identified as total huCD45⁺ leukocytes, huCD45⁺CD3⁺ T and huCD45⁺CD19⁺ B lymphocytes, respectively. Absolute cell counts were assessed by flow cytometry. N = 31 (16 CAR-Tregs, 11 UT-Tregs, 4 PBS). Two-way ANOVA test with Tukey correction for multiple comparisons. The exact p-values are reported in the graphs. All the results are expressed as mean ± standard deviation.