Figure 2|. uPAR-CAR T cells are bona fide senolytics.

(a) Cytotoxic T cell activity as determined by a 18hr-bioluminescence assay with luciferase-expressing NALM6 wild type (WT) or NALM6 overexpressing m.uPAR (NALM6-m.uPAR) as targets. Data representative of n=3 independent experiments, each performed in triplicates. (b) Cytotoxic T cell activity as determined by a 4hr-bioluminescence assay with MEK/Cdk4/6 inhibitor-induced senescent KP cells as targets; representative of n=2 independent experiments, each performed in triplicates. (c-i) NSG mice were injected with a plasmid encoding NRAS^G12V-GFP-Luciferase and treated with 0.5×10⁶ m.uPAR-h.28z CAR T cells or untransduced (UT) T cells 10 days after injection. Mice were euthanized 15 days later and livers were analyzed. (c) n fold change in luciferase signal in mice (calculated as average radiance on day 15 divided by average radiance on day −1) (n=11 mice per group). (d) Co-immunofluorescence (IF) staining of m.uPAR (red) and NRAS (green) and quantification of NRAS-positive cells (n=9 mice per group). (e) Representative stainings and quantification of SA-β-Gal positive cells (n=7 mice per group). (f) Co-IF staining of m.uPAR (red) and human CD3 (green) (n=5 mice per group). (g-i) Number of liver infiltrating T cells (g), expression of CD62L/CD45RA (h) and PD1+TIM3+LAG3+ (i) on m.uPAR-h.28z CAR T cells as determined by flow cytometry (n=4 mice per group). (c-e) Representative results of n=2 independent experiments. Data are mean± SEM. Two-tailed unpaired Student’s t-test.