Fig. 6
ABn treatment inhibits tumor growth and extends survival of mice syngrafted with WapMyc tumors. a Combination treatment protocol for orthotopically syngrafted WapMyc tumors. The mice were treated with a vehicle, navitoclax, and two concentrations of metformin alone or in combination with navitoclax for 21 days. Immunoprofiling samples were collected right after the drug treatments or after a follow-up period. In the follow-up period the samples were collected when the tumors reached Ø 2 cm and the mice had to be killed. b ABn treatment-induces BIM activation in vivo. The level of BIM immunostaining was scored in IHC samples from treated tumors using a scale of 1–4 (representative examples are shown in the figure). The samples were blinded for analysis. Student’s t-test (unpaired), SD. c ABn treatment-induced “apoptotic ponds”. Representative images of tumor samples stained for cleaved caspase-3. d ABn treatment stimulates T-cell infiltration. WapMyc tumor samples were isolated from mice killed after the 21-day AB treatment period. Representative images are shown and the data below show average and SD of CD4+ or CD8+ T cells in the tumors. Immunohistochemical stainings were performed for at least 45 tumors per antibody and 3–6 field of view (fov) per tumor were analyzed. Student’s t-test (unpaired). e ABn treatment-induced changes in proportions of peripheral lymphocytes. N (vehicle) = 7 mice, N (ABn) = 4 mice. Blood was collected after 21 days treatment with either vehicle or ABn. Student’s t-test (unpaired), SD. f Tumor growth in ABn-treated mice. Student’s t-test (unpaired), SEM. g, h ABn treatment extends survival. P-values: Difference between the vehicle and ABn-treated cohorts. i Flow cytometry-based immunoprofiling of vehicle and ABn (navitoclax+metformin)-treated tumors. The heatmaps show fold-change compared to control. N = 6 (vehicle), N = 2 Navitoclax+Metformin. j Post-treatment ratios of tumor-infiltrating PD-1+CD8+ T cells. Student’s t-test (unpaired), SD
