MgkL cell survival is maintained by senescence-associated autophagy. (A) DAPRed staining to detect autophagosomes. Cells were purified from the colony formation assay of BCR-ABL–transduced WT- or DKO-derived KLS⁺ cells on day 5. In some experiments, 5 μM SBI-0206965 was added to the culture. The cells were then subjected to DAPRed staining as described in Materials and methods. Representative results from three independent experiments are shown (original magnification, ×200; scale bar, 50 μm). (B) Flow cytometric analysis of DAPRed staining. Representative results from 4 independent experiments (left). MFIs were determined (right) (n = 4). (C) Number of MgkL and myeloid-lineage cells in the in vitro colony formation assay on day 9. Colonies were formed from 150 BCR-ABL–transduced WT KLS⁺ cells in the presence or absence of 5 µM SBI-0206965 (n = 4). (D) Number of viable MgkL and c-kit⁺ progenitor cells. Ten thousand MgkL and progenitor cells purified from the colony formation assay on day 9 were cultured in the presence or absence of 5 µM SBI-0206965 for 1 day (n = 3). (E) Schematic representation of the experimental procedure of the treatment of imatinib (150 mg/kg of body weight/d in a water, oral administration) and SBI-0206965 (20 mg/kg of body weight/d in filter-sterilized vehicle (5% Tween-80, 5% PEG400 in phosphate-buffered saline), intraperitoneal injection). (F) Proportion of CD41⁺CD150⁺ MgkL cells in BM. Total BM cells were obtained on day 8 (depicted in panel E) to determine the proportion of CD41⁺CD150⁺ MgkL cells among total BM cells (n = 8). Data are presented as the mean ± SD. Statistical significance was evaluated using the 1-way ANOVA with (B) Tukey’s multiple-comparisons test and the (C-D,F) 2-sided Student t test. N.S., not significant; *P < .05; **P < .01. (G) Survival rate of mice following combined treatment with imatinib and SBI-0206965. Survival rates were determined (n = 8). Comparison of survival rates was performed using the log-rank test.