Figure 3.

Effect of autophagy inhibition on L-asparaginase-induced cytotoxicity in ALL cells. (a, b) Apoptotic analysis in ALL cell lines REH and 697 cells with or without CQ treatment by flow cytometry (a) and western blotting (b). The proportion of dead cells was measured by flow cytometry using Annexin-V staining. Cells were treated with the indicated concentrations of L-asp and/or CQ for 48 h. (c) Cell survival assay during prolonged treatment. According to the clinical method for administering L-asp, which is repeated every 3 days in patients, ALL cells were cultured with PBS (control) or repeated (0, 72, 144 h) administration of L-asp and/or CQ. Viable cells were counted using trypan blue staining every 24 h. ALL cells were treated with repeated administration of L-asp and/or CQ. (d) Sensitivity to L-asp treatment in REH cells transfected with ATG7-siRNA. Cells transfected with control-siRNA (si-control) or ATG7-siRNA (si-ATG7) were treated with the indicated concentrations of L-asp for 48 h. Viable cells were counted by flow cytometry using Annexin-V staining. (e) Cell survival assay of parental cells and L-asp-resistant cells generated from 697 cells (697-R). (f) Apoptotic analysis of 697-R cells. (g) Western blot analysis of parental cells and 697-R cells. Data in a and d–f are represented as mean±s.d. (n=3; *P<0.05, ***P<0.001). P-values were calculated using two-sided Student's t-test (a, f), and one-way ANOVA (c).