Fig. 4: VPS34-IN1 inhibits l-asparaginase induced autophagy and bot drugs act synergistically.

A MOLM-14 cells were cultured for 6 h with l-asparaginase and/or chloroquine (10 µM) as indicated. Autophagy induction was assessed by analyzing both LC3-I and LC3-II expression by western blot. Upon autophagic induction, LC3-I is transformed to LC3-II which is then degraded in autophagolysosome. Chloroquine is able to inhibit acidification of autophagolysosome, allowing the evaluation of LC3-II formation which is an indirect marker of autophagy induction B Number of LC3B puncta per cells (right) and immunofluorescence analysis of GFP LC3 (left) in vehicle-treated or l-Asparaginase-treated (10 UI/ml) MOLM-14 cells were determined; n = 3, bars represent standard error of the mean. C MOLM-14 cells were cultured for 24 h with vehicle, chloroquine (10 µM) or l-asparaginase (10 UI/ml) and p62 accumulation was assessed by western blot. D MOLM-14 cells were cultured with increasing concentrations of VPS34-IN1 and in the presence of both l-asparaginase (10 UI/ml) and chloroquine (10 µM). Western blotting analysis of the LC3-I and LC3-II bands was performed at 6 h. E Synergy map (left) and viability matrix (right) of l-asparaginase with VPS34-IN1 for the MOLM-14 cell line. The mean viability measurement from three independent experiments was used. F Summary of synergy score from 48 h co-treatments of seven primary AML samples with l-Asparaginase and VPS34-IN1.