Abstract
As the activation of autophagy contributes to the efficacy of many anticancer therapies, deciphering the precise role of autophagy in cancer therapy is critical. Here, we report that the dual mTORC1/2 inhibitors PP242 and OSI-027 decreased cell viability but did not induce apoptosis in the non-small cell lung cancer (NSCLC) cell lines H460 and A549. PP242 induced autophagy in NSCLC cells as demonstrated by the formation of massive vacuoles and acidic vesicular organelles and the accumulation of LC3-II. JNK was activated by PP242, and PP242-induced autophagy was blocked by inhibiting JNK pathway with SP600125 or JNK siRNA, suggesting that JNK activation is required for the mTORC1/2 inhibitor-mediated induction of autophagy in NSCLC cells. Inhibiting JNK or autophagy increased the sensitivity of H460 cells to mTORC1/2 inhibitors, indicating that JNK or autophagy promoted survival in NSCLC cells treated with mTORC1/2 inhibitors. Together, these data suggest that combining mTORC1/2 inhibitors with inhibitors of JNK or autophagy might be an effective approach for improving therapeutic outcomes in NSCLC.
Highlights
PP242, respectively, for 24 h. (a,c,g) Protein levels were estimated using western blot analysis
To determine if the PP242-induced decrease in cell viability was mediated by apoptosis, H460 and A549 cells were treated with 10 μM and 5 μM of PP242, respectively, for 24, after which point, cell viability was reduced by approximately 50%
(a,b) H460 and A549 cells were treated with the indicated concentrations of PP242 for 24 h. (c) H460 and A549 cells were treated with 10 μM PP242 for the indicated amount of time. (a) Changes in cellular morphology were observed using an inverted microscope. (b,c) The percentage of vacuolated cells was calculated from 3 independent images
Summary
PP242, respectively, for 24 h. (a,c,g) Protein levels were estimated using western blot analysis. (a,c,g) Protein levels were estimated using western blot analysis. The blot shown is representative of 3 independent experiments. (b,d) Cell viability was measured using the MTT assay. The data are presented as the mean percentage of control ±SD (n = 3). (e) Apoptosis was measured as the percentage of annexin. (f) Caspase-3/7 activity was evaluated using a CaspaTag in situ Assay Kit. Data representative of 2 independent experiments are shown. (e,f) H460 and A549 cells treated with 10 μM lapatinib/100 nM bafilomycin A1 and 10 μM lapatinib/5 nM bafilomycin A1, respectively, for 24 h were used as positive controls.
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