Effect of Embelin on TRAIL Receptor 2 mAb-induced Apoptosis of TRAIL-resistant A549 Non-small Cell Lung Cancer Cells

Introduction: Some non-small cell lung cancer (NSCLC) tumor cells are insensitive to tumor necrosis factorrelatedapoptosis-inducing ligand (TRAIL) -based therapy. This study was conducted to examine the effect ofembelin on the sensitivity of the A549 NSCLC cell line to TRAIL receptor2 (TRAILR2) monoclonal antibodiesand to investigate the potential mechanisms. Materials and
Methods: A549 cells were treated with embelin,TRAILR2 mAb or a combination of both. Cell viability was measured using ATPlite assay and apoptosis rateswere determined by flow cytometry with AnnexinV-FITC and propidium iodide staining, with the expressionlevels of proteins analyzed by Western blotting.
Results: The cell survival rate of separate treatments with 100ng/ml TRAILR2 antibody or 25 uM embelin were 81.5±1.57% and 61.7±2.84%, respectively. Their combined usemarkedly decreased cell viability in A549 cells to 28.1±1.97% (P<0.05). The general caspase inhibitor Z-VADFMKcould inhibit the embelin-enhanced sensitivity of A549 cells to TRAILR2 mAb (75.97±3.17%)(P<0.05).Both flow cytometry and cell morphological analysis showed that embelin was able to increase TRAIL-inducedapoptosis in A549 cells. Combined treatment with embelin and TRAILR2 mAb augmented the activation ofinitiator caspases and effector caspase. In addition, A549 cells showed increasing levels of TRAILR2 proteinand decreasing levels of Bcl-2, survivin and c-FLIP following the treatment with embelin+TRAILR2 mAb.
Conclusions: Embelin could enhance TRAIL-induced apoptosis in A549 cells. The synergistic effect of thecombination treatment might be due to modulation of multiple components in the TRAIL receptor-mediatedapoptotic signaling pathway, including TRAILR2, XIAP, survivin, Bcl-2 and c-FLIP.