Linarin sensitizes tumor necrosis factor-related apoptosis (TRAIL)-induced ligand-triggered apoptosis in human glioma cells and in xenograft nude mice.

Tumor necrosis factor-related apoptosis-induced ligand (TRAIL) is reported as a promising anti-cancer therapeutic agent. Nevertheless, a variety of cancer cells, including human malignant glioma cells, are resistant to TRAIL treatment, indicating that it is necessary to find effective strategies to overcome the TRAIL resistance. Linarin (LIN), a natural flavonoid compound in Flos Chrysanthemi Indici (FCI), has been exhibited to exert various pharmacological activities, including anti-cancer. Here in our study, we found that non-cytotoxic doses of LIN (5μM) dramatically potentiated TRAIL (80ng/ml)-induced cytotoxicity (52.36±1.58%) and apoptosis (68.50±1.23%) using 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) and flow cytometry assays, respectively, in human glioma cells of U87MG. Apoptosis was evidenced by enhanced cleavage of Caspase-8/-9/-3 and poly (ADP-ribose) polymerase (PARP), and reduced anti-apoptotic proteins, including B-cell leukemia/lymphoma 2 (Bcl-2), mantle cell lymphoma (Mcl)-1, and Survivin. Moreover, both intrinsic and extrinsic apoptosis pathways were included in apoptosis induced by LIN and TRAIL co-treatment, along with high release of Cyto-c into cytoplasm and enhancement of fas-associated protein with death domain (FADD), death-inducing signaling complex (DISC), death receptor 4 (DR) 4 and DR5, respectively. Reactive oxygen species (ROS) generation, up to 39.86±2.32%, was also highly triggered by TRAIL and LIN combinational treatment, which was accompanied with high phosphorylation of c-Jun-N-terminal kinase (JNK). In vivo, TRAIL and LIN double treatment significantly reduced the tumor growth using xenograft tumor model through inducing apoptosis. We demonstrated that combining LIN with TRAIL treatments might be effective against TRAIL-resistant glioma cells through inducing apoptosis regulated by ROS generation.