Newly Synthesized PW06 Induced Cell Apoptosis in Human Glioblastoma Multiforme GBM 8401 Cells Through Caspase- and Mitochondria-Dependent Pathways

Abstract

Glioblastoma multiforme (GBM) is the most common, aggressive, and dangerous lethal tumor in the brain, which develops in adults. Currently, the efficiency of chemotherapy treatment for GBM patients is still unsatisfactory. PW06 was synthesized by Dr. Lien's laboratory (China Medical University, Taichung, Taiwan), and it was demonstrated to induce cancer cell apoptosis in human pancreatic carcinoma MIA PaCa-2 cells. However, the anti-cancer activities of PW06 on human GBM cancer cells are not reported. Thus, herein, PW06 was investigated on the anticancer activity on human glioblastoma multiforme GBM 8401 cells. Both PI exclusion and Annexin V/PI double staining methods were conducted for investing cell viability and apoptosis in GBM 8401 cells, respectively; they were analyzed with flow cytometer assay. Results showed that PW06 decreased total viable cell number with the process of cell apoptosis in GBM 8401 cells. Both productions of reactive oxygen species (ROS) and Ca²⁺, affect mitochondria membrane potential (ΔΨm) levels, and activities of caspase-3, -8, and -9 in GBM 8401 cells after exposure with PW06 were assayed by flow cytometer. Results showed that PW06 promoted ROS production and Ca²⁺ release from ER but lowered the levels of ΔΨm, and it also induced higher activities in caspase-3, -8, and -9 in GBM 8401 cells. Evaluation of protein expressions associated with apoptosis in GBM 8401 cells after being incubated with PW06 were conducted by Western blot analysis. Results show that PW06 increased GADD153, BiP, ATF-6α, ATF-6β, eIF2α, eIF2α^pSer51, CHOP, and caspase-4, and they are associated with ER stress-associated protein expression. However, it induced higher pro-apoptotic proteins (Bax and Bad) expression and inhibited anti-apoptotic proteins (Bcl-2, Bcl-xl, and Mcl-1) expression, even promoting higher cleaved caspase-8, -9, and -3 protein expression and increased EndoG and AIF in GBM 8401 cells. Collectively, it may suggest PW06 exits anti-GBM activity to process cell apoptosis in the human GBM 8401 cells in vitro.