Lien Jin-Cherng, Hsu Sheng-Yao, Chueh Fu-Shin, Ma Yi-Shih, Chu Yung-Lin, Chou Yu-Cheng, Lai Kuang-Chi, Chen Jaw-Chyun, Huang Yi-Ping, Wu Rick Sai-Chuen
School of Pharmacy, China Medical University, Taichung, Taiwan.
Department of Ophthalmology, Kaohsiung Show Chwan Memorial Hospital, Tainan, Taiwan.
J Biochem Mol Toxicol. 2025 May;39(5):e70264. doi: 10.1002/jbt.70264.
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 Ca, 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 Ca 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, 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.
多形性胶质母细胞瘤（GBM）是成人中最常见、侵袭性最强且危险的致命性脑肿瘤。目前，GBM患者的化疗治疗效果仍不尽人意。PW06由连博士实验室（中国台湾台中中国医药大学）合成，已证明其可诱导人胰腺癌MIA PaCa - 2细胞发生癌细胞凋亡。然而，PW06对人GBM癌细胞的抗癌活性尚未见报道。因此，本文研究了PW06对人多形性胶质母细胞瘤GBM 8401细胞的抗癌活性。分别采用PI排除法和Annexin V/PI双染法检测GBM 8401细胞的活力和凋亡情况，并用流式细胞仪进行分析。结果显示，PW06可使GBM 8401细胞的总活细胞数随着细胞凋亡过程而减少。通过流式细胞仪检测了PW06处理后GBM 8401细胞中活性氧（ROS）的产生、Ca的释放、线粒体膜电位（ΔΨm）水平以及半胱天冬酶 - 3、 - 8和 - 9的活性。结果表明，PW06促进了ROS的产生和Ca从内质网的释放，但降低了ΔΨm水平，同时还诱导GBM 8401细胞中半胱天冬酶 - 3、 - 8和 - 9的活性升高。通过蛋白质免疫印迹分析检测了PW06处理后GBM 8401细胞中与凋亡相关的蛋白质表达。结果表明，PW06增加了GADD153、BiP、ATF - 6α、ATF - 6β、eIF2α、eIF2α、CHOP和半胱天冬酶 - 4的表达，这些均与内质网应激相关蛋白表达有关。然而，它诱导了促凋亡蛋白（Bax和Bad）的高表达，抑制了抗凋亡蛋白（Bcl - 2、Bcl - xl和Mcl - 1）的表达，甚至促进了GBM 8401细胞中裂解的半胱天冬酶 - 8、 - 9和 - 3蛋白表达的升高以及EndoG和AIF的增加。总体而言，这可能表明PW06在体外对人GBM 8401细胞具有抗GBM活性并可诱导细胞凋亡。
