Li Lin, Niu Qian, Zhu Yuanmin, Fan Biao, Yang Guibin
Department of Gastroenterology, Aerospace Center Hospital, Peking University Aerospace School of Clinical Medicine, Beijing, China.
Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education), Gastrointestinal Cancer Center, Peking University Cancer Hospital and Institute, Beijing, China.
J Gastrointest Oncol. 2022 Dec;13(6):2799-2808. doi: 10.21037/jgo-22-928.
Deoxyribonucleic acid (DNA) methyltransferase inhibitors, such as decitabine, have made great advances in cancer therapy as combinational drugs. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has an obvious anti-tumor effect; however, some gastric cancer (GC) cells are resistant to TRAIL-induced cell death. This study sought to explore the synergistic anti-tumor effect of TRAIL and decitabine, and the potential synergetic mechanism.
The cell growth inhibition effect was monitored by the IncuCyte ZOOM Live-Cell Analysis System, and cell viability was determined by Cell Counting Kit-8 assays. Apoptosis was detected by Annexin V/Propidium Iodide double staining. Death receptor 4 (DR4) was knocked down by ribonucleic acid (RNA) interference, and the effect of DR4 deletion on TRAIL sensitivity was analyzed. Methylation-specific polymerase chain reaction (PCR) was applied to determine the methylation status of DR4. The messenger RNA (mRNA) and protein expression levels were detected by quantitative real-time polymerase chain reaction (qRT-PCR) and western blot. The expression of the DRs on the cell membrane surfaces was analyzed by flow cytometry.
The combined use of decitabine and TRAIL synergistically inhibited cell growth in 2 TRAIL-resistant cell lines. Further, decitabine augmented TRAIL-induced apoptosis in a caspase-dependent manner. The co-application of decitabine and TRAIL facilitated the activation of caspase-7, -8, -9, and poly ADP-ribose polymerase (PARP). Notably, decitabine increased the expression of DR4 at the transcriptional and post-transcriptional levels. DR4 expression on the cell membrane surfaces was also upregulated after decitabine exposure. The depletion of DR4 by specific inhibitors attenuated TRAIL-induced apoptosis and weakened the synergistic effects of decitabine and TRAIL. In addition, DR4 gene presented methylation status in SNU-1 cells. The low mRNA and protein expression of DR4 were also detected in SNU-1 cells.
Decitabine enhances the effect of TRAIL by inhibiting the growth and inducing the apoptosis of GC cells. This is achieved by the epigenetic modification of decitabine, which upregulates DR4. Decitabine may act as a sensitizing agent of TRAIL. The combined use of decitabine and TRAIL may provide a novel idea for GC treatment.
脱氧核糖核酸（DNA）甲基转移酶抑制剂，如地西他滨，作为联合用药在癌症治疗方面取得了重大进展。肿瘤坏死因子相关凋亡诱导配体（TRAIL）具有明显的抗肿瘤作用；然而，一些胃癌（GC）细胞对TRAIL诱导的细胞死亡具有抗性。本研究旨在探讨TRAIL与地西他滨的协同抗肿瘤作用及其潜在的协同机制。
通过IncuCyte ZOOM活细胞分析系统监测细胞生长抑制作用，并通过细胞计数试剂盒-8检测法测定细胞活力。采用膜联蛋白V/碘化丙啶双染法检测细胞凋亡。通过核糖核酸（RNA）干扰敲低死亡受体4（DR4），并分析DR4缺失对TRAIL敏感性的影响。应用甲基化特异性聚合酶链反应（PCR）确定DR4的甲基化状态。通过定量实时聚合酶链反应（qRT-PCR）和蛋白质免疫印迹法检测信使核糖核酸（mRNA）和蛋白质表达水平。通过流式细胞术分析细胞膜表面死亡受体（DRs）的表达。
地西他滨与TRAIL联合使用可协同抑制2种TRAIL抗性细胞系的细胞生长。此外，地西他滨以半胱天冬酶依赖性方式增强TRAIL诱导的细胞凋亡。地西他滨与TRAIL联合应用促进了半胱天冬酶-7、-8、-9和聚ADP核糖聚合酶（PARP）的激活。值得注意的是，地西他滨在转录和转录后水平上增加了DR4的表达。地西他滨处理后，细胞膜表面的DR4表达也上调。特异性抑制剂使DR4缺失减弱了TRAIL诱导的细胞凋亡，并削弱了地西他滨与TRAIL的协同作用。此外，DR4基因在SNU-1细胞中呈现甲基化状态。在SNU-1细胞中也检测到DR4的低mRNA和蛋白质表达。
地西他滨通过抑制GC细胞生长和诱导其凋亡来增强TRAIL的作用。这是通过地西他滨的表观遗传修饰实现的，该修饰上调了DR4。地西他滨可能作为TRAIL的增敏剂。地西他滨与TRAIL联合使用可能为GC治疗提供新思路。
