Laboratory of Biological Cancer Therapy, Xuzhou Medical College, Xuzhou, Jiangsu, PR China.
Cancer Gene Ther. 2014 Oct;21(10):416-26. doi: 10.1038/cgt.2014.45. Epub 2014 Sep 19.
Reactive oxygen species (ROS) have a crucial role in melanoma differentiation-associated gene-7 (MDA-7)/interleukin-24 (IL-24)-induced cancer cell apoptosis. However, cancer cell has a series of protective mechanisms to resist ROS damage. Nuclear factor erythroid 2-related factor 2 (Nrf2) activates antioxidant response element (ARE)-mediated gene expression involved in cellular protection against oxidative stress. As the Nrf2 repressor, Kelch-like ECH-associated protein-1 (Keap1) sequesters Nrf2 in cytoplasm to block Nrf2 nuclear translocation. In the present study, administration of MDA-7/IL-24 by means of tumor-selective replicating adenovirus (ZD55-IL-24) was used to investigate whether ZD55-IL-24 could attenuate Nrf2-mediated oxidative stress response in cancer cell. We found that ZD55-IL-24 effectively strengthened the association between Nrf2 and Keap1 to restrict Nrf2 nuclear translocation, thereby inhibiting ARE-dependent transcriptional response. To evaluate the detailed mechanism underlying the suppression of ZD55-IL-24 on Nrf2-mediated oxidative stress response, we further tested three different mitogen-activated protein kinase (MAPK) signaling pathways in A549 and HeLa cells transfected by ZD55-IL-24. Our data showed that ZD55-IL-24 inhibited extracellular signal-regulated kinase (ERK) signal pathway but activated p38 and c-Jun-NH2-kinase (JNK) signal pathways to exert the tumor-specific apoptosis. Moreover, ERK pathway inhibitor U0126 prevented Nrf2 phosphorylation at Ser40 to retard Nrf2 nuclear translocation, thus decreasing antioxidant gene transcription. In contrast, p38 pathway inhibitor SB203580 obviously promoted the dissociation of Nrf2 from Keap1 to promote antioxidant gene transcription. However, JNK pathway had no effect on Nrf2 subcellular localization or the association of Nrf2 with Keap1. Conclusively, our results indicate that ZD55-IL-24 inhibits Nrf2-mediated oxidative stress response not only by activating p38 signal pathway to potentiate the association of Nrf2 and Keap1 but also by suppressing ERK signal pathway to postpone Nrf2 nuclear translocation. Given the 'dark' side of Nrf2 on carcinoma cell survival and chemoresistance, our study provides a novel explanation about MDA-7/IL-24-induced cancer-specific apoptosis and therapeutic sensitization through suppression of the cytoprotective system.
活性氧（ROS）在黑色素瘤分化相关基因-7（MDA-7）/白细胞介素-24（IL-24）诱导的癌细胞凋亡中起着至关重要的作用。然而，癌细胞有一系列的保护机制来抵抗 ROS 损伤。核因子红细胞 2 相关因子 2（Nrf2）激活抗氧化反应元件（ARE）介导的基因表达，参与细胞对抗氧化应激的保护。Kelch 样 ECH 相关蛋白 1（Keap1）作为 Nrf2 的抑制因子，将 Nrf2 隔离在细胞质中，阻止 Nrf2 核转位。在本研究中，通过肿瘤选择性复制腺病毒（ZD55-IL-24）给予 MDA-7/IL-24，以研究 ZD55-IL-24 是否能减弱癌细胞中 Nrf2 介导的氧化应激反应。我们发现 ZD55-IL-24 能有效地增强 Nrf2 与 Keap1 的结合，限制 Nrf2 的核转位，从而抑制 ARE 依赖性转录反应。为了评估 ZD55-IL-24 对 Nrf2 介导的氧化应激反应的抑制作用的详细机制，我们进一步测试了 ZD55-IL-24 转染的 A549 和 HeLa 细胞中的三种不同的丝裂原激活蛋白激酶（MAPK）信号通路。我们的数据表明，ZD55-IL-24 抑制细胞外信号调节激酶（ERK）信号通路，但激活 p38 和 c-Jun-NH2-kinase（JNK）信号通路，从而发挥肿瘤特异性凋亡。此外，ERK 通路抑制剂 U0126 阻止 Nrf2 在 Ser40 处的磷酸化，从而阻止 Nrf2 的核转位，减少抗氧化基因的转录。相反，p38 通路抑制剂 SB203580 明显促进 Nrf2 与 Keap1 的解离，促进抗氧化基因的转录。然而，JNK 通路对 Nrf2 的亚细胞定位或与 Keap1 的结合没有影响。总之，我们的结果表明，ZD55-IL-24 不仅通过激活 p38 信号通路增强 Nrf2 与 Keap1 的结合，从而抑制 Nrf2 介导的氧化应激反应，还通过抑制 ERK 信号通路延缓 Nrf2 的核转位。鉴于 Nrf2 对癌细胞存活和化疗耐药性的“阴暗面”，我们的研究通过抑制细胞保护系统，为 MDA-7/IL-24 诱导的肿瘤特异性凋亡和治疗敏感性提供了一个新的解释。
