Department of Biological Sciences and Technology, National University of Tainan, Tainan, Taiwan.
Toxicology. 2012 Dec 16;302(2-3):221-32. doi: 10.1016/j.tox.2012.08.003. Epub 2012 Aug 15.
The development of multidrug resistance (MDR) to conventional chemoradiation therapy usually leads to failure in treating cervical cancer. This study aims to explore the effects and mechanisms of 7,3',4'-trihydroxyisoflavone (7,3',4'-THIF), one of the major metabolites of daidzein, on potentiating cytotoxicity of epirubicin (Epi), an anticancer drug in human cervical cancer HeLa cells. The cytotoxicity of Epi remarkably increased when it was combined with 7,3',4'-THIF. The cotreatment increased the reactive oxygen species (ROS) levels, including hydrogen peroxide and superoxide free radicals. 7,3',4'-THIF was shown to down-regulate the MDR1 promoter region composed of the elements of AP1, GC-box, and Y-box, as demonstrated by a luciferase assay. A negative regulation of hMDR1 gene with multiple transcription factors by this isoflavone may provide a novel molecular mechanism for MDR modulation. The mRNA expressions of MDR1, MDR-associated protein (MRP) 1, and MRP2 for the combined treatment were significantly lower than those of the Epi treatment. This result implies that MDR transporter-mediated Epi resistance is inhibited at various degrees by the addition of 7,3',4'-THIF. This isoflavone significantly enhanced intracellular Epi accumulation in HeLa cells. 7,3',4'-THIF and/or Epi triggered apoptosis through the upregulation of p53, Bax, and caspase-9. Apoptosis induction was also confirmed by the reduced mitochondrial membrane potential, increased sub-G1 and G2/M phases, nuclear DNA fragmentation, and chromatin condensation. Our findings demonstrate for the first time that 7,3',4'-THIF causes cell death in human cervical cancer cells through the ROS-dependent suppression of MDR transporters and p53-mediated activation of the intrinsic mitochondrial pathway of apoptosis. Thus, 7,3',4'-THIF has the potential to enhance the activity of a broad range of cancer chemotherapeutics in the MDR spectrum with the advantage of reducing adverse effects.
多药耐药（MDR）的发展通常导致常规放化疗治疗宫颈癌失败。本研究旨在探讨大豆苷元的主要代谢物之一 7,3',4'-三羟基异黄酮（7,3',4'-THIF）增强人宫颈癌 HeLa 细胞中抗癌药物表阿霉素（Epi）细胞毒性的作用和机制。当 Epi 与 7,3',4'-THIF 联合使用时，Epi 的细胞毒性显著增加。共处理增加了活性氧（ROS）水平，包括过氧化氢和超氧自由基。荧光素酶测定表明，7,3',4'-THIF 下调由 AP1、GC 盒和 Y 盒元件组成的 MDR1 启动子区域。这种异黄酮可能通过多个转录因子对 hMDR1 基因的负调控为 MDR 调节提供了新的分子机制。联合治疗的 MDR1、MDR 相关蛋白（MRP）1 和 MRP2 的 mRNA 表达明显低于 Epi 治疗组。这一结果表明，MDR 转运蛋白介导的 Epi 耐药在不同程度上被添加 7,3',4'-THIF 所抑制。该异黄酮显著增加了 HeLa 细胞中 Epi 的细胞内积累。7,3',4'-THIF 和/或 Epi 通过上调 p53、Bax 和 caspase-9 触发细胞凋亡。通过降低线粒体膜电位、增加亚 G1 和 G2/M 期、核 DNA 片段化和染色质浓缩也证实了细胞凋亡的诱导。我们的研究结果首次表明，7,3',4'-THIF 通过 ROS 依赖性抑制 MDR 转运蛋白和 p53 介导的内在线粒体凋亡途径的激活，导致人宫颈癌细胞死亡。因此，7,3',4'-THIF 有可能增强广谱抗癌药物在 MDR 谱中的活性，同时降低不良反应的风险。
