Zhang Q, Hou D, Luo Z, Chen P, Lv B, Wu L, Ma Y, Chu Y, Liu H, Liu F, Yu S, Zhang J, Yang D, Liu J
Department of Digestive Diseases, Huashan Hospital, Fudan University, Shanghai, China.
Institutes of Biomedical Sciences of Shanghai Medical School, Fudan University, Shanghai, China.
Cell Death Dis. 2015 Aug 27;6(8):e1867. doi: 10.1038/cddis.2015.215.
The tumor-suppressor gene cyclin-dependent kinase inhibitor 1B (P27) is downregulated in gastric cancer cells mainly through proteolytic degradation mediated by the SKP-Cullin1-F-Box (SCF) complex. But the correlation between its downregulation and gastric cancer prognosis still remains indefinite. MLN4924, an anti-tumor agent, which suppresses the SCF complex by inhibiting Cullin1 neddylation, emerges as a promising tool to elucidate its functions in gastric cancer cells. In this study, MLN4924 induced significant growth inhibition of gastric cancer cells in a dose-dependent manner, along with the simultaneous accumulation of P27 and cell cycle abnormalities such as G2/M arrest. Importantly, we found that P27 silencing in MLN4924-treated cells resulted in an enhancement of growth inhibition both in vitro and in vivo. Mechanism analysis revealed the antagonism effects of antioxidants to this excess apoptosis, suggesting reactive oxygen species (ROS) overproduction especially in the mitochondria was the principal cause of the augmentation. Moreover, the robust ROS attacked the mitochondria to initiate collapse of the mitochondrial membrane permeability and the exportation of apoptosis-inducing factor (AIF), IAP-binding mitochondrial protein (SMAC/DIABLO) and cytochrome c. Finally, we also found that P27 knockdown affected the expression profile of several critical BH3 family members to amplify the mitochondrial dysfunction and apoptosis. In summary, our findings unveiled a protective role of P27 by maintaining mitochondrial membrane permeability in MLN4924-treated gastric cancer cells, and therefore highlighted the potential combination of MLN4924 with P27 inhibition to improve its therapeutic efficacy.
肿瘤抑制基因细胞周期蛋白依赖性激酶抑制剂1B（P27）在胃癌细胞中主要通过SKP-Cullin1-F-Box（SCF）复合物介导的蛋白水解降解而下调。但其下调与胃癌预后之间的相关性仍不明确。MLN4924是一种抗肿瘤药物，通过抑制Cullin1的类泛素化修饰来抑制SCF复合物，是阐明其在胃癌细胞中功能的一种有前景的工具。在本研究中，MLN4924以剂量依赖性方式诱导胃癌细胞显著生长抑制，同时伴有P27的积累和细胞周期异常，如G2/M期阻滞。重要的是，我们发现MLN4924处理的细胞中P27沉默导致体外和体内生长抑制增强。机制分析揭示了抗氧化剂对这种过度凋亡的拮抗作用，表明活性氧（ROS）特别是线粒体中的过量产生是这种增强的主要原因。此外，大量的ROS攻击线粒体，引发线粒体膜通透性的崩溃以及凋亡诱导因子（AIF）、IAP结合线粒体蛋白（SMAC/DIABLO）和细胞色素c的释放。最后，我们还发现P27敲低影响了几个关键BH3家族成员的表达谱，从而放大了线粒体功能障碍和凋亡。总之，我们的研究结果揭示了P27在维持MLN4924处理的胃癌细胞线粒体膜通透性方面的保护作用，因此强调了MLN4924与P27抑制联合使用以提高其治疗效果的潜力。
