Array-based genome-wide RNAi screening to identify shRNAs that enhance p53-related apoptosis in human cancer cells.
Array-based genome-wide RNAi screening to identify shRNAs that enhance p53-related apoptosis in human cancer cells.
复制标题
DOI：
10.18632/oncotarget.2272
复制
发表时间：
2014-09-15
期刊：
影响因子：
--
通讯作者：
Tokino T
中科院分区：
文献类型：
--
作者：
Idogawa M;Ohashi T;Sugisaka J;Sasaki Y;Suzuki H;Tokino T
p53转导是一种潜在有效的癌症疗法，但由于对细胞凋亡的抗性，在所有人类癌症中不会产生良好的治疗反应。为了发现克服对p53诱导的细胞凋亡的抗性的因素，我们试图鉴定增强p53诱导的细胞凋亡的RNAi序列。我们在肝癌Huh-7和胰腺癌Panc-1细胞中筛选了全基因组慢病毒shRNA文库，这两种细胞都能抵抗p53诱导的凋亡。在感染表达p53或LacZ的腺病毒作为对照后，通过微阵列分析经shRNA处理的群体。我们鉴定了与对照细胞相比，p53感染细胞中显着减少的shRNAs。在这些shRNA中，shRNA-58335在测试的两种癌细胞系中显著减少。shRNA-58335在体外增强了p53相关的凋亡，并增强了腺病毒p53转导对体内肿瘤生长的抑制作用。此外，shRNA-58335增强的凋亡反应也通过用PRIMA-1处理而证实，PRIMA-1重新激活突变型p53，而不是腺病毒p53转导。我们发现，在对p53诱导的凋亡有抗性的癌细胞中，shRNA-58335在p53转导或用小分子药物恢复p53功能后引起凋亡反应。p53修复和基于RNAi的药物的组合有望成为一种有前途的新型癌症治疗方法。
p53 transduction is a potentially effective cancer therapy but does not result in a good therapeutic response in all human cancers due to resistance to apoptosis. To discover factors that overcome resistance to p53-induced apoptosis, we attempted to identify RNAi sequences that enhance p53-induced apoptosis. We screened a genome-wide lentiviral shRNA library in liver cancer Huh-7 and pancreatic cancer Panc-1 cells, both of which resist p53-induced apoptosis. After the infection of adenovirus expressing p53 or LacZ as a control, shRNA-treated populations were analyzed by microarray. We identified shRNAs that were significantly decreased in p53-infected cells compared with control cells. Among these shRNAs, shRNA-58335 was markedly decreased in both cancer cell lines tested. shRNA-58335 enhanced p53-related apoptosis in vitro and augmented the inhibitory effect of adenoviral p53 transduction on tumor growth in vivo. Furthermore, the enhanced apoptotic response by shRNA-58335 was also confirmed by treatment with PRIMA-1, which reactivates mutant p53, instead of adenoviral p53 transduction. We found that shRNA-58335 evokes the apoptotic response following p53 transduction or functional restoration of p53 with a small molecule drug in cancer cells resistant to p53-induced apoptosis. The combination of p53 restoration and RNAi-based drugs is expected to be a promising novel cancer therapy.