General Clinical Research Center, Nanjing First Hospital, Nanjing Medical University, 210006, Nanjing, China.
School of Medicine, Southeast University, 210009, Nanjing, China.
Cell Death Dis. 2018 Apr 1;9(4):417. doi: 10.1038/s41419-018-0454-8.
Mounting evidences indicate that circular RNAs (circRNAs) have a vital role in human diseases, especially cancers. More recently, circHIPK3, a particularly abundant circRNA, was proposed to be involved in tumorigenesis. However, its role in colorectal cancer (CRC) has not been explored. In this study, we found circHIPK3 was significantly upregulated in CRC tissues and cell lines, at least in part, due to c-Myb overexpression and positively correlated with metastasis and advanced clinical stage. Moreover, Cox multivariate survival analysis showed that high-level expression of circHIPK3 was an independent prognostic factor of poor overall survival (OS) in CRC (hazard ratio [HR] = 2.75, 95% confidence interval [CI] 1.74-6.51, p = 0.009). Functionally, knockdown of circHIPK3 markedly inhibited CRC cells proliferation, migration, invasion, and induced apoptosis in vitro and suppressed CRC growth and metastasis in vivo. Mechanistically, by using biotinylated-circHIPK3 probe to perform RNA pull-down assay in CRC cells, we identified miR-7 was the only one microRNA that was abundantly pulled down by circHIPK3 in both HCT116 and HT29 cells and these interactions were also confirmed by biotinylated miR-7 pull-down and dual-luciferase reporter assays. Overexpression of miR-7 mimicked the effect of circHIPK3 knockdown on CRC cells proliferation, migration, invasion, and apoptosis. Furthermore, ectopic expression of circHIPK3 effectively reversed miR-7-induced attenuation of malignant phenotypes of CRC cells by increasing the expression levels of miR-7 targeting proto-oncogenes (FAK, IGF1R, EGFR, YY1). Remarkably, the combination of circHIPK3 silencing and miR-7 overexpression gave a better effect on tumor suppression both in vitro and in vivo than did circHIPK3 knockdown or miR-7 overexpression alone. Taken together, our data indicate that circHIPK3 may have considerable potential as a prognostic biomarker in CRC, and support the notion that therapeutic targeting of the c-Myb/circHIPK3/miR-7 axis may be a promising treatment approach for CRC patients.
越来越多的证据表明，环状 RNA（circRNAs）在人类疾病中，特别是癌症中起着至关重要的作用。最近，circHIPK3，一种特别丰富的 circRNA，被提出参与肿瘤发生。然而，它在结直肠癌（CRC）中的作用尚未被探索。在这项研究中，我们发现 circHIPK3 在 CRC 组织和细胞系中显著上调，至少部分原因是 c-Myb 过表达，并与转移和晚期临床阶段呈正相关。此外，Cox 多变量生存分析显示，高水平表达 circHIPK3 是 CRC 患者总体生存（OS）不良的独立预后因素（风险比 [HR] = 2.75，95%置信区间 [CI] 1.74-6.51，p = 0.009）。功能上，circHIPK3 的敲低显著抑制 CRC 细胞的增殖、迁移和侵袭，并在体外诱导细胞凋亡，在体内抑制 CRC 的生长和转移。机制上，我们在 CRC 细胞中使用生物素化 circHIPK3 探针进行 RNA 下拉实验，鉴定出 miR-7 是唯一一种在 HCT116 和 HT29 细胞中被 circHIPK3 大量下拉的 microRNA，并且这些相互作用也通过生物素化 miR-7 下拉和双荧光素酶报告基因实验得到证实。miR-7 的过表达模拟了 circHIPK3 敲低对 CRC 细胞增殖、迁移、侵袭和凋亡的影响。此外，circHIPK3 的异位表达通过增加 miR-7 靶向原癌基因（FAK、IGF1R、EGFR、YY1）的表达水平，有效逆转了 miR-7 诱导的 CRC 细胞恶性表型的衰减。值得注意的是，与单独敲低 circHIPK3 或过表达 miR-7 相比，在体外和体内，circHIPK3 沉默和 miR-7 过表达的联合作用对肿瘤抑制的效果更好。综上所述，我们的数据表明 circHIPK3 可能作为 CRC 的一个有前途的预后生物标志物，支持靶向 c-Myb/circHIPK3/miR-7 轴的治疗可能是 CRC 患者的一种有前途的治疗方法。
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