Zheng Fei, Zhang Zhongtao, Flamini Valentina, Jiang Wen G, Cui Yuxin
Cardiff China Medical Research Collaborative, Cardiff University School of Medicine, Cardiff, U.K.
Department of General Surgery, Beijing Key Laboratory of Cancer Invasion and Metastasis Research & National Clinical Research Centre for Digestive Diseases, Beijing Friendship Hospital, Capital Medical University, Beijing, P.R. China.
Anticancer Res. 2017 Aug;37(8):4361-4369. doi: 10.21873/anticanres.11830.
BACKGROUND/AIM: Colorectal cancer (CRC) is the third most common cancer in the world. The high mortality of this tumor is mainly due to its invasive properties, as it forms metastases in multiple organs, preferentially in the liver. There has evidence showing that C-X-C chemokine receptor type 4 (CXCR-4) and its ligand, stromal cell-derived factor-1 (SDF-1), plays an important role in cancer progression and metastasis. However, the molecular mechanism underling the CRCR4-mediated CRC metastasis has not been well characterized. In this study, we aimed to investigate the roles of CXCR4 in colorectal cancer using the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9)-based genomic editing technique.
We knocked-down CXCR4 using specific guide-RNA linked CRISPR/Cas9 in HT115 and COLO201 colon cancer cell lines which exhibited high levels of endogenous CXCR4 gene expression. Stable HT115 cells with CXCR4 knock-down were established by CRISPR plasmid transfection and validation was confirmed using T7 endonuclease 1 (T7EN1), flow cytometry (FACS) and western blotting assays.
Knock-down of CXCR4 did not decrease proliferation of HT115 cells, but decreased the adhesion potential of cells to the human umbilical vein endothelial cells (HUVEC) and extracellular matrix. We further demonstrated that the AKT and type 1 insulin-like growth factor receptor (IGF1R) signalling pathways may be involved in the alteration of adhesion in CRC cells when CXCR4 is knocked down.
Our data suggest that CXCR4 plays a key role in colorectal cancer progression via the mediation of tumor cell adhesion.
背景/目的：结直肠癌（CRC）是全球第三大常见癌症。该肿瘤的高死亡率主要归因于其侵袭性，因为它会在多个器官形成转移灶，尤其是在肝脏。有证据表明，C-X-C趋化因子受体4（CXCR-4）及其配体基质细胞衍生因子-1（SDF-1）在癌症进展和转移中起重要作用。然而，CXCR4介导的结直肠癌转移的分子机制尚未完全明确。在本研究中，我们旨在使用基于成簇规律间隔短回文重复序列（CRISPR）/CRISPR相关蛋白9（Cas9）的基因组编辑技术，研究CXCR4在结直肠癌中的作用。
我们在HT115和COLO201结肠癌细胞系中使用与CRISPR/Cas9相连的特异性向导RNA敲低CXCR4，这两种细胞系内源性CXCR4基因表达水平较高。通过CRISPR质粒转染建立CXCR4敲低的稳定HT115细胞系，并使用T7核酸内切酶1（T7EN1）、流式细胞术（FACS）和蛋白质印迹分析进行验证。
敲低CXCR4并未降低HT115细胞的增殖，但降低了细胞与人脐静脉内皮细胞（HUVEC）及细胞外基质的黏附能力。我们进一步证明，当CXCR4被敲低时，AKT和1型胰岛素样生长因子受体（IGF1R）信号通路可能参与了结直肠癌细胞黏附的改变。
我们的数据表明，CXCR4通过介导肿瘤细胞黏附在结直肠癌进展中起关键作用。
