Lane Jane, Martin Tracey A, Jiang Wen G
Metastasis and Angiogenesis Research Group, Cardiff University School of Medicine, Cardiff, UK.
World J Oncol. 2010 Feb;1(1):7-13. doi: 10.4021/wjon2010.01.1202. Epub 2010 Feb 1.
Cell motility and migration are known to be regulated by the Rho family of GTPases through their effects on the actin cytoskeleton. In breast cancer studies, RhoC has been identified as a highly specific marker in detecting tumors that developed metastases. This study aims to investigate the impact of targeting RhoC in human breast cancer cells by utilising ribozyme transgene technology and to assess its effect on cancer cell invasion.
Retroviral hammerhead ribozyme transgenes, regulated by doxycycline, were designed to specifically target human RhoC mRNA. The breast cancer cell line MDA-MB-231 was transfected with either a retroviral RhoC transgene or a control retroviral transgene. Stably transfected cells were tested for their invasiveness and migratory properties .
testing of the invasiveness of wild type, plasmid control and the RhoC knockdown cells showed that MDA-MB-231 cells had significantly reduced invasiveness compared with MDA-MB-231 (p < 0.038 RHOC2 knockdown cells; p < 0.006 RHOC3 knockdown cells) and MDA-MB-231 control plasmid cells (p < 0.07 RHOC2 knockdown cells; p < 0.002 RHOC3 knockdown cells). An even greater reduction in invasiveness of the MDA-MB-231 cells compared with the MDA-MB-231 cells was seen in response to hepatocyte growth factor (HGF/SF) (p < 0.009 RHOC1 knockdown; p = 0.004 RHOC2 knockdown; p = 0.00007 RHOC3 knockdown). The addition of doxycycline significantly improved the effectiveness of the ribozyme transgenes (p < 0.04 for all three Rho ribozymes), but did not improve the effectiveness of these knockdown cells when treated with HGF/SF (p > 0.1 for all three ribozymes).
This data would indicate that targeting RhoC may be an effective way to reduce the invasive potential of human breast cancer cells.
已知细胞运动性和迁移受Rho家族小GTP酶调控，通过其对肌动蛋白细胞骨架的作用发挥影响。在乳腺癌研究中，RhoC已被确定为检测发生转移肿瘤的高度特异性标志物。本研究旨在利用核酶转基因技术研究靶向RhoC对人乳腺癌细胞的影响，并评估其对癌细胞侵袭的作用。
设计由强力霉素调控的逆转录病毒锤头状核酶转基因，特异性靶向人RhoC mRNA。用逆转录病毒RhoC转基因或对照逆转录病毒转基因转染乳腺癌细胞系MDA-MB-231。对稳定转染的细胞进行侵袭性和迁移特性检测。
对野生型、质粒对照和RhoC基因敲低细胞的侵袭性检测显示，与MDA-MB-231细胞相比，MDA-MB-231细胞侵袭性显著降低（RhoC2基因敲低细胞p < 0.038；RhoC3基因敲低细胞p < 0.006），与MDA-MB-231对照质粒细胞相比也显著降低（RhoC2基因敲低细胞p < 0.07；RhoC3基因敲低细胞p < 0.002）。与MDA-MB-231细胞相比，MDA-MB-231细胞对肝细胞生长因子（HGF/SF）反应时侵袭性降低更明显（RhoC1基因敲低p < 0.009；RhoC2基因敲低p = 0.004；RhoC3基因敲低p = 0.00007）。添加强力霉素显著提高了核酶转基因的有效性（三种Rho核酶均p < 0.04），但在用HGF/SF处理时这些基因敲低细胞的有效性未提高（三种核酶均p > 0.1）。
这些数据表明靶向RhoC可能是降低人乳腺癌细胞侵袭潜能的有效方法。
