Stable Knockdown of Clusterin by Vector-Based RNA Interference in a Human Breast Cancer Cell Line Inhibits Tumour Cell Invasion and Metastasis

Abstract

Overexpression of the clusterin (CLU) gene occurs in breast cancer and is associated with lymph node metastasis. The present study explored the effect of CLU silencing on invasion and metastasis, and the relationship between CLU expression and the extracellular signal-regulated kinase (ERK) / matrix metalloproteinase-9 (MMP) signalling pathway in human breast cancer cells. A pcDNA3.1-based RNA interference approach was used to knockdown the CLU gene in MDA-231 cells (MDA-231-CLUi); control MDA-231 cells were transfected with an empty vector (MDA-231-Vec). Reverse transcription-polymerase chain reaction was used to assess CLU and MMP-9 mRNA levels, and Western blotting was used to analyse CLU, MMP-9 and ERK protein levels. Metastatic potential was evaluated using in vitro and in vivo models of invasion and metastasis. Compared with MDA-231-Vec cells, the MDA-231-CLUi cells demonstrated reduced migration and invasion in vitro and decreased metastatic potential in vivo. Reintroduction and reexpression of the CLU gene into the MDA-231-CLUi cells restored the invasive phenotype. MMP-9 mRNA and protein levels were reduced in MDA-231-CLUi cells, and there was a correlation between activated ERK and CLU and MMP-9 protein levels. CLU may regulate the aggressive behaviour of human breast cancer cells through modulation of ERK signalling and MMP9 expression.