title: Small interfering RNA targeting S100A4 sensitizes non-small-cell lung cancer cells (A549) to radiation treatment.
abstract: OBJECTIVE
  This study aimed to investigate the impact of S100A4-small interfering RNA (S100A4-siRNA) on apoptosis and enhanced radiosensitivity in non-small-cell lung cancer (A549) cells. We also explored the mechanisms of radiosensitization and identified a new target to enhance radiosensitivity and gene therapy for non-small-cell lung cancer.
  
  
  METHODS
  RNA interference is a powerful tool for gene silencing. In this study, we constructed an effective siRNA to knock down S100A4. A549 cells were randomly divided into three groups: blank, negative control, and S100A4-siRNA. To investigate the effect of S100A4-siRNA, the expression of S100A4, E-cadherin, and p53 proteins and their messenger RNA (mRNA) was detected by Western blot and quantitative real-time polymerase chain reaction. Transwell chambers were used to assess cell invasion. Cell cycle and apoptosis were analyzed by flow cytometry. Radiosensitivity was determined by colony formation ability.
  
  
  RESULTS
  Our results demonstrate that S100A4-siRNA effectively silenced the S100A4 gene. When siRNA against S100A4 was used, S100A4 protein expression was downregulated, whereas the expressions of E-cadherin and p53 were upregulated. In addition, a clear reduction in S100A4 mRNA levels was noted compared with the blank and negative control groups, whereas E-cadherin and p53 mRNA levels increased. Transfection with S100A4-siRNA significantly reduced the invasiveness of A549 cells. S100A4 silencing induced immediate G2/M arrest in cell cycle studies and increased apoptosis rates in A549 cells. In clonogenic assays, we used a multitarget, single-hit model to detect radiosensitivity after S100A4 knockdown. All parameters (D0, Dq, α, β) indicated that the downregulation of S100A4 enhanced radiosensitivity in A549 cells. Furthermore, S100A4-siRNA upregulated p53 expression, suggesting that S100A4 may promote A549 cell proliferation, invasion, and metastasis by regulating the expression of other proteins. Therefore, siRNA-directed S100A4 knockdown may represent a viable clinical therapy for lung cancer.
  
  
  CONCLUSIONS
  S100A4 downregulation potentially enhances the sensitivity of human A549 cells to radiotherapy.
authors.affiliation: Department of Oncology, Affiliated Jinshan Hospital, Fudan University, Shanghai, People's Republic of China.
authors.forename: Ruixue
authors.initials: R
authors.lastname: Qi
authors.affiliation: Department of Oncology, Affiliated Jinshan Hospital, Fudan University, Shanghai, People's Republic of China.
authors.forename: Tiankui
authors.initials: T
authors.lastname: Qiao
authors.affiliation: Department of Oncology, Affiliated Jinshan Hospital, Fudan University, Shanghai, People's Republic of China.
authors.forename: Xibing
authors.initials: X
authors.lastname: Zhuang
keywords: A549 cells; S100A4; lung cancer; small interfering RNA