Secreted clusterin gene silencing enhances chemosensitivity of a549 cells to cisplatin through AKT and ERK1/2 pathways in vitro.

BACKGROUND/AIMS: Several studies have shown secreted clusterin (sCLU) silencing directed against sCLU mRNA in sCLU-rich lung cancer cell lines sensitized cells to chemotherapy. However, the molecular mechanisms underlying the effect of sCLU silencing on lung cancer cell chemosensitivity is not known. In the present study, we aimed to determine that vector expressing short hairpin RNA against sCLU RNA (sCLU-shRNA) enhances the chemosensitivity in human small cell lung cancer A549 cells in vitro by inhibition of phosphorylated ERK1/2 (p-ERK1/2) and Akt (p-Akt). METHODS: The pCDNA3.1-sCLU and control scrambled pCDNA3.1 plasmid was constructed. We investigated the effects of sCLU overexpression by pCDNA3.1-sCLU transfection on chemosensitivity to cisplatin (DDP) in A549 cells in vitro. We down-regulated sCLU expression by short hairpin RNA against sCLU RNA (sCLU-shRNA) and investigated the effects on chemosensitivity to DDP in A549 cells and A549(DDP)in vitro. In order to confirm the correlation between sCLU and AKT and ERK1/2 signals, cells were treated with wortmannin and U0126. RESULTS: We found the chemotherapeutic agent DDP activated sCLU. Overexpression of sCLU increased cellular DDP chemoresistance in the A549(DDP) and pCDNA3. 1-sCLU transfected A549 cells via inhibition DDP-induced apoptosis. Whereas sCLU knockdown induced chemosensitization in the S549 and A549(DDP) cells via increase of DDP-induced apoptosis. sCLU overexpression activated pAKT Ser(473) and pERK1/2(Thr202/Tyr204), and vice versa. Inhibition of pAKT Ser(473) and pERK1/2(Thr202/Tyr204) was sufficient to induce significant recover y in chemosensitivity to DDP in A549(DDP) in the presence of sCLU overexpression. The DDP activated sCLU, which directly regulated pAKT and pERK1/2. CONCLUSIONS: This novel finding suggests that therapies directed against sCLU and its downstream signaling targets pAKT and pERK1/2 may have the potential to enhance the efficacy of DDP-based chemotherapy.