RESUMEN
Neuroblastoma is a solid tumor that occurs mainly in children. Malignant neuroblastomas have a poor prognosis because conventional chemotherapeutic agents are not very effective. Survivin, a member of the inhibitor of the apoptosis protein family, plays a significant role in cell division, inhibition of apoptosis, and promotion of cell proliferation and invasion. Previous studies found that survivin is highly expressed in some malignant neuroblastomas and is correlated with poor prognosis. The aim of this study was to investigate whether survivin could serve as a potential therapeutic target of human neuroblastoma. We employed RNA interference to reduce survivin expression in the human neuroblastoma SH-SY5Y cell line and analyzed the effect of RNA interference on cell proliferation and invasion in vitro and in vivo. RNA interference of survivin led to a significant decrease in invasiveness and proliferation and increased apoptosis in SH-SY5Y cells in vitro. RNA interference of survivin inhibited tumor growth in vivo by 68±13% (P=0.002) and increased the number of apoptotic cells by 9.8±1.2% (P=0.001) compared with negative small interfering RNA (siRNA) treatment controls. Moreover, RNA interference of survivin inhibited the formation of lung metastases by 92% (P=0.002) and reduced microvascular density by 60% (P=0.0003). Survivin siRNA resulted in significant downregulation of survivin mRNA and protein expression both in vitro and in vivo compared with negative siRNA treatment controls. RNA interference of survivin was found to be a potent inhibitor of SH-SY5Y tumor growth and metastasis formation. These results support further clinical development of RNA interference of survivin as a treatment of neuroblastoma and other cancer types.
Asunto(s)
Animales , Humanos , Apoptosis/efectos de los fármacos , Inhibidores de Cisteína Proteinasa/farmacología , Proteínas Inhibidoras de la Apoptosis/efectos de los fármacos , Neoplasias Pulmonares/secundario , Neuroblastoma/patología , ARN Interferente Pequeño/farmacología , Antineoplásicos/farmacología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Regulación hacia Abajo/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Inmunohistoquímica , Etiquetado Corte-Fin in Situ , Proteínas Inhibidoras de la Apoptosis/genética , Proteínas Inhibidoras de la Apoptosis/metabolismo , Ratones Desnudos , Invasividad Neoplásica , Neuroblastoma/secundario , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , ARN Neoplásico/efectos de los fármacos , ARN Neoplásico/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Cytotoxic effect of either cisplatin or p53 gene transfection of lung cancer cells may be different depending on the p53 status of cells. We investigated cytotoxic effects on the combined treatment of cisplatin and adenovirus mediated p53 gene transfer (Avp53) in both H460 and H1299 cells in vitro. The results showed the highest numbers of apoptotic cells in both H460 and H1299 cells following the combined treatment regardless of p53 status in comparison with either cisplatin or Avp53 alone. The expression levels of p53, p21, Bax and ICE were examined to understand a possible cellular signal path of the combined treatment. In western analyses, the patterns of phosphorylated p53 protein were different between Avp53 and combined treatment. The expressions of p21 and Bax were increased in combined treatment, whereas the cleaved form of ICE (20 kD) was not detected. These results suggest that cisplatin induced p53 protein phosphorylation and may activate the downstream of p53 gene expression such as p21 and Bax. The enhanced apoptosis of lung cancer cells by the combined treatment may be useful in the development of clinical therapeutic modality of lung tumors.