Department of Thoracic Oncology, Tianjin Lung Cancer Center, Tianjin Cancer Institute & Hospital, Tianjin Medical University, Tianjin 300060, PR China.
Biochem Biophys Res Commun. 2011 Jul 1;410(2):364-70. doi: 10.1016/j.bbrc.2011.06.020. Epub 2011 Jun 12.
Neuroblastoma is the most common childhood solid tumor, yet current treatment approaches have not been able to effectively control this cancer. Amplification and overexpression of MYCN have been shown to be closely related with high risk and poor prognosis in neuroblastoma. This suggests that MYCN is an important target for the antitumor therapy. Recently, vector-based RNA interference (RNAi) systems have been successfully used to eliminate gene expression, but knockdown of MYCN by vector-based RNAi as a therapeutic model for neuroblastoma has not been fully established. In this study, we used a lentivirus vector-based RNAi approach which expresses short hairpin RNA (shRNA) to knockdown MYCN in neuroblastoma cell lines IMR-32 and LAN-1. Western blotting analysis showed that expressions of MYCN were efficiently downregulated after infection with MYCN shRNA expression vector. The stable suppression of MYCN expression induced differentiation and apoptosis in neuroblastoma cell lines. Furthermore, we demonstrated that these changes were associated with caspase-3 activation, p27 upregulation as well as Bcl-2 and MDM2 downregulation. Finally, we demonstrated that downregulation of MYCN expression significantly reduced colony formation in vitro and tumor growth in nude mice. Our data indicate that lentivirus vector-mediated silencing of MYCN in neuroblastoma cells could efficiently and significantly inhibit tumor growth both in vitro and in vivo. Therefore we demonstrate the therapeutic potential of lentivirus-delivered shRNA as a novel approach for treatment of neuroblastoma and other malignant tumors with MYCN overexpression.
神经母细胞瘤是最常见的儿童实体肿瘤，但目前的治疗方法还不能有效地控制这种癌症。研究表明，MYCN 的扩增和过表达与神经母细胞瘤的高风险和预后不良密切相关。这表明 MYCN 是抗肿瘤治疗的一个重要靶点。最近，基于载体的 RNA 干扰（RNAi）系统已被成功用于消除基因表达，但基于载体的 RNAi 敲低 MYCN 作为神经母细胞瘤的治疗模型尚未得到充分确立。在本研究中，我们使用了一种基于慢病毒载体的 RNAi 方法，该方法表达短发夹 RNA（shRNA），以敲低神经母细胞瘤细胞系 IMR-32 和 LAN-1 中的 MYCN。Western blot 分析表明，感染 MYCN shRNA 表达载体后，MYCN 的表达被有效地下调。MYCN 表达的稳定抑制诱导神经母细胞瘤细胞系分化和凋亡。此外，我们证明这些变化与 caspase-3 激活、p27 上调以及 Bcl-2 和 MDM2 下调有关。最后，我们证明下调 MYCN 表达显著减少了体外集落形成和裸鼠体内肿瘤生长。我们的数据表明，慢病毒载体介导的神经母细胞瘤细胞中 MYCN 的沉默可以有效地显著抑制体外和体内肿瘤的生长。因此，我们证明了慢病毒递送的 shRNA 作为治疗 MYCN 过表达的神经母细胞瘤和其他恶性肿瘤的新方法的治疗潜力。
