靶向控释siRNA调控S6K1选择性剪接逆转非小细胞肺癌放疗抵抗的作用及机制研究

Non-small cell lung cancer (NSCLC) is one of the leading causes of cancer-related mortality worldwide, and radiotherapy plasys an important part in its treatment. Recently it was found that activation of mammalian target of rapamycin (mTOR) pathway was closely associated with radioresistance of NSCLC. Ribosomal S6 kinase 1 (S6K1), a major mTOR downstream signaling molecule, has two alternatively spliced short isoforms, S6K1(L) and S6K-1(S), which present with opposite functions by different feedback effects on the expressions of mTOR. Our previous work demonstrated an increase expression of S6K1(S) and a decrease expression of S6K1(L) in NSCLC radioresistant cell lines compared with that of parental cell lines. In addition, we also found there were sequences of exonic splicing enhancers located within downstream of exon 6 of Pre-mRNA of S6K1, indicating the possibility of modulating alternative splicing by targeting specific siRNA to reverse radioresistance. In this project, we will further study the molecular basis of S6K1 alternative splicing in the radioresistance of NSCLC, explore the role of siRNA in modulating S6K1 alternative splicing and its related mechanisms and analyze the effects of targeting S6K1 alternative splicing by controlled release of siRNA on the radiosensitivity of NSCLC. Successful completion of this study should provide new insights into the pathogenesis of radioresistance of NSCLC and provide new strategies to overcome its radioresistance.

非小细胞肺癌（NSCLC）是目前全球死亡率最高的癌症之一，放疗是其重要治疗手段。新近发现mTOR信号通路的过度活化与NSCLC放疗抵抗密切相关，其主要作用靶点S6K1的编码基因经选择性剪接可产生两种功能相反的蛋白异构体S6K1(L)和S6K-1(S)，反馈性影响mTOR表达。我们前期发现：在NSCLC放疗抵抗细胞株中S6K-1(S)表达升高，S6K1(L)表达下降；S6K1 Pre-mRNA在6号外显子下游存在促剪接因子结合的剪接增强子序列，提示可通过设计序列特异性siRNA靶向调控该位点剪接，逆转放疗抵抗。本课题拟进一步解析S6K1选择性剪接参与NSCLC放疗抵抗的分子基础，探讨siRNA干预S6K1选择性剪接的作用及机制，并在细胞和动物水平观测PLGA纳米材料靶向控释siRNA调控S6K1选择性剪接对NSCLC放疗敏感性的影响，为临床肺癌的放疗抵抗提供更具针对性的新策略理论和实验基础。

非小细胞肺癌（NSCLC）是目前全球死亡率最高的癌症之一，放疗是其重要治疗手段。如何减少肺癌放疗抵抗、增加放疗敏感性已成为改善其临床疗效的关键问题。本项目通过探讨mTOR信号通路分子S6K1及其剪接异构体在非小细胞肺癌及其放疗抵抗中的作用，证实了S6K1及其剪接异构体在非小细胞肺癌的发生发展中发挥着重要作用，且通过特异性阻断S6K1可下调PDCD4蛋白的表达，逆转非小细胞肺癌的放疗抵抗，为临床非小细胞肺癌的放疗抵抗提供了新的治疗靶点。溴结构域蛋白4（bromodomain-containing protein 4,BRD4）是溴结构域和超末端结构家族成员，在整个细胞周期中通过溴结构域结合组蛋白上的乙酰化赖氨酸残基，然后招募不同的转录调节因子从而调节靶基因的表达。本项目发现通过阻断非小细胞肺癌BRD4，可通过增加P21的表达明显增加非小细胞肺癌的放疗敏感性。另外，研究还发现二甲双胍可以通过下调磷酸化AMPKα来降低大鼠放射性肺炎的发生率。这些成果为非小细胞肺癌的放疗增敏及保护提供了极具前景的治疗靶点。