NR_038340在乳腺癌辐射抵抗中的作用及其机制探讨

We have been demonstrated that overexpression of NR_038340 correlated with radioresistance and poor prognosis in patients with breast cancer. Our data have been proved that miR-200c could bind to NR_038340 and decrease its expression level through reducing its half-life. Interaction of NR_038340 and USP7 leads to stabilization expression of CHK1 through enhancing USP7 ability of deubiquitylation to CHK1, which in turn promotes the breast cancer cell’s DNA damage repair and radioresistance. Based on our preliminary data, we first present that NR_038340 is one of the key genes in breast cancer radioresistance. To test the hypothesis, firstly, we will explore the physiological and functional roles of NR_038340 in breast cancer radioresistance by in vitro, mouse xenograft and transgene zebrafish model; secondly, we will deeply detect the mechanism of NR_038340 in breast cancer radioresistance by using RIA-seq，RNA pull down assay and mass spectrometry; last but not least, the effect of NR_038340 and its related effectors will be identified by clinical multicenter sample of breast cancer. From this work, the functional role of NR_038340 in breast cancer radioresistance will be explored and this will be helpful to discover NR_038340 as a clinical prognostic factor and a potential therapeutic target in patients with breast cancer.

前期我们发现，NR_038340是乳腺癌辐射抵抗和预后不良的分子标志物；与 miR-200c直接结合，半衰期被缩短从而表达水平降低；与USP7直接结合，对CHK1发挥去泛素化作用，进而稳定CHK1来介导辐射诱发的DNA损伤修复，以促进乳腺癌细胞的辐射抵抗过程。因此我们首次提出：NR_038340是乳腺癌辐射抵抗的关键分子之一。本项目拟通过体内外实验及转基因动物实验探讨NR_038340在乳腺癌辐射抵抗中的生物学功能；并利用RNA pull down、质谱、RIA-Seq等技术探讨其促进乳腺癌辐射抵抗的分子机制；再通过临床多中心大样本检测NR_038340及其相关蛋白质、miRNA 在预警乳腺癌辐射抵抗及判断预后方面的作用。本项目完成将揭示NR_038340在乳腺癌辐射抵抗中的作用机制，对乳腺癌预后判断具有重要的理论及临床价值，为寻找乳腺癌治疗的新靶点提供充分的理论依据。

放疗抵抗是乳腺癌放射治疗中的一大难题，往往直接导致乳腺癌治疗的失败。MiR-200c是被广泛报道的具有放疗增敏作用的miRNA，可明显增加乳腺癌的辐射敏感性，但具体作用机制尚不完全清楚。该课题中，我们在乳腺癌细胞中过表达miR-200c，然后行lncRNA芯片检测差异表达的lncRNA，发现表达miR-200c可以诱导一系列的lncRNA发生改变。在众多差异表达的lncRNA中，我们发现LINC02582（NR_038340）是miR-200c调控的靶基因；在乳腺癌细胞中过表达LINC02582可以诱导乳腺癌细胞辐射抵抗，而抑制LINC02582则增强乳腺癌细胞辐射敏感性。进一步的机制研究发现LINC02582可特异性地结合去泛素化蛋白酶USP7，抑制CHK1蛋白的泛素化降解，从而提高CHK1的蛋白水平，进而促进乳腺癌细胞的辐射抵抗。最后分析临床乳腺癌标本发现miR-200c、LINC02582及CHK1三者的表达水平存在相关性，LINC02582的高表达是乳腺癌患者放疗后预后不良的因素之一。我们证明了miR-200c/LINC02582/USP7/CHK1信号通路轴是调节乳腺癌细胞辐射敏感性的重要信号通路，为探讨乳腺癌放疗抵抗机制开辟了一条新的研究途径，也为临床防治乳腺癌放疗抵抗提供了新的理论依据。