STAT转录因子家族与新型STAT3抑制剂在调控乳腺癌放射敏感性与辐射耐受性中的作用及分子机制研究

Radiation therapy is the preferred option for breast cancer before and after surgery, as well as for advanced stage breast cancer. Key element to improve therapeutic effect for better outcome is to enhance radiosensitivity and reduce radioresistance in tumors. In preliminary studies, we found that STAT family of transcription factors are induced and activated in breast cancer cells receiving radiation. Particularly, STATs (mainly STAT3) are consistently expressed and constitutively activated in radioresistant breast cancer cells, indicating that STAT factors involve in the regulation of radiosensitivity and radioresistance of cancer cells, while molecular mechanisms remain un-elucidated. We therefore hypothesized that STAT factors regulate radiosensitivity and radioresistance via regulating cancer stem cell function and self-renewal; the novel STAT3 inhibitors we developed may block STAT3 activity and thus regulate the function of cancer stem cells to achieve enhanced radiosensitivity and reduced radioresistance in breast cancer cells. We will alter expression of STATs and then characterize the effect on radiosensitivity / radioresistance , cancer stem cell function and self-renewal, and cell cycle arrest. We will also screen to identify the signaling pathways and related genes involved in radiosensitivity / radioresistance. In vivo, xenograft and transgenic animal model studies will clarify the value of STAT3 as a target gene for improving radiosensitivity, and whether the novel STAT3 inhibitors we develop could be used as radiosensitizer during radiation therapy.

放射治疗是乳腺癌手术前后以及中晚期乳腺癌治疗的首选方法，为放疗增敏从而减低肿瘤的辐射耐受性是提高放疗效果和治愈率的关键。在前期研究中我们发现放射照射后乳腺癌细胞中STAT因子被激活，在耐辐射乳腺癌细胞中STAT（STAT3等）的表达和激活显著上调，表明STAT转录因子家族参与了调节肿瘤细胞放射敏感性和辐射耐受性，但分子机理并未阐明。我们假设STAT通过影响肿瘤干细胞功能来调控放射敏感性和辐射耐受性；使用新型STAT3抑制剂即可阻断STAT3活性从而调控肿瘤干细胞的功能以达到增强放射敏感性并减低辐射耐受性。体外实验将改变表达水平来验证STAT对放射敏感性/辐射耐受性，肿瘤干细胞功能与自我更新和细胞周期的影响。我们也将确定参与介导辐射反应/辐射耐受性的信号传导通路和相关基因。移植瘤和转基因动物模型研究将明确STAT3是否为放射敏感性靶基因，以及新研发STAT3抑制剂可否作为乳腺癌放疗增敏剂。

放射治疗是乳腺癌手术前后以及中晚期乳腺癌治疗的首选方法，为放疗增敏从而减低肿瘤的辐射耐受性是提高放疗效果和治愈率的关键。在前期研究中我们发现放射照射后乳腺癌细胞中STAT因子被激活，在耐辐射乳腺癌细胞中STAT（STAT3等）的表达和激活显著上调，表明STAT转录因子家族参与了调节肿瘤细胞放射敏感性和辐射耐受性，但分子机理并未阐明。我们假设STAT通过影响肿瘤干细胞功能来调控放射敏感性和辐射耐受性，使用新型STAT3抑制剂即可阻断STAT3活性从而调控肿瘤干细胞的功能以达到增强放射敏感性并减低辐射耐受性。本研究开展的体内体外实验建立了三阴性乳腺癌辐射耐受细胞系并发现电离辐射(IR)诱导野生型和辐射耐受型的三阴性乳腺癌细胞系MDA-MB-231和MDA-MB-468中STAT3转录因子的Tyr-705残基磷酸化，诱导细胞活性氧（ROS）水平升高， 激活了STAT3/Bcl-2通路和活性氧生成。与辐射敏感的亲代三阴性乳腺癌细胞相比，辐射耐受乳腺癌细胞ROS水平明显较低，且磷酸化STAT3和Bcl-2的蛋白水平较高。在具有获得性抗辐射性的TNBC细胞中，STAT3/Bcl-2通路高度持续激活，而较低的活性氧水平与TNBC细胞获得性抗辐射性有关。Niclosamide，一种STAT3抑制剂，通过抑制STAT3和Bcl-2并诱导ROS水平升高，有效地抑制了乳腺癌细胞活性和STAT3活性， 并与IR协同诱导抗辐射TNBC细胞凋亡。通过诱导ROS生成， Niclosamide增强抗辐射MDA-MB-231/R细胞的辐射敏感性，而用shRNA特异性敲除STAT3可恢复抗辐射TNBC细胞的放射敏感性。本研究表明Niclosamide对异种乳腺癌肿瘤的体内增敏作用，联合放射治疗具有良好的安全性。本项目完成了计划书中的既定目标，证明了STAT3调节乳腺癌细胞放射敏感性的机制，并明确STAT3抑制剂可作为乳腺癌放射增敏剂, 阐明了乳腺癌放射耐受性分子生物学机制，拓展了临床乳腺癌放射个体化治疗的基础研究，为放射治疗临床应用研究提供了新的思路。