RIZ1基因对宫颈腺癌放疗敏感性的影响及作用机制研究

The incidence of cervical adenocarcinoma has increased gradually in recent years, and radiation therapy is the preferred program for patients with advanced cervial cancer, however the radiosensitivity and prognosis of cervical adenocarcinoma is poorer compared with squamous cell carcinoma. Therefore, there is an urgent need to predict and improve the radiosensitivity of cervical adenocarcinoma. The retinoblastoma protein-interacting zinc finger gene RIZ1 is a new important tumor suppressor gene. In our preliminary experiments, we found that the expression of RIZ1 was significantly down-regulated in cervical cancer tissues, especially in cervical adenocarcinoma, after treatment with 5-aza-dC,the mRNA expression of RIZ1 was recovered and cell cycle was arrested at the G2/M phase,and also the sensitivity enhancement ratio (SER) was increased. In this project, we will examine the association between the expression of RIZ1 and its relationship with radiosensitivity in cervical adenocarcinoma tissues by using clinical specimen. And also we will construct HeLa cell lines and the model of athymic mouse with high stabilized expression of RIZ1, and to investigate whether the radiosensitivity of cervial adenocarcinoma can be improved after upregulation of RIZ1 in vitro and in vivo. At the same time, the application of Affymetrix company expression microarray screening the differentially expressed genes and related signal pathway, clarify the key molecular and function mechanism of RIZ1 gene improving the radiosensitivity of cervial adenocarcinoma. Based on this study, the conclusion will provide a theoretical and experimental basis for clinic to predict the prognosis and improve the curative effect for clinical radiotherapy and provide new molecular targets for individualization radiation.

宫颈腺癌的发病率逐年上升，放射治疗是中晚期宫颈癌患者的首选方案，而宫颈腺癌的放疗敏感性较低，所以预后较差。寻找临床上可预测和提高宫颈腺癌放射敏感性的靶基因，将对指导临床治疗具有重要意义。我们前期研究发现，抑癌基因RIZ1由于异常甲基化在宫颈腺癌组织中低表达，应用甲基化抑制剂可恢复RIZ1的表达，并使细胞阻滞在G2/M期，细胞凋亡增加，肿瘤细胞的放射敏感性也随之增强。本项目中，我们将结合临床资料重点分析RIZ1表达与放疗疗效的关联；建立稳定高表达RIZ1的HeLa细胞和裸鼠体内移植瘤模型，体内外实验探讨RIZ1表达增强后能否提高宫颈腺癌的放疗敏感性。同时应用表达谱芯片筛查这一过程中差异表达基因及相关信号通路,阐明RIZ1基因提高放射敏感性的作用机制。从临床患者-细胞系-动物模型三个层次，明确RIZ1基因在预测和提高放疗敏感性的作用，为临床个体化放疗及提高放疗疗效提供理论依据和实验基础。

宫颈癌是妇科常见的恶性肿瘤之一，在世界范围内，其发病率和死亡率分别位列女性恶性肿瘤的第四位。放射治疗是既是中晚期宫颈癌的主要方法，也是早期宫颈癌术后辅助治疗的主要手段，然而有一部分患者疗效不满意。在宫颈癌中，肿瘤放疗敏感性及患者预后受许多因素的影响，放射敏感性差异与细胞乏氧、细胞周期、增殖活性、DNA损伤修复和细胞凋亡等密切相关。如果在放射治疗前可预测肿瘤的放射敏感性，制定个体化方案，则可提高疗效，避免不必要的毒副作用。在本项目资助下我们围绕RIZ1与宫颈癌的侵袭转移及放疗敏感性的关系进行了一系列的研究。首先我们应用免疫组织化学方法，检测159例FIGO分期为Ⅱb及Ⅲa 期的宫颈癌组织中RIZ1的表达水平，研究发现RIZ1在宫颈鳞癌组织中表达下调，与放疗抵抗组相比，放疗敏感组中RIZ1的表达水平升高，而且Logistics回归分析结果提示RIZ1表达水平（OR：0.313；95%CI，0.101-0.974；P=0.045）是宫颈癌患者是否出现放疗抵抗的独立预测因子，即RIZ1高表达的患者出现放疗抵抗的风险降低。同时我们通过体外细胞实验证实RIZ1过表达也抑制宫颈癌SiHa和HeLa细胞的增殖、迁移及侵袭能力，诱导宫颈癌细胞周期发生G2-M期阻滞，诱导宫颈癌细胞发生凋亡。此外，我们通过体外及体内实验评估了RIZ1对宫颈癌放疗敏感性的影响。结果显示，RIZ1的过表达显着降低了HeLa和SiHa细胞暴露于电离辐射后的存活率，RIZ1的过表达增强了电离辐射诱导的细胞凋亡及DNA损伤，动物实验中RIZ1的过表达抑制了裸鼠体内的肿瘤生长并增强了放疗敏感性，射线照射后RIZ1过表达的肿瘤组织中形态学变化明显，凋亡明显，提示增殖能力的PCNA蛋白表达降低。综上所述，我们通过临床患者-细胞系-动物模型三个层次，明确RIZ1表达增强后能否提高宫颈癌的放疗敏感性，改善宫颈癌患者的预后。本项目将会为临床通过基因水平预测和提高宫颈癌放疗敏感性提供新的思路，对于最终指导宫颈癌的临床个体化治疗具有重要的现实意义和良好的应用价值。