Annexin A2过表达在鼻咽癌放射抗拒的作用及其分子机制研究

Annexin A2 is a candidate oncogene which is closely related to proliferation, differentiation, and apoptosis of tumor cells. In our previous study, Annexin A2 was found significantly up-regulated in the radioresistant nasopharyngeal carcinoma (NPC) tissues and cells， and it may be involved in the radioresistance of NPC. In order to reveal the role and molecular mechanism of the Annexin A2 overexpression on radioresistance of NPC，the radiosensitive NPC cell line CNE-2 with Annexin A2 expression stably up-regulated and the radioresistant NPC cell line CNE-2(R743) with Annexin A2 expression stably down-regulated will be established,in our project, meanwhile the role of Annexin A2 in NPC radioresistance will be performed by establishing the model of nude mouse xenograft in vivo and the model of cell in vitro,and then，the effect of Annexin A2 on the biological activities of p53, and its relationship with NPC radioresistance will be analysed using the established model of NPC cell in vitro. For the purpose of study the mechanism of Annexin A2 on NPC radioresistance，the Annexin A2 interacting protein will be identified, and the regulatory network of Annexin A2 in NPC radioresistance will be built by using the co-immunoprecipitation coupled with two-dimensional gel electrophoresis(2DE) and mass spectrometry analysis, thus the key target protein and signal path involved in Annexin A2 regulation will be identified ,and then the role and molecular mechanism of NPC radioresistance will be clarified. The result may be provide scientific evidence for NPC radiosensitivity prediction, targeted therapy, and personalized radiotherapy in NPC patient.

Annexin A2是一个候选癌基因，与肿瘤细胞增殖、分化和凋亡密切相关。我们前期研究发现：在放射抗拒的鼻咽癌组织和细胞中，Annexin A2表达均上调，可能与鼻咽癌放射抗拒有关。为揭示Annexin A2过表达在鼻咽癌放射抗拒的作用及分子机制，本项目拟建立Annexin A2表达稳定上调的CNE-2细胞株和稳定下调的CNE-2(R743)细胞株，通过体内和体外实验研究Annexin A2在鼻咽癌放射抗拒的作用。以建立的细胞株为样本，分析Annexin A2对p53生物学活性的影响及与鼻咽癌放射抗拒的关系。采用免疫共沉淀、双向电泳和质谱，鉴定Annexin A2的相互作用蛋白，构建Annexin A2在鼻咽癌放射抗拒的调控网络，明确Annexin A2调控的关键靶蛋白及信号通路，阐明Annexin A2在鼻咽癌放射抗拒的作用机制，为鼻咽癌放射敏感性预测、靶向治疗和个体化放疗提供科学依据。

采用siRNA干扰技术成功建立Annexin A2 表达下调的CNE-2(R743)细胞模型，通过集落形成实验、流式细胞术和TUNEL分别分析了Annexin A2 表达变化对鼻咽癌细胞放射敏感性、细胞周期和细胞凋亡的影响；应用免疫组化技术检测了131例鼻咽癌组织标本Annexin A2的表达，通过临床及随访资料分析了Annexin A2的表达与放射治疗后局部复发和远处转移的关系。细胞实验结果表明，下调Annexin A2基因表达可明显增强放射抗拒细胞CNE-2(R743)的放射敏感性，提高该细胞G2+M期的比例和细胞凋亡率。临床实验结果表明，Annexin A2表达与鼻咽癌的分期无相关性；Annexin A2表达与鼻咽癌放射治疗后的局部复发和远处转移具有正相关趋势，但尚无统计学意义，可能是由于病例数较少和随访时间短，因此为得到更加客观可靠的结果，需在后续的研究中增加病例数和延长随访时间，进一步明确Annexin A2的表达与鼻咽癌放射敏感性的关系。