LCN2在鼻咽癌放射抗拒性中的作用与机制

We screened out LCN2 from microarray data of radiation induced resistant-CNE2 cells and radiosensitive-CNE2 cells, on further examination we found that LCN2 involved in radiation resistance and DNA repair of nasopharyngeal carcinoma cells, and it could affect the phosphorylation of AKT. Previous studies have indicated that AKT plays an important role in tumor radiation resistance through the interaction with DNAPKcs. Thus, we proposed that, LCN2 can induce the radiation resistance of nasopharyngeal carcinoma by AKT/DNAPKcs. According to this hypothesis, through gene chip analysis, as well as a variety of in vitro and in vivo experiments, the project intends to identify the contribution of Lcn2 on nasopharyngeal carcinoma radioresistance, and confirmed that the underlying mechanism is associated with the effect of Akt phosphorylation on DNAPKcs kinase activity and its binding ability with Ku dimer, which will provide a new explication for NPC radioresistance-related study, and a new target for the diagnosis and treatment of NPC.

我们前期从鼻咽癌射线诱导抗拒的CNE2细胞及野生CNE2基因组表达谱芯片筛选出放射抗拒性相关基因LCN2，随后验证结果提示LCN2与鼻咽癌细胞放射抗拒性及DNA修复有关，并可影响AKT磷酸化。前人的研究表明AKT可作用于DNAPKcs，从而在肿瘤放射抗拒性方面起重要作用。基于此，本项目提出以下假设：LCN2可以通过AKT/DNAPKcs介导鼻咽癌的放射抗拒性。针对这一假设，本项目拟通过基因芯片分析、及各种体内外实验，明确LCN2诱导鼻咽癌放射抗拒性的作用，并证实LCN2介导鼻咽癌放射抗拒性的作用机制与通过AKT磷酸化影响DNAPKcs激酶活性及其与Ku二聚体的结合有关。这将为鼻咽癌放射抗拒性的预测与干预，从而提高鼻咽癌患者预后提供一个潜在的靶点。

放疗抵抗引起的复发是目前鼻咽癌治疗失败的主要原因之一，我们筛选GEO数据库中的芯片数据发现LCN2高表达于放射抗拒的鼻咽癌细胞，然而其在鼻咽癌放射抗拒中的作用与机制尚未明确。本项目通过实时定量PCR（RT-qPCR）和Western blotting检测了LCN2在NPC细胞系中的表达，通过免疫组织化检测了LCN2组织标本表达，并对生存结局进行了分析。通过克隆形成及免疫荧光实验研究了敲低和过表达LCN2对鼻咽癌细胞放射敏感性的影响。 通过Western blotting和组织芯片检查了LCN2和缺氧诱导因子1-alpha（HIF-1A）之间的可能相关性。结果表明，LCN2在放射抗拒的鼻咽癌细胞系中高表达。免疫组化分析显示，LCN2表达在耐放射的鼻咽癌组织中显着上调，并且与NPC复发相关。敲除LCN2会损害鼻咽癌细胞修复DNA损伤的能力，从而增强了NPC细胞的放射敏感性，而LCN2过表达会增加鼻咽癌细胞的放射抗性。此外，LCN2和HIF-1A的表达呈明显的正相关。综上，本项目研究提示，LCN2与鼻咽癌的放射抵抗和复发有关，并可能通过与HIF-1A相互作用促进放射抵抗表型的发展，有望成为鼻咽癌放射抗拒性的预测指标和治疗靶点。