氨基酸转运蛋白LAT1调控mTOR信号通路对鼻咽癌放射敏感性的影响及其机制研究

Radio-resistance is a critical reason for the failure of nasopharyngeal carcinoma (NPC) treatment. Previous studies from our and other groups indicated that the activation of mTOR signaling pathway correlated closely with NPC radio-resistance. However, the upstream factors which induce activation of mTOR signaling pathway in NPC remain unclear. Our unpublished data revealed that there were significantly higher expression levels of LAT1, which positively correlate with mTOR signaling activities in NPC cells. We also found that LAT1 knockdown leaded to decreased mTORC1 activity and elevated radio-sensitivity in NPC cells. Thus, we proposed that LAT1 regulates the radio-sensitivity of NPC through mTOR signaling pathway. To test this hypothesis, we will use CRISPR/Cas9 to genetically knockout LAT1 based on NPC cell lines with different radio-sensitivities. Using those LAT1 knockout cells, accomplished with nude-mice NPC xenograft models as well as clinical NPC tissue microarrays, we try to elucidate the role of LAT1/mTOR in regulating NPC radio-sensitivity. Furthermore, we will perform an initiative pharmacologic evaluation of LAT1 inhibitor as a radiosensitizer in NPC. We believe that this project will provide novel insights to NPC radio-resistance, which will facilitate the NPC radiosensitization research and therapy.

放疗抵抗是鼻咽癌治疗失败的重要原因，其他学者和我们的前期研究表明mTOR信号通路的激活与鼻咽癌放疗抵抗密切相关，但其上游诱发因素尚不甚明确。我们前期研究发现放疗抵抗的鼻咽癌组织和细胞系中氨基酸转运蛋白LAT1表达水平明显增高，并与mTOR信号通路活性呈正相关；沉默LAT1表达，mTORC1活性降低，伴鼻咽癌细胞放射敏感性增加。在此基础上，我们提出高表达LAT1通过mTOR信号通路诱导鼻咽癌放疗抵抗的假说。本项目在放射敏感性不同的鼻咽癌细胞系基础上，拟采用CRISPR/Cas9技术建立LAT1基因敲除的鼻咽癌细胞株，细胞学实验结合鼻咽癌裸鼠移植瘤模型和人体鼻咽癌组织芯片阐明LAT1/mTOR信号通路与鼻咽癌放疗抵抗的关系，并进行初步的LAT1抑制剂放射增敏药效评价，力求为鼻咽癌的放射增敏治疗提供新的理论依据和分子靶点。

本实验研究方向在实验过程中有高度类似的科研文献发表，因此调整方向为MEX3在鼻咽癌发病中的机制研究..经过生物信息学分析和组织芯片，发现：1) MEX3基因在鼻咽癌的癌组织中的表达明显高于癌旁组织；并且与肿瘤的分级分期显著相关；2)通过比较鼻咽癌细胞系，发现EBV低负荷的肿瘤细胞株CNE-2Z中MEX3表达较低；3)构建敲除MEX3的慢病毒shRNA，发现shMEX3能明显降低鼻咽癌细胞的存活，增加鼻咽癌细胞的凋亡，并具有更强的克隆形成能力，更慢的划痕愈合，并具有明显增强的侵袭迁移能力；4）MEX3在鼻咽癌中的作用与SCIN和NK-kb信号通路有关。