PGE2及Slit2/Robo1相关信号途径在放疗后死亡鼻咽癌细胞促进残癌细胞加速转移的分子机制研究

Metastasis is the major failure cause of nasopharyngeal carcinoma(NPC). "Tumor bed effect" indicates that radiotherapy (RT) could promote tumor metastasis by alterng tumor microenvironment, but the underlying mechanisms are still unknown. It remains unclear whether lethally irradiated cancer cells could promote survival cancer cells metastasis by secreting some importment signal molecules. Our previous work showed that the supernatant of lethally irradiated nasopharyngeal carcinoma cells could promote metastasis of unirradiated cells, indicating dead caner cells after RT could bona fide promote survival cancer cells metastasis. We also found that PGE2 and Slit2 were elevated in the supernatant of irradiated NPC cells. As we previously found that Slit2/miR-218/Robo1 pathway could promote cancer metastasis, we propose that dead NPC cells after RT could promote survival cells metastasis partially by Slit2/Robo1 and PGE2 related pathway. This project will establshed coculture cell model based on radioresistant and radiosensitive NPC cells and study the mechanisms of PGE2 and Slit2 secretaion by dead NPC cells after RT, as well as the the role of Slit2/Robo1 and PGE2 related pathway in the promotion of cancer metastasis. We believe this research will pave the way for the eluciation of the underlying molecular mechnisms by which dead cancer cells could promote survival cancer cells metastasis after RT.

转移是鼻咽癌治疗失败的主要原因。瘤床效应表明放疗可改变肿瘤微环境促进肿瘤的转移，但机制不明。放疗后庞大的"死亡癌细胞群体"是否可以旁分泌一些信号分子促进残癌转移目前还不清楚。我们前期工作发现，致死剂量照射的鼻咽癌细胞的培养上清可显著的促进照射后存活癌细胞的转移，提示的确存在放疗致死癌细胞促进残癌转移的现象。我们发现照射后死亡鼻咽癌细胞的培养上清中PGE2和 Slit2的表达显著上调，而我们前期工作曾首次发现Slit2-miR-218-Robo1环路可促进肿瘤转移，提示Slit2/Robo1和PGE2相关信号通路可能介导了死亡癌细胞促残癌细胞转移这一过程。本项目在构建致死剂量照射的放疗敏感鼻咽癌细胞和放疗抵抗细胞共培养模型基础上，探讨死亡癌细胞分泌PGE2和Slit2的机制、PEG2和Slit2-Robo1及其下游信号促残癌转移的机制，为全面阐明放疗后死亡癌细胞促残癌转移的分子机制奠定基础。

转移是鼻咽癌治疗失败的主要原因。瘤床效应表明放疗可改变肿瘤微环境促进肿瘤的转移，但机制不明。放疗后庞大的“死亡癌细胞群体”是否可以旁分泌一些信号分促进残癌转移目前还不清楚。我们建立共培养模型，并通过不同剂量照射鼻咽癌细胞确定12Gy为致死剂量，Transwell 和划痕实验检测致死剂量照射的鼻咽癌细胞的培养上清可显著的促进照射后存活癌细胞的转移，提示的确存在放疗致死癌细胞促进残癌转移的现象。并通过RT-PCR，Western blot 和 ELISA发现照射后死亡鼻咽癌细胞的培养上清中 PGE2 和 Slit2 的表达显著上调，为了进一步验证sli2和PGE2参与放疗致死癌细胞促进残癌转移的作用， 构建sli2和PGE2的shRNA慢病毒载体，以及加入sli2以及PGE2的蛋白因子上调表达，进一步通过迁移侵袭实验验证sli2和PGE2促进鼻咽癌的转移。随后探讨sli2和PGE2参与鼻咽癌转移的机制，采用Western Blot 检测PI3K/AKT/β-catenin的表达变化以及加入PI3K以及AKT 抑制剂检测对sli2和PGE2促进鼻咽癌转移的影响，通过C0-IP验证PI3K与slit2和PGE2具有相互作用，结果表明Slit2/Robo1，PGE2/EP2通过PI3K/AKT信号通路促进β-catenin的核转位增强放疗后残存鼻咽癌细胞的转移作用。为全面阐明放疗后死亡癌细胞促残癌转移的分子机制奠定基础，为鼻咽癌的临床治疗提供了干预靶点。