MicroRNA-607靶向RKIP调控鼻咽癌放疗敏感性的机制研究

Our previous studies showed that Raf kinase inhibitor protein (RKIP) downregulation/deletion closely associated with the invasion and metastasis of nasopharyngeal carcinoma (NPC), and played an important role in the radioresistance of NPC cells. But the microRNAs about targeted regulation of RKIP are unknown, and the mechanism in radiotherapy sensitivity is unclear. In the preliminary experiments, we predicted RKIP may be the direct target gene of miR-607 with bioinformatic methods. The project will use the miR-607 overexpression and knockdown lentivector stable transfection, quantitative real-time PCR, western-blot, flow cytometry, luciferase reporter gene system and H2AX Foci methods to demonstrate targeted regulation of RKIP and role of radiotherapy resistance by miR-607 and to elucidate the molecular mechanisms. The project will not only help to clarify the radioresistant mechanisms in NPC, and provide a theoretical basis on miR-607/RKIP/Keap1-Nrf2 signaling pathway as a new strategy on individual special radiotherapy of NPC cells .

我们前期研究表明Raf激酶抑制蛋白(Raf kinase inhibitor protein，RKIP)在鼻咽癌侵袭转移和放疗敏感性中挥重要作用，但是靶向调控RKIP的microRNA及其对鼻咽癌放疗敏感性作用尚不清楚。我们通过生物信息学方法预测RKIP可能是miR-607的直接靶基因。本项目以人低分化鼻咽癌细胞CNE-2和放射抗拒细胞亚系CNE-2R为研究对象，采用过表达miR-607和miR-607沉默慢病毒载体稳定转染、实时定量PCR、western-blot、流式细胞术、荧光素酶报告基因系统和H2AX Foci免疫荧光等方法，旨在获得miR-607通过靶向调控RKIP在鼻咽癌放疗抵抗中的可靠证据并阐明其分子机制。项目研究结果将有助于发展miR-607/RKIP/Keap1-Nrf2信号通路成为新的肿瘤细胞特异性放疗增敏策略之一。

我们前期研究表明Raf激酶抑制蛋白(Raf kinase inhibitor protein，RKIP)在鼻咽癌侵袭转移和放疗敏感性中挥重要作用，但是靶向调控RKIP的microRNA及其对鼻咽癌放疗敏感性作用尚不清楚。本项目以人低分化鼻咽癌细胞CNE-2和放射抗拒细胞亚系CNE-2R为研究对象，采用过表达miR-607和miR-607沉默慢病毒载体稳定转染、实时定量PCR、western-blot、流式细胞术、荧光素酶报告基因系统和H2AX Foci免疫荧光等方法，旨在获得miR-607通过靶向调控RKIP在鼻咽癌放疗抵抗中的可靠证据并阐明其分子机制。结果表明miR-607在鼻咽癌放疗抵抗细胞系表达上调，并与RKIP表达负性相关，并与放疗敏感性相关。miR-607直接靶向RKIP mRNA3’UTR结合模序。过表达miR-607促进CNE-2细胞增殖、抑制凋亡和G2期阻滞；下调miR-607表达，抑制CNE-2R细胞增殖、促进凋亡和G2期阻滞抑制。miR-607/RKIP通过调控Keap1-Nrf2信号通路来发挥放疗敏感性作用。项目研究结果将有助于发展miR-607/RKIP /Keap1-Nrf2信号通路成为新的肿瘤细胞特异性放疗增敏策略之一。