抗氧化酶PRDX1调控结直肠癌转移和耐药性的机制及生物学意义

Our previous study found peroxiredoxins 1(PRDX1) expression showed a gradual increasing trend in the early stages of colorectal cancer. On the contrary, PRDX1 expression level decreasing gradually in the metastatic process.According to our preliminary experimental results and the literature review, we speculate that PRDX1 is an important regulatory aspects in the colorectal cancer metastasis and drug resistance. To test this hypothesis, this project is to carry out functional and mechanism studies in order to gain further insight into PRDX1 from clinical, cellular and animal models levels. First, we aim to clarify the dynamic adjustment of PRDX1 at each stage of the evolution of colorectal cancer using large samples testing. Combined with clinical data, we will reveal the relationship between PRDX1 and colorectal cancer metastasis and drug resistance. Second, we plan to build orthotopic xenograft colorectal cancer models with PRDX1 gene silencing and overexpression, and we will also develop DMH induced colorectal cancer model on the basis of transgenic mice. The effect of PRDX1 on colorectal cancer metastasis and drug resistance will be contrastively analyzed in vitro and in vivo. Then, we intend to screen PRDX1 interacting protein by gene chip, proteomics and online bioinformatics analysis, and the key molecules and signaling networks of PRDX1 play the regulation role will be clear. The succseeful completion of this project will provide a new target and theoretical basis for early warning and enhancing effect of chemotherapy in treating colorectal cancer.

我们前期研究发现在结直肠癌演进的前期阶段PRDX1表达呈逐级递增趋势，但后期转移过程中PRDX1表达水平发生逆转。前期实验结果结合文献回顾，我们推测PRDX1很可能是结直肠癌晚期发生转移及化疗耐药的重要调控环节。为验证这一假设，本项目拟从临床、细胞和动物模型三个层面深入剖析PRDX1的功能及作用机制：大样本鉴定PRDX1在结直肠癌演进各个阶段的动态调节，结合临床资料揭示PRDX1与结直肠转移及耐药的相关性；应用基因过表达及干扰技术，结合可视化结直肠癌原位移植瘤模型及PRDX1转基因小鼠DMH诱发结直肠癌模型，体内外系统双向验证PRDX1过表达在结直肠癌转移及耐药中的功能；利用基因芯片、蛋白质组学技术并生物信息学分析，筛选验证PRDX1相关差异基因/蛋白，揭示PRDX1在结直肠癌转移及耐药控中的关键分子和信号网络，为结直肠癌转移的早期预警、改善临床化疗效果提供新的有效靶点和理论依据。

随着现代分子生物学的发展，人类迈入了功能基因组的研究时代。本研究从临床、细胞和动物模型三个层面深入剖析PRDX1的功能及作用机制。收集人结直肠癌的石蜡标本及冻存标本，分别用免疫组织化学及qPCR、Western Blot技术检测PRDX1表达情况，结果显示，在结直肠癌的发生与演进过程中，PRDX1的表达与其呈正相关；而在结直肠癌发展后期，尤其是转移阶段，PRDX1的表达趋势发生改变甚至逆转；构建稳定过表达及干扰PRDX1的人结直肠癌细胞株SW620及SW480，细胞体、内外功能研究结果显示， PRDX1能够抑制结直肠癌的增殖和运动迁移能力，其抗氧化功能可能发挥抑制作用，同时PRDX能够促进结直肠癌细胞对化疗药物耐药性；通过原核注射方法构建结直肠特异性高表达PRDX1的转基因小鼠，结合腹腔注射二甲基肼（DMH）诱导结直肠癌发生，结果显示PRDX1在活体内整体水平能够抑制肿瘤的发生和发展；基因芯片分析获得与PRDX1密切相关的上调基因452个，下调基因644个，聚类筛选出多个肿瘤相关的信号转导通路；应用CO-IP及Western blot鉴定部分差异基因，结果显示PRDX1能够通过与MAPK9结合激活MAPK信号通路进而在结肠癌演进过程中发挥重要作用。本研究在活体内接近“真实”地再现PRDX1过表达在结直肠癌发生发展中的作用并初步探讨其机制，完善了PRDX1基因的基础研究背景资料，为结直肠癌的临床诊治提供新的靶点和直接有力的理论依据，并为活体内PRDX1基因的分子调控机制研究提供理想的动物模型。