miR-26a抑制炎症相关性结直肠癌的发生发展及其机制研究

Patients with inflammatory bowel disease are at increased risk for colitis-associated colorectal cancer (CAC). The key anti-CAC therapeutic regimens include controlling colitis and inhibiting colorectal cancer cell proliferation. Our pilot studies demonstrated that miR-26a impaired macrophage-mediated inflammatory response in vitro and in vivo, and suppressed the proliferation of colon cancer cells. This leads to the hypothesis that miR-26a inhibits colorectal tumorigenesis by inhibiting colitis, and inhibits colorectal tumor development by suppressing colorectal cancer proliferation. To investigate the suppressive role of miR-26a in CAC, we will i) use whole body, enterocyte-specific and myeloid cells-specific transgenic mice as a model to determine the role of miR-26a in AOM/DSS induced CAC initiation and promotion; and ii) use a combination of bioinformatics and Illumina mRNA sequencing to screen the potential targets of miR-26a associated with inflammation and proliferation. Leading targets from the screening will be further confirmed by a psiCHECK2.2 dual-luciferase reporter assay, as well as in vitro and in vivo experiments; iii) establish the correlation between miR-26a distribution, expression and clinical pathological features of colorectal cancer by using clinical samples from patients with colorectal cancer. The proposed studies are innovative because it will investigate a novel miRNA-mediated pathway that is potentially involved in CAC suppression. miR-26a may be a novel target for the diagnosis and treatment of colitis-associated colorectal cancer.

肠炎与肠癌密切相关，控制肠炎及抑制肠癌细胞增殖为治疗炎症相关性结直肠癌的关键。我们前期研究发现miR-26a可以在体内外实验中抑制免疫细胞的炎症反应及抑制肠癌细胞株的增殖，由此我们假设miR-26a一方面通过抑制肠炎而抑制肠癌发生，另一方面通过抑制肿瘤细胞增殖而抑制肠癌发展。为证实此假设，并深入研究机理，本研究将开展以下工作：1）用已获得的全身、髓系细胞及肠上皮细胞特异性过表达的转基因小鼠证实过表达miR-26a可以抑制AOM/DSS诱导的炎症相关性结直肠癌的发生及发展。2）应用软件预测及转录组测序方法筛选可促进炎症反应及细胞增殖的miR-26a靶基因，并在psiCHECK2.2双荧光素酶系统及体内外实验中验证。3）用临床样本分析miR-26a的分布表达情况与结直肠癌的发生、发展的关系。本研究深入解析了miR-26a在炎症相关性结直肠癌中的作用，为临床诊断及治疗此疾病提供了新思路。

肠炎与肠癌密切相关，控制肠炎及抑制肠癌细胞增殖为治疗炎症相关性结直肠癌（CAC）的关键。我们前期研究发现miR-26a可以在体内外实验中抑制免疫细胞的炎症反应及抑制肠癌细胞株的增殖，由此我们假设miR-26a一方面通过抑制肠炎而抑制肠癌发生，另一方面通过抑制肿瘤细胞增殖而抑制肠癌发展。本项目开展了以下工作：1）构建了全身、髓系细胞及肠上皮细胞特异性过表达miR-26a的转基因小鼠，在AOM/DSS诱导的小鼠CAC模型上证实过表达miR-26a可以抑制小鼠肠炎，并且抑制肠癌细胞增殖。2）我们用生物软件预测及 Illumina mRNA 测序分析的方法筛选并鉴定了与炎症反应有关的 miR-26a 靶基因，包括IL6，TLR3, PTEN, PKCδ, GSK3β, TAK1, TAB3，并在 psiCHECK2.2 双荧光素酶系统中验证。3）我们建立了一种体内靶向传递miRNA 的技术(CpG-miRNA 技术)，能够特异性地传递miRNA至TLR9高表达的肿瘤细胞和巨噬细胞。利用该方法合成了CpG- miR26a，并在体内外实验中证实CpG-miR26a可以显著抑制肠癌细胞HCT116增殖。4）我们证实了TGR5的激动剂可以诱导miR26a的表达，因此我们进一步评估了TGR5激动剂DY284对于CAC的治疗效果，结果发现相比对照组，DY284治疗组小鼠肠道不典型增生和腺癌数目减少、肿瘤直径减小、并且肠道炎症评分降低。5）收集了300余例结直肠癌临床样本，初步分析发现miR-26a在肿瘤组织中显著低表达。本研究在细胞、动物及临床水平分析了miR-26a在炎症相关性结直肠癌中的作用，为临床诊断及治疗此疾病提供了新思路。