DMBT1基因在胆囊癌发生发展中的作用及分子机制研究

DMBT1 has been considered as a candidate tumor suppressor gene, whose expression is down-regulated or depleted in multiple types of malignant tumor, and yet DMBT1 is usually over-expressed when chronic inflammation occurs. Research overseas concentrates on the effect of EMBT1 during tumor development, while rarely did scientists mention of its role and regulation mechanism in early stage of tumor. In previous studies, we find that expression of DMBT1 dramatically down-regulated in gallbladder carcinoma, while the converse is true in normal adjacent tissues, usually with chronic inflammation. In vitro, high-expression of DMBT1 remarkably inhibits proliferation and migration in gallbladder carcinoma cells, and induces apoptosis. Therefore, we presumed that the expression of DMBT1 has been induced in the early stage of gallbladder carcinoma for immunological protection. Down-regulation or depletion of DMBT1, occurring as tumor advances, contributes to tumor growth. On the basis of prior experiments, this study is intended to illustrate the role and molecular mechanism of DMBT1 in process of tumorigenesis and development from in vitro to in vivo by establishing DMBT1-gene knockout mouse model and human gallbladder carcinoma orthotopic grafting model, taking advantage of RNA interference technology and microarray technology. Moreover, we intend to primarily indicate the regulation mechanisms involved in the process of tumorigenesis and development. The scientific meaning of this study lays emphases on enriching the understanding of tumorigenesis and development of gallbladder carcinoma, providing new target with high-effecacy for clinical treatment.

候选抑癌基因DMBT1在多种恶性肿瘤中表达下调或缺失，但在多数慢性炎症中却呈高表达。国外研究主要集中于DMBT1对肿瘤进展的影响,而对其在肿瘤发生早期的作用及调控机制却鲜有报道。我们前期研究发现：胆囊癌组织中DMBT1表达明显下调，而癌旁组织(多合并慢性炎症)中呈高表达；体外实验证明DMBT1高表达能显著抑制胆囊癌细胞增殖、迁移，并促进其凋亡。故推测DMBT1在胆囊癌发生早期被诱导表达，参与机体免疫防护；随着肿瘤进展其表达下调或缺失，转而有利于肿瘤生长。本研究将在前期工作基础上，利用DMBT1基因敲除鼠模型和人胆囊癌裸鼠原位移植瘤模型，通过RNA干扰、基因芯片等技术，从体外到体内，旨在探讨DMBT1在胆囊癌发生、发展过程中所起的作用及其分子机制；并对DMBT1在两个不同表达阶段的调控机制进行初步研究。其科学意义在于：丰富胆囊癌发生发展的理论体系，为临床治疗提供新的高效靶点。

脑恶性肿瘤缺失基因1（DMBT1）因其在脑恶性肿瘤中广泛缺失而得名，被发现在多种不同类型的恶性肿瘤进展过程中存在表达下调或缺失，被认为是一个潜在的抑癌基因，其作用机制可能与炎症向癌症转化及肿瘤的侵袭转移密切相关。但其在胆囊癌中研究极少，通过前期实验我们发现DMBT1在胆囊癌中低表达，可能同样是胆囊癌的一个潜在抑癌基因。因此希望通过深入的研究了解DMBT1在胆囊癌侵袭转移中的作用及其分子机制。. 我们在胆囊癌组织中发现DMBT1低表达且存在拷贝数缺失。体外和体内功能实验均显示DMBT1的低表达能促进胆囊癌细胞侵袭转移。运用转录组测序和基因探针富集分析进行深入的DMBT1分子机制探索，通过核糖核酸拉下实验和RNA结合蛋白免疫共沉淀实验验证了长链非编码RNA结直肠肿瘤差别表达基因（CRNDE）能够特异性的结合DMBT1和细胞凋亡抑制蛋白1（C-IAP1），CRNDE在DMBT1与C-IAP1两者之间起到了脚手架的作用。CRNDE、C-IAP1均在胆囊癌中高表达，是DMBT1的直接靶基因。DMBT1表达的下调可使得CRNDE及C-IAP1的表达上调。同时DMBT1表达下调可显著影响PI3K/AKT信号通路中基因的表达，最终激活PI3K/AKT信号通路促进胆囊癌的致癌性。. 通过临床、基因、生化、功能方面的研究，发现DMBT1是胆囊癌的一个抑癌基因，能够通过结合CRNDE和C-IAP1，影响PI3K/AKT信号通路，发挥其抑癌作用。而针对这条通路的靶向治疗，也许能为胆囊癌的治疗打开一个新的方向，从而改善胆囊癌患者的预后。