PTTG 对miRNAs的调控在胃癌中的作用及其机制的研究

Overexpression of PTTG, pituitary tumor transforming gene, reportedly occurs in a wide variety of tumors, and is associated with tumor stage, lymph node metastasis and prognosis. PTTG overexpression might be a powerful biomarker of tumor diagnosis and a potential target for gene therapy. miRNAs are a class of small noncoding RNA, which is aberrant expression in a variety of tumors, and participated in tumorigenesis and tumor progression as cancer suppressor genes or oncogenes. However, the functions and mechanism of PTTG regulating miRNAs expression in gastric cancer remain unclear. Our previous studies showed that PTTG overexpression in gastric tissues and cells, and this phenomenon can promote the proliferation, invasion and migration of gastric cancer cells. Further studies have shown that PTTG could participate in the transcriptional regulation of miR200c and miR21. However, the specific regulation mechanisms of PTTG on miR200c and miR21 are unclear. On the basis of previous work, this project will perform a in-depth study on specific mechanisms of PTTG regulating miRNAs expression and the downstream signaling pathways of PTTG-miRNAs, and study the effection of PTTG-miRNAs on the malignant behavior of gastric cancer cells. The project is to elucidate the mechanism of PTTG on occurrence and development of gastric cancer, and will provide a theoretical basis for a new target of gastric cancer gene therapy.

PTTG即垂体瘤转化基因，在许多肿瘤中都呈高表达，且与肿瘤临床分期、转移、预后等密切相关，可能成为肿瘤诊断的分子标志及靶向治疗的靶点。miRNAs是一类小分子非编码RNA，在多种肿瘤中异常表达，并参与了多种肿瘤的发生发展过程。然而关于PTTG对miRNAs的调控机制及其在胃癌中的作用目前鲜有研究。本课题的前期研究显示，PTTG在胃癌组织及细胞中呈现高表达并参与调控胃癌细胞的恶性生物学行为，进一步通过大量的筛选证实在胃癌细胞中PTTG可能参与了对miR21及miR200c的转录调控，然而关于其具体的机制尚不清楚，亟待解决。本课题拟在前期工作的基础上更加深入的探讨PTTG对miRNAs表达调控的具体机制及其对下游信号通路的影响，并系统的研究其对胃癌细胞的恶性生物学行为的影响。该项目对阐明PTTG在胃癌发生发展的作用机制，并为其作为胃癌靶向治疗的新靶点提供新的理论依据。

多项研究显示PTTG为致癌基因，在多种肿瘤中呈高表达，且与肿瘤分期、淋巴结转移以及临床预后密切相关，甚至可以作为独立预后因素。PTTG可以作为转录因子调控下游一系列基因的表达从而导致肿瘤的发生、发展。miRNAs是一类小分子非编码RNA，在多种肿瘤中异常表达。然而在胃癌中PTTG对miRNAs的作用机制研究甚少。在本课题中，我们首先通过免疫组化分析显示胃癌组织中PTTG表达量明显高于癌旁组织，且胃癌组织高表达的PTTG都倾向于胞核分布。Western blot结果也显示PTTG在胃癌细胞系中的表达明显高于HEK-293。在多种胃癌细胞系中过表达或抑制PTTG，通过CCK-8、EdU以及Transwell等实验证实PTTG可以促进细胞的增殖、侵袭及迁移等恶性生物学行为。在BGC-823和MGC-803中过表达及抑制PTTG后进行RT-PCR，结果发现miR200c与PTTG表达呈负相关，miR21与PTTG的表达呈正相关，CHIP实验证实PTTG能与miR200c和miR21启动子结合。在MKN-45和BGC-823中过表达PTTG，再分别共转染miR200c mimics或miR21 inhibitor，证实二者能逆转PTTG对细胞恶性生物学行为的促进作用。最后我们在BGC-823中过表达PTTG后再过表达miR200c或抑制miR21，通过Western blot实验发现pRb、c-Myc、Vimentin可能是PTTG-miR200c/miR21调控的下游分子。本项目胃癌组织及细胞为研究对象，深入探讨PTTG在胃癌中的表达情况、胃癌中PTTG对miRNAs的调控机制以及二者对胃癌生物学行为的影响，为阐明PTTG在胃癌中的作用机制及其作为临床预后因子、肿瘤治疗靶点提供新的理论依据。