PSME3基因在口腔黏膜癌变发生发展中的作用及其机制研究

Oral carcinogenesis is characterized by its multi-stage process of development, easiness to be observed and convenience of obtaining the tissue sample for research. That provides us with a good research model to clarify the biological effect and clinical significance of new biomarkers during the course of carcinogenesis which is a hotspot of study at present. Previous research have revealed that protein REGγ encoded by the PSME3 gene had a significantly high expression in OSCC tissues. Prospective regular follow up study of two independent large cohort of OSCC patient have also confirmed that REGγ were correlates with malignant transformation as well as recurrence and disease-free survival of patients with OSCC. There is significantly negative correlation between the expression of REGγ and survival probability of the patients. However, its specific biological effect on OSCC and molecular mechanism, such as related signaling pathways and underlying key factors involved in the biological process of oral carcinogenesis, is unclear. So gene silence and overexpression of PSME3 will be used in order to observe its effect on biological behaviors related to regulating proliferation, apoptosis, migration, and invasion in vitro. Moreover, a human OSCC xenograft-nude mouse mode will be established to detect tumor cell proliferation, intratumoral angiogenesis and tumor apoptosis, in order to explore whether PSME3 have biological effects on OSCC tumor growth in vivo. Furthermore, bioinformatics approach, gene expression profiling and Co-Immunoprecipitation would be employed to identify its related signaling pathways and key molecules. In addition, we will carry out functional verification to explore the molecular mechanism of its anti-tumor activity, along with the feasibility of PSME3 and key molecules as the target to provide a new strategy for tumor prevention. All those findings will be further confirmed by the carcinogenic 4-nitroquinoline-1-oxide (4NQO) model of genomic DNA of PSME3 (-/-) knock-out mice and PSME3 overexpression transgenic mice (Tg-PSME3 mice). That might be helpful to enucleate the role and the molecular mechanism of PSME3 gene in carcinogenesis of oral mucosa or other tissue so as to provide a new strategy for tumor prevention and treatment.

口腔黏膜癌变阶段明显、易观察、可重复取材，利用该模型研究肿瘤发生过程中新分子的作用及临床意义是目前热点。本课题组前期研究发现，PSME3基因编码的蛋白酶体激活因子REGγ蛋白在口腔鳞癌（OSCC）组织明显增高；两个独立的大队列随访都证实其表达与患者生存概率显著负相关。但该基因在其中发挥的生物学效应、具体机制、参与环节及涉及关键分子均待阐明。故本项目拟①采用PSME3基因沉默与过表达方法，分别从细胞和动物水平考察该基因对OSCC细胞在体外体内生长增殖、侵袭转移的影响；②应用基因芯片、生物信息学技术和蛋白质免疫共沉淀等方法，寻找该基因参与口腔黏膜癌变的信号通路和关键节点蛋白；③探索利用REGγ蛋白及信息通路关键节点蛋白靶向阻断口腔黏膜癌变的可行性；④利用敲除和过表达该基因小鼠的化学诱癌模型从遗传学上印证上述发现。结果有助于阐明PSME3在人类肿瘤中的作用机制，为靶向防治口腔黏膜癌变提供依据。

PSME3基因编码的蛋白是口腔鳞癌（OSCC）预后的警示因子，但是该基因在OSCC发生发展及转移中的生物学效应、具体机制仍不清楚。本课题研究发现：①稳定靶向沉默PSME3基因后，OSCC细胞体外生长增殖、侵袭转移等恶性生物学行为被部分逆转，而过表达该基因则可促进OSCC细胞的恶性生物学行为。② 使用PSME3稳定沉默的OSCC细胞构建移植瘤模型，肿瘤的生长速度减缓，转移瘤形成的比率降低，瘤内微血管的密度减少。反之亦然。③PSME3基因通过VEGF等12条相互关联的信号通路影响OSCC细胞体外生长和侵袭能力，还可通过IL-6/CCL2信号轴参与口腔鳞癌血管生成。④PSME3编码蛋白与C1QBP蛋白可直接相互作用，二者在口腔黏膜癌变各阶段表达趋势一致，均可作为口腔黏膜癌变预后的警示因子。⑤化学致癌剂诱导的Psme3 +/+（过表达）小鼠癌变进程显著缩短，从遗传学上印证了PSME3 基因在口腔黏膜癌变过程中的重要作用。综上，本研究得到结论：PSME3基因可能通过与C1QBP互作，调控VEGF、IL-6/CCL2等信号通路，进而参与OSCC发生发展及转移的恶性进程。