miR-205下调ASPP2表达促进食管癌转移和EMT的作用及机制研究

ASPP2, a member of the ankyrin-repeat-, SH3-domain- and proline- rich-region-containing protein (ASPP) family is frequently decreased in various cancers. Clinical data reveals that reduced expression of ASPP2 is related with poor clinical outcomes in diffuse large B-cell lymphoma and tumor metastasis and poor recurrence-free survival in breast cancer patients. The effects and underlying mechanism of ASPP2 on tumor metastasis are remains unknown. In current study; we analyzed ASPP2 expression in 150 esophageal carcinomas (ESC) samples and revealed that ASPP2 expression was down-regulated in primary tumor and lymph metastasis compared with the normal esophageal tissue. Further, decreased ASPP2 expression correlated with poor prognosis in ESC patients. MicroRNAs (miRNAs) inhibit translation or induce mRNA degradation in general by binding to the 3' untranslational region (3'UTR) of target mRNAs. Since ASPP2 was regulated at protein level in ESC, we analyzed potential miRNA target ASPP2 through targetscan，Microlnspector and targetscan. We found miR-205 was matched with ASPP2 3'UTR and the expressions of miR-205 were sharply increased in ESC cell lines compared with the normal esophageal cell line. Further, we try to investigate the underlying molecular mechanism of ASPP2 down-regulated by miR-205. The expression of ASPP2 and miR-205 were deceased by RNA interference or increased by transected with over expression plasmid, then tumorigenicity and metastasis potentials of ESC cells were studied in vitro and in nude mice. Apoptosis were examined by TUNEL assay and Annexin V-FITC kit. Wound-healing and Matrigel invasion assays were used to determine migration and invasion potentials of ESC cells. RT-PCR,Western blot and Immunofluorscence were used to analysis EMT. We also try to investigated the molecular mechanism of ASPP2 regulated EMT and tumor metastasis by interaction with YAP or p53. Our findings will present functional and mechanistic insight into the critical role of ASPP2 in the progression and metastasis of cancer.

ASPP2作为潜在的肿瘤抑制基因，在多种肿瘤中存在表达失调，乳腺癌和血液系统肿瘤的临床分析提示ASPP2与肿瘤转移和复发相关，但ASPP2的表达调控机制及其抑制肿瘤转移的作用和机制有待进一步研究。我们首次检测ASPP2在食管癌中的表达水平，发现ASPP2在食管癌细胞系中，原发瘤及淋巴转移灶中表达水平均显著下降；提示它在食管癌的转移中发挥了重要的作用。这里我们想通过临床病例分析、细胞生物学实验、分子机制探讨及体内裸鼠实验四个方面，阐明miR-205抑制ASPP2表达的分子机制，揭示ASPP2的全新调控方式；以食管癌为模型从EMT的角度，阐明ASPP2抑制肿瘤转移的作用和机制；同时分析ASPP2 和 miR-205与食管癌转移和预后的关系，评价血清miR-205作为食管癌预后指标的价值，为临床诊断提供帮助。

ASPP2作为潜在的肿瘤抑制基因，在多种肿瘤中存在表达失调，乳腺癌和血液系统肿瘤的临床分析提示ASPP2与肿瘤转移和复发相关，但ASPP2的表达调控机制及其抑制肿瘤转移的作用和机制有待进一步研究。我们利用免疫组化技术检测了ASPP2在食管癌组织中的表达情况，分析了ASPP2与各项临床指标的关系，发现ASPP2表达下调与肿瘤转移和预后不良密切相关；通过生物信息学预测和分子生物学实验证实，阐明了miR-205抑制ASPP2转录后翻译下调ASPP2表达的分子机制；通过体内外功能试验揭示了ASPP2表达缺失促进食管癌转移及EMT的作用。进一步分析了ASPP2调控EMT的分子机制，利用免疫组化技术检测了YAP在食管癌组织中的表达情况，分析了ASPP2与YAP表达的关系；阐明了ASPP2调控YAP信号通路的分子机制；揭示了ASPP2表达缺失促进食管癌转移及EMT的作用