循环血液中长链非编码RNA在前列腺癌复发转移中的预测作用及其机制研究

Emerging evidence indicates that a large number of specific long non-coding RNA (lncRNA) involve in both tumourigenesis and tumour progression of prostate cancer (PCa). Specific lncRNA expression profiles have been found and analyzed in peripheral blood of prostate cancer patients. However, the model of circulating lncRNA and the role of circulating lncRNA in the metastasis of PCa remains elusive. Our previous studies found that the expression levels of MALAT-1 are significantly elevated in supernatants of prostate cancer cell lines compared to normal prostate cell line and the detectable rate of MALAT-1 in the microvesicles of peripheral blood of metastatic PCa patients is much higher than that of localized PCa patients. Further functional experiments in vitro demonstrated that co-culture of PCa cell and microvesicles which contain over-expressed MALAT-1 can enhance the capability of the invasion of PC3 cell. Based on the results of previous experiments, our study takes circulating lncRNA as the starting point and will be divided into three parts: (1) to reveal the dysregulated circulating lncRNA and establish the circulating lncRNA panel to predict the recurrence and metastasis of PCa; (2) The biological function and molecular mechanism of lncRNA in supernatants of prostate cancer cell lines to promote the metastasis of prostate cancer; (3) the roles of circulating lncRNA in promoting the metastasis of prostate cancer in vivo. With experiments on clinical level, cellular level and mouse model of prostate cancer, we intend to elucidate the role of lncRNA in circulation in regulating the metastasis of prostate cancer and explore its clinical application, which might provide new gene targets and intervention strategies for the diagnosis and treatment of metastatic prostate cancer.

大量特异的长链非编码RNA（lncRNA）参与了前列腺癌（PCa）的发生发展，PCa患者循环血中含有特定的lncRNA表达谱。但循环lncRNA在PCa转移中的机制不明。我们前期研究发现，PCa细胞培养液微囊泡中MALAT-1表达量显著高于正常前列腺细胞，且转移性PCa循环血微囊泡中MALAT-1的阳性率明显高于局限性PCa。功能实验证实，PC3细胞与高表达MALAT-1微囊泡共培养后，其侵袭能力明显增加。本研究以循环lncRNA为切入点开展如下研究：⑴ 找到差异性表达的lncRNA，建立预测PCa转移的lncRNA模型；⑵ 体外实验研究细胞培养液微囊泡中lncRNA促进PCa转移的生物学功能和分子机制；（3）体内实验研究循环lncRNA在PCa转移中的作用并证明其分子机制。在临床、体外和体内三个层面阐明循环lncRNA在PCa复发和转移中其分子机制，为转移性PCa的诊断和治疗提供新策略。

前列腺癌（prostate cancer, PCa）严重威胁着老年男性的健康。PCa患者循环血中含有特定的lncRNA表达谱，大量特异的长链非编码RNA（lncRNA）参与了PCa的发生发展。但循环lncRNA在PCa转移中的机制不明。我们前期研究发现，PCa细胞培养液细胞外囊泡中MALAT-1表达量显著高于正常前列腺细胞，且转移性PCa循环血细胞外囊泡中MALAT-1的阳性率明显高于局限性PCa。功能实验证实，PC3细胞与高表达MALAT-1细胞外囊泡共培养后，其侵袭能力明显增加。我们以前期研究为基础，以循环lncRNA为切入点，开展了如下研究：（1）首先找到差异性表达的lncRNA，建立预测PCa转移的lncRNA模型；（2）体外实验研究细胞培养液细胞外囊泡中lncRNA促进PCa转移的生物学功能和分子机制；（3）体内实验研究循环lncRNA在PCa转移中的作用并证明其分子机制。.通过本项目的研究，我们首先建立了PCa相关外泌体RNA表达谱和转移相关RNA表达谱。为了能够更加针对性的检测循环细胞外囊泡RNA，解决将组织RNA检测应用于循环细胞外囊泡RNA检测效率低的问题，我们建立了循环外泌体RNA靶基因最优化片段检测法，并建立基于循环外泌体RNA组合的PCa预测模型。此后，我们继续深入研究了前期发现的两条lncRNA——PlncRNA-1和MALAT1，发现PlncRNA-1通过驯化转移灶的环境，使PCa细胞更容易定植；MALAT1通过激活基质细胞中IL6-RANKL信号通路促进PCa的骨转移。我们进一步分析PCa转移相关的循环细胞外囊泡RNA表达谱，找到了3个循环外泌体中PCa转移相关的lncRNA，即TTTY15，lncAPP和lncAMPC。基于此，我们进一步阐述了TTTY15，lncAPP和lncAMPC通过提高PCa“种子细胞”的恶性程度在促进PCa进展和转移中的分子机制。.本项目基于前期基础进行创新性研究，通过研究PCa循环血中lncRNA的表达谱，建立PCa的预测模型，为临床提供PCa的复发转移预警；通过着重阐述循环血中lncRNA 在PCa复发与转移的作用机制，完善了细胞外囊泡lncRNA通过“恶化种子”及“驯化土壤”双重分子机制促进PCa进展转移的学说，为临床转移性PCa治疗及预后提供有力的理论依据。