lncRNA DATOC1影响microRNA成熟促进卵巢癌转移的分子机制研究

Metastasis is one of the most intractable problems for the treatment of ovarian cancer. The research of molecular mechanism on ovarian cancer metastasis will provide theoretical basis for developing pharmaceuticals used for the novel diagnostic biomarkers and intervention of ovarian cancer. Based on previous work, lncRNA DATOC-1 was screened by lncRNA microarray and DICER RIP-seq. In vitro ovarian cancer cell line knockdown of DATOC-1 demonstrated declined proliferation and migration capability. Subsequent small RNA sequencing of microRNAs illustrate that a small cohort of microRNAs with tumor-suppressive characteristics named as mDDSM expression was significantly high in the mature form and premature counterpart. Based on previous results, novel theory of up-regulated lncRNA DATOC-1 interacting with DICER to participate in the process of microRNAs maturation was put forward. The project will explore the in vivo and in vitro function of DATOC-1 during ovarian cancer metastasis and downward molecular mechanism underlying the oncogenic function and evaluate the biomarker performance of DATOC-1 on the early diagnosis of ovarian cancer metastasis and prognosis of patients’ survival. The project will provide a new way of ovarian cancer diagnosis and intervention.

转移是卵巢癌诊治中的棘手问题，研究lncRNA在转移中的作用对转移性卵巢癌的诊治具有重要意义。. 我们发现：卵巢癌转移样本中高表达的lncRNA DATOC-1能与DICER酶结合从而调控一类具有抑癌表型的microRNA成熟体表达显著下调，这些抑癌microRNA多数靶向MMP家族蛋白。于是，申请者提出DATOC-1通过与DICER酶特异性的结合、抑制DICER酶功能，从而下调mDDSM系列microRNA成熟，提高MMP家族蛋白表达，进而促进卵巢癌转移的假说。项目后续将通过动物模型观察DATOC-1对卵巢癌转移的影响,通过RIP-seq、蛋白截断等手段研究DATOC-1与DICER酶的结合以及对mDDSM系列microRNA及MMP家族的调控。. 该项目创新性地提出lncRNA分子调控DICER酶功能的理论假说，所得研究结果有望为转移性卵巢癌的诊治提供新的方向。

卵巢癌干细胞（OCSCs）具有高度致癌性并且对传统化疗药物有很强的抵抗力，因此，它们是卵巢癌转移和复发的重要因素。据报道，树突状细胞-细胞因子诱导的杀伤细胞（DC-CIK细胞）对多系统的癌细胞都具有显着的杀伤作用，包括血液系统，消化系统，呼吸系统，泌尿系统和生殖系统。但是，目前尚不清楚DC-CIK细胞是否可以选择性地杀死OCSC。在这项研究中，我们收集了卵巢癌患者的经血样本，以获得单核细胞和DC-CIK细胞，并分离自体CD44 + / CD133 + OCSC用作靶细胞。实验结果表明，当DC-CIK细胞和OCSCs在体外分别以5：1,10：1和50：1的比例混合和培养时，DC-CIK细胞杀死了大量的OCSCs，抑制了它们的体外侵袭， 促进了细胞凋亡。qPCR和Western结果显示DC-CIK细胞通过释放TNF-α刺激OCSC中的TNFR1、ASK1、AIP1、JNK高表达并磷酸化。在使用CRISPR / Cas9技术敲除OCSC中内源性TNFR1基因后，DC-CIK细胞对OCSC的杀伤力显着减弱。临床样本分析结果表明，TNFR1表达水平与卵巢癌分期和预后呈负相关。 因此，我们创新地证实来自卵巢癌患者经血的DC-CIK细胞可以通过分泌TNF-α来激活OCSC中TNFR1-ASK1-AIP1-JNK通路的表达并杀死自体OCSC。