COPB2与NUPR1互相"对话"在去势抵抗前列腺癌进展中的作用机制研究

The incidence and mortality of prostate cancer in our country has been increasing. Prostate cancer, especially castration-resistant prostate cancer (CRPC), is a difficult about clinical treatment. Based on our previous studies, the membrane vesicle COPB2 indicated significant higher expression in CRPC patients. Interference with COPB2 can inhibit the proliferation, invasion of CRPC cells and promote its apoptosis. Furthermore, COPB2 inhibited the expression of NUPR1 in PC-3 cells, the expression of COPB2 and NUPR1 was negatively correlated with each other both in tissues and cells. It is presumed that COPB2 participates in the prostate cancer process as an oncoprotein, and plays an important role in the progress of CRPC through mutual regulating NUPR1. To further study: (1) to test the expression of COPB2 and NUPR1 in tissue and assess their correlation with the clinical prognosis of CRPC patients; (2) interaction for COPB2 and NUPR1 to regulate the proliferation and invasion of prostate cancer in vivo, vitro and rescue experiments; (3) to explore the key downstream signals of COPB2 and NUPR1 in CRPC. By clarifying the mechanism of the interaction between COPB2 and NUPR1 in CRPC, our present study will provide new markers for the diagnosis of prostate cancer, and a basis for the development of effective drugs.

我国前列腺癌的发病率及死亡率逐年上升，前列腺癌尤其是去势抵抗性前列腺癌（CRPC）是临床治疗的难点。课题组前期发现膜泡蛋白COPB2在CRPC患者中显著高表达，干扰COPB2显著抑制CRPC细胞的增殖和侵袭并促进其凋亡。进一步发现PC-3细胞中COPB2抑制NUPR1的表达，两者在组织和细胞中的表达呈显著负相关。推测COPB2作为癌蛋白参与前列腺癌进程，与NUPR1互相调控在CRPC进展中发挥重要作用，为此，拟进行（1）检测临床标本COPB2及NUPR1的表达，评价其与CRPC进程及预后相关性；（2）采用体内外及功能回补实验证实COPB2与NUPR1互相调控对前列腺癌增殖和侵袭的影响；（3）探究COPB2与NUPR1在CRPC中互相调控激活/抑制的关键下游信号。本研究通过阐明COPB2与NUPR1在CRPC中互相调控的作用机制，有望为诊断前列腺癌提供新指标，为开发有效药物提供新依据。

前列腺癌，特别是去势抵抗前列腺癌是目前临床研究热点。本课题基于蛋白质组学鉴定的COPB2蛋白，对其进行临床、功能和机制研究。.1、COPB2在前列腺癌增殖侵袭中的作用和机制。.通过临床-细胞，动物-分子机制研究证实：①通过前列腺癌细胞株（PC-3/DU-145/CWR22RV1）研究，利用CCK8、平板克隆、细胞周期、细胞凋亡、划痕愈合及Transwell侵袭实验，证实COPB2能促进前列腺癌增殖和侵袭；②通过裸鼠皮下成瘤实验，注射PC-3细胞，证实COPB2能促进瘤体的生长；③通过表达谱芯片分析及Western blot，证实COPB2促进前列腺癌增殖是通过MEK-ERK/p38-CREB/c-fos信号通路实现。④通过miRNA预测网站、临床新鲜组织检测及细胞实验，证实COPB2是miRNA-4318的靶基因，并受其调控；⑤通过表达谱芯片分析预测COPB2互作蛋白，细胞实验及Co-IP证实NUPR1与COPB2相互作用，相互调控。⑥此部分研究内容已发表中华系列文章1篇，投稿SCI文章1篇。.2、COPB2表达与前列腺癌临床特征指标的相关性。.①COPB2在前列腺癌组织中高表达；②COPB2高表达与患者年龄、Gleason评分及PSA术后生化复发呈正相关；③COPB2可分泌至血清，在前列腺癌患者外周血中表达增高。④此部分研究内容投稿SCI文章1篇。.3、KDM5A通过ETS1依赖途径抑制miR-330-3p并激活COPB2/PI3K/AKT信号参与前列腺癌进展。.①KDM5A在前列腺癌组织中高表达，且与骨和淋巴结转移、Gleason评分、临床分期正相关，且与更差预后相关；②通过前列腺癌细胞株（PC-3/DU-145）研究，利用CCK8、平板克隆、EdU增殖、细胞凋亡、划痕愈合、Transwell侵袭、血管形成实验，证实KDM5A能促进前列腺癌增殖和侵袭；③KDM5A通过H3K4me2组蛋白去甲基化增强ETS1表达；④ETS1抑制miR-330-3p转录；⑤miR-330-3p与COPB2 3’ UTR区结合抑制其表达；⑥KDM5A促进前列腺癌进展通过下游COPB2/PI3K/AKT信号通路实现。⑦此部分研究内容投稿SCI文章1篇（已接收）。.申请一项发明专利，拟设计COPB2与NUPR1的联合检测试剂盒，用于临床检测。