靶向OVOS2及其关键信号通路调控皮肤恶性黑素瘤的发生及其网络机制解析

Cutaneous malignant melanoma (CMM) is characterized by its early metastasis and high mortality, and the discovery of key genes related to CMM occurrence may pose a promise for gene therapy in future. In our previous studies, we found a protease inhibitor gene OVOS2 is dramatically up-regulated in CMM, and in vitro experiments showed that it plays an extremely important role in promoting cancer, rather than tumor suppressor role as most of the protease inhibitors usually have. However, nothing has been known whether OVOS2 actually has the promoting cancer in vivo, as well as the regulatory mechanism of the key intermediate signaling pathways in OVOS2 promoted CMM occurrence. In this project, we intends to firstly clarify the cancer promoting effects of OVOS2 in vivo by using the nude mice model, and further reveal the key signaling pathways regulated by OVOS2 in CMM by comparatively analyzing the differentially expression profilings of genes and proteins between the normal and OVOS2-RNAi A375 cell lines; furthermore, we want to observe their cellular behaviors (proliferation, apoptosis, invasion and metastasis) and in vitro tumor promoting effects in nude mice by using the specific inhibitors targeting at the key identified signal pathways. These results would coordinatedly dissect the molecular networks of CMM occurrence regulated by OVOS2. This project will not only give new insights into the molecular regulatory mechanism of CMM occurrence, but also provide an effective target gene for early diagnosis and gene therapy of CMM in future.

皮肤恶性黑素瘤（CMM）具转移早、致死率高等特点，而致病关键基因的发掘则是未来基因治疗的关键所在。在前期项目中，我们发掘了一个在CMM中超高表达的编码蛋白酶抑制剂的基因OVOS2，并通过体外实验发现其在CMM中发挥着极为重要的促癌作用，而不是蛋白酶抑制剂所通常具有的抑癌作用。然而OVOS2在体内是否确实具有促癌作用及关键的中间信号通路调控机制则一无所知。本项目拟首先通过裸鼠成瘤模型探讨其促癌作用，进而比较分析正常和OVOS2-RNAi的A375细胞株的基因和蛋白差异表达谱，揭示OVOS2调控CMM发生的关键信号通路，并进一步应用靶向关键信号通路的特异性抑制剂处理细胞株并观察其细胞学行为（增殖、凋亡、侵袭、转移）及裸鼠成瘤效应，协同解析OVOS2调控CMM发生的网络分子机制。本项目将为深入解析CMM发生的分子调控机制提供新视角，并有望为CMM的早期诊断和基因治疗提供高效的关键靶标。

皮肤恶性黑素瘤（cutaneous malignant melanoma，CMM）是皮肤科致死率最高的恶性肿瘤，具有进展快、易转移、放化疗不敏感等特点。目前CMM发生的分子机制尚不完全清楚，而致病关键基因的发掘则是未来基因治疗的关键所在。在前期项目中，我们发掘了一个在CMM中超高表达的转录本OVOS2，与正常组织相比上调了33倍，然而目前关于OVOS2的确切功能还知之甚少。本项目通过构建OVOS2的RNAi载体、稳定转染CMM细胞株及裸鼠成瘤实验，揭示OVOS2在CMM的发生中起到了关键的促癌作用；获得了OVOS2调控CMM发生的转录组、蛋白组和磷酸化蛋白组表达响应谱，分析了OVOS2所可能调控CMM发生的关键生物学过程及信号通路；通过转录组、蛋白组及关联分析和western揭示了在黑素瘤发生中OVOS2很大程度上对PI3K-AKT和RAS/MAPK这两个重要的肿瘤发生信号通路起着关键的调控作用，可能与其过度激活信号通路有关，继而诱导细胞周期相关蛋白表达，参与细胞周期和细胞增殖的调控；并结合基因转录表达调控的ceRNA机制及NCBI数据库对OVOS2的功能注释变化情况和转录本情况，进一步探讨了OVOS2在CMM中发挥调控作用的其他可能途径。本项目为深入解析CMM发生的分子调控机制提供了新视角，并有望为CMM的早期诊断和基因治疗提供高效的关键靶标。