骨肉瘤转移发生率高，疗效差。我们前期研究发现Ⅷ型胶原蛋白NC1结构域vastatin抑制血管内皮细胞形成血管，降低荷瘤鼠肿瘤微血管密度，抑制转移，先后下调JAG2和Pck1的表达，不影响VEGF表达；同时还能抑制骨肉瘤细胞的迁移侵袭。有转移的骨肉瘤患者标本中vastatin表达显著下降，并和JAG2和Pck1表达水平以及肿瘤微血管密度呈负相关，和VEGF表达水平则无明显相关性，提示vastatin可通过独立于VEGF外的JAG2/Pck1通路直接抑制骨肉瘤转移和抑制血管生成而间接抑制转移。本项目拟采用pathway array的实验方法，筛选vastatin 调控JAG2和Pck1的重要中间分子，并通过Co-Ip 等方法验证vastatin与这些分子之间的结合特异性及结合序列，阐明其在骨肉瘤转移中的分子机制，并通过大样本量验证、功能回复实验和PDX模型来寻找潜在的骨肉瘤肺转移新治疗靶点。

Osteosarcoma (OS) has high incidence of metastases, especially lung metastasis, and the metastasis is difficult to be cured. Inhibition of angiogenesis is an important strategy for the treatment of metastasis. Our previous study found that type VIII collagen NC1 domain vastatin could inhibit vascular endothelial cells to form blood vessels, reduce the tumor bearing mice tumor microvessel density (MVD) and suppress metastasis, down-regulate the expression of JAG2 and Pck1, and do not change expression of vascular endothelial growth factor (VEGF). Meanwhile, we found that vastatin inhibited the invasiveness of OS cell line Saos-2. Specimens of OS patients with metastasis showed that vastatin expression was significantly decreased. In the contrast, the expression of JAG2 and Pck1 and tumor MVD was increased in specimens of OS patients with metastasis. Additionally, the expression level of VEGF had no obvious relation to the vastatin expression. The above results indicated that vastatin could inhibit blood vessel generation, thereby inhibiting osteosarcoma metastasis by JAG2/Pck1 signaling pathway independent of VEGF. This project intends to adopt pathway array to screen the important intermediate molecular between vastatin and JAG2 and Pck1. Furthermore, through Co-Ip assay, we will validate the molecular and sequence which specifically bind to vastatin, so as to clarify the molecular mechanisms of inhibition effect of vastatin to metastasis of OS. Then the large sample verification and functional recovery experiment will be administrated to find potential new therapeutic targets for metastasis of osteosarcoma.