GOLPH3调节Wls促进Wnt分泌调控脑胶质瘤增殖的机制研究

Many studies showed that several Wnt proteins were up-regulated in brain gliomas and the level of β-catenin, the main downstream effect molecule of Wnt pathway, is associated with the poor prognosis of patients, suggesting that the close relationship between Wnt pathway and brain gliomas progression. Although the signaling pathways of Wnt proteins are almost clear, the molecular mechanism of Wnt protein secretion is largely unknown. GOLPH3 is a brand new and also the first oncogene at Golgi apparatus discovered in 2009. Previously, we found that GOLPH3 down-regulation decreased the proliferation of glioma cells. Recently, we found that GOLPH3 down-regulation could decrease the protein level of Wls, the chaperon protein of Wnt secretion, and inhibit its being trafficked to Golgi apparatus and plasma membrane. On the contrary, Wls was sorted into the lysosome to be degraded after GOLPH3 down-regulation. Whether GOLPH3 promote glioma cell growth by regulating Wls retrograde trafficking and then Wnt secretion? Thus, by using the cultured glioma cells, nude mice and brain glioma tissues, we intended to study and clarify the mechanism that GOLPH3 promoted glioma growth by regulating Wls retrograde trafficking and Wnt secretion at molecular, cellular and system levels. Our results will not only identify that GOLPH3 may be a new gene up-regulated accompanying Wnt in gliomas, but also provide valuable idea for potential targets for molecular targeting therapy of brain gliomas.

多种Wnt蛋白在脑胶质瘤组织中明显升高，且其下游分子β-catenin水平与脑胶质瘤较差预后明显相关，提示Wnt信号通路与脑胶质瘤的发生密切相关。虽然Wnt信号通路已较为清楚，但有关Wnt分泌机制的研究则较少。高尔基蛋白GOLPH3是2009年新发现的、也是第一个被发现的位于高尔基体的癌基因蛋白，我们发现下调GOLPH3可抑制胶质瘤细胞增殖，抑制Wnt分泌的分子伴侣Wls循环转运到高尔基体和细胞膜，促进其在溶酶体降解。是否GOLPH3通过促进Wls的循环转运促进Wnt的分泌导致肿瘤细胞增殖？因此，本项目拟采用培养的胶质瘤U251和U87细胞、裸鼠原位脑胶质瘤模型与胶质瘤标本，从分子、细胞与整体水平研究、阐明GOLPH3通过调节Wls的循环转运调控Wnt的分泌，促进脑胶质瘤生长的机制。预期结果将揭示GOLPH3可能是伴随着Wnt高表达的全新基因，为脑胶质瘤的分子靶向治疗提供潜在的靶点。

Wnt 信号通路失调与脑胶质瘤的进展密切相关，虽然 Wnt 信号通路级联反应已较为清楚，但有关Wnt分泌机制的研究则较少。高尔基蛋白 GOLPH3 是 2009 年新发现 的、也是第一个被发现的位于高尔基体的癌基因蛋白，我们发现下调 GOLPH3抑制Wnt分泌的分子伴侣Wls循环转运到高尔基体和细胞膜，促进其在溶酶体降解。因此，本项目拟从分子、细胞与整体水平研究、阐明GOLPH3通过调节Wls的循环转运调控 Wnt 的分泌，促进脑胶质瘤生长的机制，为脑胶质瘤的分子靶向治疗提供潜在的靶点。 .研究发现：1）GOLPH3 和Wls在脑胶质瘤组织中的表达水平升高，呈正相关关系；下调GOLPH3抑制Wls循环转运到细胞膜和高尔基体，促进其到溶酶体降解；Wls可部分介导GOLPH3 促进脑胶质瘤细胞增殖的作用；下调GOLPH3抑制Wnt2b的分泌，进而抑制β-catenin信号。. 2）脑胶质瘤中EGFR上调是促进其进展的一个重要因素，EGFR与配体结合后可以通过PI3K-AKT-mTOR，Ras-Raf-MAPK，JAK2-STAT3三条途径发挥促进细胞增殖的作用，我们上一个国自然项目研究发现GOLPH3可以通过抑制EGFR的内吞降解，持续激活PI3K-AKT-mTOR，而不是Ras-Raf-MAPK通路促进脑胶质瘤进展，但对于JAK2-STAT3通路没有研究。本研究发现脑胶质瘤组织中，GOLPH3的表达水平与JAK2及STAT3正相关；作为脚手架蛋白，GOLPH3促进JAK2与STAT3的结合与激活，进而促进脑胶质瘤生长。.3）用angiopep-2 (A2-CL)作为靶头靶向血脑屏障和脑胶质瘤组织，发现纳米脂质体包载GOLPH3的siRNA可以抑制脑胶质瘤生长，说明GOLPH3的小干扰可以用纳米材料递送达到治疗效果。.综上，除了通过抑制EGFR的内吞降解导致PI3K-AKT-mTOR 通路持续激活，GOLPH3还可以通过调节Wls的再循环与Wnt的分泌，或是通过易化JAK2-STAT3通路等多种机制促进脑胶质瘤的进展，展现了一个分子在体内的多途径作用机制。作为第一个发现的高尔基体癌基因蛋白，GOLPH3通过参与蛋白的翻译后修饰，循环转运再利用等途径参与肿瘤的进展，是一个有潜力的分子治疗靶点。