Hu Jie, Dong Aiwen, Fernandez-Ruiz Veronica, Shan Juanjuan, Kawa Milosz, Martínez-Ansó Eduardo, Prieto Jesus, Qian Cheng
Division of Gene Therapy and Hepatology, Center for Applied Medical Research, University of Navarra, Pamplona, Spain.
Cancer Res. 2009 Sep 1;69(17):6951-9. doi: 10.1158/0008-5472.CAN-09-0541. Epub 2009 Aug 18.
Aberrant activation of Wnt signaling plays an important role in hepatocarcinogenesis. In addition to direct effects on tumor cells, Wnt signaling might be involved in the organization of tumor microenvironment. In this study, we have explored whether Wnt signaling blockade by exogenous expression of Wnt antagonists could inhibit tumor angiogenesis and control tumor growth. Human Wnt inhibitory factor 1 (WIF1) and secreted frizzled-related protein 1 (sFRP1) were each fused with Fc fragment of human IgG1 to construct fusion proteins WIF1-Fc and sFRP1-Fc. The recombinant adenoviral vectors carrying WIF1-Fc and sFRP1-Fc driven by cytomegalovirus promoter were constructed. Ad-WIF1-Fc or Ad-sFRP1-Fc induced the expression and correct conformation of WIF1-Fc and sFRP1-Fc fusion proteins. These molecules caused down-regulation of E2F1, cyclin D1, and c-myc and promoted cell apoptosis in hepatocellular carcinoma cells. Treatment of established hepatocellular carcinoma tumors with Ad-WIF1-Fc and/or Ad-sFRP1-Fc resulted in significant inhibition of tumor growth and prolonged animal survival. The antineoplastic effect was associated with increased apoptosis of tumor cells, reduced microvessel density, and decreased expression of vascular endothelial growth factor and stromal cell-derived factor-1. Tube formation and migration of human microvascular endothelial cells and mouse endothelial progenitor cells (EPC) were significantly inhibited by both WIF1-Fc and sFRP1-Fc. In addition, these molecules blocked EPC differentiation and caused EPC apoptosis. Our data indicate that Wnt antagonists WIF1-Fc and sFRP1-Fc inhibit Wnt signaling and exert potent antitumor activity by increasing the apoptosis rate in tumor cells and by impairing tumor vascularization.
Wnt信号通路的异常激活在肝癌发生过程中起着重要作用。除了对肿瘤细胞的直接影响外，Wnt信号通路可能还参与肿瘤微环境的构建。在本研究中，我们探讨了通过外源性表达Wnt拮抗剂来阻断Wnt信号通路是否能够抑制肿瘤血管生成并控制肿瘤生长。人Wnt抑制因子1（WIF1）和分泌型卷曲相关蛋白1（sFRP1）分别与人IgG1的Fc片段融合，构建融合蛋白WIF1-Fc和sFRP1-Fc。构建了由巨细胞病毒启动子驱动的携带WIF1-Fc和sFRP1-Fc的重组腺病毒载体。Ad-WIF1-Fc或Ad-sFRP1-Fc诱导了WIF1-Fc和sFRP1-Fc融合蛋白的表达及正确构象。这些分子导致E2F1、细胞周期蛋白D1和c-myc的下调，并促进肝癌细胞的凋亡。用Ad-WIF1-Fc和/或Ad-sFRP1-Fc处理已建立的肝癌肿瘤，可显著抑制肿瘤生长并延长动物存活时间。抗肿瘤作用与肿瘤细胞凋亡增加、微血管密度降低以及血管内皮生长因子和基质细胞衍生因子-1表达减少有关。WIF1-Fc和sFRP1-Fc均显著抑制人微血管内皮细胞和小鼠内皮祖细胞（EPC）的管腔形成和迁移。此外，这些分子阻断EPC分化并导致EPC凋亡。我们的数据表明，Wnt拮抗剂WIF1-Fc和sFRP1-Fc通过提高肿瘤细胞凋亡率和损害肿瘤血管生成来抑制Wnt信号通路并发挥强大的抗肿瘤活性。
