Design,Synthesis Of Pyrimidine-based Derivatives And Its Inhibition Effects On Angiogenesis And Epithelial-to-Mesenchymal Transition

Cancer metastasis has been considered to be a hallmark in the transition of tumors from dormant to malignant state and accounts for 90% of cancer deaths. This condition is a multiple and intricate process, in which EMT plays an essential role.During EMT, epithelial cells lose their polarity and junctions and are transformed into mesenchymal-like cells with motile and invasive properities in order to promote the malignancy of tumor. Furthermore, angiogenesis has been demonstrated to be a basic prerequisite in tumor proliferation, invasion, and metastasis which can supply nutrients, oxygen and growth factors through new vessels and promote tumor growth rapidly. Hence, developing small molecular compounds that inhibit the processes of EMT and angiogenesis could serve as an effective strategy against cancer progression.Based on the concept of molecular hybridization, we designed, synthesized a series of novel 1,2,3-triazole-pyrimidine-urea hybrids and evaluated their anticancer activity against several tumor cell lines including EC109, MGC-803, MCF-7 and B16-F10. Majority of the compounds exhibited intensely inhibition activity against B16-F10 melanoma cell line which is widely used in the metastasis research of tumor cells, suggesting that the compounds may interfere with the process of EMT.Meanwhile, recent research on the compounds comprising pharmacophores of pyrimidine and urea became more appealing and promising for the inhibition of tyrosine kinases, especially for the blockage of the VEGF/VEGFR signaling pathway,which indicated that the synthesized compounds could inhibit the tumor angiogenesis.Therefore, we chose compound 38, the most potent one against B16-F10, as the tested compound and investigated the underlying molecular mechanisms of compound 38 on EMT and angiogenesis.First of all, wound healing experiment and transwell assay in B16-F10 and MGC-803 cell lines demonstrated that compound 38 could significantly inhibit the tumor cells migration and invasion. To further investigate the molecular mechanism,we detected the protein expression related to EMT in MGC-803. The results showed that compound 38 reduced the expression of Snail、Slug and ZEB-1, which resulted in the upregulation of E-Cadherin and ZO-1 as well as downregulation of N-Cadherin and Vimentin. In addition, compound 38 could inhibit the protein levels of MMP-2and MMP-7 and the translocation of β-catenin to the nucleus. All these data indicated that compound 38 could interfere with the process of EMT in MGC-803 to exert the function of anti-metastasis. Meanwhile, compound 38 also can induce MGC- 803 cells apoptosis relating to the mitochondrial apoptotic pathway and death receptor signaling pathway.Angiogenesis is critical to growth and metastasis of solid tumors. In this process,blood vessels will allow easier access of tumor cells to circulation and proliferation at distant sites. Currently, angiogenesis represents a rational target for therapeutic intervention in clinical. Thus, we selected HUVEC as the tested cell to carry out the research on angiogenesis. Cell viability assay, wound healing assay, transwell assay and tube formation experiment indicated that compound 38 could inhibit the proliferation, motility, invasion and vascular tube formation of endothelial cells in vitro. Further chicken chorioallantoic membrane assay proved that compound 38 can inhibit angiogenesis in vivo. Subsequent analyses revealed that compound 38 reduced the expression of VEGF and Bcl-2 and increased the expression of Bax and p53 in concentration dependent manner in HUVEC which elucidated that compound 38 as a potential antiangiogenic agent not only induced HUVEC apoptosis but also inhibited VEGF/VEGFR2 signaling pathway.In conclusion, based on the concept of molecular hybridization, we obtained a novel series of pyrimidine derivatives and some synthesized compounds exhibited intensively antitumor activity. Among them, compound 38 has the potent to inhibit EMT and angiogenesis, induces apoptosis and inhibits tumor cells migration and invasion. The present work preliminarily elucidates the molecular mechanism of the compound and provides a significant support for the further study on structure modification and drug targets.