Reasearch On Function And Molecular Mechanisms Of A Novel Small Molecule YF-452 Inhibiting Angiogenesis

Angiogenesis is a sophisticated multistep process including endothelial cell proliferation,migration,degradation of basement membrane and new tube-like structure formation.lt is essential for the body’s growth and development.Hower,angiogenesis also plays important roles in some disease states,especially in cancer.The new blood vessels grow and infiltrate into the solid tumor,providing essential oxygen and nutrients for it and facilitating the growth and speading of tumors.Thus,antiangiogensis has been thought as one of the most important anticancer therapies.Compared with chemotherapy directed at cancer cells,which often mutate rapidly and are prone to acquire"drug resistance" to treatment,the antiangiogenic therapy is obviously advantageous.Angiogenesis is tightly regulated by a series of stimulators and inhibitors. Among the numerous growth factors and cytokines which show angiogenic effects,vascular endothelial growth factor(VEGF) is one of the most compelling critical angiogenic factors regulating tumor angiogenesis and is frequently highly expressed in cancers. VEGF exerts its biological actions by binding to its receptors which expressed on the surface of endothelial cells,namely,VEGFRl(Flt-1),required mainly for mitogenic and chemotactic responses and VEGFR2(KDR/Flk-1),contributing to endothelial cell morphogenesis.VEGFR2 activation lead to the activation of diverse intracellular signaling molecules,including extraellular signal-regulated kinase(ERKs), focal adhesion(FAK),Src family kinase and so on,which promote the proliferation, migration,differentiatiom of endothelial cells and survival,also regulate vascular permeability. So,VEGFR2 and those intracellular signaling molecules were thought to be important targets for the inhibition of tumor angiogenesis.Currently,many VEGFR2 inhibitors have been applied in the cancer treatment in clinic,however,most of them suffer some adverse effects.Thus,it is still needed to discover some novel small molecular angiogenesis inhibitors with self-owned intellectual property right as anticancer candidates.In this investigation,obtained a potent compound YF-452 through screening from the compound libraries of our laboratory and analyzed its inhibitory effects on HUVEC in vitro. The activity of YF-452 also was examined in models ex-vivo and in vivo. And then we investigated the mechanisms blocking angiogenesis.YF-452 could significantantly inhibit HUVEC migration,invasion and tube-like structure formation.Ex-vivo(rat aortic ring spouting assay)and In vivo models(CAM model and mouse corneal micropocket model)showed that YF-452 suppresses the spouting of micro vessels and the new blood vessel formation. And further studies were found that YF-452 inhibits tumor growth in human PC-3 xenograft in athymic mice and intumoral angiogenesis,compared with the solvent control,administration of 40mg/kg/d YF-452 reached about 70% inhibition.CD31 immumofluorescence assay of tumor sections indicated the antiangiogenic effect of YF-452.To evaluate the health status of mice administrated YF-452, toxic pathologic changes in organs were detected by H&E staining and body weights were monitored once every 2 days throughout the whole experiment.No significant differences in vehicle and treated group were found,which indicated YF-452 with limited toxicity. The results of western blot analysis indicated that YF-452 suppressed VEGF-induced phosphorylation VEGFR2 kinase and the downstream protein kinase,including Src,FAK and ERK.Collectively,our findings suggest that small molecular compound YF-452 inhibits angiogenesis by targeting VEGFR2 activation and may be a candidate in antiangiogensis and anticancer therapies.In addition,we established HFD/STZ mouse model of type 2 diabetes,and hindlimb ischemia mouse model and wound healing mouse model based on it for other members of our laboratory to evaluate the effect of the angiogenic compound which is used for diabetic complications treatment.