Regulation Mechanism Of Legumain In Neuroblastoma And Research Of Prodrug Based On Legumain

ObjectiveTo explore expression of legumain in neuroblastoma and the correlation between the expression and clinic histopathological classifications.And explore the role of legumain in invasion and migration of neuroblastoma and its new possible regulatory mechanism.Test effect of prodrug EMC-AANL-DOX by means of hydrolysis activity of legumain on NB tumor models.MethodsNeuroblastoma(NB)is highly malignant solid tumor in children.Legumain is expressed in adult solid tumors,but its expression in NB has not been examined.In this study,first legumain expression in NB cell lines and in microarrays of tumor tissues collected from 46 children with undifferentiated neuroblastoma,differentiated neuroblastoma and ganglioneuroblastoma were examined.Second,by the way of gene gene interference and gene transfection,effect of legumain on tumor invasion and migration was also tested,especially effect of legumain on epithelial mesenchymal transition of neuroblastoma,by adding activated legumain(also called asparagine endopeptidase and specific asparagine endopeptidase inhibitor,effect of legumain on EMT was examined.Third,a targeted prodrug EMC-AANL-DOX based on hydrolysis activity of legumain was synthesized and examined.The efficacy,specificity,and toxicity of EMC-AANL-DOX were then evaluated in a tumor bearing mouse model of neuroblastoma.Tumor volumes and lifetime was evaluated.Damage to blood cells,renal functions and cardiac functions was also evaluated by biochemical criterion and histopathology analysis.ResultsLegumain is extensively expressed in neuroblastoma.Correlation ananlysis of legumain and microarrays of tumor tissues collected from 46 children with undifferentiated neuroblastoma,differentiated neuroblastoma and ganglioneuroblastoma showed that legumain expression is correlated with differentiation of NB.Legumain is highly expressed in undifferentiated NB and NB with metastasis,its expression is a little lower in GNBi or GNBn,and expression is the lowest in differentiated NB.Legumain can promote invasion and migration of neuroblastoma by the way of epithelial mesenchymal transition,and this transition is proceeded by asparagine endopeptidase activity of legumain.The target prodrug EMC-AANL-DOX,a novel doxorubicin-based prodrug activated by legumain,showed lower inhibition of bone marrow,no damage to cardic and renal function,which proved it as a effective target for tumor therapy.ConclusionsDegrees of differentiation in neuroblastoma is correlated with expression of legumain,Legumain can promote invasion and migration of NB by epithelial mesenchymal transition,which is proceed by its asparagine endopeptidase activity.Prodrng based on legumain has proved better effect and lower toxicity.Modification of traditional drugs based on target enzymatic activity provides a new choice for tumor therapy.