Screening And Anti-tumor Activity Study Of Legumain Small Molecule Inhibitors

Cancer has become one of the most harmful chronic diseases to human beings,and it incidence rate increases year by year.The lysosomal protease Legumain is highly expressed in many human tumors and promotes the invasive activity of cancer cells.Therefore,screening small-molecule inhibitors of Legumain is expected to be a noval way for tumor treatment.The results of this study including:(1)Recombinant human Legumain was expressed in E.coli,and the result indicates that the expressed Legumain has no activity.Subsequently we produced the recombinant protein in Pichia pastoris.The results showed that Legumain was most highly expressed under pH 6 culture condition,which adjusted using ammonia.The optimum pH for the autocatalytic activation of the recombinant Legumain is pH3.(2)Based on Legumain specific cleavage of fluorescent substrate(Z-Ala-Ala-Asn-AMC),twe established the compounds screening system of the Legumain inhibitors,and six Legumain inhibitors were selected from a compounds library.Among them,Esomeprazole magnesium has the best inhibition effect on Legumain activity.(3)MTT assay showed that the cytotoxicity of the small molecule inhibitors to MDA-MB-231 were very low.(4)Transwell assay showed that Esomeprazole magnesium inhibited the invasion ability of MDA-MB-231 cells in a concentration-dependent manner.(5)Finally,the inhibition of breast cancer by Esomeprazole magnesium is tested with a nude mouse transfer model.The metastasis of cancer cells in nude mice study showed that treatment of Esomeprazole magnesium decreased the mice lung nodule numbers.HE stain also confirmed that Esomeprazole magnesium inhibited the lung metastasis of breast cancer cells in nude mice.In conclusion,Esomeprazole magnesium ignificantly inhibits the activity of Legumain and the metastasis of breast cancer cells.This study provides a new strategy for the development of drugs to inhibit tumor metastasis.