A Preliminary Study On The Anti-Cholangiocarcinoma Effect And Mechanism Of Norcantharidin In Vitro

Cholangiocarcinoma is a relatively rare malignant tumor,but when diagnosed it is usually difficult to cure in the late stage,and due to its strong heterogeneity and invasion,there are few effective treatment drugs.Cantharidin is a sesquiterpene compound extracted from traditional Chinese medicine Blister beetle,which has good anti-cancer effect,but Cantharidin has greater side effects.This greatly limits the clinical application of Cantharidin.It was found that artificial removal of 1 and 2methyl groups from Cantharidin can form a new derivative Norcantharidin,which retains the anti-cancer activity of Cantharidin and reduces its clinical side effects.However,there are few reports on Norcantharidin for treating cholangiocarcinoma at present.This study uses network pharmacology to predict and combine with cholangiocarcinoma cell model experiment to preliminarily explore the mechanism of Norcantharidin against cholangiocarcinoma.This study first explored the inhibitory effect of Norcantharidin on cholangiocarcinoma in vitro based on cell biology and other related research methods,then used network pharmacology combined with molecular docking to predict the targets and mechanisms of Norcantharidin against cholangiocarcinoma,and finally used cell biology and molecular biology methods for further prediction to provide basic theoretical basis for the clinical application of Norcantharidin in treating cholangiocarcinoma.(1)This study used CCK8 assay,colony formation assay and scratch healing assay to confirm that Norcantharidin can inhibit the proliferation of Cholangiocarcinoma cell lines QBC939 and Hu CCT1 in vitro,and showed concentration-and time-dependence.It can also inhibit the migration ability and colony formation ability of QBC939 and Hu CCT1 cell lines,which is concentration dependent.Flow cytometry analysis showed that Norcantharidin could regulate G2/M phase arrest of QBC939 and Hu CCT1 cell lines.(2)This study used network pharmacology to predict the important target of Norcantharidin against Cholangiocarcinoma and verified EGFR,PTGS2,and CASP3 by molecular docking and Western blot.Combined with the results of network pharmacology prediction,it indicates that Norcantharidin may have an anti-Cholangiocarcinoma effect by inducing apoptosis.(3)This study further verified the above results by fluorescence staining and flow cytometry: Norcantharidin induces apoptosis of cholangiocarcinoma cells,and Western blotting shows that Norcantharidin can regulate the expression of proteins related to endogenous apoptotic pathways,indicating that Norcantharidin may induce apoptosis of cholangiocarcinoma cells through Caspase-dependent endogenous apoptotic pathways,thus exerting an anti-cholangiocarcinoma effect.