Synthesis Of Formononetin-Dithiocarbamate Derivative 1121 And Anticancer Study Against Prostate Cancer

It is reported that isoflavone distributed widely in nature has high antitumor activities.Also dithiocarbamate is a privileged scaffold in drug discovery with anticancer activity.Molecular hybridization is a useful strategy of rational design of new ligands based on the recognition of pharmacophoric subunits in the molecular structure of two or more known bioactive derivatives.Our group has designed and synthesizeda series of formononetin-dithiocarbamate hybrids based on the molecular hybridization principle.We assessed their anticancer activities in several cancer cells.The results showed the compound 1121 was the most promising candidate among the derivatives.So we explored the specific anticancer mechanism of 1121 in prostate cancer cell?PC3?.The findings are divided as following:1.We used MTT assay to assess the anticancer activities of derivatives preliminarily and compound 1121 was the most outstandingone.Compound 1121 showed potent cytotoxicity better than 5-FU on PC3 cells.Its IC₅₀ value on PC3 cells was 1.04±0.047.2.We used flow cytometry to detect if compound 1121 induced apoptosis in PC3 cell.And the portion of Annexin V-FITC+ cells and PI+ cells didn't get changed after 1121 treatment.And neither did the expression of apoptosis-related proteins.3.We used MTT assay to monitor the proliferation of PC3 cells to explore the anticancer mechanism of1121.The growth curve displayed that 1121 could induced significant growth arrest.We use crystal violet staining to value the ability of clone formation.Results showed compound 1121 decreased the clone formation rate dramatically.4.One of the characters of cancer cells is infinite proliferation and cell cycle arrest can induce growth delay.Since the staining PI can bind to intracellular DNA which change accompanied with cell cycle,we used PI staining and flow cytometry to detect the different cell cycle phages of PC3 cells.The results showed 1121 induced significant G0/G1 phage arrest.CDK4 and Cyclin D1 are vital proteins that regulate cell cycle to transit from G1 phage to S phage.We also detected decrease in their expression levels.5.MAPK signaling pathway which make a series of tyrosine kinase phosphorylation its symbol is closely related to cell proliferation.We used Western Blotting to detect the expression of ERK,c-Jun?p38 and their phosphorylation.Results displayed 1121 inhibited the phosphorylation of ERK,c-Jun?p38.Therefore,1121 can inhibit cell proliferation through MAPK signaling pathway to a degree.6.Metastasis is among the reasons that make tumor difficult to cure.We used Transwell assay to assess the ability of cell metastasis.Our founding showed that the metastasized cells number decreased with the treatment of 1121.Wnt signaling pathway activation often leads to a high metastasis rate accompanied with lower expression of proteins Axin,GSK3?,APC and TCF4 and higher of protein ?-Catenin.And our work made them proved.Therefore the anti-metastasis effect of compound 1121 was related with inhibition of Wnt signaling pathway.Sum it up,we found that compound 1121 had significant antiproliferation effects on PC3 cells.In addition,compound 1121 arrested cell cycle accompanied with cell cycle-related proteins decreased expression and impaired the metastasis ability of PC3 cells which was related with Wnt signaling pathway.Therefore we consider that compound 1121 a promising candidate of treatment on prostate cancer.