Antitumor Effects And Mechanism Of WYX72,a Novel Imide-modified Flavonoid Compound,on Hepatocellular Carcinoma In Vitro And In Vivo

Objective:Flavonoids are a class of natural compounds which are low toxicity and multi-target and exert many pharmacological activities,such as anti-tumor effects,antiinflammatory and antioxidation.The imide compounds have also shown potent antitumor activity.Therefore,the conjugates of flavonoids and imide may exert favorable antitumor activity.In this study,the antitumor activity and molecular mechanism of WYX72,a novel imidazole-modified flavonoid compound,were evaluated in vitro and in vivo.Methods:The antiproliferative effects of WYX72 was detected by MTT assay.on hepatocellular carcinoma Hep G2,SMMC-7721 cells,colon cancer HCT-116,HT-29 cells,breast cancer MCF-7 and MDA-MB-231 cells.The cell death type was detected by AO/EB/Hoechst 33342 staining,Annexin V/PI staining,Rh123staining,PI staining and DCFH-DA staining.The anti-migration effect of WYX72 on tumor cell was examined by cell scratches.The protein expression of Caspase family,cytochrome C,Bcl-2 family and cell cycle was detected by Western Blot.Furthermore,the antitumor growth and metastasis effects of WYX72were also investigated by mouse H22 solid tumor,H22 lung metastasis and CT-26 lung metastasis model in vivo.Results:MTT assay showed that WYX72 significantly inhibited the proliferation of Hep G2,SMMC-7721,HCT-116,HT-29,MCF-7 and MDA-MB-231 cells,the IC₅₀ value of these cells after treated with WYX72for 48 h was?7.80±2.76??M,?1.54±0.45??M,?8.22±0.21??M,?4.85±0.99??M and?2.91±1.04??M,respectively.Our data demonstrated that WYX72 could induce Hep G2 cells and SMMC-7721 cells apoptosis,G₂/M phase arrest and decrease of mitochondrial membrane potential in a dose-dependent manner.DCFH-DA staining showed that WYX72 increased the content of reactive oxygen species in HepG2 and SMMC-7721 cells in a concentration-dependent.The results of cell scratch test showed that WYX72 could significantly inhibit the migration of tumor cells in a dose-and time-dependent manner.Western Blot results demonstrated that WYX72 up-regulated the protein expression of Bax,p53,Cyclin-B1,down-regulated the protein expression of Bcl-2?Cyclin-D1,activated Caspase-8?Caspase-3?PARP-1,and also promoted the release of cytochrome C from mitochondria to cytoplasm.Mouse H22 solid tumor and lung metastasis model results show that WYX72 significantly inhibited tumor growth and metastasis,and the inhibition rate was 70.11%and 64.18%,respectively.Furthermore,WYX72 also evidently inhibited CT-26 lung metastasis,and the inhibition rate was 78.04%.Conclusions:WYX72 exerted potent antitumor growth and metastasis effects in vivo and in vitro,and these effects were related to mitochondrial and death receptors signal pathway.