β-elemene Tryptophan Methyl Ester Synergizes With Arsenic Trioxide Anti-tumor Effects And Mechanisms Research In Vitro

Objective:β-Elemene is an active component of herb medicine Curcuma Wenyujin Y.H. withantitumor activities, and it has generally antitumor activities and lowly cell toxicity. However,the β-elemene has poor water-solubility. The investigation found that introduction of tryptophaninto β-elemene synthesized β-elemene amino acid conjugates not only could increased itswater-solubility evidently, but it can promote antitumor activities synergizes with arsenic trioxidein human acute pro-myelocytic leukemia. In this study, we investigated the mechanisms ofβ-elemene tryptophan methyl ester with arsenic trioxide antitumor activities in vitro.Methods:The anti-proliferative action of ETME on human hepatocellular carcinoma HepG2,SMMC-7721, human colon cancer HCT-116cells and human leukemia K562cells wasdetermined by MTT assays. Only chose human hepatocellular carcinoma SMMC-7721as thefurther investigation object, MTT assays was used to determine the antitumor activities of EMTEcombination with As2O3or ADM. The anti-proliferative action of ETME and As2O3combinationon human hepatocellular carcinoma HepG2, SMMC-7721was determined by MTT assays.Synergistic interaction between the two drugs was assessed using the combination index (CI)method. The staining of AO/EB, PI/Hoechst33342, Rh123/Hoechst33342and the cell cycledistribution were analyzed by high content screening (HCS) analysis respectively. And westernblotting was used to investigate the correlatively protein expression of p53, the members ofCaspase and Bcl-2family. Results:The result of MTT assays show that, ETME action on human HepG2, SMMC-7721,human colon HCT-116cells and human leukemia K562were all exerted potent anti-proliferativeeffects, and displayed dose dependent manner. After48hours, the IC50is (22.92±1.2) μM,(34.48±0.84) μM, and (21.92±1.24) μM, respectively. ETME combination with A2O3or ADMaction on SMMC-7721cell was all disappeared growth inhibition at a manner of dose-dependent.At the20μM concentration of ETME and5μM concentration of A2O3, the combination index(CI) is1.25.The results of AO/EB and PI/Hoechst33342staining indication that, after ETME andA2O3combination, they can induced human hepatocellular carcinoma SMMC-7721apoptosis,and at a dose-and time-dependent manner. After combination, nuclei hyper chromatic densegranular can be visible in morphology, nuclear fission is reduced, formation of apoptotic bodies.The apoptotic cell ratio was increased to27.7%after combination.ETME combination with A2O3reduced the mitochondrial membrane potential,deregulated the expression of pro-caspase9, pro-caspase3, pro-caspase8, Bcl-2proteins, andthese were associated with activation of p53protein. That can be reversed by PFT-α. Theinhibition of casapse was only reversed the down regulation of caspase family expression, andhave no influence to the expression of Bcl-2family.ETME combination with A2O3detection of the cell cycle, the result of HCS shown thatcan be blocked at G2/M stage. The Western Blot detected that, the protein expression of cyclinB1and CDK2increased, and down regulated cyclinD1and p27protein, that can be reversed byPFT-α also. Conclusions:β-elemene tryptophan methyl ester expert evidence anti proliferative activity on humanhepatocellular carcinoma HepG2, SMMC-7721, human colon HCT-116cells and humanleukemia K562. ETME combination with ATO or ADM was all displayed anti proliferativeactivity. β-elemene tryptophan methyl ester with arsenic trioxide synergistic induced theapoptotic of SMMC-7721cell by the mitochondrial apoptosis pathway of p53protein. Thiscombination was arrested human hepatocellular carcinoma SMMC-7721cell cycle at G2/M.