The Antitumor Activity Of Imidazoles

Objective:This study aims to explore the antitumor activity of four new imidazole derivatives and analyze the underlying mechanisms,and thus provide an experimental basis for future studies.Methods:Human cervical cancer HeLa cells were used to determine the antitumor effects of the imidazoles by MTT assay.The compounds used were 2-phenyl-5-benzhydryl-1-?3-bromobenzyl?-1H-imidazole?C2?,2-phenyl-5-benzhydryl-1-?2-bromobenzyl?-1H-imidazole?C5?,2-?3-methyphenyl?-5-benzhydryl-1-?3-bromobenzyl?-1H-imidazole?C8?and(1,2-di-?3-methyphenyl?-5-benzhydryl-1H-imidazole?C10?,respectively.The effects of imidazoles on morphology of HeLa cells were detected by AO/EB staining method,and the DNA damage was analyzed by comet assay.The effects of imidazoles on cell apoptosis and cell cycle were also investigated by flow cytometry.Mitochondrial membrane potential,reactive oxygen species?ROS?and ATP content were detected to further unravel apoptosis-related events.Western Blotting was used to detect the expressions of Bcl-2,Bax,Caspase 3 and PARP.The autophagy of HeLa cells was determined by MDC staining method and Western Blotting analysis?p62,Beclin-1 and LC3-II/LC3-I?.Results:?1?The MTT assay results showed that the imidazole derivatives C2,C5,C8 and C10 were able to significantly inhibit the cell viabilities of HeLa cells as compared to the control;the IC₅₀0 values were 36.2±1.0,28.3±1.0,29±1.0 and 33.8±1.1?g/mL,respectively.?2?The results from AO/EB staining showed that the control cells were normal--the nuclei were intact and the green fluorescence was observed.After treatment with the imidazole derivatives,the nucleoplasm of the HeLa cells was condensed and showed orange fluorescence,demonstrating that the compounds induced apoptosis.?3?The results from comet electrophoresis showed that the cells treated with the imidazole compounds were tailed as compared with the control,indicating that these compounds were able to induce DNA damage in HeLa cells.?4?Flow cytometry experiments reveal a significant increase in the proportion of apoptosis in the cells treated with the compounds.The cells were also found to be blocked in the G2/M phase after treatment with the compounds.?5?After treatment with the compounds,the reactive oxygen species levels were increased,the mitochondrial membrane potential was decreased,and the ATP content was decreased.?6?MDC staining experiments reveal that the cells treated with the compounds showed clear green fluorescence as compared to the control,indicating that the compounds were able to promote autophagy in HeLa cells.The expressions of p62 and Beclin-1 proteins were decreased,and the expressions of LC3-II/LC3-I were increased.?7?Western blotting data demonstrate that Bcl-2 and Bcl-2/Bax expressions were down regulated while Bax,PARP and Caspase 3 expression were up-regulated.Conclusion:These results indicate that the imidazole compounds C2,C5,C8 and C10 can increase reactive oxygen species level,reduce mitochondrial membrane potential and ATP content of HeLa cells.The compounds are also shown to initiate apoptosis by inhibiting Bcl-2,increasing Bax and promoting Caspase 3.In addition,these compounds can also promote autophagy by inhibiting p62,reducing Beclin-1 and increasing LC3-II/LC3-I.In summary,the four new imidazole derivatives C2,C5,C8 and C10 can promote apoptosis and autophagy of HeLa cells.