Study On The Nonsense Mutation Readthrough Activity Of 5-methoxyflavone

Objective: To investigate the read-through activity of 5-methoxyflavone on gene nonsense mutations and study the potential of this compound to treat tumors and other diseases caused by nonsense mutations.Methods: Through the cell model of the nonsense mutant luciferase established earlier in this research group,a positive read-through agent 5-methoxyflavone capable of reading through the nonsense mutant luciferase was preliminarily screened.Next,HEK293 MUT cells containing nonsense mutant luciferase were treated with5-methoxyflavone,the luciferase activity was detected by chemiluminescence,and the protein level of luciferase was detected by Western blot;finally,the treatment was performed with 5-methoxyflavone Human colon cancer cell HT-29 containing nonsense mutant APC gene was detected by Western blot expression of APC protein and downstream protein ?-catenin;Analysis of cytotoxicity of 5-methoxyflavone by CCK-8 experiment.The same method was used to analyze the compounds with similar structure to 5-methoxyflavone.Results:After preliminary confirmation that 5-methoxyflavone has nonsense mutation read-through activity,further compound detection and analysis revealed that5-methoxyflavone exhibited higher readthrough efficiency in HEK293 MUT cells when compared with G418(P<0.05).In HT-29 cells,the protein level of nonsense mutant APC was increased obviously along with the increase of dose of5-methoxyflavone.In addition,the readthrough activity of 5-methoxyflavone on nonsense mutant APC gene was stronger than that of G418(P<0.05).It was also found that ?-catenin decreased with the recovery of APC expression.Finally,the results showed that the toxicity of 5-methoxyflavone to normal cells was obviously less than that of G418(P<0.05).For compounds with a similar structure to5-methoxyflavonoids,good read-through activity for nonsense mutations was also found,and was superior to G418 in various indicators(P <0.05).Conclusions:5-methoxyflavone has good nonsense mutation readthrough activity,low toxicity and has a significant improvement in the malignant characterization of tumor cells.It may be a promising candidate for further study to develop nonsense read through pharmaceuticals.