Study On Anti-lung Cancer Activity Of Triterpenoids From Momordica Charantia L.vines

Objective:Lung cancer is one of the malignant tumors with the fastest increase in morbidity and mortality,and the greatest threat to the health and life of the population.The current treatment for lung cancer is mainly chemotherapy.Chemotherapy drugs usually have relatively large side effects.It is imperative to develop low-toxic and effective anti-lung cancer drugs.Therefore,this study preliminarily explores the anti-lung cancer effect of bitter gourd through two aspects.The first part uses network pharmacology and bioinformatics methods to screen out the anti-tumor active ingredients triterpenes in bitter gourd.The second part verifies the inhibitory effect of bitter gourd triterpenes on lung cancer cells through in vitro cell experiments.The mechanism of action and the development of lung cancer drugs provide a theoretical basis.The first part: Screen the active ingredients of bitter gourd for the treatment of lung cancer by means of network pharmacology.Method:1.Literature search collects all the active ingredients in bitter melon,and predicts the effective targets of the active ingredients in bitter melon through Swiss ADME database and Pharm Mapper server.2.TCGA database predicts the differentially expressed genes of lung cancer,and crosses with the effective target of the active ingredient of Momordica charantia in the first step,and obtains the potential target of Momordica charantia for the treatment of lung cancer.3.Build a co-expression network for potential targets to analyze modules with similar expression characteristics,and analyze the correlation between different modules and clinical traits.4.Use the DAVID database to explore the biological process of the GO function of potential targets,and use the cluster Profiler package in the R language to analyze the KEGG signal pathway.5.Construct protein-protein interaction network,"compound-target" network and "path-target" network,and conduct topological analysis to determine key compounds and core targets.6.The docking status of the core target by the molecular docking verification and the key small molecule compound.Results:1.Through the analysis of network pharmacology combined with bioinformatics methods,26 candidate active ingredients in bitter melon were initially screened and 1129 effective targets were obtained.2.A total of 2,999 gene targets related to lung cancer were obtained in the TCGA database,and 133 drug and disease co-action targets were obtained by intersecting with the effective targets of bitter gourd.3.Co-expression network analysis shows that the cyan module has a higher positive correlation with T staging and gender,and the green module has a higher negative correlation with T staging.4.The results of the PPI network showed that the core target genes of Momordica charantia in the treatment of lung cancer were ALB,MMP9,SLC6A4,ACE,CCNB1,F2,CCNA2,OPRM1,CASR,CHRM1.5.Construct a "compound-target" network to determine the treatment of lung cancer by Momordica charantia The key compounds are Kuguacin A,Kuguacin C,Kuguacin J,(+)-Catechin,Feurlic Acid and Momordicine.6.The biological process involved in GO function annotation includes cell proliferation,gluconeogenesis,regulation of reactive oxygen metabolism,regulation of vasoconstriction,positive regulation of inflammatory response,synthesis of DNA biological processes,hypoxia response,and vascular remodeling and other functions.7.KEGG pathway analysis shows that the main pathways for the treatment of lung cancer by bitter melon include p53 signaling pathway,glycolysis/gluconeogenesis,chemical carcinogenesis,cell cycle,oocyte meiosis and other pathways.8.The results of molecular docking show that all six key compounds can dock spontaneously with ten core targets.Kuguacin J,Kuguacin C,Kuguacin A,Momordicine ? have higher docking free energies,all of which are triterpenoids in Momordica charantia.The second part: combined with the results of network pharmacology,through in vitro cell experiments,preliminary research on the inhibitory effect of Momordica charantia triterpenoids on lung cancer A549 cells.method:1.CCK8 method is used to detect the inhibitory ability of Momordica charantia triterpenes on the proliferation of A549 cells,and the appropriate dosage concentration is selected to carry out follow-up experiments.2.Angiogenesis experiment to observe the effect of Momordica charantia triterpenes on the angiogenesis of A549 cells.3.The clone formation experiment detects the effect of Momordica charantia triterpenes on the malignant proliferation ability of A549 cells.4.Transwell experiment to detect the influence of Momordica charantia triterpenes on the invasion ability of A549 cells.5.Detect the effect of Momordica charantia triterpenes on the migration ability of A549 cells by plate scratch test.6.Finally,the effect of Momordica charantia triterpenes on the glucose uptake capacity of A549 cells was tested by in vitro glucose consumption experiment.Results:1.CCK8 method detects the proliferation inhibition test of Momordica charantia triterpene on A549 cells,by setting different concentrations(10,20,40,80,160μg/m L),the treatment time is 24,48,72 h respectively,the results show that Momordica charantia triterpenes Terpenes can inhibit the proliferation of A549 cells,and the highest inhibition rate is 99.21±0.08%.Momordica charantia triterpenes have a good time-effect and dose-effect relationship on A549 cells.2.The clone formation experiment proved that Momordica charantia triterpenes can inhibit the malignant proliferation of A549 cells.The greater the concentration of the drug,the stronger the inhibition rate.3.Angiogenesis experiments showed that Momordica charantia triterpenes can inhibit the angiogenesis of A549 cells,and as the concentration of Momordica charantia triterpenes increases,the number of angiogenesis decreases.4.Transwell experiment found that Momordica charantia triterpenes can inhibit the invasion ability of A549 cells.The greater the concentration of the drug,the fewer cells A549 enters the lower chamber,and the stronger the inhibitory effect.5.Scratch repair experiments found that Momordica charantia triterpenes can inhibit the migration ability of A549 cells,and the inhibitory effect increases with the increase of drug concentration.6.Glucose consumption experiment found that Momordica charantia triterpenes can effectively inhibit the glucose consumption ability of A549 cells.Conclution:The anti-lung cancer active ingredients in bitter gourd are screened by network pharmacology as triterpenoids,and they have anti-lung cancer effects through multiple pathways and multiple targets.Further in vitro experiments proved that bitter gourd vine triterpenoids can effectively inhibit the proliferation,angiogenesis,invasion and migration of A549 cells,and can reduce the ability of A549 cells to consume grapes.These studies provide new ideas for in-depth exploration of the anti-lung cancer mechanism of bitter melon triterpenoids and the development of new drugs for lung cancer treatment.