Bioinformatics Analysis And Mechanism Study Of Anti-Human Breast Cancer Targets Of Atorvastatin

Background and objective:Breast cancer(BC)is the most common tumor in women and the leading cause of cancer death.Now BC treatments including surgery,radiotherapy,chemotherapy and endocrine therapy and targeted therapy,although the above method can reduce the mortality of BC,but consider the BC incidence increased year by year,gradually younger onset age,looking for new treatment methods and means to reduce the recurrence rate of BC,improve the patient's survival rate is particularly important.Statins are 3-hydroxy-3-methylglutaryl-CoA(HMG-CoA)reductase inhibitors that inhibit endogenous cholesterol production in the liver and reduce abnormal blood lipid levels,which can effectively reduce the incidence of cardiovascular events.Clinically,statins are often used in combination with other lipid-lowering drugs for primary and secondary prevention of coronary heart disease.Statins are often divided into hydrophilic statins and lipopilic statins due to their different physical and chemical properties,and Atorvastatin(ATO)belongs to lipopilic statins.ATO not only has lipid-lowering effect,but also inhibits proliferation,promotes apoptosis,inhibits neovasculogenesis,reduces metastasis,and strengthens chemotherapy activities on tumor cells.However,the mechanism of how ATO inhibits the occurrence and progression of BC is still unclear.This project aims to use bioinformatics method,analyze drug database and a variety of public gene expression profile database to predict and analyze the potential targets of ATO anti-BC,and further explore the influence of ATO on the occurrence and development of BC and the mechanism of action by molecular biology experiment.Research Contents and Methods:1.Prediction of the anti-tumor target proteins of atorvastatin.1)The direct target proteins of atorvastatin were identified using the public database DrugBank;2)The STRING online database was used to construct the interaction network and signal pathway analysis between atorvastatin target proteins;3)Through cBioPortal atorvastatin platform analysis target protein in breast cancer genomics data,and uses OncoPrint visualization.2.Analysis of differentially expressed genes in breast cancer and normal breast tissue.1)Using GEO for breast cancer expression profile chip data variance analysis,using R language map volcanic visualization;2)Genetic variations with Wayne figure integration,genetic ID from David online conversion tool to convert the ID;3)Venn diagram was used to integrate the atorvastatin target protein data with the breast cancer differential gene data.3.Verify the effect of ATO on the malignant biological characteristics of BC.1)CCK-8,EdU,scratch healing test,Transwell invasion test and flow cytometry were used to investigate the effects of different concentrations of ATO on cell proliferation,migration and invasion,cell cycle and apoptosis of MDA-MB-231 cells;2)The mRNA and protein expressions of the potential target after ATO intervention in MDA-MB-231 cells were detected by qRT-PCR and Western Blot.Results:1.Five direct targets of ATO were identified by DrugBank database,and 114 indirect targets of ATO were found by protein interaction network and KEGG signaling pathway analysis.2.The expression profiles of breast cancer and normal breast tissue were obtained from the GEO database,and 75 differentially expressed genes were obtained by using R language for differential analysis,among which 21 genes were up-regulated and 54 genes were down-regulated.After gene ID conversion using David database,29 BC differentially expressed genes were obtained.3.The intersection of ATO indirect action target and BC differential expression gene was obtained,and the target gene CCNA2 was found.Seven genes associated with CcNA2 in BC were obtained by STRING and GEPIA database analysis,and the relationship between them and the prognosis of breast cancer was analyzed.4.The results of CCK-8 and EDU experiments showed that the proliferation ability of MDA-MB-231 cells decreased significantly with the increase of ATO concentration.The scratch healing assay and Trans well invasion assay showed that the migration and invasion ability of MDA-MB-231 cells decreased significantly with the increase of ATO concentration.Flow cytometry results showed that ATO could block MDA-MB-231 cells in the G0/G1 phase,and with the increase of ATO concentration,the apoptosis rate of MDA-MB-231 cells increased significantly.5.qRT-PCR and Western Blot results showed that ATO could significantly reduce the protein expression level of CCNA2 in MDA-MB-231 cells,and also significantly reduce the mRNA expression level of CCNA2 and its associated 7 targets.Conclusion:Atorvastatin can inhibit the proliferation,migration,invasion and other malignant biological behaviors of breast cancer cells.CCNA2 is the target of Atorvastatin to inhibit the occurrence and development of breast cancer,and Atorvastatin can affect the occurrence and development of breast cancer by inhibiting the expression of CCNA2.