A Study On The Identification Of Key Protein-coding Genes In Adenocarcinomas And The Action Mechanism Of Extracellular Vesicles In The Development Of Adenocarcinoma

Background: Cancer is one of the most serious diseases that endanger human health.Its morbidity and mortality are increasing in recent years.Invasion and metastasis are the most prominent features of malignant tumors,which seriously threaten the health of patients.Surgical resection and chemotherapy are still the best options for treatment.However,most cancer have metastasized to important organs(such as lung,liver,and brain),when they are first diagnosed.Metastasis impairs the physiological functions of multiple systems and organs in patients.Adenocarcinoma is an important subtype of malignant tumors,which can originate in a variety of organs and is prone to metastasize.Lung cancer has the highest morbidity and mortality not only in China,but also in the world.Adenocarcinoma accounts for 50%of all lung cancers and is not easy to diagnose.This type of cancer has high invasiveness and an increasing incidence in recent years.Mostly importantly,its the overall survival is low.Pancreatic cancer is one of the deadliest malignant tumors,and 80% of clinical cases are adenocarcinoma,ranking first among asymptomatic cancers.Pancreatic cancer is difficult to diagnose at an early stage,due to its lack of reliable markers.The 5-years survival rate of.patients suffering from pancreatic cancer is only about 8%.Therefore,it is important to clarify the mechanism of adenocarcinoma occurrence and development for effective prevention and treatment.Pancreatic stellate cells are the main regulatory cells for the production and degradation of epithelial mesenchyme in pancreatic cancer.They are key participants in the formation of pancreatic cancer microenvironment,and have complicated interactions to pancreatic cancer.Our previous studies have suggested that SDF-1/CXCR4 is involved in the regulation of the interaction between pancreatic stellate cells and pancreatic cancer.In recent years,many studies have found that extracellular vesicles(EVs)play an important role in the interaction between tumors and the microenvironment.However,the role of extracellular vesicles derived from pancreatic stellate cells(PSCs-EVs)in the interaction between pancreatic cancer and its microenvironment and its related molecular mechanism are not clear.In this study,bioinformatical methods were used to screen the key genes associated with patient survival and prognosis,and then,PSCs-EVs were extracted and tested for its effects on invasion,metastasis and secretion of pancreatic cancer cells.The goal of our study is to explore the role of extracellular vesicles in the occurrence and development of pancreatic cancer.Objective: In the first part of our study,we discovered novel protein-coding genes related to the survival and prognosis of cancers(lung adenocarcinoma and pancreatic cancer)through screening GEO database;In the second part,we exploreed the molecular mechanism by which extracellular vesicles derived from pancreatic stellate cells affect the novel key protein-coding genes and cancer development.Methods:In the first part,we searched for the shared key protein genes of lung adenocarcinoma and pancreatic cancer).In details,we used bioinformatical methods and GEO database to find the differentially expressed genes between cancerous and normal tissues in lung adenocarcinoma cancer and pancreatic cancer.Then,we defined biological pathways and biological function related with these differentially expressed genes by KEGG and GO analyses.Moreover,we constructed the PPI network for these genes to determine the central gene sets,and performed survival analysis on these identified key genes.Finally,q RT-PCR experiment was conducted to confirm the difference in m RNA level expression of the key genes between tumors and normal tissues from clinical samples.In the second part,we extracted EVs from PSCs,and then,used scratching and Transwell experiments to explore the effect of EVs on the migration and invasion of pancreatic cancer cells.q RT-PCR and Western Blot were used to detect their effect on the secretion of CXCR4 and the selected key genes in pancreatic cancer cells.In order to further explore the role of selected key genes in the effect of extracellular vesicles on progression of pancreatic cancer,we knocked down the selected key genes in pancreatic cancer cell lines,and then,detected its effect on the behavioral function of pancreatic cancer cells by scratch and Transwell experiments.Western Blot was performed to detect whether extracellular vesicles promoted the selected key genes expression by activating PI3K/AKT pathway.Results:1.In the search for common key protein genes in tumors,we found that TOP2 A is a key coding gene in the two highly aggressive adenocarcinomas: ASPM,CCNB2,CDC20,CDC45,MELK,TOP2 A and UBE2 T are associated with lung adenocarcinoma cancer;ANLN,CCNB1,CDK1,CEP55,ECT2,NUSAP1,RACGAP1,TOP2 A and ZWINT are key genes related to the progression of pancreatic cancer.2.In the part of studying the effect of PSCs-EVs on pancreatic cancer,we found that PSCs-EVs can be taken up by pancreatic cancer cells,PANC-1,and promoted their migration and the expression of CXCR4 and TOP2 A in the cells.Scratching and Transwell experiments showed that knocking down of TOP2 A in pancreatic cancer cell lines reduced the tumor-promoting effect of extracellular vesicles on the migration and invasion of pancreatic cancer cells,.Also,Western Blot showed that extracellular vesicles activate PI3K/AKT pathway,and reduce the expression of TOP2 A via this signalin pathway.Conclusions:1.TOP2 A is a newly defined key protein-encoding gene both in the progression of lung adenocarcinoma cancer and pancreatic cancer,and is related to the survival of lung adenocarcinoma and pancreatic cancer patients.2.PSCs-EVs enhance the migration and invasion of PANC1 pancreatic cancer cells bypromoting the expression of TOP2 A,which is mediated by and the activation of CXCR4/PI3K/AKT pathway.