Study On Exosomes Proteome In Lung Cancer Metastasis And Kinase Regulatory Network Construction

Exosomes are a type of extracellular vesicles secreted by cells,which carry a large number of biological molecules such as proteins,nucleic acids and lipids into the cellular microenvironment.They can transmit information to the remote organs through body fluids,resulting in phenotypic changes of recipient cells.Recent studies have shown that exosomes play an important role in tumorigenesis,development and distant metastasis,but the role of exosomal proteins in cancer metastasis remains unclear.At the same time,phosphoproteins of the exosome are often associated with signaling,and the kinase regulatory network behind abnormal protein phosphorylation will contribute to the diagnosis and treatment of cancer.In response to these problems,this research uses quantitative proteomics technology to systematically study exosomes in lung cancer cells,lung cancer tissues,lung cancer plasma and mouse models.The main research achievements are as follows.First,based on quantitative proteomic analysis of exosomes,the mechanism that exosomes secreted by highly metastatic lung cancer cells significantly enhance the matastasis of recipient cells.Exosomes derived from highly/poorly metastatic lung cancer cells were compared by label-free quantitative proteomics techniques,and more than 1,400 exosomal proteins were identified.Through bioinformatic analysis of difference proteins,hepatocyte growth factor(HGF)was found to be related to cancer metastasis and was verified in plasma exosomes of lung cancer mouse models and lung cancer patients.It was found that HGF can activite c-Met pathway inducing epithelialmesenchymal transition in recipient cells,and further confirmed by HGF knockdown,kinase inhibition and protease pretreatment experiments.Therefore,exosomes secreted by highly metastatic lung cancer cells promote metastasis of cells by activating HGF/cMet signaling pathway,which reveals new targets for cancer metastasis and therapeutic studies.Second,based on the quantitative analysis of exosomal phosphorylation proteome of lung cancer tissue,a multi-omics intergration analysis strategy was established to reveal the lung cancer-associated kinase regulatory network.The exosomal protein was isolated from clinical cancer tissues and adjacent cancer tissues from patients with nonsmall cell lung cancer,and the differential expression of exosomal phosphorylated peptides and phosphorylated proteins were comprehensively analyzed by phosphopeptide specific enrichment and quantitative proteomic analysis.Based on the Tied diffusion(Tie DIE)algorithm,we integrated transcriptome and genomic datasets from non-small cell lung cancer patients in the Cancer Genome Project(TCGA),and combined analysis with phosphorylated proteome datasets.We constructed the kinase signaling networks involving exosomal phosphorylated proteins and validated the key phosphoproteins in the network in lung cancer tissue exosomes samples,which revealing reliable targets for the treatment of cancer and drug discovery.In summary,based on quantitative proteomics technology,this thesis has carried out in-depth research on proteome and phosphorylated proteome of various exosomes.The key exosomal proteins involved in lung cancer cell metastasis were discovered and verified,and their functions were explored,revealing a new way for exosomal proteins to promote lung cancer metastasis.Integrating multi-omics datasets analysis of phosphorylation proteins constructs a corresponding kinase regulatory network in exosomes,provides new strategies for cancer research and precision medicine,and is of great significance for new drug development and clinical treatment.