The Role Of NAP1 Overexpression Promotes Invasion And Metastasis Of Non-Small Cell Lung Cancer

Background and aims:Non-small-cell lung cancer(NSCLC)accounts for 85%-90%of all lung cancer cases,and is the most common cause of cancer-related deaths worldwide.With modern cancer therapies,the 5-year survival rate for all lung cancer stages is around16%.Metastatic NSCLC is an incurable disease and the most common sites of metastases are the contralateral lung(40-50%),liver(20%),adrenal gland,bones and brain.High mortalities of NSCLC are the result of metastasis;therefore,reducing metastasis is the key to curtailing the rate of death from NSCLC.However,not much is known about specific molecular mechanisms underpinning NSCLC metastasis.In recent years,the understanding of the etiology of malignant tumors has become more and more profound,but the process of regulating tumor cell,especially the invasiveness and metastasis ability has not been elucidated.Changes in cytoskeletal protein actin are critical for cancer invasion and metastasis.Previous study had shown that NCK-associated protein 1(NCKAP1/NAP1)is an important regulator of cytoskeletal actin dynamics,which plays an important role in tumorigenesis and development.NAP1 is widely expressed in the body and is a non-catalytical tyrosine kinase adaptor protein that mediates the activation of receptors from the membrane into effectors in the cell in order to regulate the formation of the agonist cytoskeleton.Under the stimulation of extracellular signals,cell surface receptors are associated with a variety of intracellular signaling molecules,which in turn activate a range of cellular responses,such as migration,proliferation,and differentiation.Studies have shown that NAP1protein binds to cell surface receptors to regulate cytoskeletal morphology and thus affect cell migration.In summary,NAP1 is likely to act as a new transfer driver to regulate tumor cell invasion and metastasis,but there are few related studies,and its regulation mechanism is poorly understood.So far,no research has clarified the function of NAP1 in the lung cancer.Based on clinical data,the up-regulation of NAP1 expression in lung was found to be associated with tumor metastasis.Aim to study its role in NSCLC,we knock down and overexpress NAP1 protein by gene technology.To comprehensively evaluate the biological significance of NAP1 protein expression in NSCLC.Focus on the role of NAP1 high expression in the invasion and metastasis of NSCLC and its possible mechanism,in order to provide a new target for clinical prevention and treatment of advanced lung cancer.Methods:1.Clinical data study:Immunohistochemical analysis of NSCLC tumor samples and matched adjacent tissues,combined with Ocomine database data analysis,screening differential proteins.2.In vitro cell experiments:RNA interference technology and gene overexpression were used to positively and negatively regulate NAP1 gene expression,and to study the effects of NAP1 on tumor cell growth pattern,morphology,proliferation,migration and metastasis.Western blotting,immunoprecipitation and gene chip technology were used to explore its regulatory mechanisms.3.To generate a xenotransplantation model,5×10~5 cancer cells were suspended in 100?l PBS and injected subcutaneously into the right flanks of mice.The tumor-bearing mice were sacrificed on treatment day 42,and the lungs were removed and processed for histopathological analysis with H&E staining.Results:1.Immunohistochemical analysis revealed that NAP1 is highly expressed in NSCLC tissues and its expression is associated with tumor metastasis.This is consistent with the database data results.2.In vitro results:Both knock down and overexpression of NAP1/HSP90 can affect the migration and invasion of no small cell lung caner.3.The protein complex formed by the NAP1-HSP90 interaction is essential for maintaining the structural stability of NAP1.The interaction between NAP1 and HSP90 weakens the degradation of NAP1 by the ubiquitin-proteasome system.4.NAP1 participates in HSP90-mediated invasion and metastasis by activating MMP9 and epithelial-mesenchymal transition in non-small cell lung cancer cellsConclusion:NAP1 plays an important role in the invasion and metastasis of NSCLC.NAP1 can enhance the migration and invasion of no small cell lung caner.Mechanistic studies further reveal that the binding of NAP1 to the cellular chaperone heat shock protein 90(HSP90)is required for its protein stabilization,and NAP1plays an essential role in HSP90-mediated invasion and metastasis by provoking MMP9 activation and the epithelial-mesenchymal-transition in NSCLC cells.