TCF7L2 Promotes Anoikis Resistance And Metastasis Of Gastric Cancer By Transcriptionally Activating PLAUR

BackgroundGastric cancer(GC)is a high-incidence solid tumor in China and the Hexi Corridor Area of Gansu Province.With the improvement of surgical methods,the development of new anti-tumor drugs,and the continuous improvement of comprehensive tumor treatment models,the prognosis of GC patients has been effectively improved.However,the five-year survival rate of patients with advanced GC is still not satisfactory,and the long-term survival of GC patients whose tumor recurrence and metastasis seriously hinder.Therefore,exploring the mechanism of GC cell metastasis from the molecular level and finding effective therapeutic targets is a scientific problem that needs to be solved urgently.The abnormal proliferation of tumor cells and the resistance to anoikis lay the biological foundation for their distant metastasis.Among them,anoikis resistance is a key pre-step,and the colonization of tumor cells in distant target organs creates favorable conditions.The research on the molecular regulation mechanism of GC cell anoikis resistance will help to understand the mechanism of GC cell metastasis.Urokinase-type plasminogen activator receptor(PLAUR)is almost not expressed in normal tissues,but is significantly up-regulated in tumor tissues and is closely related to tumor cell invasion and migration.In previous experiments,we confirmed that PLAUR is up-regulated in GC and may promote anoikis resistance in GC cells.The upstream molecular regulation mechanism is still unclear.Therefore,through bioinformatics and in vivo and in vitro experiments,we clarified that transcription factor 7-like 2(TCF7L2)transcription activates PLAUR,thereby promoting GC cell proliferation,anoikis resistance and migration.Methods(1)The effects of TCF7L2 and PLAUR on the proliferation of GC cells were detected by CCK8 and Ed U kits;a cell model of anoikis resistance was constructed using ultralow adsorption 6-well plate.Then,the effects of TCF7L2 and PLAUR on anoikis resistance of GC cells were analyzed by flow cytometry and western blot;transwell andwound healing experiments were used to evaluate the effect of TCF7L2 and PLAUR on GC cell migration.(2)Use nude mice to inject tumor cells subcutaneously or tail vein to construct subcutaneous tumor formation and lung metastasis models to observe the effects of TCF7L2 and PLAUR on tumor growth and metastasis in vivo.(3)Use bioinformatics methods to find potential transcription factors that regulate PLAUR expression;use online databases to analyze the expression of transcription factor TCF7L2 in GC and its prognosis,and explore the binding site of TCF7L2 in the PLAUR promoter region;Immunofluorescence staining was used to analyze the localization and quantitative expression of TCF7L2 and PLAUR in GC cells and tissues;q RT-PCR and western blot were used to analyze the regulatory effect of TCF7L2 on PALUR;the dual-luciferase reporter gene assay and chromatin immunoprecipitation experiment confirmed that TCF7L2 is positive for PLAUR It has a positive transcriptional regulation effect.(4)Through the functional recovery experiment(Rescue),it is confirmed that TCF7L2 promotes GC cell proliferation,anoikis resistance and migration in a PLAURdependent manner.(5)Immunohistochemistry and western blot were used to analyze the expression of TCF7L2 protein in GC and adjacent tissues;based on the results of immunohistochemistry,the relationship between TCF7L2 and the prognosis of GC patients was analyzed.Results(1)In vitro,TCF7L2 and PLAUR improved the proliferation,anoikis resistance and migration ability of GC cells;(2)In vivo,TCF7L2 and PLAUR promote tumor growth and metastasis in nude mice.(3)It is predicted that there are binding sites between the transcription factor TCF7L2 and the PLAUR promoter region.The former is mainly located in the nucleus and cytoplasm,and the latter is mainly located in the cytoplasm;TCF7L2 as an upstream molecule positively regulates the expression of PLAUR in GC;TCF7L2 activates PLAUR through transcription,prompting the latter to be up-regulated in GC,(4)Through the Rescue experiment,it was confirmed that TCF7L2 promotes GC cell proliferation,anoikis resistance and migration in a PLAUR-dependent manner.(5)TCF7L2 is highly expressed in GC tissues,which is closely related to the prognosis of patients.It is an independent risk factor for predicting the poor prognosis of GCpatients.ConclusionTCF7L2 is highly expressed in GC and is an independent risk factor for poor prognosis of patients.TCF7L2 is used as a transcription factor in combination with the PALUR promoter to induce the up-regulation of PLAUR expression in GC.At the same time,TCF7L2 promotes GC cell proliferation,anoikis resistance and migration in a PLAUR-dependent form.TCF7L2 and PLAUR are potential targets for GC therapy.