Effect Of HECW1 On Biological Function And Prognostic Value Of Clear Cell Renal Cell Carcinoma

BackgroundKidney cancer is one of the most common malignant tumors of urinary system,and its morbidity and mortality are increasing year by year in the world.Clear cell renal cell carcinoma(ccRCC)is the most common pathologic type.Although the prevalence of imaging has increased the detection rate of sporadic renal cancer,most patients have already been locally advanced when they are detected,and 17%of patients have distant metastasis at the time of initial diagnosis.Surgical resection is the main treatment for early localized renal cancer,but about 20%of patients will have recurrence and metastasis after surgery.However,partially advanced or advanced renal cancer is not sensitive to radiotherapy and chemotherapy,so targeted therapy is usually given to such patients.But after 6-18 months of treatment,targeted drug resistance often occurs,resulting in treatment failure.The progression of renal cancer after treatment is one of the important reasons for the low survival rate of patients.Therefore,it is of great clinical significance to explore the molecular mechanism of renal cancer progression,seek new targets for renal cancer treatment and find effective prognostic factors for renal cancer.HECW1(HECT,C2 And WW Domain Containing E3 Ubiquitin Protein Ligase 1),also known as NEDL1,is a member of E3 Ubiquitin Ligase HECT family.Ubiquitin proteins exist widely in integrated cells and can be linked to each other through enzymatic reactions to mediate the degradation of target proteins,which is called the ubiquitination degradation pathway of proteins.The ubiquitination pathway mainly involves three kinds of ubiquitination enzymes:ubiquitin activase E1,ubiquitin binding enzyme E2 and ubiquitin ligase E3.Among them,ubiquitin ligase E3 can recognize the target protein that needs to be ubiquitinated and catalyze the transfer of ubiquitin molecules from ubiquitin binding enzyme E2 to the target protein,making this system with substrate specificity.A large number of different ubiquitin ligases E3 exist in the human body.Ubiquitin plays an important role in regulating protein function,and the loss of ubiquitin system leads to a variety of human diseases.Previous studies have reported that HECW1 is highly expressed in neuronal cells and can promote the ubiquitination of mutant SUPERoxide dismutase-1(SUPEROXIDE dismutase-1),leading to neurodegenerative diseases.On the other hand,HECW1 can promote the transcription of wild-type p53 gene and enhance the role of wild-type tumor suppressor gene p53 in promoting tumor cell apoptosis.In addition,THYROID transcription factor-1(TTF-1)is closely related to thyroid function and normal function of lung epithelial cells,and its specific site ubiquitination and degradation is also dependent on HECW1.In recent years,a number of studies have demonstrated the role of HECW1 in a variety of tumors,but the effect of HECW1 on the biological function of renal cancer has not been reported at home and abroad.In the early stage,the research group led by my tutor used data-independent Acquisition mass spectrometry(DIA-MS)to analyze and screen samples of 230 patients with renal clear cell carcinoma.HECW1 expression was significantly down-regulated in ccRCC samples.Combined with previous literature reports and previous studies of our group,HECW1 may play an important role in the occurrence and development of renal cancer.ObjectiveTo explore the effect of HECW1 on the biological function of renal carcinoma.To explore the expression difference of HECW1 in renal cancer tissues and cell lines,and to establish an evaluation system for predicting the prognosis of renal cancer patients by HECW1 expression level.MethodsTo explore the expression difference of HECW1 in renal carcinoma and normal tissues,as well as its expression difference in paired renal carcinoma/paracancer tissues,by using TCGA data analysis.Qrt-pcr was used to compare the RNA expression of HECW1 in renal cancer and its adjacent tissues.The cell lines 786-O and 769-P were transfected with SI-RNA to interfere with the expression of HECW1,or HECW1 was overexpressed in these cell lines by transfection with overexpressed plasmid,and then CCK8 proliferation assay was used to observe the effect of HECW1 on the proliferation of renal cell lines.Transwell assay was used to detect the effect of HECW1 on the migration and invasion of renal carcinoma cells.The effect of HECW1 on apoptosis of renal carcinoma cells was detected by flow cytometry using Annexin V-FITC apoptosis detection kit.Immunohistochemical staining and corresponding H&E staining were performed on the tissue samples collected from patients with renal cancer,and the staining results were quantified by immunohistochemical Score(H-Score).Clinical data and prognostic information of patients with renal cancer were collected during follow-up,combined with immunohistochemical H-score,and statistical analysis(ROC,Kawa-Meier survival curve,PROPORTIONAL risk regression model COX,consistency index C-index)was conducted.To establish a survival prognosis prediction system based on HECW1 expression level in clinical renal cancer patients.ResultsTCGA data showed that the expression level of HECW1 in renal carcinoma tissues(n=508)was lower than that in normal tissues(n=72).The expression level of HECW1 in renal carcinoma tissues was lower than that of its adjacent tissues(n=69).HECW1 mRNA expression was higher in 49 of a total of 60 matched samples of clinical renal cancer patients than in their para-cancer counterparts.HECW1 inhibited the proliferation,invasion and migration of renal carcinoma cells.Promotes apoptosis of renal carcinoma cells.Immunohistochemical staining combined with H-score showed that HECW1 expression was low in renal carcinoma tissues and positively correlated with TNM stage.Low HECW1 expression predicts the progression of renal cancer and is associated with poor prognosis of renal cancer patients.The combination of HECW1 expression level with existing clinicopathological indicators of renal cancer(WHO/ISUP grading,TNM staging and SSIGN score)can more accurately predict the clinical prognosis of patients with renal cancer.ConclusionThrough the collection and analysis of TCGA data from the public database and the detection of tissue samples from clinical patients,this study found that the expression level of HECW1 in renal cancer tissues was lower than that in normal tissues.HECW1 inhibited the proliferation,migration and invasion of renal carcinoma cells and promoted their apoptosis.Immunohistochemical score combined with survival analysis showed that low HECW1 expression predicted poor prognosis in patients with renal cancer.COX regression analysis and C-index consistency index analysis showed that the combination of HECW1 expression level with existing clinicopathological evaluation indicators(WHO/ISUP grading,TNM staging and SSIGN score)could more accurately predict the clinical prognosis of patients with renal cancer.