The Study Of YBOX-1's Effects In Progression And Prognosis Of Cholangiocarcinoma

BackgroundCholangiocarcinoma(CCA)is a disease entity comprising diverse epithelial tumours,it is a hepatic malignancy with high morbidity.Over the past four decades,the incidence of CCA has increased gradually.The preferred treatment for patients with CCA is surgery.However,only approximately 35% of patients who suffer from early-stage disease can be subjected to surgical resection.Given the aggressiveness of CCA tumor,strong invasiveness,and early metastasis,most patients who suffer from advanced-stage disease at presentation show high mortality.The effectiveness of surgery decreases and prognosis worsens due to lymph node or distant metastases.Less than 20% patients survive for more than 2 years.The available systemic therapies for patients with unresectable or advanced-stage CCA are almost ineffective.Cisplatin and gemcitabine are the standard chemotherapy drugs.The median overall survival of chemotherapy is less than 1 year.Diagnosing patients with earliest-stage asymptomatic CCA is challenging.New biomarkers must be identified because no efficiency marker can be clinically used for CCA.Y-box binding protein-1(YBOX-1)levels are highly correlated with malignancies,such as melanoma,osteosarcoma,and lung and breast cancers.The role of YBOX-1 levels in CCA caught our interest.Cold shock domain(CSD)is a large family of proteins contain broad nucleic acid binding properties.YBOX-1 is the most important member of CSD in oncology.As a nucleic-acid-binding protein,YBOX-1 is implicated in nearly all cellular processes.YBOX-1 also plays an important role in mRNA complexes and microRNA processing.In cancer cells,YBOX-1 plays important roles in translation and other processes,multi-drug resistance,and EMT(epithelial-mesenchymal transition).YBOX-1 could be a marker for predicting poor prognosis and a potential therapeutic target.Despite some reports on the prognostic value of YBOX-1 in oncology,the effects of YBOX-1 in CCA remain indistinct.Objective1.we wish to find out the effect of YBOX-1 in CCA invasion,migration,and proliferation.2.We want to know the relationship between the expression of YBOX-1 and overall survival of 91 resected patients with CCA through the expression of YBOX-1 in tissue samples.Methods1.Through the analysis of protein which bind with GATA6 by mass spectrometry,the tumor related molecules were screened out.YBOX-1 was selected through literature review and pre experiment.The binding of YBOX-1 and RPSA promoter was detected by double luciferase reporter gene.2.The invasiveness capacity of cell was determined by Transwell assay.The migration capacity of cell was determined by wound-healing assay.3.The effect of YBOX-1 on CCA cell in vivo was performed by the subcutaneous xenotransplantation injection of cells into nude mice.After 4 weeks,the mice were sacrificed after transplantation.The weight of masses was measured and stained by H.E..4.We enrolled tissue samples of 91 patients with CCA between 2011 and 2016.IHC was used to investigate the expression of YBOX-1 in tissues of patients with clinical CCA.YBOX-1 was considered weak(+)or strongly positive(++)if less and more than 25% of CCA cells show positive cytoplasmic staining.A total of 91 resected tissue samples and 31 peritumoral samples were collected from 91 patients with CCA.Follow ups were made with all patients for 5 years.The mean age was 56.20 ± 9.83 years.Patients were composed of 43 females and 48 males.Recurrence-free survival time was calculated from the time of surgery to tumor palindromia.The overall survival time was obtained from the date of surgery to the date of death or last contact.Results1.Protein mass spectrometry showed that YBOX-1 was bound to GATA6.Detection of double luciferase reporter gene showed that YBOX-1 combined with RPSA promoter.YBOX-1 regulates the expression of RPSA by combining with GATA6 to form a transcription complex,thus affecting the biological characteristics of cholangiocarcinoma cells.2.In the Transwell assay,the number of QBC939 sh-YBOX-1 cells is less than QBC939 and QBC939 sh-Control.The number of RBE ex-YBOX-1 cells increased.In wound-healing assay,the established knockdown of YBOX-1 protein cell strain(QBC939 sh-YBOX-1)decreased.RBE ex-YBOX-1 cells show an increased migration pattern compared with the RBE and RBE ex-Control.The result showed the important role of YBOX-1 in CCA cell invasion and migration.3.The weight of masses shows less proliferation of QBC939 sh-YBOX-1 than QBC939 sh-Control.RBE ex-YBOX-1 cells showed remarkably increased masses compared with the RBE ex-Control cells.The results confirmed that YBOX-1 promoted CCA proliferation.4.A total of 55 of CCA cancerous tissues(60.4%)showed strongly positive YBOX-1 cytoplasmic staining,whereas a total of 36(39.6%)show weak positive staining.A total of 31 matched paracancerous samples show negative cytoplasmic YBOX-1 expression.Strongly positive YBOX-1 expression is associated with poor overall survival(P = 0.02)and early recurrence(P = 0.03).ConclusionYBOX-1 overexpression plays an important role in enhancing tumor growth,migration,and invasion in CCA.YBOX-1 is correlated with poor overall survival and early recurrence in CCA.The YBOX-1 may be an important factor of overall survival and a promising pathological marker for predicting poor prognosis.It could be a target for prognostic in the future.