The Function And Phenotype Of DAND5 In Pan-cancer And Gastric Cancer

Part 1 Insights into the oncogenic,prognostic and immunological role of DAND5 in pan-cancer:A comprehensive data-mining based studyObjectivesDAND5,as a secreted protein,is an antagonist of TGF-β,Nodal and Wnt signaling pathway.It plays an important role in regulating organogenesis and tissue differentiation.Up to now,several studies have elucidated its role in breast cancer and colorectal cancer.However,its role in pan-cancer remains largely unknown.This study aims to conduct a comprehensive data-mining based analysis to further explore the oncogenic,prognostic and immunological role of DAND5 in pan-cancer.MethodsDAND5 expression data of tumor and normal tissues as well as related clinical data were downloaded from The Cancer Genome Atlas(TCGA),Genotype-Tissue Expression(GTEx),Human Protein Atlas(HPA),Tumor Immune Evaluation Database(TIMER),cBioPortal and GeneMANIA databases.Correlation between DAND5 expression and prognosis,genomic alterations as well as RNA modification were analyzed.Protein-Protein interaction(PPI)network was performed to identify the potential pathways and functions of DAND5.Besides,DAND5 correlation with tumor microenvironment(TME),tumor infiltrates immune cells(TIICs)and immune-related genes was also explored in pan-cancer.ResultsDifferential analyses showed an up-regulation of DAND5 expression in 19 cancer types and a down-regulation of DAND5 expression in 11 cancer types.Its aberrant expression could be an indicator for overall survival in 9 cancer types.Among the various mutation types of DAND5 in pan-cancer,amplification was the most common type.DAND5 expression had a significant correlation with most of the RNA modified genes in pan-cancer and was involved in the regulation of a variety of signaling pathways.In addition,the expression of DAND5 and its correlation with TME,tumor immune-infiltrating cells(TIICs)and immune-related genes was also confirmed.ConclusionThe study indicates that DAND5 plays an imperative role in the tumorigenesis and immunity of various tumors.It may not only serve as a potential diagnostic and prognostic biomarker,but also can be a novel potential target for predicting the effect of immunotherapy.Part 2 The expression level of DAND5 and its relationship with related clinical features in gastric cancer patientsObjectivesIn this study,immunohistochemistry(IHC)staining was conducted to clarify the expression of DAND5 in paired tumor and para-tumor tissues of intestinal gastric cancer(GC)patients.Relationships between expression of DAND5 in GC and related clinical features were further explored.MethodsA total of 48 consecutive patients with intestinal GC who directly underwent surgery without cancer-related treatment before the operation from January to December,2017 were enrolled in the study.IHC staining and scoring evaluation of their paired tumor and para-tumor tissue samples were carried out.Related demographic and clinical data were collected via the hospital’s medical electronic database(Donghua Hospital Information System,HIS)and retrospectively analyzed.ResultsDAND5,with a granular staining,was mainly located in the cytoplasm,consisting with its secreted characteristics.The expression of DAND5 in GC tissues was significantly higher than that of the paired para-tumor tissues.High expression level of DAND5 was significantly correlated with lymph node metastasis.However,there was no significant correlation between DAND5 and gender,age,smoking or alcohol consumption condition,tumor location,tumor size,ulcerative lesion,differentiation,tumor infiltration depth,metastasis,tumor stage or increased level of other tumor biomarkers.ConclusionExpression of DAND5 is up-regulated in intestinal GC and DAND5 can be regarded as a novel potential biomarker in the diagnosis of intestinal GC.Higher expression level of DAND5 is significantly correlated with lymph node metastasis and attention should be paid to lymph node dissection when GC patients with high expression level of DAND5 is in face of choosing treatment options.Part 3 DAND5 and its function in gastric cell lines——an in vitro studyObjectivesBy carrying out in vitro experiment,function of DAND5 in GC cell lines including proliferation,migration and apoptosis was investigated.MethodsAfter the quality control by performing RT-qPCR to confirm the knockdown and overexpression of DAND5 in two GC cell lines,the functional phenotype(proliferation,migration and apoptosis)of DAND5 in GC was determined by carrying out in vitro experiments,including CCK8,colony formation assay,transwell and flow cytometry.ResultsIn GC cell lines(AGS and HGC27 cell line),the ability of cell proliferation,colony formation and cell migration were significantly inhibited after the knockdown of DAND5 while the apoptotic cells were significantly increased.On the contrary,after the overexpression of DAND5,cell proliferation,colony formation and cell migration were significantly enhanced while the apoptotic cells were significantly reduced.ConclusionDAND5 is a potential oncogene in GC as it can promote the proliferation and migration of GC cells while inhibit the apoptosis of GC cells.The carcinogenic mechanism of DAND5 in GC needs to be further clarified in vitro and in vivo.