Study On The Enhanced Cancer Stem Cell Properties And Chemoresistance Of Ovarian Cancer Cells Induced By NID1

Nidogen-1(NID1)has been identified as a novel candidate diagnostic biomarker of ovarian cancer in our previous study.We have found that NID1 was a mesenchymal-associated protein,of which the high expression level indicated the poor prognosis of ovarian cancer patients.The subsequently functional analysis confirmed that NID1 induced the partial epithelial-mesenchymal transition(EMT)in ovarian cancer cells by activating ERK/MAPK signaling pathway and thus enhanced the invasive and migratory capabilities of ovarian cancer cells.It has been well known that EMT is closely related to cancer stem cell(CSC).Sepcifically,EMT remodels the phenotypes of cancer cells,such as the loss of epithelial traits and the acquirement of mesenchymal traits and CSC characteristics,thereby increasing their abilities of invasiveness,migration,anti-apoptosis and chemoresistance.For this,we speculated that NID1 probably induced the CSC status of ovarian cancer cells via EMT,followed by the enhancement of the migration and chemoresistance of these cells.In the present study,we not only focus on the potential effects of NID1 on theCSC characteristics and chemoresistances of ovarian cancer cells,but also the prognostic value of NID1 combined with the CSC markers in ovarian cancer.Collectively,this study will shed light on the development of anticancer therapy targeting both NID1 and CSC.Part ?: Effects of NID1 on the CSC status of ovarian cancer cells.The self-renewal ability of OVCAR-3-NID1-MC cells with stable etopic expression of NID1 was detected by tumor sphere formation assay.It revealed that the number and the diameter of tumor spheres formed by OVCAR-3-NID1-MC cells were both significantly greater,compared to those of tumor spheres formed by control cells.Accordingly,the expression levels of CD44 and ABCG2(two well-known CSC markers of ovarian cancer)were remarkably increased in OVCAR-3-NID1-MC cells.These results indicated that NID1 might promote the entrance of OVCAR-3 cells into the CSC status.Part ?: Effects of NID1 on cisplatin resistance in ovarian cancer cells.One gene expression microarray dataset(GSE51373)concerning the cisplatin resistance in ovarian cancer was downloaded from the public GEO database.We found that the expression level of NID1 in the resistant group was obviously higher than that in the sensitive group,and its level was positively correlated with the expression of MDR1 and ABCG2(two well-characterized drug efflux transporters).Moreover,OVCAR-3-NID1-MC cells indeed showed higher cisplatin resistantance andcorrespondingly higher expression levels of MDR1 and ABCG2.Conversely,the cisplatin-resistance of NID1-silencing HEY cells was slightly decreased,and their MDR1 and ABCG2 expression levels were reduced as well.These results suggested that NID1 might enhance the cisplatin resistance of ovarian cancer cells.Part ?: Effects of NID1 on CSC status,motility and cisplatin resistance of ovarian cancer cells were dependent on ERK/MAPK pathway.We treated OVCAR-3-NID1-MC cells with an ERK/MAPK inhibitor U0126.Inhibition of ERK1/2 phosphorylation results in less malignant phenotypes of OVCAR-3-NID1-MC cells,including reduced levels of CD44 and ABCG2 weaker migratory capability and more sensitivity to cisplatin.These results demonstrated that NID1 activates ERK/MAPK pathway whereby it contributes to the CSC status of ovarian cancer cells and thus their abilities of migration and cisplatin resistance.Part ?: NID1-mediated EMT in ovarian cancer cells was independent on integrin-FAK signaling pathway.We previously found that integrin-FAK pathway was activated in OVCAR-3-NID1-MC cells.Here we treated OVCAR-3-NID1-MC cells with a FAK inhibitor PF573228,resulting in inhibition of ERK/MAPK signaling pathway without the alteration of EMT-relevant markers.These results revealed that integrin-FAK pathway didn't involve in the NID1-mediated EMT in ovarian cancer cells,although it indeed was an upstream signaling ofERK/MAPK pathway.Part ?: NID1 and CSC markers acted as the combinatorial prognostic pattern in ovarian cancer.Two gene expression microarray datasets(GSE26712 and GSE14764)based on the tumor tissues from ovarian cancer were downloaded from the public GEO database.We found that the NID1 level was positively correlated with the expression level of CD44 and ABCG2.Furthermore,the overall survival of ovarian cancer patients harboring both of high NID1 level and positive CSC markers was shorter than that of other ovarian cancer patients.These results implied that NID1 and CSC markers acted as the combinatorial prognostic pattern in ovarian cancer.As the extension of the previous work concerning ‘NID1 induce EMT through the ERK/MAPK pathway in ovarian cancer cells',this study further demonstrated that NID1 might contribute to the CSC characteristics and the cisplatin resistance of ovarian cancer cells via ERK/MAPK pathway as well.Therefore,we speculated that NID1 stimulated ERK/MAPK pathway to induce EMT of ovarian cancer cells,and thus enabled them entering the CSC status,accompanied by the greater abilities of migration and chemoresistance.Additionally,the combinatorial prognostic pattern of NID1 and CSC markers provided an rationale for the development of novel NID1-and CSC-targeted therepies in ovarian cancer.