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Free CA125 promotes ovarian cancer cell migration and tumor metastasis by binding Mesothelin to reduce DKK1 expression and activate the SGK3/FOXO3 pathway.

ABSTRACT: Objective: CA125/MUC16 is an O-glycosylated protein that is expressed on the surfaces of ovarian epithelial cells. This molecule is a widely used tumor-associated marker for diagnosis of ovarian cancer. Recently, CA125 was shown to be involved in ovarian cancer metastasis. The purpose of this study was to investigate the mechanism of CA125 during ovarian cancer metastasis. Methods: We analyzed the Oncomine and CSIOVDB databases to determine the expression levels of DKK1 in ovarian cancer. DKK1 expression levels were upregulated or downregulated and applied with CA125 to Transwell and Western blot assays to ascertain the underlying mechanism by which CA125 stimulates cell migration via the SGK3/FOXO3 pathway. Anti-mesothelin antibodies (anti-MSLN) were used to block CA125 stimulation. Then the expression levels of DKK1were tested by enzyme-linked immunosorbent assay (ELISA) to eliminate the blocking effect of anti-MSLN to CA125 stimulation. Xenograft mouse models were used to detect the effects of CA125 and anti-MSLN on ovarian cancer metastasis in vivo. Results: DKK1 levels were downregulated in ovarian tumor tissues according to the analyses of two databases and significantly correlated with FIGO stage, grade and disease-free survival in ovarian cancer patients. DKK1 levels were downregulated by CA125 stimulation in vitro. Overexpression of DKK1 reversed the ability of exogenous CA125 to mediate cell migration by activating the SGK3/FOXO3 signaling pathway. Anti-MSLN abrogated the DKK1 reduction and increased the apoptosis of ovarian cancer cells. The use of anti-MSLN in xenograft mouse models significantly reduced tumor growth and metastasis accelerated by CA125. Conclusions: These experiments revealed that the SGK3/FOXO3 pathway was activated, wherein decreased expression of DKK1 was caused by CA125, which fuels ovarian cancer cell migration. Mesothelin is a potential therapeutic target for the treatment of ovarian cancer metastasis.

SUBMITTER: Huo Q

PROVIDER: S-EPMC7893585 | biostudies-literature | 2021

REPOSITORIES: biostudies-literature

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Free CA125 promotes ovarian cancer cell migration and tumor metastasis by binding Mesothelin to reduce DKK1 expression and activate the SGK3/FOXO3 pathway.

Huo Qianyu Q Xu Chen C Shao Yanhong Y Yu Qin Q Huang Lunhui L Liu Yunde Y Bao Huijing H

International journal of biological sciences 20210114 2

<b>Objective:</b> CA125/MUC16 is an O-glycosylated protein that is expressed on the surfaces of ovarian epithelial cells. This molecule is a widely used tumor-associated marker for diagnosis of ovarian cancer. Recently, CA125 was shown to be involved in ovarian cancer metastasis. The purpose of this study was to investigate the mechanism of CA125 during ovarian cancer metastasis. <b>Methods:</b> We analyzed the Oncomine and CSIOVDB databases to determine the expression levels of DKK1 in ovarian ...[more]

PMID: 33613114

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Project description:The mucin MUC16 and the glycosylphosphatidylinositol anchored glycoprotein mesothelin likely facilitate the peritoneal metastasis of ovarian tumors. The biochemical basis and the kinetics of the binding between these two glycoproteins are not clearly understood. Here we have addressed this deficit and provide further evidence supporting the role of the MUC16-mesothelin interaction in facilitating cell-cell binding under conditions that mimic the peritoneal environment.In this study we utilize recombinant-Fc tagged human mesothelin to measure the binding kinetics of this glycoprotein to MUC16 expressed on the ovarian tumor cell line OVCAR-3. OVCAR-3 derived sublines that did not express MUC16 showed no affinity for mesothelin. In a flow cytometry-based assay mesothelin binds with very high affinity to the MUC16 on the OVCAR-3 cells with an apparent Kd of 5-10 nM. Maximum interaction occurs within 5 mins of incubation of the recombinant mesothelin with the OVCAR-3 cells and significant binding is observed even after 10 sec. A five-fold molar excess of soluble MUC16 was unable to completely inhibit the binding of mesothelin to the OVCAR-3 cells. Oxidation of the MUC16 glycans, removal of its N-linked oligosaccharides, and treatment of the mucin with wheat germ agglutinin and erythroagglutinating phytohemagglutinin abrogates its binding to mesothelin. These observations suggest that at least a subset of the MUC16-asscociated N-glycans is required for binding to mesothelin. We also demonstrate that MUC16 positive ovarian tumor cells exhibit increased adherence to A431 cells transfected with mesothelin (A431-Meso+). Only minimal adhesion is observed between MUC16 knockdown cells and A431-Meso+ cells. The binding between the MUC16 expressing ovarian tumor cells and the A431-Meso+ cells occurs even in the presence of ascites from patients with ovarian cancer.The strong binding kinetics of the mesothelin-MUC16 interaction and the cell adhesion between ovarian tumor cells and A431-Meso+ even in the presence of peritoneal fluid strongly support the importance of these two glycoproteins in the peritoneal metastasis of ovarian tumors. The demonstration that N-linked glycans are essential for mediating mesothlein-MUC16 binding may lead to novel therapeutic targets to control the spread of ovarian carcinoma.

Project description:Super Enhancers are a class of DNA Cis-regulatory elements that can regulate cell identity, cell fate, stem cell pluripotency, and even tumorigenesis.Increasing evidence shows that epigenetic modifications play an important role in the pathogenesis of various types of cancer. However, current research is far from sufficient to reveal the complex mechanisms behind epigenetic modifications.The aim of this study is to explore the function and mechanism of a new Super Enhancer in Pancreatic ductal adenocarcinoma (PDAC) tumors. By using epigenetic modification and chromatin interaction information, we found a new Super Enhancer enriched with abnormal active histone modifications in PDAC tumors and cells, called DKK1-super-enhancer (DKK1-SE).DKK1-SE was grouped according to the activity modification information, and the e1 component was identified as the main active unit by dual luciferase reporter gene system and dCas9-KRAB system.DNA motif analysis and core site deletion assay revealed that the e1 component possessed AP1 transcription factor family binding sites.Mechanically, AP1 activates DKK1 transcriptional activity by recruiting JUND and FOSL2 to induce enhancer chromatin remodeling.CRISPR/ Cas9-mediated deletion of DKK1-SE resulted in significant downregulation of its target gene DKK1.Analysis of TCGA data of PDAC tumors combined with immunohistochemical results of PDAC patients demonstrated that DKK1 was closely related to malignant clinical features of PDAC.Bioinformatics analysis of RNA-seq data showed that DKK1 was mainly related to tumor-related biological functions such as extracellular matrix, cell adhesion, angiogenesis, and cell proliferation.Cell malignant phenotype assay showed that deletion or interference of DKK1-SE significantly inhibited the proliferation, colony formation, migration and invasion of PANC-1, and these phenomena were partially alleviated after restoring DKK1 expression in DKK1-SE deficient cells.Experiments of Subcutaneous transplantation tumor in nude mice and Orthotopic transplantation tumor model showed that DKK1-SE deletion not only inhibited tumor proliferation, but also reduced the complexity of tumor microenvironment, resulting in less angiogenesis of tumor tissues and reduced fibrosis degree.In conclusion, DKK1-SE drives the expression of DKK1 by recruiting AP1 family transcription factors, which plays a carcinogenic role in PDAC tumors and enhances the complexity of the tumor microenvironment.

Dataset Information

Free CA125 promotes ovarian cancer cell migration and tumor metastasis by binding Mesothelin to reduce DKK1 expression and activate the SGK3/FOXO3 pathway.

Publications

Free CA125 promotes ovarian cancer cell migration and tumor metastasis by binding Mesothelin to reduce DKK1 expression and activate the SGK3/FOXO3 pathway.

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