Establishment and characterization of a novel human cholangiocarcinoma cell line with high metastatic activity

Uthaisar, Kwuntida; Vaeteewoottacharn, Kulthida; Seubwai, Wunchana; Talabnin, Chutima; Sawanyawisuth, Kanlayanee; Obchoei, Sumalee; Kraiklang, Ratthaphol; Okada, Seiji; Wongkham, Sopit

doi:10.3892/or.2016.4974

Cited by 28 publications

(30 citation statements)

References 40 publications

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“…KKU-213 and KKU-214, the CCA cell lines were provided from the Japanese Collection of Research Bioresources (JCRB) Cell Bank, Osaka, Japan. Highly metastatic CCA sublines; KKU-213L5 and KKU-214L5 were established from the parental cells, KKU-213 [ 49 ] and KKU-214 [ 50 ]. All cells were cultured in HAM’s F-12 (Gibco, NY) containing 10% fetal bovine serum (Gibco) and 1% antibiotic-antimycotic (Gibco), under the standard protocol at 37 °C and 5% CO 2 .…”

Section: Methodsmentioning

confidence: 99%

O-GlcNAcylation mediates metastasis of cholangiocarcinoma through FOXO3 and MAN1A1

et al. 2018

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The leading cause of death in cancer patients is metastasis, for which an effective treatment is still necessary. During metastasis, cancer cells aberrantly express several glycans that are correlated with poor patient outcome. This study was aimed toward exploring the effects of O-GlcNAcylation on membranous N-glycans that are associated with the progression of cholangiocarcinoma (CCA). Global O-GlcNAcylation in CCA cells was depleted using specific siRNA against O-GlcNAc transferase (OGT), which transfers GlcNAc to the acceptor proteins. Using an HPLC-Chip/Time-of-Flight (Chip/TOF) MS system, the N-glycans associated with O-GlcNAcylation were identified by comparing the membranous N-glycans of siOGT-treated cells with those of scramble siRNA-treated cells. In parallel, the membranous N-glycans of the parental cells (KKU-213 and KKU-214) were compared with those of the highly metastatic cells (KKU-213L5 and KKU-214L5). Together, these data revealed that high mannose (Hex9HexNAc2) and biantennary complex (Hex5HexNAc4Fuc1NeuAc1) N-linked glycans correlated positively with metastasis. We subsequently demonstrate that suppression of O-GlcNAcylation decreased the expression of these two N-glycans, suggesting that O-GlcNAcylation mediates their levels in CCA. In addition, the ability of highly metastatic cells to migrate and invade was reduced by the presence of Pisum Sativum Agglutinin (PSA), a mannose-specific lectin, further indicating the association of high mannose type N-glycans with CCA metastasis. The molecular mechanism of O-GlcNAc-mediated progression of CCA was shown to proceed via a series of signaling events, involving the activation of Akt/Erk (i), an increase in FOXO3 phosphorylation (ii), which results in the reduction of MAN1A1 expression (iii) and thus the accumulation of Hex9HexNAc2 N-glycans (iv). This study demonstrates for the first time the association between O-GlcNAcylation, high mannose type N-glycans, and the progression of CCA metastasis, suggesting a novel therapeutic target for treatment of metastatic CCA.

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Section: Methodsmentioning

confidence: 99%

O-GlcNAcylation mediates metastasis of cholangiocarcinoma through FOXO3 and MAN1A1

et al. 2018

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“…Cells were obtained from the Japanese Collection of Research Bioresources (JCRB) Cell Bank, Osaka, Japan. The highly metastatic CCA sublines; KKU-213L5 and KKU-214L5 were established from the parental cells, KKU-213 and KKU-214, as described elsewhere36.…”

Section: Methodsmentioning

confidence: 99%

High glucose levels boost the aggressiveness of highly metastatic cholangiocarcinoma cells via O-GlcNAcylation

Phoomak

Vaeteewoottacharn

Silsirivanit

et al. 2017

Sci Rep

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Increased glucose utilization is a feature of cancer cells to support cell survival, proliferation, and metastasis. An association between diabetes mellitus and cancer progression was previously demonstrated in cancers including cholangiocarcinoma (CCA). This study was aimed to determine the effects of high glucose on protein O-GlcNAcylation and metastatic potentials of CCA cells. Two pairs each of the parental low metastatic and highly metastatic CCA sublines were cultured in normal (5.6 mM) or high (25 mM) glucose media. The migration and invasion abilities were determined and underlying mechanisms were explored. Results revealed that high glucose promoted migration and invasion of CCA cells that were more pronounced in the highly metastatic sublines. Concomitantly, high glucose increased global O-GlcNAcylated proteins, the expressions of vimentin, hexokinase, glucosamine-fructose-6-phosphate amidotransferase (GFAT) and O-GlcNAc transferase of CCA cells. The glucose level that promoted migration/invasion was shown to be potentiated by the induction of GFAT, O-GlcNAcylation and an increase of O-GlcNAcylated vimentin and vimentin expression. Treatment with a GFAT inhibitor reduced global O-GlcNAcylated proteins, vimentin expression, and alleviated cell migration. Altogether, these results suggested the role of high glucose enhanced CCA metastasis via modulation of O-GlcNAcylation, through the expressions of GFAT and vimentin.

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“…To establish a highly metastatic CCA subpopulation, cells that formed lung metastasis were isolated after five successive i.v. passages into NOD/scid/Jak3 null (NOJ) mice (26). This protocol was performed with two types of intrahepatic CCA with different aggressive scales (Fig.…”

Section: Increases In Extended High-mannose Glycans Are Metastasis-spmentioning

confidence: 99%

Metastasis of cholangiocarcinoma is promoted by extended high-mannose glycans

Park

Phoomak

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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Membrane-bound oligosaccharides form the interfacial boundary between the cell and its environment, mediating processes such as adhesion and signaling. These structures can undergo dynamic changes in composition and expression based on cell type, external stimuli, and genetic factors. Glycosylation, therefore, is a promising target of therapeutic interventions for presently incurable forms of advanced cancer. Here, we show that cholangiocarcinoma metastasis is characterized by down-regulation of the Golgi α-mannosidase I coding geneMAN1A1, leading to elevation of extended high-mannose glycans with terminating α-1,2-mannose residues. Subsequent reshaping of the glycome by inhibiting α-mannosidase I resulted in significantly higher migratory and invasive capabilities while masking cell surface mannosylation suppressed metastasis-related phenotypes. Exclusive elucidation of differentially expressed membrane glycoproteins and molecular modeling suggested that extended high-mannose glycosylation at the helical domain of transferrin receptor protein 1 promotes conformational changes that improve noncovalent interaction energies and lead to enhancement of cell migration in metastatic cholangiocarcinoma. The results provide support that α-1,2-mannosylatedN-glycans present on cancer cell membrane proteins may serve as therapeutic targets for preventing metastasis.

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Establishment and characterization of a novel human cholangiocarcinoma cell line with high metastatic activity

Cited by 28 publications

References 40 publications

O-GlcNAcylation mediates metastasis of cholangiocarcinoma through FOXO3 and MAN1A1

O-GlcNAcylation mediates metastasis of cholangiocarcinoma through FOXO3 and MAN1A1

High glucose levels boost the aggressiveness of highly metastatic cholangiocarcinoma cells via O-GlcNAcylation

Metastasis of cholangiocarcinoma is promoted by extended high-mannose glycans

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