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Cholangiocarcinoma (CCA) is an aggressive tumor of the biliary epithelium with poor survival that shows limited response to conventional chemotherapy. Increased expression of glucosylceramide synthase (GCS) contributes to drug resistance and the progression of various cancers; the expression profiles of GCS (UGCG) and the genes for glucocerebrosidases 1, 2, and 3 (GBA1, GBA2, and GBA3) were therefore studied in CCA. The biological functions of GCS for cell proliferation and cisplatin sensitivity in CCA were explored. GCS expression was higher in CCA tumor tissues than that of GBA1, GBA2, and GBA3. Verification of GCS expression in 29 paired frozen CCA tissues showed that 8 of 29 cases (27.6%) had high GCS expression. The expression of GCS and GBA2 was induced in CCA cell lines following low-dose cisplatin treatment. Suppression of GCS by either palmitoylamino-3-morpholino-1-propanol (PPMP), GCS knockdown or a combination of the two resulted in reduced cell proliferation. These treatments enhanced the effect of cisplatin-induced CCA cell death, increased the expression of apoptotic proteins and reduced phosphorylation of ERK upon cisplatin treatment. Taken together, inhibition of the GCS increased cisplatin-induced CCA apoptosis via the inhibition of the ERK signaling pathway. Thus, targeting GCS might be a strategy for CCA treatment. [ABSTRACT FROM AUTHOR]

Alteration of mucin-type O-glycosylation is implicated in tumor progression and metastasis of cholangiocarcinoma (CCA). Core 1 β1-3 Galactosyltransferase (C1GALT1) is a primary enzyme that regulates the elongation of core 1-derived mucin-type O-glycans. Dysregulation of C1GALT1 has been documented in multiple cancers and is associated with aberrant core 1 O-glycosylation and cancer aggressiveness; however, the expression of C1GALT1 and its role in CCA progression remains unknown. Our study demonstrated that C1GALT1 was downregulated in CCA tissues at both the mRNA and protein levels. The biological function of C1GALT1 using siRNA demonstrated that suppression of C1GALT1 in the CCA cell lines (KKU-055 and KKU-100) increased CCA progression, evidenced by: (i) Induction of CCA cell proliferation and 5-fluorouracil resistance in a dose-dependent manner; (ii) up-regulation of growth-related genes, ABC transporter genes, and anti-apoptotic proteins; and (iii) an increase in the activation/phosphorylation of AKT and ERK in silencing C1GALT1 cells. We demonstrated that silencing C1GALT1 in CCA cell lines was associated with immature core 1 O-glycosylation, demonstrated by high expression of VVL-binding glycans and down-regulation of other main O-linked glycosyltransferases β1,3-N-acetylglucosaminyltransferase 6 (B3GNT6) and ST6 N-Acetylgalactosaminide Alpha-2,6-Sialyltransferase 1 (ST6GALNAC1) in C1GALT1 knockdown. Our findings demonstrate that down-regulation of C1GALT1 in CCA increases the expression of immature core 1 O-glycan, enhancing CCA progression, including growth and 5-fluorouracil resistance via the activation of the AKT/ERK signaling pathway. [ABSTRACT FROM AUTHOR]

RING finger protein 43 (RNF43) is a ubiquitin E3 ligase that negatively regulates Wnt/β-catenin signalling. Mutation, inactivation and downregulation of RNF43 in cholangiocarcinoma (CCA) are associated with a less favourable prognosis. Since the functional role of RNF43 in CCA has not yet been demonstrated, the present study aimed to assess the effect of its overexpression in mediating CCA suppression via Wnt/β-catenin signalling pathway inhibition. Accordingly, RNF43 was overexpressed, and various malignant phenotypic changes studied, including cell proliferation, cell migration, chemotherapeutic sensitivity and the expression of several Wnt/β-catenin target genes. Overexpression of RNF43 in the CCA cell-line KKU-213B hindered activation of Wnt/β-catenin signalling, evidenced by: i) Accumulation of β-catenin in the cytoplasmic fraction and downregulation of several known Wnt target genes at the mRNA level [AXIN2, survivin (BIRC5), CCND1, MMP-7, c-MYC and ABCB1 (MDR1)]; ii) a reduction of cell proliferation; iii) a significant decrease in KKU-213B cell migration with RNF43 overexpression via upregulation of E-cadherin (CDH1); and iv) a reduction in N-cadherin (CDH2), MMP-2, MMP-7 and MMP-9. In addition, overexpression of RNF43 increased 5-fluorouracil sensitivity and downregulation of ABC transporter genes [including ABCB1 and ABCC1 (MRP1)]. The current results demonstrate a functional role for RNF43 in CCA by: i) Blocking β-catenin nuclear translocation; and ii) the subsequent downregulation of Wnt/β-catenin target genes (the latter being involved in the progression of CCA and chemotherapeutic drug susceptibility). Therefore, the present findings suggest that RNF43 could serve a tumour suppressive role in CCA. [ABSTRACT FROM AUTHOR]