Overexpression of oncogenic spliced-AGR2vH potentiates cholangiocarcinoma cell tumorigenicity with proteomic alterations

Abstract Upregulated expression of Anterior gradient 2 (AGR2) has been observed in cells, highly metastatic mouse models, nras-mutant zebrafish and cholangiocarcinoma (CCA) specimens derived from patients. Our previous study reported that AGR2 splicing into AGR2vH can promote CCA cell metastasis and survivability. This present study aimed to investigate the molecular mechanisms underlying AGR2vH tumorigenicity in vitro and in vivo. AGR2vH was determined in patient tissues and presented the upregulation in CCA tumor tissues compared with matched-normal adjacent tissues. During the in vitro studies, AGR2vH was ectopically overexpressed in KKU-213A cells. Established AGR2vH-overexpressing CCA cells were found to exhibit increased proliferative and clonogenic ability. For in vivo tumorigenicity, a higher tumorigenic potential was identified in AGR2vH-overexpressing cells xenograft mice. Moreover, liquid chromatography-mass spectrometry with protein bioinformatics was used to examine the proteomic alteration. The CCA cell proteome was altered, and it was indicated that AGR2vH may be associated with CCA cell proliferation via the activation of Wnt/β-catenin signaling pathway, which was verified via the comparative immunoblotting of β-catenin in cytoplasmic and nuclear fractionated proteins. These present results provided evidence that the upregulation of AGR2vH promotes the tumorigenicity of CCA cells, which was associated with an alteration of the CCA cell proteome.