LOXL1 exerts oncogenesis and stimulates angiogenesis through the LOXL1-FBLN5/αvß3 integrin/FAK-MAPK axis in ICC.

Aberrant expression of lysyl oxidase-like 1 (LOXL1) reportedly leads to fibrous diseases. Recent studies have revealed its role in cancers. In this study, we observed an elevated level of LOXL1 in the tissues and sera of patients with intrahepatic cholangiocarcinoma (ICC) compared with levels in nontumor tissues and sera of unaffected individuals. Overexpression of LOXL1 in RBE and 9810 cell lines promoted cell proliferation, colony formation, and metastasis in vivo and in vitro and induced angiogenesis. In contrast, depletion of LOXL1 showed the opposite effects. We further showed that LOXL1 interacted with fibulin 5 (FBLN5), which regulates angiogenesis, through binding to the αvß3 integrin in an arginine-glycine-aspartic (Arg-Gly-Asp) domain-dependent mechanism and enhanced the focal adhesion kinase (FAK)-mitogen-activated protein kinase (MAPK) signaling pathway inside vascular endothelial cells. Our findings shed light on the molecular mechanism underlying LOXL1 regulation of angiogenesis in ICC development and indicate that the LOXL1-FBLN5/αvß3 integrin/FAK-MAPK axis might be the critical pathological link leading to angiogenesis in ICC.

Role of Sciellin in gallbladder cancer proliferation and formation of neutrophil extracellular traps.

Apart from primary tumor development and metastasis, cancer-associated thrombosis is the second cause of cancer death in solid tumor malignancy. However, the mechanistic insight into the development of gallbladder cancer (GBC) and cancer-associated thrombosis remains unclear. This study aimed to investigate the mechanistic role of Sciellin (SCEL) in GBC cell proliferation and the development of venous thromboembolism. The expression level of SCEL was determined by immunohistochemical staining. Roles of SCEL in gallbladder cancer cell were determined by molecular and cell biology methods. SCEL was markedly upregulated in GBC and associated with advanced TNM stages and a poor prognosis. Furthermore, SCEL interacted with EGFR and stabilized EGFR expression that activates downstream PI3K and Akt pathway, leading to cell proliferation. In addition, SCEL induces tumor cell IL-8 production that stimulates the formation of neutrophil extracellular traps (NETs), accelerating thromboembolism. In xenografts, SCEL-expressing GBCs developed larger tumors and thrombosis compared with control cells. The present results indicate that SCEL promotes GBC cell proliferation and induces NET-associated thrombosis, thus serving as a potential therapeutic target.

Genomic ERBB2/ERBB3 mutations promote PD-L1-mediated immune escape in gallbladder cancer: a whole-exome sequencing analysis.

OBJECTIVES: Patients with gallbladder carcinoma (GBC) lack effective treatment methods largely due to the inadequacy of both molecular characterisation and potential therapeutic targets. We previously uncovered a spectrum of genomic alterations and identified recurrent mutations in the ErbB pathway in GBC. Here, we aimed to study recurrent mutations of genes and pathways in a larger cohort of patients with GBC and investigate the potential mechanisms and clinical significance of these mutations. DESIGN: We performed whole-exome sequencing (WES) in 157 patients with GBC. Functional experiments were applied in GBC cell lines to explore the oncogenic roles of ERBB2/ERBB3 hotspot mutations, their correlation with PD-L1 expression and the underlying mechanisms. ERBB inhibitors and a PD-L1 blocker were used to evaluate the anticancer activities in co-culture systems in vitro and in vivo. RESULTS: WES identified ERBB2 and ERBB3 mutations at a frequency of 7%-8% in the expanded cohort, and patients with ERBB2/ERBB3 mutations exhibited poorer prognoses. A set of in vitro and in vivo experiments revealed increased proliferation/migration on ERBB2/ERBB3 mutation. Ectopic expression of ERBB2/ERBB3 mutants upregulated PD-L1 expression in GBC cells, effectively suppressed normal T-cell-mediated cytotoxicity in vitro through activation of the PI3K/Akt signalling pathway and contributed to the growth and progression of GBC in vivo. Treatment with an ERBB2/ERBB3 inhibitor or a PD-L1 monoclonal antibody reversed these immunosuppressive effects, and combined therapy revealed promising therapeutic activities. CONCLUSIONS: ERBB2/ERBB3 mutations may serve as useful biomarkers in identifying patients who are sensitive to ERBB2/ERBB3 inhibitors and PD-L1 monoclonal antibody treatment. TRIAL REGISTRATION NUMBER: NCT02442414;Pre-results.

Chloride intracellular channel 1 regulates the antineoplastic effects of metformin in gallbladder cancer cells.

Metformin is the most commonly used drug for type 2 diabetes and has potential benefit in treating and preventing cancer. Previous studies indicated that membrane proteins can affect the antineoplastic effects of metformin and may be crucial in the field of cancer research. However, the antineoplastic effects of metformin and its mechanism in gallbladder cancer (GBC) remain largely unknown. In this study, the effects of metformin on GBC cell proliferation and viability were evaluated using the Cell Counting Kit-8 (CCK-8) assay and an apoptosis assay. Western blotting was performed to investigate related signaling pathways. Of note, inhibition, knockdown and upregulation of the membrane protein Chloride intracellular channel 1 (CLIC1) can affect GBC resistance in the presence of metformin. Our data demonstrated that metformin apparently inhibits the proliferation and viability of GBC cells. Metformin promoted cell apoptosis and increased the number of early apoptotic cells. We found that metformin can exert growth-suppressive effects on these cell lines via inhibition of p-Akt activity and the Bcl-2 family. Notably, either dysfunction or downregulation of CLIC1 can partially decrease the antineoplastic effects of metformin while upregulation of CLIC1 can increase drug sensitivity. Our findings provide experimental evidence for using metformin as an antitumor treatment for gallbladder carcinoma.

MYBL2 is a Potential Prognostic Marker that Promotes Cell Proliferation in Gallbladder Cancer.

BACKGROUND: Gallbladder cancer (GBC) is an aggressive and highly lethal biliary tract malignancy, with extremely poor prognosis. In the present study, we analyzed the potential involvement of MYBL2, a member of the Myb transcription factor family, in the carcinogenesis of human GBC. METHODS: MYBL2 expression levels were measured in GBC and cholecystitis tissue specimens using quantitative real-time PCR (qRT-PCR) and immunohistochemical (IHC) assays. The effects of MYBL2 on cell proliferation and DNA synthesis were evaluated using Cell Counting Kit-8 assay (CCK-8), colony formation, and 5-ethynyl-2'-deoxyuridine (EdU) retention assay, flow cytometry analysis, western blot, and a xenograft model of GBC cells in nude mice. RESULTS: MYBL2 expression was increased in GBC tissues and associated with histological differentiation, tumour invasion, clinical stage and unfavourable overall survival in GBC patients. The downregulation of MYBL2 expression resulted in the inhibition of GBC cell proliferation, and DNA replication in vitro, and the growth of xenografted tumours in nude mice. Conversely, MYBL2 overexpression resulted in the opposite effects. CONCLUSIONS: MYBL2 overexpression promotes GBC cell proliferation through the regulation of the cell cycle at the S and G2/M phase transitions. Thus, MYBL2 could serve as a potential prognostic and therapeutic biomarker in GBC patients.