RESUMO
Cholangiocarcinoma (CCA) is often diagnosed late, leading to incomplete tumor removal, drug resistance and reduced chemotherapy efficacy. Curcumin has the potential for anti-cancer activity through various therapeutic properties and can improve the efficacy of chemotherapy. We aimed to investigate the synergistic effect of a combination of curcumin and gemcitabine against CCA, targeting the LAT2/glutamine pathway. This combination synergistically suppressed proliferation in gemcitabine-resistant CCA cells (KKU-213BGemR). It also resulted in a remarkable degree of CCA cell apoptosis and cell cycle arrest, characterized by a high proportion of cells in the S and G2/M phases. Knockdown of SLC7A8 decreased the expressions of glutaminase and glutamine synthetase, resulting in inhibited cell proliferation and sensitized CCA cells to gemcitabine treatment. Moreover, in vivo experiments showed that a combination curcumin and gemcitabine significantly reduced tumor size, tumor growth rate and LAT2 expression in a gemcitabine-resistant CCA xenograft mouse model. Suppression of tumor progression in an orthotopic CCA hamster model provided strong support for clinical application. In conclusion, curcumin synergistically enhances gemcitabine efficacy against gemcitabine-resistant CCA by induction of apoptosis, partly via inhibiting LAT2/glutamine pathway. This approach may be an alternative strategy for the treatment of gemcitabine-resistant in CCA patients.
Assuntos
Apoptose , Proliferação de Células , Colangiocarcinoma , Curcumina , Desoxicitidina , Resistencia a Medicamentos Antineoplásicos , Sinergismo Farmacológico , Gencitabina , Glutamina , Desoxicitidina/análogos & derivados , Desoxicitidina/farmacologia , Animais , Colangiocarcinoma/tratamento farmacológico , Colangiocarcinoma/metabolismo , Colangiocarcinoma/patologia , Humanos , Curcumina/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Camundongos , Glutamina/metabolismo , Linhagem Celular Tumoral , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto , Transdução de Sinais/efeitos dos fármacos , Neoplasias dos Ductos Biliares/tratamento farmacológico , Neoplasias dos Ductos Biliares/metabolismo , Neoplasias dos Ductos Biliares/patologia , Glutaminase/metabolismo , Glutaminase/antagonistas & inibidores , MasculinoRESUMO
BACKGROUND/AIM: Forkhead box O4 (FOXO4) has been demonstrated to be a tumor suppressor and proposed as target for treatment of a variety of cancer types. However, the role of FOXO4 in cholangiocarcinoma (CCA), a dangerous cancer of bile-duct epithelium, has rarely been explored. MATERIALS AND METHODS: The proliferative rate of CCA cell lines KKU-213B, KKU-055 and KKK-D068 was investigated using the sulforhodamine B (SRB) assay. Levels of FOXO4, cyclin E1 (CCNE1), CCNE2, cyclin-dependent kinase 2 (CDK2) and cell division cycle 25A (CDC25A) expression were measured using reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR). The cell-cycle profile was explored using flow cytometry. RESULTS: The SRB assay demonstrated that KKU-213B expressed low levels of FOXO4 but its proliferative rate was highest of all cell lines tested. Interestingly, ectopic expression of FOXO4 significantly suppressed proliferation of KKU-213B cells. Cell-cycle analysis revealed that the cell population in the G0/G1 phase was significantly higher in FOXO4-transfected KKU-213B cells than in controls. RT-qPCR analysis demonstrated that the levels of expression of genes that play a role in the G1/S transition, namely CCNE1, CCNE2, CDK2 and CDC25A, were significantly lower in FOXO4-transfected KKU-213B cells compared to controls. CONCLUSION: FOXO4 suppressed CCA cell proliferation partly via down-regulating the expression of genes involved in the G1/S transition, leading to G0/G1 arrest. Our findings suggest that induction of FOXO4 expression might be an alternative approach for the treatment of CCA.
Assuntos
Neoplasias dos Ductos Biliares/genética , Proteínas de Ciclo Celular/genética , Colangiocarcinoma/genética , Fatores de Transcrição Forkhead/genética , Pontos de Checagem da Fase G1 do Ciclo Celular/genética , Expressão Gênica , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Expressão Ectópica do Gene , Fatores de Transcrição Forkhead/metabolismo , HumanosRESUMO
Cholangiocarcinoma (CCA) is a cancer of bile duct, considered to be an incurable and lethal cancer. High mortality rate of CCA patients is underlined by cancer metastasis, an ability of the cancer cells that spread to secondary organs. Recently, we have identified Anterior Gradient-2 (AGR2), from a pair of non-metastatic/metastatic cell lines (KKU-213/KKU-213L5), as a gene that is highly and specifically upregulated in the metastatic cell line. AGR2 encodes for a disulfide isomerase enzyme, ubiquitously detected in mucus-secreting tissues. Overexpression of AGR2 has been reported in several types of human cancer. Role of the overexpressed AGR2 in cancer is still unclear. Here, we found that upregulation of AGR2 in metastatic CCA cells coincides with an aberrant splicing of AGR2 mRNA, and that isoforms of AGR2 RNA, such as AGR2vE, AGR2vF, and AGR2vH are specific to the metastatic cells. We demonstrated that the AGR2vH isoform enables metastatic-associated phenotypes in CCA cells. Depletion of AGR2vH by an isoform-specific interfering RNA in metastatic KKU-213L5 cell results in significant reduction of cancer cell migration and invasion, and a slight decrease of cell adhesion. Overexpression of AGR2vH in non-metastatic KKU-213 cells promotes cancer cell migration, invasion, adhesion, and moderate cell proliferation. Moreover, we found that expression of a metastasis-associated gene, vimentin, positively correlates with expression of AGR2vH. Our results support the notion that aberrant alternative splicing of AGR2 facilitates an accumulation of the oncogenic AGR2vH isoform, in turn, contributes to the pathogenesis and severity of CCA.
