RESUMO
BACKGROUND: Tumor resistance is a global challenge for tumor treatment. Cancer stem cells (CSCs) are the main population of tumor cells for drug resistance. We have reported that high aldehyde dehydrogenase (ALDH) activity represents a functional marker for cervical CSCs. Here, we aimed at disulfiram (DSF), an ALDH inhibitor, that has the potential to be used for cervical cancer treatment. METHODS: MTT assay, western blot, vector construction and transfection, cell sorting and in vivo anti-tumor assays were performed using cervical cancer cell lines SiHa and HeLa. Cell cycle distribution and cell apoptosis were carried out by flow cytometry. The cytotoxicity of DSF was detected by MTT assay and cervical cancer xenograft models. RESULTS: DSF was cytotoxic to cervical cancer cell lines in a copper (Cu)-dependent manner. Disulfiram/copper (DSF/Cu) complex induced deregulation of S-phase and inhibited the expression of stemness markers in cervical cancer cells. Furthermore, DSF/Cu could also reduce the cancer stem cell-like LGR5+ cells which lead to cisplatin resistance in cervical cancer cells. DSF/Cu complex had the greater antitumor efficacy on cervical cancer than cisplatin in vitro and in vivo. CONCLUSION: Our findings indicate that the cytotoxicity of DSF/Cu complex may be superior to cisplatin because of targeting LGR5-positive cervical cancer stem-like cells in cervical cancer. Thus, the DSF/Cu complex may represent a potential therapeutic strategy for cervical cancer patients.
Assuntos
Antineoplásicos , Cobre , Dissulfiram , Neoplasias do Colo do Útero , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Cisplatino/farmacologia , Cobre/farmacologia , Dissulfiram/farmacologia , Feminino , Humanos , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Neoplasias do Colo do Útero/patologiaRESUMO
Cancer stem cells (CSCs), also known as tumor-initiating cells, contribute to tumorigenesis, resistance to chemoradiotherapy and recurrence in human cancers, suggesting targeting CSCs may represent a potential therapeutic strategy. Leucine-rich repeat-containing G-protein-coupled receptor 5 (LGR5) has recently been found to be a bona fide marker of colorectal CSCs. Our previous study showed that LGR5 functions as a tumor promoter in cervical cancer by activating the Wnt/ß-catenin pathway. However, very little is known about the function or contribution of LGR5 to cervical CSCs. Here, we have modulated the expression of LGR5 using an overexpression vector or short hairpin RNA in cervical cancer cell lines. We demonstrated that elevated LGR5 expression in cervical cancer cells increased tumorsphere-forming efficiency; conferred chemoresistance to cisplatin treatment; augmented cell migration, invasion and clonogenicity; and elevated the levels of stem cell-related transcription factors in vitro. Furthermore, modulated LGR5+ cells, unlike LGR5- cells, were highly tumorigenic in vivo. In addition, the modulated LGR5+ cells could give rise to both LGR5+ and LGR5- cells in vitro and in vivo, thereby establishing a cellular hierarchy. Finally, we found that the increased tumorsphere-forming efficiency induced by LGR5 could be regulated through the inhibition or activation of the Wnt/ß-catenin pathway in cervical cancer cells. Taken together, these results indicate that LGR5 has a vital oncogenic role by promoting cervical CSC traits and may represent a potential clinical target.
Assuntos
Resistencia a Medicamentos Antineoplásicos/genética , Regulação Neoplásica da Expressão Gênica , Células-Tronco Neoplásicas/metabolismo , Receptores Acoplados a Proteínas G/genética , Neoplasias do Colo do Útero/genética , Animais , Antígenos CD , Antineoplásicos/farmacologia , Caderinas/genética , Caderinas/metabolismo , Morte Celular/efeitos dos fármacos , Morte Celular/genética , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Cisplatino/farmacologia , Feminino , Genes Reporter , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Células HeLa , Humanos , Camundongos , Camundongos Endogâmicos NOD , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/patologia , Receptores Acoplados a Proteínas G/metabolismo , Transdução de Sinais , Esferoides Celulares/efeitos dos fármacos , Esferoides Celulares/metabolismo , Esferoides Celulares/patologia , Análise de Sobrevida , Carga Tumoral/efeitos dos fármacos , Neoplasias do Colo do Útero/tratamento farmacológico , Neoplasias do Colo do Útero/mortalidade , Neoplasias do Colo do Útero/patologia , Vimentina/genética , Vimentina/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Leucine-rich repeat-containing G protein-coupled receptor 5 (LGR5), a seven transmembrane receptor known as a potential stem cell marker for intestinal crypts and hair follicles, has recently been found to be overexpressed in some types of human cancers. However, the role of LGR5 in cervical cancer remains unclear. In this study, the expression of LGR5 gradually increases from normal cervix to cervical cancer in situ and to cervical cancers as revealed by immunohistochemistry and western blot analyses. Through knocking down or overexpressing LGR5 in SiHa and HeLa cells, the expression level of LGR5 was found to be positively related to cell proliferation in vitro and to tumor formation in vivo. Further investigation indicated that LGR5 protein could significantly promote the acceleration of cell cycle. Moreover, the TOP-Flash reporter assay and western blot for ß-catenin, cyclinD1, and c-myc proteins, target genes of the Wnt/ß-catenin pathway, indicated that LGR5 significantly activated Wnt/ß-catenin signaling. Additionally, the blockage of Wnt/ß-catenin pathway resulted in a significant inhibition of cell proliferation induced by LGR5. Taken together, these results demonstrate that LGR5 can promote proliferation and tumor formation in cervical cancer cells by activating the Wnt/ß-catenin pathway.
Assuntos
Transformação Celular Neoplásica/genética , Receptores Acoplados a Proteínas G/metabolismo , Neoplasias do Colo do Útero/patologia , Via de Sinalização Wnt/genética , beta Catenina/genética , Animais , Ciclo Celular/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Sobrevivência Celular , Colo do Útero/metabolismo , Colo do Útero/patologia , Ciclina D1/genética , Feminino , Células HeLa , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Transplante de Neoplasias , Proteínas Proto-Oncogênicas c-myc/genética , Receptores Acoplados a Proteínas G/biossíntese , Receptores Acoplados a Proteínas G/genética , Transplante Heterólogo , Neoplasias do Colo do Útero/genética , Proteínas Wnt/antagonistas & inibidores , Proteínas Wnt/genética , Proteínas Wnt/metabolismo , beta Catenina/antagonistas & inibidores , beta Catenina/metabolismoRESUMO
OBJECTIVE: Leucine-rich repeat-containing G protein-coupled receptor 5 (LGR5) has recently been reported to be a marker of cancer stem cells (CSCs) in colorectal cancer (CRC), and the prognostic value of LGR5 in CRC has been evaluated in several studies. However, the conclusions remain controversial. In this study, we aimed to evaluate the association between the expression of LGR5 and the outcome of CRC patients by performing a meta-analysis. METHODS: We systematically searched for relevant studies published up to February 2014 using the PubMed, Web of Science, EMBASE and Wangfang databases. Only articles in which LGR5 expression was detected by immunohistochemistry were included. A meta-analysis was performed using STATA 12.0, and pooled hazard ratios (HRs) with 95% confidence intervals (CIs) were used to estimate the strength of the association between LGR5 expression and the prognosis of CRC patients. RESULTS: A total of 7 studies comprising 1833 CRC patients met the inclusion criteria, including 6 studies comprising 1781 patients for overall survival (OS) and 3 studies comprising 528 patients for disease-free survival (DFS). Our results showed that high LGR5 expression was significantly associated with poor prognosis in terms of OS (HR: 1.87, 95% CI: 1.23-2.84; Pâ=â0.003) and DFS (HR: 2.44, 95% CI: 1.49-3.98; P<0.001). Further subgroup analysis revealed that many factors, including the study region, number of patients, follow-up duration and cutoff value, affected the significance of the association between LGR5 expression and a worse prognosis in patients with CRC. In addition, there was no evidence of publication bias, as suggested by Begg's and Egger's tests. CONCLUSIONS: The present meta-analysis indicated that high LGR5 expression was associated with poor prognosis in patients with CRC and that LGR5 is an efficient prognostic factor in CRC.
