RESUMEN
Several signaling pathways are aberrantly activated in head and neck squamous cell carcinoma (HNSCC), including the Hedgehog-Gli (HH-GLI), WNT, EGFR, and NOTCH pathways. The HH-GLI pathway has mostly been investigated in the context of canonical signal transduction and the inhibition of the membrane components of the pathway. In this work we investigated the role of downstream inhibitors GANT61 and lithium chloride (LiCl) on cell viability, wound closure, and colony forming ability of HNSCC cell lines. Five HNSCC cell lines were treated with HH-GLI pathway inhibitors affecting different levels of signal transduction. GANT61 and LiCl reduce the proliferation and colony formation capabilities of HNSCC cell lines, and LiCl has an additional effect on wound closure. The major effector of the HH-GLI signaling pathway in HNSCC is the GLI3 protein, which is expressed in its full-length form and is functionally regulated by GSK3ß. LiCl treatment increases the inhibitory Ser9 phosphorylation of the GSK3ß protein, leading to increased processing of GLI3 from full-length to repressor form, thus inhibiting HH-GLI pathway activity. Therefore, downstream inhibition of HH-GLI signaling may be a promising therapeutic strategy for HNSCC.
Asunto(s)
Biomarcadores de Tumor/metabolismo , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Neoplasias de Cabeza y Cuello/tratamiento farmacológico , Cloruro de Litio/farmacología , Proteínas del Tejido Nervioso/metabolismo , Piridinas/farmacología , Pirimidinas/farmacología , Proteína Gli3 con Dedos de Zinc/metabolismo , Antimaníacos/farmacología , Apoptosis , Biomarcadores de Tumor/genética , Proliferación Celular , Glucógeno Sintasa Quinasa 3 beta/genética , Neoplasias de Cabeza y Cuello/metabolismo , Neoplasias de Cabeza y Cuello/patología , Humanos , Proteínas del Tejido Nervioso/antagonistas & inhibidores , Proteínas del Tejido Nervioso/genética , Células Tumorales Cultivadas , Proteína Gli3 con Dedos de Zinc/antagonistas & inhibidores , Proteína Gli3 con Dedos de Zinc/genéticaRESUMEN
Malignant transformation is associated with loss of cell differentiation, anaplasia. Transcription factors gli, required for embryonic development, may be involved in this process. We studied the activity of transcription factors gli in high-grade gliomas and their role in maintenance of stem cell state and glioma cell survival. 20 glioma cell lines and a sample of a normal adult brain tissue were used in the present study. We found the expression of gli target genes, including GLI1 and FOXM1, in all tested glioma cell lines, but not in the normal tissue. Interestingly, the expression of gli target genes in some glioma cell lines was observed together with a high level of their transcriptional repressor, Gli3R. Knockdown of GLI3 in one of these lines resulted in decrease of gli target gene expression. These data suggest that Gli3R does not prevent the gli target genes transcription, and gli3 acts in glioma cells more as an activator, than a repressor of transcription. We observed that gli regulated the expression of such genes, as SOX2 or OCT4 that maintain stem cell state, and TET1, involving in DNA demethylation. Treatment with GANT61 or siRNA against GLI1, GLI2, or GLI3 could result in complete glioma cell death, while cyclopamine had a weaker and line-specific effect on glioma cell survival. Thus, the gli transcription factors are abnormally active in high-grade gliomas, regulate expression of genes, maintaining the stem cell state, and contribute to glioma cell survival.
Asunto(s)
Neoplasias Encefálicas/genética , Glioma/genética , Células Madre Neoplásicas/metabolismo , Piridinas/farmacología , Pirimidinas/farmacología , Proteínas Represoras/genética , Neoplasias Encefálicas/metabolismo , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Proteína Forkhead Box M1/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Técnicas de Silenciamiento del Gen , Glioma/metabolismo , Células HeLa , Humanos , Células Madre Neoplásicas/efectos de los fármacos , Proteínas del Tejido Nervioso/antagonistas & inhibidores , Proteínas del Tejido Nervioso/genética , Proteínas Nucleares/antagonistas & inhibidores , Proteínas Nucleares/genética , Proteínas Represoras/antagonistas & inhibidores , Proteína con Dedos de Zinc GLI1/antagonistas & inhibidores , Proteína con Dedos de Zinc GLI1/genética , Proteína Gli2 con Dedos de Zinc/antagonistas & inhibidores , Proteína Gli2 con Dedos de Zinc/genética , Proteína Gli3 con Dedos de Zinc/antagonistas & inhibidores , Proteína Gli3 con Dedos de Zinc/genéticaRESUMEN
Although important progresses have been made in the diagnosis and treatment of bladder cancer (BCa), the overall survival for patients with advanced BCa remains poor. It is necessary to uncover the molecular mechanism underlying the initiation and progression of bladder cancer. According to previous reports, mircoRNAs (miRNAs) can regulate tumorigenesis by targeting their downstream mRNAs. This study aims to explore and analyze a novel miRNA-mRNA axis which can regulate the progression of bladder cancer. Based on the microarray analysis, 182 mRNAs were found to be upregulated in BCa tissues. Gene oncology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that these upregulated mRNAs are related with hedgehog pathway. Gli3, an important factor of hedgehog pathway, belongs to these 182 upregulated mRNAs. Therefore, Gli3 was chosen to do further study. Kaplan-Meier analysis revealed that highly expressed Gli3 predicted unfavorable prognosis for patients with BCa. Results of functional experiments indicated the inhibitory effects of silenced Gli3 on cell proliferation, migration and EMT progress. Mechanically, Gli3 was the target mRNA of miR-7-5p in BCa cells. Finally, rescue assays were performed to validate the specific function of miR-7-5p/Gli3 axis in BCa progression. According to all data, we concluded that miR-7-5p acts as a tumor suppressor in BCa by downregulating Gli3.
