BDNF/TrkB Signaling as a Potential Novel Target in Pediatric Brain Tumors: Anticancer Activity of Selective TrkB Inhibition in Medulloblastoma Cells

Medulloblastoma (MB) is the most common malignant pediatric brain tumor. Deregulation of brain-derived neurotrophic factor (BDNF)/tropomyosin-related kinase B (TrkB) signaling has been associated with increased proliferative capabilities, invasiveness, and chemoresistance in several types of cancer. However, the relevance of this pathway in MB remains unknown. Here, we show that the selective TrkB inhibitor N-[2-[[(hexahydro-2-oxo-1H-azepin-3-yl)amino]carbonyl]phenyl]-benzo[b]thiophene-2-carboxamide (ANA-12) markedly reduced the viability and survival of human cell lines representative of different MB molecular subgroups. These findings provide the first evidence supporting further investigation of TrkB inhibition as a potential novel strategy for MB treatment

Avaliação do efeito sinérgico do butirato de sodio e tyrphostin AG1478 na proliferação de glioblastoma multiforme

Introdução: Gliomas são os tumores cerebrais mais frequentes em pacientes com neoplasias de Sistema Nervoso Central (SNC), sendo o Glioblastoma Multiforme (GBM) o mais agressivo e letal deles. Apesar dos esforços na melhoria dos tratamentos atuais, o prognóstico para os pacientes com GBM continua sendo incerto. Sendo necessário o uso de novas estratégias terapêuticas que visem melhorar o manejo dos gliomas malignos. A combinação de terapias que agem nas principais vias de sinalização celular envolvidas na progressão do câncer poderia potencializar o efeito antitumoral das monoterapias. Métodos: As linhagens celulares U-87 e A-172 foram tratadas com o anti-EGFR tyrphostin AG1478, o inibidor de histonas deacetilases butirato de sódio (NaB) ou a combinação de ambos, por 72 horas. Tanto a viabilidade avaliada em 72 horas quanto a proliferação celular a longo prazo foram medidas através do ensaio de exclusão com azul de tripan em câmara de Neubauer. A influência do tratamento no ciclo celular e a capacidade de formar colônias foram avaliadas através da marcação com iodeto de propídeo e ensaio clonogênico, respectivamente. Resultados: Foi possível observar que o tratamento combinado com AG1478 e NaB foi capaz de reduzir a viabilidade e a proliferação celular na linhagem U-87 de GBM. Conclusão: Nosso trabalho mostrou que a inibição da via do receptor do fator de crescimento epidérmico (EGFR) combinada com a inibição das histonas deacetilases foi mais efetiva que as monoterapias na inibição da viabilidade e a proliferação celular. Esta redução foi significativa na linhagem U-87. Futuros estudos devem ser feitos para descobrir as possíveis interações entre as duas vias de sinalização em GBM.Introduction: Gliomas are the most frequent brain tumors, in patients with Central Nervous system (NCS) malignancies, being the Glioblastoma Multiforme the most aggressive and lethal of all. Despite current multimodality treatment efforts, the prognosis for GBM patients remains poor. New therapeutic strategies that target these pathways to improve the treatment of malignant gliomas are needed. Combination of therapies with synergistic effects in the cellular signaling pathways of cancer could potentiate the anti-tumor effect of monotherapy alone. Methods: U87 and A172 cell lines were treated with the anti-EGFR Thyrphostin AG1478, the Histone Deacetylase inhibitor (HDACi) Sodyum Butyrate (NaB), or combination of both, for 72 hours. The cellular proliferation in short and in a long time was measured through the trypan-blue assay on neubauer chamber, the influence on the cell cycle and the capability of form colonies was evaluated by nuclear staining with propidium iodide and clonogenic assay respectively. Results: We found that combined treatment with AG1478 and NaB, are able to reduce the viability and proliferation in U-87. Conclusion: Our work show that combined inhibition of both epidermal growth factor receptor and histone deacetylases was able to reduce cell proliferation in GBM cell lines. This reduction was considerably significant in U-87 cell lines when compared with individual treatments. Further studies should be performed to discover the possible crosstalk between the signaling pathways of both targets in GBM

Avaliação do efeito sinérgico do butirato de sodio e tyrphostin AG1478 na proliferação de glioblastoma multiforme

Introdução: Gliomas são os tumores cerebrais mais frequentes em pacientes com neoplasias de Sistema Nervoso Central (SNC), sendo o Glioblastoma Multiforme (GBM) o mais agressivo e letal deles. Apesar dos esforços na melhoria dos tratamentos atuais, o prognóstico para os pacientes com GBM continua sendo incerto. Sendo necessário o uso de novas estratégias terapêuticas que visem melhorar o manejo dos gliomas malignos. A combinação de terapias que agem nas principais vias de sinalização celular envolvidas na progressão do câncer poderia potencializar o efeito antitumoral das monoterapias. Métodos: As linhagens celulares U-87 e A-172 foram tratadas com o anti-EGFR tyrphostin AG1478, o inibidor de histonas deacetilases butirato de sódio (NaB) ou a combinação de ambos, por 72 horas. Tanto a viabilidade avaliada em 72 horas quanto a proliferação celular a longo prazo foram medidas através do ensaio de exclusão com azul de tripan em câmara de Neubauer. A influência do tratamento no ciclo celular e a capacidade de formar colônias foram avaliadas através da marcação com iodeto de propídeo e ensaio clonogênico, respectivamente. Resultados: Foi possível observar que o tratamento combinado com AG1478 e NaB foi capaz de reduzir a viabilidade e a proliferação celular na linhagem U-87 de GBM. Conclusão: Nosso trabalho mostrou que a inibição da via do receptor do fator de crescimento epidérmico (EGFR) combinada com a inibição das histonas deacetilases foi mais efetiva que as monoterapias na inibição da viabilidade e a proliferação celular. Esta redução foi significativa na linhagem U-87. Futuros estudos devem ser feitos para descobrir as possíveis interações entre as duas vias de sinalização em GBM.Introduction: Gliomas are the most frequent brain tumors, in patients with Central Nervous system (NCS) malignancies, being the Glioblastoma Multiforme the most aggressive and lethal of all. Despite current multimodality treatment efforts, the prognosis for GBM patients remains poor. New therapeutic strategies that target these pathways to improve the treatment of malignant gliomas are needed. Combination of therapies with synergistic effects in the cellular signaling pathways of cancer could potentiate the anti-tumor effect of monotherapy alone. Methods: U87 and A172 cell lines were treated with the anti-EGFR Thyrphostin AG1478, the Histone Deacetylase inhibitor (HDACi) Sodyum Butyrate (NaB), or combination of both, for 72 hours. The cellular proliferation in short and in a long time was measured through the trypan-blue assay on neubauer chamber, the influence on the cell cycle and the capability of form colonies was evaluated by nuclear staining with propidium iodide and clonogenic assay respectively. Results: We found that combined treatment with AG1478 and NaB, are able to reduce the viability and proliferation in U-87. Conclusion: Our work show that combined inhibition of both epidermal growth factor receptor and histone deacetylases was able to reduce cell proliferation in GBM cell lines. This reduction was considerably significant in U-87 cell lines when compared with individual treatments. Further studies should be performed to discover the possible crosstalk between the signaling pathways of both targets in GBM

Combined Inhibition of HDAC and EGFR Reduces Viability and Proliferation and Enhances STAT3 mRNA Expression in Glioblastoma Cells

Changes in expression of histone deacetylases (HDACs), which epigenetically regulate chromatin structure, and mutations and amplifications of the EGFR gene, which codes for the epidermal growth factor receptor (EGFR), have been reported in glioblastoma (GBM), the most common and malignant type of brain tumor. There are likely interplays between HDACs and EGFR in promoting GBM progression, and HDAC inhibition can cooperate with EGFR blockade in reducing the growth of lung cancer cells. Here, we found that either HDAC or EGFR inhibitors dose-dependently reduced the viability of U87 and A-172 human GBM cells. In U87 cells, the combined inhibition of HDACs and EGFR was more effective than inhibiting either target alone in reducing viability and long-term proliferation. In addition, HDAC or EGFR inhibition, alone or combined, led to G0/G1 cell cycle arrest. The EGFR inhibitor alone or combined with HDAC inhibition increased mRNA expression of the signal transducer and activator of transcription 3 (STAT3), which can act either as an oncogene or a tumor suppressor in GBM. These data provide early evidence that combining HDAC and EGFR inhibition may be an effective strategy to reduce GBM growth, through a mechanism possibly involving STAT3

Combined treatments with a retinoid receptor agonist and epigenetic modulators in human neuroblastoma cells

Neuroblastoma (NB) is the most common extracranial solid childhood tumor accounting for around 15% of pediatric cancer deaths and most probably originates from a failure in the development of embryonic neural crest cells. Retinoids can inhibit the proliferation and stimulate differentiation of NB cells. In addition, epigenetic events involving changes in chromatin structure and DNA methylation can mediate the effects of retinoids; hence, the scope of this study is to investigate the use of retinoids and epigenetic drugs in NB cell lines. Here, we demonstrate that the combination of retinoid all trans-retinoic acid (ATRA) with inhibitors of either histone deacetylases (HDACs) or DNA methyltransferase is more effective in impairing the proliferation of human SH-SY5Y and SK-N-BE(2) NB cells than any drug given alone. Treatments also induced differential changes on the messenger RNA (mRNA) expression of retinoid receptor subtypes and reduced the protein content of c-Myc, the neuronal markers NeuN and β-3 tubulin, and the oncoprotein Bmi1. These results suggest that the combination of retinoids with epigenetic modulators is more effective in reducing NB growth than treatment with single drugs