RESUMO
To determine the role of BRAFV600E mutation and MAPK signaling as well as the effects of BRAF and MEK directed therapy in gastroenteropancreatic neuroendocrine neoplasia (GEP-NEN), with a focus on highly aggressive gastroenteropancreatic neuroendocrine carcinoma (GEP-NEC). Using Sanger sequencing of BRAF exon 15 we determined the frequency of BRAFV600E mutations in 71 primary GEP-NENs. MEK phosphorylation was examined by immunohistochemistry in corresponding tissue samples. To evaluate the biological relevance of BRAFV600E mutation and MAPK signaling in GEP-NECs, effects of a pharmacological BRAF and MEK inhibition were analyzed in NEC cell lines both in vitro and in vivo. BRAFV600E mutation was detected in 9.9% of all GEP-NENs. Interestingly, only NECs of the colon harbored BRAFV600E mutations, leading to a mutation frequency of 46.7% in this subgroup of patients. In addition, a BRAFV600E mutation was significantly associated with high levels of MEK phosphorylation (pMEK) and advanced tumor stages. Pharmacological inhibition of BRAF and MEK abrogated NEC cell growth, inducing G1 cell cycle arrest and apoptosis only in BRAFV600E mutated cells. BRAF inhibitor dabrafenib and MEK inhibitor trametinib prevented growth of BRAFV600E positive NEC xenografts. High frequencies of BRAFV600E mutation and elevated expression levels of pMEK were detected in biologically aggressive and highly proliferative colorectal NECs. We provide evidence that targeting BRAF oncogene may represent a therapeutic strategy for patients with BRAF mutant colorectal NECs.
Assuntos
Antineoplásicos/farmacologia , Carcinoma Neuroendócrino/genética , Neoplasias Colorretais/genética , Neoplasias Intestinais/genética , Tumores Neuroendócrinos/genética , Neoplasias Pancreáticas/genética , Proteínas Proto-Oncogênicas B-raf/genética , Neoplasias Gástricas/genética , Idoso , Animais , Antineoplásicos/uso terapêutico , Apoptose/efeitos dos fármacos , Apoptose/genética , Carcinoma Neuroendócrino/tratamento farmacológico , Carcinoma Neuroendócrino/mortalidade , Carcinoma Neuroendócrino/patologia , Linhagem Celular Tumoral , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/mortalidade , Neoplasias Colorretais/patologia , Éxons/genética , Feminino , Seguimentos , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem da Fase G1 do Ciclo Celular/genética , Humanos , Imidazóis/farmacologia , Imidazóis/uso terapêutico , Neoplasias Intestinais/tratamento farmacológico , Neoplasias Intestinais/mortalidade , Neoplasias Intestinais/patologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/genética , Masculino , Camundongos , Camundongos Endogâmicos NOD , Pessoa de Meia-Idade , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Mutação , Tumores Neuroendócrinos/tratamento farmacológico , Tumores Neuroendócrinos/mortalidade , Tumores Neuroendócrinos/patologia , Oximas/farmacologia , Oximas/uso terapêutico , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/mortalidade , Neoplasias Pancreáticas/patologia , Fosforilação/genética , Piridonas/farmacologia , Piridonas/uso terapêutico , Pirimidinonas/farmacologia , Pirimidinonas/uso terapêutico , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/mortalidade , Neoplasias Gástricas/patologia , Análise de Sobrevida , Análise Serial de Tecidos , Vemurafenib/farmacologiaRESUMO
BACKGROUND: Gastroenteropancreatic neuroendocrine carcinomas (GEP-NECs) are biologically aggressive tumors, associated with a very poor survival. Due to their rarity, our knowledge on GEP-NEC biology is very limited. The aim of this study was to establish a GEP-NEC cell line model that might contribute to a better understanding of this rare malignant disease to further develop novel therapeutic approaches in preclinical studies. METHODS: Small cell neuroendocrine cancer cell line NEC-DUE3 was derived from a lymph node metastasis of a neuroendocrine carcinoma (NEC) located at the anal canal. Morphological characteristics and the expression of neuroendocrine markers were comprehensively investigated. For genetic profiling, NEC-DUE3 cells were analyzed by DNA fingerprinting. Chromosomal aberrations were mapped by array comparative genomic hybridization. NEC-DUE3 cell tumorigenicity was evaluated in vivo and the sensitivity to chemotherapeutic agents was assessed in vitro. RESULTS: NEC-DUE3 cells were characterized by the expression of molecular markers that are commonly observed in GEP-NECs, were sensitive to treatment with cisplatin, and able to form tumors in immunodeficient mice. CONCLUSION: We established and characterized the first small cell GEP-NEC cell line that may serve as a valuable tool to create a better understanding of the biology of these rare tumors and to develop novel treatment strategies.
