To determine the role of BRAF 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 BRAF mutations in 71 primary GEP-NENs. MEK phosphorylation was examined by immunohistochemistry in corresponding tissue samples. To evaluate the biological relevance of BRAF 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. BRAF mutation was detected in 9.9% of all GEP-NENs. Interestingly, only NECs of the colon harbored BRAF mutations, leading to a mutation frequency of 46.7% in this subgroup of patients. In addition, a BRAF 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 BRAF mutated cells. BRAF inhibitor dabrafenib and MEK inhibitor trametinib prevented growth of BRAF positive NEC xenografts. High frequencies of BRAF 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.
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.
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