期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2015
卷号：112
期号：6
页码：E536-E545
DOI：10.1073/pnas.1418163112
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：SignificanceUsing Sleeping Beauty transposon mutagenesis in a melanoma model driven by oncogenic BRAF (B-Raf proto-oncogene, serine/threonine kinase), we identified both known and novel candidate genes that mediate resistance to the BRAF inhibitor PLX4720. We validate ES-cell expressed Ras as a novel promoter of BRAF inhibitor resistance and propose that AKT (v-akt murine thymoma viral oncogene homolog 1)-mediated inactivation of BAD (BCL2-associated agonist of cell death) constitutes a pathway that may contribute to hepatocyte growth factor-mediated therapy resistance. Our work establishes Sleeping Beauty mutagenesis as a powerful tool for the identification of novel resistance genes and mechanisms in genetically modified mouse models. BRAF (v-raf murine sarcoma viral oncogene homolog B) inhibitors elicit a transient anti-tumor response in [~]80% of BRAFV600-mutant melanoma patients that almost uniformly precedes the emergence of resistance. Here we used a mouse model of melanoma in which melanocyte-specific expression of BrafV618E (analogous to the human BRAFV600E mutation) led to the development of skin hyperpigmentation and nevi, as well as melanoma formation with incomplete penetrance. Sleeping Beauty insertional mutagenesis in this model led to accelerated and fully penetrant melanomagenesis and synchronous tumor formation. Treatment of BrafV618E transposon mice with the BRAF inhibitor PLX4720 resulted in tumor regression followed by relapse. Analysis of transposon insertions identified eight genes including Braf, Mitf, and ERas (ES-cell expressed Ras) as candidate resistance genes. Expression of ERAS in human melanoma cell lines conferred resistance to PLX4720 and induced hyperphosphorylation of AKT (v-akt murine thymoma viral oncogene homolog 1), a phenotype reverted by combinatorial treatment with PLX4720 and the AKT inhibitor MK2206. We show that ERAS expression elicits a prosurvival signal associated with phosphorylation/inactivation of BAD, and that the resistance of hepatocyte growth factor-treated human melanoma cells to PLX4720 can be reverted by treatment with the BAD-like BH3 mimetic ABT-737. Thus, we define a role for the AKT/BAD pathway in resistance to BRAF inhibition and illustrate an in vivo approach for finding drug resistance genes.
关键词：melanoma ; drug resistance ; BRAF inhibitors ; mouse models
