RESUMEN
The cascade comprising Raf, mitogen-activated protein kinase kinase (MEK) and extracellular signal-regulated kinase (ERK) is a therapeutic target in human cancers with deregulated Ras signalling, which includes tumours that have inactivated the Nf1 tumour suppressor. Nf1 encodes neurofibromin, a GTPase-activating protein that terminates Ras signalling by stimulating hydrolysis of Ras-GTP. We compared the effects of inhibitors of MEK in a myeloproliferative disorder (MPD) initiated by inactivating Nf1 in mouse bone marrow and in acute myeloid leukaemias (AMLs) in which cooperating mutations were induced by retroviral insertional mutagenesis. Here we show that MEK inhibitors are ineffective in MPD, but induce objective regression of many Nf1-deficient AMLs. Drug resistance developed because of outgrowth of AML clones that were present before treatment. We cloned clone-specific retroviral integrations to identify candidate resistance genes including Rasgrp1, Rasgrp4 and Mapk14, which encodes p38alpha. Functional analysis implicated increased RasGRP1 levels and reduced p38 kinase activity in resistance to MEK inhibitors. This approach represents a robust strategy for identifying genes and pathways that modulate how primary cancer cells respond to targeted therapeutics and for probing mechanisms of de novo and acquired resistance.
Asunto(s)
Resistencia a Antineoplásicos , Leucemia Mieloide Aguda/tratamiento farmacológico , Leucemia Mieloide Aguda/metabolismo , Quinasas de Proteína Quinasa Activadas por Mitógenos/antagonistas & inhibidores , Proteínas ras/metabolismo , Animales , Benzamidas/farmacología , Resistencia a Antineoplásicos/efectos de los fármacos , Resistencia a Antineoplásicos/genética , Genes ras , Factores de Intercambio de Guanina Nucleótido/genética , Factores de Intercambio de Guanina Nucleótido/metabolismo , Leucemia Mieloide Aguda/enzimología , Leucemia Mieloide Aguda/genética , Ratones , Proteína Quinasa 14 Activada por Mitógenos/genética , Proteína Quinasa 14 Activada por Mitógenos/metabolismo , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Proteínas ras/genéticaRESUMEN
Monosomy 7 and deletions of 7q are recurring leukemia-associated cytogenetic abnormalities that correlate with adverse outcomes in children and adults. We describe a 2.52-Mb genomic DNA contig that spans a commonly deleted segment of chromosome band 7q22 identified in myeloid malignancies. This interval currently includes 14 genes, 19 predicted genes, and 5 predicted pseudogenes. We have extensively characterized the FBXL13, NAPE-PLD, and SVH genes as candidate myeloid tumor suppressors. FBXL13 encodes a novel F-box protein, SVHis a member of a gene family that contains Armadillo-like repeats, and NAPE-PLD encodes a phospholipase D-type phosphodiesterase. Analysis of a panel of leukemia specimens with monosomy 7 did not reveal mutations in these or in the candidate genes LRRC17, PRO1598, and SRPK2. This fully sequenced and annotated contig provides a resource for candidate myeloid tumor suppressor gene discovery.
Asunto(s)
Neoplasias de la Médula Ósea/genética , Deleción Cromosómica , Cromosomas Humanos Par 7/genética , Genes Supresores de Tumor , Proteínas de Neoplasias/genética , Transactivadores/genética , Secuencia de Bases , Northern Blotting , Clonación Molecular , Biología Computacional , Mapeo Contig , ADN Complementario/genética , Bases de Datos de Ácidos Nucleicos , Humanos , Hibridación Fluorescente in Situ , Datos de Secuencia Molecular , Polimorfismo Conformacional Retorcido-Simple , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Análisis de Secuencia de ADNRESUMEN
The PTPN11 gene encodes SHP-2 (Src homology 2 domain-containing protein tyrosine Phosphatase), a nonreceptor tyrosine protein tyrosine phosphatase (PTPase) that relays signals from activated growth factor receptors to p21Ras (Ras) and other signaling molecules. Mutations in PTPN11 cause Noonan syndrome (NS), a developmental disorder characterized by cardiac and skeletal defects. NS is also associated with a spectrum of hematologic disorders, including juvenile myelomonocytic leukemia (JMML). To test the hypothesis that PTPN11 mutations might contribute to myeloid leukemogenesis, we screened the entire coding region for mutations in 51 JMML specimens and in selected exons from 60 patients with other myeloid malignancies. Missense mutations in PTPN11 were detected in 16 of 49 JMML specimens from patients without NS, but they were less common in other myeloid malignancies. RAS, NF1, and PTPN11 mutations are largely mutually exclusive in JMML, which suggests that mutant SHP-2 proteins deregulate myeloid growth through Ras. However, although Ba/F3 cells engineered to express leukemia-associated SHP-2 proteins cells showed enhanced growth factor-independent survival, biochemical analysis failed to demonstrate hyperactivation of the Ras effectors extracellular-regulated kinase (ERK) or Akt. We conclude that SHP-2 is an important cellular PTPase that is mutated in myeloid malignancies. Further investigation is required to clarify how these mutant proteins interact with Ras and other effectors to deregulate myeloid growth.
Asunto(s)
Leucemia Mielomonocítica Crónica/genética , Mutación Missense , Proteínas Serina-Treonina Quinasas , Proteínas Tirosina Fosfatasas/genética , Secuencia de Aminoácidos , Células Cultivadas , Niño , Humanos , Péptidos y Proteínas de Señalización Intracelular , Leucemia Mieloide/genética , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Datos de Secuencia Molecular , Neurofibromina 1/genética , Fosforilación , Proteína Tirosina Fosfatasa no Receptora Tipo 11 , Proteínas Tirosina Fosfatasas/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas c-akt , Proteínas ras/metabolismoRESUMEN
Abnormal hedgehog signaling, most commonly caused by loss of PTCH1 inhibitor activity,drives tumorigenesis of basal cell carcinomas (BCCs). To assess whether other tumors also have abnormal hedgehog signaling, we have assayed RNA from common cancers at nine different sites for levels of expression of hedgehog target genes that are up-regulated uniformly in BCCs. We report here that such dysregulation appears not to be common in the types of non-BCC cancers studied, indicating that the molecular pathogenesis of BCCs, like their frequency and behavior, differs markedly from that of most other cancers.
