RESUMO
The oncogenic BCR/ABL kinase activity induces and maintains chronic myelogenous leukemia (CML). We show here that, in BCR/ABL-transformed cells and CML blast crisis (CML-BC) progenitors, the phosphatase activity of the tumor suppressor PP2A is inhibited by the BCR/ABL-induced expression of the PP2A inhibitor SET. In imatinib-sensitive and -resistant (T315I included) BCR/ABL+ cell lines and CML-BC progenitors, molecular and/or pharmacological activation of PP2A promotes dephosphorylation of key regulators of cell proliferation and survival, suppresses BCR/ABL activity, and induces BCR/ABL degradation. Furthermore, PP2A activation results in growth suppression, enhanced apoptosis, restored differentiation, impaired clonogenic potential, and decreased in vivo leukemogenesis of imatinib-sensitive and -resistant BCR/ABL+ cells. Thus, functional inactivation of PP2A is essential for BCR/ABL leukemogenesis and, perhaps, required for blastic transformation.
Assuntos
Crise Blástica/metabolismo , Proteínas Cromossômicas não Histona/fisiologia , Proteínas de Fusão bcr-abl/fisiologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Fosfoproteínas Fosfatases/metabolismo , Fosfoproteínas Fosfatases/fisiologia , Fatores de Transcrição/fisiologia , Animais , Antineoplásicos/farmacologia , Benzamidas , Linhagem Celular Transformada , Colforsina/farmacologia , Proteínas de Ligação a DNA , Inibidores Enzimáticos/metabolismo , Chaperonas de Histonas , Humanos , Mesilato de Imatinib , Técnicas In Vitro , Células K562 , Leucemia/prevenção & controle , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Camundongos , Camundongos SCID , Transplante de Neoplasias , Fosfoproteínas Fosfatases/antagonistas & inibidores , Piperazinas/farmacologia , Proteína Fosfatase 2 , Pirimidinas/farmacologia , Células Tumorais Cultivadas , Proteínas Supressoras de Tumor/antagonistas & inibidores , Proteínas Supressoras de Tumor/fisiologiaRESUMO
Inactivation of DNA mismatch repair genes (MRG) is a recently described pathway of cancer development and progression resulting in genetic instability. Germline mutations in MRG have been studied predominantly in patients with hereditary non-polyposis colorectal cancer (HNPCC) where it is associated with microsatellite instability (MSI). The expression of MRG in primary breast cancer is still largely unexplored. The hMSH2 MRG encodes a protein that recognizes and binds to mismatch sequences of DNA. We investigated the relation-ship between hMSH2 expression and clinicopathological and biological characteristics, including p53 and p185 expression, in 44 primary invasive breast cancers. hMSH2 was not expressed in 11 cases (25%). Interestingly, p53 (p=0.05), p185 and steroidal receptor expression (p=0.07) were more frequent in tumors without hMSH2 expression. Furthermore, in 30 of 44 cases we analyzed hMSH2 expression in relation to MSI at 9 dinucleotide loci, and found that MRG expression was not significantly related to MSI. The presence of hMSH2 and p53 alterations in the same tumor suggests that the two oncoproteins act through a common mutational pathway, whereas the absence of a correlation between hMSH2 and MSI suggests that oncogenetic mechanisms of progression in primary breast cancer differ from those in HNPCC.
Assuntos
Neoplasias da Mama/metabolismo , Proteínas de Ligação a DNA/metabolismo , Genes p53 , Proteínas Proto-Oncogênicas/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias da Mama/diagnóstico , Neoplasias da Mama/genética , Neoplasias Colorretais Hereditárias sem Polipose/genética , Neoplasias Colorretais Hereditárias sem Polipose/metabolismo , Reparo do DNA , Proteínas de Ligação a DNA/genética , Feminino , Humanos , Imunoquímica , Masculino , Repetições de Microssatélites/genética , Pessoa de Meia-Idade , Proteína 2 Homóloga a MutS , Mutação , Proteínas Proto-Oncogênicas/genética , Receptor ErbB-2/metabolismo , Proteína Supressora de Tumor p53/metabolismoRESUMO
We analysed the differential expression pattern of the three distinct TAG-72 carbohydrate epitopes detected by monoclonal antibodies (MAbs) B72.3, CC83 and CC49 in a consecutive series of 114 patients with primary breast cancer and in 39 synchronous lymph node metastases. B72.3, CC83 and CC49 were expressed in respectively 81 (71%), 68 (60%) and 96 (84%) of the 114 cases. Interestingly, MAb B72.3 was significantly expressed in a subgroup of patients characterised by larger tumour size (p=0.013), lymph node metastasis (p=0.0002), high histopathological grade (p=0.006), high cell kinetics (p=0.04) and advanced clinical stage (p=0.0019). In 20 (51%) of the 39 pairs of matched primary breast cancers and synchronous lymph node metastases, TAG-72 was expressed in the tumour but not in the corresponding metastatic lymph node; tumours with TAG-72-negative lymph nodes appeared to be clinicopathologically more aggressive. CC49, the most immunoreactive and widely used MAb, was not detected in 34% of the metastases of expressing primary tumours. All three MAbs were found in a significantly lower per cent of synchronous metastases with respect to primary tumours (p<0.001).
Assuntos
Adenocarcinoma Mucinoso/metabolismo , Antígenos de Neoplasias/metabolismo , Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/metabolismo , Carcinoma Ductal de Mama/metabolismo , Carcinoma Lobular/metabolismo , Carcinoma Medular/metabolismo , Glicoproteínas/metabolismo , Adenocarcinoma Mucinoso/secundário , Adulto , Idoso , Anticorpos Antineoplásicos , Neoplasias da Mama/patologia , Carcinoma Ductal de Mama/secundário , Carcinoma Lobular/secundário , Carcinoma Medular/secundário , Epitopos/imunologia , Epitopos/metabolismo , Feminino , Humanos , Técnicas Imunoenzimáticas , Linfonodos/metabolismo , Metástase Linfática/patologia , Pessoa de Meia-Idade , Invasividade Neoplásica , Estadiamento de Neoplasias , Prognóstico , Timidina/metabolismoRESUMO
The oncogenic fusion tyrosine kinase nucleophosmin/anaplastic lymphoma kinase (NPM/ALK) induces cellular transformation in anaplastic large-cell lymphomas (ALCLs) carrying the t(2;5) chromosomal translocation. Protein-protein interactions involving NPM/ALK are important for the activation of downstream signaling pathways. This study was aimed at identifying novel NPM/ALK-binding proteins that might contribute to its oncogenic transformation. Using a proteomic approach, several RNA/DNA-binding proteins were found to coimmunoprecipitate with NPM/ALK, including the multifunctional polypyrimidine tract binding proteinassociated splicing factor (PSF). The interaction between NPM/ALK and PSF was dependent on an active ALK kinase domain and PSF was found to be tyrosine-phosphorylated in NPM/ALK-expressing cell lines and in primary ALK(+) ALCL samples. Furthermore, PSF was shown to be a direct substrate of purified ALK kinase domain in vitro, and PSF Tyr293 was identified as the site of phosphorylation. Y293F PSF was not phosphorylated by NPM/ALK and was not delocalized in NPM/ALK(+) cells. The expression of ALK fusion proteins induced delocalization of PSF from the nucleus to the cytoplasm and forced overexpression of PSF-inhibited proliferation and induced apoptosis in cells expressing NPM/ALK. PSF phosphorylation also increased its binding to RNA and decreased the PSF-mediated suppression of GAGE6 expression. These results identify PSF as a novel NPM/ALK-binding protein and substrate, and suggest that PSF function may be perturbed in NPM/ALK-transformed cells.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Linfoma Difuso de Grandes Células B/metabolismo , Proteínas Tirosina Quinases/metabolismo , Proteínas de Ligação a RNA/metabolismo , RNA/metabolismo , Animais , Apoptose , Linhagem Celular , Humanos , Linfoma Difuso de Grandes Células B/genética , Linfoma Difuso de Grandes Células B/patologia , Camundongos , Fator de Processamento Associado a PTB , Fosforilação , Fosfotirosina/metabolismo , Ligação Proteica , Proteínas Tirosina Quinases/química , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Transcrição Gênica/genéticaRESUMO
Altered mRNA translation is one of the effects exerted by the BCR/ABL oncoprotein in the blast crisis phase of chronic myelogenous leukemia (CML). Here, we report that in BCR/ABL+ cell lines and in patient-derived CML blast crisis mononuclear and CD34+ cells, p210(BCR/ABL) increases expression and activity of the transcriptional-inducer and translational-regulator heterogeneous nuclear ribonucleoprotein K (hnRNP K or HNRPK) in a dose- and kinase-dependent manner through the activation of the MAPK(ERK1/2) pathway. Furthermore, HNRPK down-regulation and interference with HNRPK translation-but not transcription-regulatory activity impairs cytokine-independent proliferation, clonogenic potential, and in vivo leukemogenic activity of BCR/ABL-expressing myeloid 32Dcl3 and/or primary CD34+ CML-BC patient cells. Mechanistically, we demonstrate that decreased internal ribosome entry site (IRES)-dependent Myc mRNA translation accounts for the phenotypic changes induced by inhibition of the BCR/ABL-ERK-dependent HNRPK translation-regulatory function. Accordingly, MYC protein but not mRNA levels are increased in the CD34+ fraction of patients with CML in accelerated and blastic phase but not in chronic phase CML patients and in the CD34+ fraction of marrow cells from healthy donors. Thus, BCR/ABL-dependent enhancement of HNRPK translation-regulation is important for BCR/ABL leukemogenesis and, perhaps, it might contribute to blast crisis transformation.
Assuntos
Proteínas de Fusão bcr-abl/fisiologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/etiologia , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Biossíntese de Proteínas , Proteínas Proto-Oncogênicas c-myc/genética , Ribonucleoproteínas/metabolismo , Antígenos CD34 , Crise Blástica/etiologia , Linhagem Celular Tumoral , Ribonucleoproteínas Nucleares Heterogêneas Grupo K , Humanos , RNA Mensageiro/genética , Células Tumorais CultivadasRESUMO
Imatinib (STI571 or CGP57148B) is an innovative treatment for tumours with a constitutively activated form of c-ABL, c-KIT, or PDGFR. Such tumours include Philadelphia-chromosome-positive (Ph-positive) leukaemias, gastrointestinal stromal tumours, and PDGFR-positive leukaemias. Diseases such as primary hypereosinophilia and dermatofibrosarcoma protuberans also seem to respond to imatinib. Clinical trials assessing the therapeutic effects of imatinib have shown that the drug is highly effective with few associated side-effects, achieving durable cytogenetic responses in many patients with chronic-phase BCR-ABL-positive leukaemias. However, the emergence of resistance, particularly in patients with acute leukaemias, has prompted intense research, and many are concerned about the future prospects for imatinib. The resistance has been found in patients with acute-phase disease, but may also occur in patients with chronic-phase disease. Two cellular mechanisms for resistance to imatinib have been identified: amplification of BCR-ABL gene and mutations in the catalytic domain of the protein. In addition, suboptimum inhibition of BCR-ABL in vivo could contribute to the selection of resistant cells. We have summarised all currently available data on resistance to imatinib, both published and unpublished, including the mechanisms of resistance identified so far, and their clinical relevance to the different forms of Ph-positive leukaemias is discussed. Furthermore, we discuss strategies to overcome or prevent the development of resistance.