RESUMO
Protein phosphatase 2A (PP2A) is a human tumor suppressor that inhibits cellular transformation by regulating the activity of several signaling proteins critical for malignant cell behavior. PP2A has been described as a potential therapeutic target in chronic myeloid leukemia, Philadelphia chromosome-positive acute lymphoblastic leukemia and B-cell chronic lymphocytic leukemia. Here, we show that PP2A inactivation is a recurrent event in acute myeloid leukemia (AML), and that restoration of PP2A phosphatase activity by treatment with forskolin in AML cells blocks proliferation, induces caspase-dependent apoptosis and affects AKT and ERK1/2 activity. Moreover, treatment with forskolin had an additive effect with Idarubicin and Ara-c, drugs used in standard induction therapy in AML patients. Analysis at protein level of the PP2A activation status in a series of patients with AML at diagnosis showed PP2A hyperphosphorylation in 78% of cases (29/37). In addition, we found that either deregulated expression of the endogenous PP2A inhibitors SET or CIP2A, overexpression of SETBP1, or downregulation of some PP2A subunits, might be contributing to PP2A inhibition in AML. In conclusion, our results show that PP2A inhibition is a common event in AML cells and that PP2A activators, such as forskolin or FTY720, could represent potential novel therapeutic targets in AML.
Assuntos
Colforsina/uso terapêutico , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/patologia , Proteína Fosfatase 2/metabolismo , Apoptose/efeitos dos fármacos , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Western Blotting , Medula Óssea , Estudos de Casos e Controles , Caspases/metabolismo , Proliferação de Células/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Feminino , Perfilação da Expressão Gênica , Humanos , Técnicas Imunoenzimáticas , Leucemia Mieloide Aguda/metabolismo , Masculino , Pessoa de Meia-Idade , Análise de Sequência com Séries de Oligonucleotídeos , Fosforilação/efeitos dos fármacos , Prognóstico , Proteína Fosfatase 2/antagonistas & inibidores , Proteína Fosfatase 2/genética , RNA Mensageiro/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células Tumorais CultivadasRESUMO
BACKGROUND: The EVI1(ecotropic virus integration site 1) gene codes for a zinc-finger transcription factor, whose transcriptional activation leads to a particularly aggressive form of acute myeloid leukaemia (AML). Although, EVI1 interactions with key proteins in hematopoiesis have been previously described, the precise role of this transcription factor in promoting leukaemic transformation is not completely understood. Recent works have identified specific microRNA (miRNA) signatures in different AML subgroups. However, there is no analysis of miRNAs profiles associated with EVI1 overexpression in humans. METHODS: We performed QT-RT-PCR to assess the expression of 250 miRNAs in cell lines with or without EVI1 overexpression and in patient samples. We used ChIP assays to evaluated the possible binding of EVI1 binding to the putative miRNA promoter. Proliferation of the different cell lines transfected with the anti- or pre-miRs was quantified by MTT. RESULTS: Our data showed that EVI1 expression was significantly correlated with the expression of miR-1-2 and miR-133-a-1 in established cell lines and in patient samples. ChIP assays confirmed that EVI1 binds directly to the promoter of these two miRNAs. However, only miR-1-2 was involved in abnormal proliferation in EVI1 expressing cell lines. CONCLUSIONS: Our data showed that EVI1 controls proliferation in AML through modulation of miR-1-2. This study contributes to further understand the transcriptional networks involving transcription factors and miRNAs in AML.
