Assuntos
Proteínas de Ligação a DNA/genética , Células Precursoras Eritroides/enzimologia , Interferon-alfa/uso terapêutico , Janus Quinase 2/genética , Mutação , Polietilenoglicóis/uso terapêutico , Proteínas Proto-Oncogênicas/genética , Dioxigenases , Células Precursoras Eritroides/citologia , Humanos , Interferon alfa-2 , Proteínas RecombinantesRESUMO
High-throughput DNA sequence analysis was used to screen for TET2 mutations in bone marrow-derived DNA from 239 patients with BCR-ABL-negative myeloproliferative neoplasms (MPNs). Thirty-two mutations (19 frameshift, 10 nonsense, 3 missense; mostly involving exons 4 and 12) were identified for an overall mutational frequency of approximately 13%. Specific diagnoses included polycythemia vera (PV; n=89), essential thrombocythemia (ET; n=57), primary myelofibrosis (PMF; n=60), post-PV MF (n=14), post-ET MF (n=7) and blast phase PV/ET/MF (n=12); the corresponding mutational frequencies were approximately 16, 5, 17, 14, 14 and 17% (P=0.50). Mutant TET2 was detected in approximately 17 and approximately 7% of JAK2V617F-positive and -negative cases, respectively (P=0.04). However, this apparent clustering of the two mutations was accounted for by an independent association between mutant TET2 and advanced age; mutational frequency was approximately 23% in patients > or =60 years old versus approximately 4% in younger patients (P<0.0001). The presence of mutant TET2 did not affect survival, leukemic transformation or thrombosis in either PV or PMF; a correlation with hemoglobin <10 g per 100 ml in PMF was noted (P=0.05). We conclude that TET2 mutations occur in both JAK2V617F-positive and -negative MPN, are more prevalent in older patients, display similar frequencies across MPN subcategories and disease stages, and hold limited prognostic relevance.
Assuntos
Proteínas de Ligação a DNA/genética , Mutação/genética , Policitemia Vera/genética , Mielofibrose Primária/genética , Proteínas Proto-Oncogênicas/genética , Trombocitemia Essencial/genética , Idoso , Idoso de 80 Anos ou mais , Dioxigenases , Feminino , Humanos , Janus Quinase 2/genética , Masculino , Pessoa de Meia-Idade , Taxa de SobrevidaRESUMO
Telomerase catalytic subunit (hTERT) exerts important cellular functions including telomere homeostasis, genetic stability, cell survival and perhaps differentiation. However, the nature of external or internal signals, which regulate hTERT expression in tissues, remains poorly understood. Thus, whereas it has been described that hTERT gene is regulated along the differentiation of primitive myeloid progenitors, the effect of specific cytokines on telomerase expression in each myeloid lineage is currently unknown. Based on these considerations, we have investigated hTERT expression in erythroid cells treated with erythropoietin (EPO) and transforming growth factor beta (TGFbeta), as putative positive and negative regulators, respectively. We describe here that EPO activates hTERT gene transcription in in vitro-expanded primary erythroid precursors as well as in UT7 erythroleukemia cells. In UT7 cells, this study shows also that EPO acts through a JAK2/STAT5/c-myc axis. In contrast, TGFbeta blocks EPO signaling downstream of c-myc induction through a Smad3-dependent mechanism. Finally, hTERT appears to be efficiently regulated by EPO and TGFbeta in an opposite way in erythropoietic cells, arguing for a role of telomerase in red blood cell production.
Assuntos
Células Precursoras Eritroides/metabolismo , Eritropoetina/metabolismo , Regulação Leucêmica da Expressão Gênica , Telomerase/biossíntese , Fator de Crescimento Transformador beta/metabolismo , Antígenos CD34/biossíntese , Apoptose , Linhagem Celular , Linhagem Celular Tumoral , Proliferação de Células , Sobrevivência Celular , Humanos , Modelos Biológicos , Plasmídeos/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismoRESUMO
Myeloproliferative disorders (MPDs) are clonal haematopoietic malignancies involving the abnormal proliferation of myeloid lineages. The World Health Organisation (WHO) classification of haematopoietic malignancies distinguishes MPDs from myelodysplastic/ myeloproliferative disorders and systemic mastocytosis. These malignancies frequently involve constitutive tyrosine kinase activity, resulting from either oncogenic fusion protein production or from point mutations. Chronic myelogenous leukaemia is the model used for studies of the consequences of such molecular defects. However, the heterogeneity of the clinical course of MPDs should be seen in a more rationale conceptual framework, including the many molecular events associated with these diseases. This review focuses on the various tyrosine kinase-related molecular mechanisms underlying both MPDs and rare diseases with myeloproliferative features. We pay particular attention to the newly identified JAK2 V617F mutation in polycythaemia vera, essential thrombocythaemia and idiopathic myelofibrosis and deal with disease heterogeneity and putative additional molecular mechanisms.
Assuntos
Genes abl , Transtornos Mieloproliferativos/genética , Proteínas Tirosina Quinases/genética , Transdução de Sinais , Predisposição Genética para Doença , Humanos , Janus Quinase 2/genética , Janus Quinase 2/metabolismo , Mastocitose/genética , Transtornos Mieloproliferativos/diagnóstico , Transtornos Mieloproliferativos/metabolismo , Mutação Puntual , Proteínas Tirosina Quinases/metabolismo , Proteínas Proto-Oncogênicas c-kit/genética , Doenças Raras/genética , Doenças Raras/metabolismo , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética , Receptor beta de Fator de Crescimento Derivado de Plaquetas/genética , Fatores de Poliadenilação e Clivagem de mRNA/genéticaAssuntos
Fase G1/fisiologia , Transtornos Mieloproliferativos/enzimologia , Transtornos Mieloproliferativos/genética , Proteínas Quinases/genética , Proteínas Quinases/metabolismo , Perfilação da Expressão Gênica , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Humanos , Transtornos Mieloproliferativos/patologiaAssuntos
Policitemia Vera/diagnóstico , Policitemia Vera/genética , Policitemia/diagnóstico , Policitemia/genética , Proteínas Tirosina Quinases/genética , Proteínas Proto-Oncogênicas/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Autoanálise , Linhagem Celular Tumoral , Feminino , Humanos , Janus Quinase 2 , Masculino , Pessoa de Meia-Idade , Mutação , Transtornos Mieloproliferativos/diagnóstico , Transtornos Mieloproliferativos/genética , Policitemia Vera/classificação , Polimorfismo de Nucleotídeo Único , Valor Preditivo dos Testes , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Sensibilidade e Especificidade , TemperaturaRESUMO
Human telomerase, a cellular reverse transcriptase (hTERT), is a nuclear ribonucleoprotein enzyme complex that catalyzes the synthesis and extension of telomeric DNA. This enzyme is specifically activated in most malignant tumors but is usually inactive in normal somatic cells, suggesting that telomerase plays an important role in cellular immortalization and tumorigenesis. Terminal maturation of tumor cells has been associated with the repression of telomerase activity. Using maturation-sensitive and -resistant NB4 cell lines, we analyzed the pattern of telomerase expression during the therapeutic treatment of acute promyelocytic leukemia (APL) by retinoids. Two pathways leading to the down-regulation of hTERT and telomerase activity were identified. The first pathway results in a rapid down-regulation of telomerase that is associated with retinoic acid receptor (RAR)-dependent maturation of NB4 cells. Furthermore, during NB4 cell maturation, obtained independently of RAR by retinoic X receptor (RXR)-specific agonists (rexinoids), no change in telomerase activity was observed, suggesting that hTERT regulation requires a specific signaling and occurs autonomously. A second pathway of hTERT regulation, identified in the RAR-responsive, maturation-resistant NB4-R1 cell line, results in a down-regulation of telomerase that develops slowly during two weeks of all-trans retinoic acid (ATRA) treatment. This pathway leads to telomere shortening, growth arrest, and cell death, all events that are overcome by ectopic expression of hTERT. These findings demonstrate a clear and full dissociation between the process of tumor cell maturation and the regulation of hTERT mRNA expression and telomerase activity by retinoids. We propose telomerase expression as an efficient and selective target of retinoids in the therapy of tumors.
Assuntos
Leucemia Promielocítica Aguda/tratamento farmacológico , RNA , Telomerase/metabolismo , Tretinoína/farmacologia , Morte Celular/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Proteínas de Ligação a DNA , Regulação para Baixo , Humanos , Leucemia Promielocítica Aguda/patologia , RNA Mensageiro/análise , Telomerase/genética , Telômero , Células Tumorais CultivadasRESUMO
Early biochemical studies defined 4 functional domains of the erythroid protein 4.1 (4.1R). From amino-terminal to carboxy-terminal, these are 30 kd, 16 kd, 10 kd, and 22/24 kd in size. Although the functional properties of both the 30-kd and the 10-kd domain have been demonstrated in red cells, no functional activities have been assigned to either the 16-kd or the 22/24-kd domain in these cells. We here describe new mutations in the sequence encoding the C-terminal 22/24-kd domain that are associated with hereditary elliptocytosis. An unusually mild phenotype observed in heterozygous and homozygous members of 1 family suggested heterogeneity in the pattern of expression of 4.1R deficiency. Using a variety of protein and messenger RNA (mRNA) quantification strategies, we showed that, regardless of the alteration in the C-terminal primary sequence, when the protein is produced, it assembles at the cell membrane. In addition, we found that alterations in red cell morphologic features and membrane function correlate with the amount of membrane-associated protein-and therefore with the amount of mRNA accumulated-rather than with the primary structure of the variant proteins. These data suggest that an intact sequence at exons 19 through 21 encoding part of the C-terminal 22/24-kd region is not required for proper protein 4.1R assembly in mature red cells. (Blood. 2000;95:1834-1841)
Assuntos
Proteínas do Citoesqueleto , Eliptocitose Hereditária/genética , Proteínas de Membrana/genética , Neuropeptídeos , RNA Mensageiro/genética , Adulto , Sequência de Aminoácidos , Clonagem Molecular , Análise Mutacional de DNA , Eritrócitos/metabolismo , Feminino , Humanos , Masculino , Proteínas de Membrana/análise , Proteínas de Membrana/química , Dados de Sequência Molecular , Peso Molecular , Reação em Cadeia da Polimerase , Polimorfismo Conformacional de Fita Simples , Conformação Proteica , Estrutura Terciária de Proteína , Splicing de RNA , RNA Mensageiro/análise , RNA Mensageiro/metabolismo , Relação Estrutura-AtividadeRESUMO
Although hereditary spherocytosis (HS) is a common disorder of the red cell membrane, its clinical and biologic expression at birth and in early infancy has received little attention. In order to obtain insights into the natural history of HS during infancy, we studied 46 neonates, 39 from families in which 1 of the parents had previously been given a diagnosis of HS and 7 presenting with nonimmune hemolytic anemia and no family history of HS. Of these 46 neonates, 23 were subsequently confirmed to have HS and 23 were found to be healthy. The hematologic and biologic analyses carried out in this cohort of 46 newborns enabled us to develop guidelines for early diagnosis of HS. A careful clinical follow-up of 34 HS patients during the first year of life allowed us to define several important clinical features of HS during this period. Hemoglobin values are usually normal at birth but decrease sharply during the subsequent 20 days, which leads, in many cases, to a transient and severe anemia. The anemia is severe enough to warrant blood transfusions in a large number of infants with HS (26 of 34 in our series). The aggravation of anemia appears to be related to the inability of these infants to mount an appropriate erythropoietic response to anemia and to the development of splenic filtering function. These findings indicate that careful monitoring of infants with HS during the first 6 months of life is important for appropriate clinical management. (Blood. 2000;95:393-397)
Assuntos
Esferocitose Hereditária/sangue , Transfusão de Sangue , Membrana Eritrocítica/patologia , Sangue Fetal/química , Hemoglobinas/metabolismo , Humanos , Recém-Nascido , Estudos Longitudinais , Valores de Referência , Contagem de Reticulócitos , Esferocitose Hereditária/genética , Esferocitose Hereditária/fisiopatologia , Esferocitose Hereditária/terapiaRESUMO
INTRODUCTION: In hereditary spherocytosis, erythropoiesis has been described as 'sluggish' during the first months of life. The lack of appropriate erythropoietic response to compensate for increased red cell destruction necessitates blood transfusions in 70-80% of hereditary spherocytosis-affected infants during their first year of life. After this period, less than 30% require regular transfusion support. This transient requirement for transfusion led us to wonder whether anemic hereditary spherocytosis infants, like anemic premature infants, could benefit from recombinant erythropoietin therapy (rHu-Epo). MATERIAL AND METHODS: In 16 hereditary spherocytosis infants (age range 16-119 days) with severe anemia, a compassionate open preliminary study was performed. rHu-Epo treatment (1000 IU/kg/week) was instituted together with iron supplementation. Hemoglobin values and reticulocyte counts were repeatedly assessed. RESULTS: In 13 out of 16 infants, prompt increases in reticulocyte counts were noted after the first week of treatment with 1000 IU/kg/week of rHu-Epo. During treatment with Epo these infants maintained clinically acceptable levels of hemoglobin and did not require blood transfusions. As the infants grew and began to mount an adequate erythropoietic response, the rHu-Epo dose could be tapered and the treatment could be discontinued before the age of nine months. CONCLUSION: Epo treatment in most hereditary spherocytosis infants appears to be effective in the management of anemia and could serve as a valuable alternative to packed RBC transfusions.