RESUMO
Asparaginase, which depletes asparagine and glutamine, activates amino-acid stress response. Oxidative stress mediated by excessive reactive oxygen species (ROS) causes enhanced mitochondrial permeabilization and subsequent cell apoptosis and is considered as a plausible mechanism for drug-induced hepatotoxicity, a common toxicity of asparaginase in adults with acute lymphoblastic leukemia (ALL). Studies investigating the pharmacogenetics of asparaginase in ALL are limited and focused on asparaginase-induced allergic reaction common in pediatric patients. Here, we sought to determine a potential association between the variant rs4880 in SOD2 gene, a key mitochondrial enzyme that protects cells against ROS, and hepatotoxicity during asparaginase-based therapy in 224 patients enrolled on CALGB-10102, a treatment trial for adults with ALL. We report that the CC genotype of rs4880 is associated with increased hepatotoxicity following asparaginase-based treatment. Thus, rs4880 likely contributes to asparaginase-induced hepatotoxicity, and functional studies investigating this single-nucleotide polymorphism (SNP) are needed to develop therapeutic approaches that mitigate this toxicity.
Assuntos
Antineoplásicos/efeitos adversos , Asparaginase/efeitos adversos , Doença Hepática Induzida por Substâncias e Drogas/genética , Variantes Farmacogenômicos , Polimorfismo de Nucleotídeo Único , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Superóxido Dismutase/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Doença Hepática Induzida por Substâncias e Drogas/enzimologia , Doença Hepática Induzida por Substâncias e Drogas/patologia , Feminino , Frequência do Gene , Predisposição Genética para Doença , Humanos , Masculino , Pessoa de Meia-Idade , Testes Farmacogenômicos , Fenótipo , Leucemia-Linfoma Linfoblástico de Células Precursoras/enzimologia , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Medição de Risco , Fatores de Risco , Resultado do Tratamento , Adulto JovemRESUMO
CpG islands frequently contain gene promoters or exons and are usually unmethylated in normal cells. Methylation of CpG islands is associated with delayed replication, condensed chromatin and inhibition of transcription initiation. The investigation of aberrant CpG-island methylation in human cancer has primarily taken a candidate gene approach, and has focused on less than 15 of the estimated 45,000 CpG islands in the genome. Here we report a global analysis of the methylation status of 1,184 unselected CpG islands in each of 98 primary human tumours using restriction landmark genomic scanning (RLGS). We estimate that an average of 600 CpG islands (range of 0 to 4,500) of the 45,000 in the genome were aberrantly methylated in the tumours, including early stage tumours. We identified patterns of CpG-island methylation that were shared within each tumour type, together with patterns and targets that displayed distinct tumour-type specificity. The expression of many of these genes was reactivated by experimental demethylation in cultured tumour cells. Thus, the methylation of particular subsets of CpG islands may have consequences for specific tumour types.
Assuntos
Metilação de DNA , Fosfatos de Dinucleosídeos/análise , Neoplasias/genética , Adenocarcinoma/genética , Sequência de Bases , Neoplasias Encefálicas/genética , Neoplasias da Mama/genética , Carcinoma Intraductal não Infiltrante/genética , Carcinoma Lobular/genética , Neoplasias do Colo/genética , Fosfatos de Dinucleosídeos/genética , Feminino , Genoma Humano , Humanos , Masculino , Dados de Sequência Molecular , Mapeamento por RestriçãoRESUMO
Although cancer classification has improved over the past 30 years, there has been no general approach for identifying new cancer classes (class discovery) or for assigning tumors to known classes (class prediction). Here, a generic approach to cancer classification based on gene expression monitoring by DNA microarrays is described and applied to human acute leukemias as a test case. A class discovery procedure automatically discovered the distinction between acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL) without previous knowledge of these classes. An automatically derived class predictor was able to determine the class of new leukemia cases. The results demonstrate the feasibility of cancer classification based solely on gene expression monitoring and suggest a general strategy for discovering and predicting cancer classes for other types of cancer, independent of previous biological knowledge.
Assuntos
Perfilação da Expressão Gênica , Leucemia Mieloide/classificação , Leucemia Mieloide/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/classificação , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Doença Aguda , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Adesão Celular/genética , Ciclo Celular/genética , Proteínas de Homeodomínio/genética , Humanos , Leucemia Mieloide/tratamento farmacológico , Proteínas de Neoplasias/genética , Neoplasias/classificação , Neoplasias/genética , Análise de Sequência com Séries de Oligonucleotídeos , Oncogenes , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Valor Preditivo dos Testes , Reprodutibilidade dos Testes , Resultado do TratamentoRESUMO
B and T lymphocytes in 37 untreated patients with malignant lymphoma and Hodgkin's disease were studied. B cells in the peripheral blood were investigated with respect to surface immunoglobulins and in a few patients with respect to intracytoplasmic immunoglobulins by means of immunofluorescence. T cell function was studied by direct phytohemagglutinin (PHA) microtest (from the same sample of whole blood), mixed lymphocyte culture (MLC), and by delayed hypersensitivity to various antigens. In the 13 patients with Hodgkin's disease the histologic subtype was nodular sclerosis in nine, lymphocyte predominant in two, mixed cellularity in two. Only one of these patients had disseminated disease (stage IV); he showed impaired cellular immunity, a very low percentage of B cells and low levels of serum immunoglobulins. Of the remaining patients with Hodgkin's disease, with one exception, normal percentages but rather low absolute numbers of B lymphocytes per mm(3) of blood were found. One patient with a low percent and low absolute number of B lymphocytes showed very high serum IgG. Of 24 patients with non-Hodgkin's malignant lymphoma, seven (29%) showed monoclonal B cell proliferation in the peripheral blood (five mukappa, two gammakappa). By morphologic criteria, 14 patients had involvement of bone marrow, five of these had involvement of peripheral blood. Four of the latter five patients showed marked increases in percentages and absolute numbers of B lymphocytes in the peripheral blood reflecting the monoclonal proliferation. In three additional patients monoclonal proliferation of lymphocytes was found by immunofluorescence although the blood smears appeared morphologically normal. Serum immunoglobulin abnormalities without monoclonal B cell proliferation in the peripheral blood were observed in six patients.
Assuntos
Linfócitos B/imunologia , Doença de Hodgkin/imunologia , Linfoma/imunologia , Adolescente , Adulto , Idoso , Membrana Celular/imunologia , Criança , Feminino , Imunofluorescência , Teste de Histocompatibilidade , Doença de Hodgkin/sangue , Humanos , Imunoglobulinas/análise , Lectinas , Ativação Linfocitária , Linfoma/sangue , Masculino , Pessoa de Meia-Idade , Testes Cutâneos , Linfócitos T/imunologia , Macroglobulinemia de Waldenstrom/imunologiaRESUMO
Core-binding factor acute myeloid leukemia (CBF-AML) is defined by the presence of either t(8;21)(q22;q22)/RUNX1-RUNX1T1 or inv(16)(p13.1q22)/t(16;16)(p13.1;q22)/CBFB-MYH11. The resulting fusion genes require a 'second hit' to initiate leukemogenesis. Mutation assessment of 177 adults with CBF-AML, including 68 with t(8;21) and 109 with inv(16)/t(16;16), identified not only mutations well known in CBF-AML but also mutations in the CCND1 and CCND2 genes, which represent novel frequent molecular alterations in AML with t(8;21). Altogether, CCND1 (n=2) and CCND2 (n=8) mutations were detected in 10 (15%) patients with t(8;21) in our cohort. A single CCND2 mutation was also found in 1 (0.9%) patient with inv(16). In contrast, CCND1 and CCND2 mutations were detected in only 11 (0.77%) of 1426 non-CBF-AML patients. All CCND2 mutations cluster around the highly conserved amino-acid residue threonine 280 (Thr280). We show that Thr280Ala-mutated CCND2 leads to increased phosphorylation of the retinoblastoma protein, thereby causing significant cell cycle changes and increased proliferation of AML cell lines. The identification of CCND1 and CCND2 mutations as frequent mutational events in t(8;21) AML may provide further justification for cell cycle-directed therapy in this disease.
Assuntos
Cromossomos Humanos Par 21 , Cromossomos Humanos Par 8 , Ciclina D1/genética , Ciclina D2/genética , Leucemia Mieloide Aguda/genética , Mutação , Translocação Genética , Adolescente , Adulto , Idoso , Biomarcadores Tumorais/genética , Feminino , Seguimentos , Humanos , Leucemia Mieloide Aguda/patologia , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Prognóstico , Taxa de Sobrevida , Adulto JovemRESUMO
In this prospective phase 2 clinical trial conducted by Cancer and Leukemia Group B (CALGB, now the Alliance), we studied decitabine as maintenance therapy for younger adults with acute myeloid leukemia (AML) who remained in first complete remission (CR1) following intensive induction and consolidation. Given that decitabine is clinically active in AML and with hypomethylating activity distinct from cytotoxic chemotherapy, we hypothesized that 1 year of maintenance therapy would improve disease-free survival (DFS) for AML patients <60 years, who did not receive allogeneic stem cell transplantation in CR1. After blood count recovery from final consolidation, patients received decitabine at 20 mg/m2 intravenously daily for 4-5 days, every 6 weeks for eight cycles. One hundred and thirty-four patients received decitabine and 85 (63%) had favorable risk AML. The median number of cycles received was 7 (range: 1-8) and the primary reason for discontinuation was relapse. DFS at 1 year and 3 years was 79% and 54%, respectively. These results are similar to the outcomes in the historical control comprising similar patients treated on recent CALGB trials. Thus, maintenance with decitabine provided no benefit overall. Standard use of decitabine maintenance in younger AML patients in CR1 is not warranted. This trial was registered at www.clinicaltrials.gov as NCT00416598.
Assuntos
Azacitidina/análogos & derivados , Leucemia Mieloide Aguda/tratamento farmacológico , Quimioterapia de Manutenção/métodos , Adolescente , Adulto , Azacitidina/administração & dosagem , Decitabina , Intervalo Livre de Doença , Feminino , Humanos , Leucemia Mieloide Aguda/mortalidade , Masculino , Pessoa de Meia-Idade , Indução de Remissão , Taxa de Sobrevida , Adulto JovemRESUMO
Recurrent chromosomal abnormalities and gene mutations detected at the time of diagnosis of acute myeloid leukemia (AML) are associated with particular disease features, treatment response and survival of AML patients, and are used to denote specific disease entities in the World Health Organization classification of myeloid neoplasms and acute leukemia. However, large studies that integrate cytogenetic and comprehensive mutational information are scarce. We created a comprehensive oncoprint of mutations associated with recurrent cytogenetic findings by combining the information on mutational patterns of 80 cancer- and leukemia-associated genes with cytogenetic findings in 1603 adult patients with de novo AML. We show unique differences in the mutational profiles among major cytogenetic subsets, identify novel associations between recurrent cytogenetic abnormalities and both specific gene mutations and gene functional groups, and reveal differences in cytogenetic and mutational features between patients younger than 60 years and those aged 60 years or older. The identified associations between cytogenetic and molecular genetic data may help guide mutation testing in AML, and result in more focused application of targeted therapy in patients with de novo AML.
Assuntos
Aberrações Cromossômicas , Ontologia Genética , Genes Neoplásicos , Leucemia Mieloide Aguda/genética , Mutação , Adulto , Fatores Etários , Idoso , Análise Mutacional de DNA , DNA de Neoplasias/genética , Feminino , Humanos , Cariotipagem , Masculino , Pessoa de Meia-IdadeRESUMO
BACKGROUND: Epidemiologic studies of acute myeloid leukemias (AMLs) show small increases in risk of disease associated with certain occupations and chemical exposures. PURPOSE: This study was designed to determine whether the presence of mutationally activated ras oncogenes in AML are associated with occupational and chemical exposures. METHODS: We interviewed 62 patients with newly diagnosed AML (or their next-of-kin), all of whom were enrolled in a national multicenter clinical trial, and 630 healthy control subjects. DNA extracted from patients' pretreatment bone marrow samples was amplified by using the polymerase chain reaction and probed with allele-specific oligonucleotides for activating point mutations at the 12th, 13th, and 61st codons of three protooncogenes: H-ras (also known as HRAS), K-ras (also known as KRAS2), and N-ras (also known as NRAS). RESULTS: Patients with ras mutation-positive AML had a higher frequency (six of 10 patients) of working 5 or more years in an a priori high-risk occupation than did patients with ras mutation-negative AML (eight of 52; odds ratio [OR] = 6.8; 95% confidence interval [CI] = 1.3-36). Patients with ras mutation-positive AML were more likely than patients with ras mutation-negative AML to have breathed chemical vapor on the job (OR = 9.1; 95% CI = 1.3-64) or to have had skin contact with chemicals (OR = 6.9; 95% CI = 1.3-37). When ras-positive patients were compared with healthy control subjects, the ORs for occupation and occupational exposures remained elevated, while patients with ras mutation-negative AML showed no increased risk when compared with control subjects. CONCLUSION: Activation of ras proto-oncogenes may identify an etiologic subgroup of AML caused by occupation and chemical exposure. IMPLICATION: Disease etiology may be better understood if epidemiologic measures of exposure are integrated with molecular assays of the genetic defects responsible for cancer initiation and promotion.
Assuntos
Regulação Leucêmica da Expressão Gênica/genética , Genes ras/genética , Leucemia Mieloide/genética , Exposição Ocupacional , Doença Aguda , Adulto , Idoso , Estudos de Casos e Controles , Códon/efeitos dos fármacos , Códon/genética , Feminino , Genes ras/efeitos dos fármacos , Humanos , Leucemia Eritroblástica Aguda/induzido quimicamente , Leucemia Eritroblástica Aguda/epidemiologia , Leucemia Eritroblástica Aguda/genética , Leucemia Monocítica Aguda/induzido quimicamente , Leucemia Monocítica Aguda/epidemiologia , Leucemia Monocítica Aguda/genética , Leucemia Mieloide/induzido quimicamente , Leucemia Mieloide/epidemiologia , Leucemia Mieloide Aguda/induzido quimicamente , Leucemia Mieloide Aguda/epidemiologia , Leucemia Mieloide Aguda/genética , Leucemia Mielomonocítica Aguda/induzido quimicamente , Leucemia Mielomonocítica Aguda/epidemiologia , Leucemia Mielomonocítica Aguda/genética , Masculino , Pessoa de Meia-Idade , MutaçãoRESUMO
Lymphoid cells contain specific receptors for glucocorticoids. We have used [3H]dexamethasone-21-mesylate to label covalently glucocorticoid receptors in rat thymic lymphocytes and in neoplastic cells obtained from patients with acute lymphoblastic leukemia and malignant lymphoma. The covalently labeled glucocorticoid receptors were identified by polyacrylamide gel electrophoresis (in the presence of 0.1% sodium dodecyl sulfate). In cytosolic fractions prepared from rat thymic lymphocytes, [3H]-dexamethasone-21-mesylate labels a protein (Mr approximately equal to 95,000) which was identified as the glucocorticoid receptor by the following criteria: (a) labeling of this moiety is inhibited by treatment with a 100-fold molar excess of glucocorticoids, such as dexamethasone and triamcinolone acetonide; and (b) the covalently labeled Mr approximately equal to 95,000 protein is activated (by heating at 20 degrees for 30 min) to a form that binds to DNA-cellulose. When intact thymocytes are treated with [3H]dexamethasone-21-mesylate, an Mr approximately equal to 95,000 moiety is also labeled covalently. Approximately 35% of the glucocorticoid receptors can be labeled covalently when intact thymocytes are treated with 100 nM [3H]dexamethasone-21-mesylate for 30 min at 4 degrees. Neoplastic cells from acute lymphoblastic leukemia and malignant lymphoma were treated with [3H]dexamethasone-21-mesylate. In all samples, an Mr approximately equal to 95,000 moiety was labeled covalently; labeling was inhibited by excess glucocorticoid. Smaller moieties were also identified by competition experiments; these may represent proteolytic fragments of the Mr approximately equal to 95,000 receptor. Thus, in rat and human lymphoid cells, [3H]dexamethasone-21-mesylate can be used to label covalently the glucocorticoid receptor.
Assuntos
Marcadores de Afinidade/metabolismo , Dexametasona/análogos & derivados , Leucemia Linfoide/metabolismo , Linfócitos/metabolismo , Linfoma/metabolismo , Receptores de Glucocorticoides/metabolismo , Receptores de Esteroides/metabolismo , Animais , Linhagem Celular , Dexametasona/metabolismo , Humanos , Cinética , Linfonodos/metabolismo , Masculino , Peso Molecular , Ratos , Receptores de Glucocorticoides/isolamento & purificação , Timo/metabolismoRESUMO
Glucocorticoid-receptor complexes in cytoplasm from normal lymphoid cells incubated with [3H]dexamethasone can be resolved into three different components. Two of these correspond to the well-established activated and nonactivated forms, while the third appears similar to the mero-receptor complex first described by Sherman et al. (Fed. Proc., 37: 167-173, 1978). Based on their differential affinities for DNA- and DEAE-cellulose in buffers of low ionic strength (the activated complex binds to DNA- and DEAE-cellulose; the nonactivated complex binds to only DEAE-cellulose; the mero-receptor-like complex binds to neither), we have developed a rapid minicolumn chromatographic procedure for separating these forms, and have applied it to examine the relative proportion of different complexes in cytosols from cells of leukemia patients. All samples from nine patients with chronic lymphocytic leukemia contained these three complexes in proportions similar to those seen with normal lymphoid tissue. Cytosols from six of eight specimens from patients with acute nonlymphocytic leukemia contained a lower proportion of activated complexes and a higher proportion of mero-receptor-like complexes than cytosols from normal or chronic lymphocytic leukemia cells. Whether such differences in the properties of cytosolic complexes are due to degradation taking place after the cells are broken, and whether they can be correlated to in vivo therapeutic response, is not known, but studies are in progress to answer these questions. The minicolumn procedure described here offers a simple and reliable method for these purposes.
Assuntos
Dexametasona/metabolismo , Leucemia/metabolismo , Linfócitos/metabolismo , Receptores de Glucocorticoides/metabolismo , Receptores de Esteroides/metabolismo , Adolescente , Adulto , Idoso , Cromatografia de Afinidade/métodos , Cromatografia DEAE-Celulose/métodos , Citosol/metabolismo , DEAE-Celulose , DNA , Feminino , Humanos , Indicadores e Reagentes , Cinética , Leucemia Linfoide/metabolismo , Masculino , Pessoa de Meia-Idade , Receptores de Glucocorticoides/isolamento & purificaçãoRESUMO
Glucocorticoid-receptor complexes in cytoplasm from normal lymphoid and leukemia cells incubated with glucocorticoid can be resolved into three different components, activated, nonactivated, and mero-receptor complexes, in relative amounts, dependent on the conditions to which the cells or cytosols are exposed. Recently, we reported that cytosols of acute nonlymphocytic leukemia (ANLL) cells contained high levels of mero-receptor complexes relative to those of chronic lymphocytic leukemia (CLL) or normal lymphoid cells. In the present study, we examined the cause for the lability of cytosolic complexes of ANLL cells. Mero-receptor accumulated rapidly in ANLL cytosols in a time-dependent fashion. The accumulation was most rapid in cytosols which contained activated receptor complexes, but it also occurred in cytosols containing only nonactivated receptor forms. Molybdate (20 mM) slowed but did not prevent the conversion to mero-receptor. Cytosols of ANLL specimens of the M4 French-American-British class (with monocytoid differentiation properties), in general, contained more stable complexes than did specimens of the M1 to M3 French-American-British classes (primarily myelocytic differentiation) suggesting that lability may in part be related to the state or direction of differentiation of the leukemic cells. In keeping with this hypothesis, cytosols of polymorphonuclear cells isolated from normal blood were much more labile than were those of monocytes. Mixing experiments with ANLL and CLL cells showed that the lability of ANLL complexes is not due simply to a higher content of proteolytic enzymes in these cells, because addition of ANLL cells or cytosols to CLL specimens did not result in increased mero-receptor. To the contrary, addition of CLL cells to ANLL specimens greatly stabilized the cytosolic complexes. These findings indicate the presence of an endogenous factor, present in CLL but lacking in ANLL cells, which is capable of stabilizing cytosolic complexes.
Assuntos
Dexametasona/metabolismo , Leucemia Linfoide/metabolismo , Leucemia/metabolismo , Proteínas de Neoplasias/fisiologia , Receptores de Glucocorticoides/metabolismo , Receptores de Esteroides/metabolismo , Triancinolona Acetonida/metabolismo , Doença Aguda , Medula Óssea/metabolismo , Citosol/metabolismo , Estabilidade de Medicamentos , Humanos , Cinética , Monócitos/metabolismo , Ligação ProteicaRESUMO
Cytogenetic study of short-term cultures from 10 adipose tissue tumors (eight lipomas, one myxoid liposarcoma, and one mixed liposarcoma) have revealed clonal chromosome abnormalities in seven cases. In both malignant tumors, translocation (12;16) was the sole aberration, and in the mixed liposarcoma, the breakpoints could be sublocalized to bands 12q13.3 and 16p11.2, thus confirming findings of Eneroth et al., Cancer Genet. Cytogenet., 48: 101-107, 1990. Three lipomas displayed predominantly normal karyotypes; in a fourth case, the karyotype 44,XX,-6,der (7)t(6;7)(p21.3-22;p22)ins(7)(p22q11.2q22),-13 was found. Four remaining lipomas were characterized by structural rearrangements of chromosome 12. We were able to achieve high resolution banding patterns in two tumors with translocations (3;12)(q28;q15) and (1;2;12)(p36.;q13;q15). In both of these cases, the chromosome 12 breakpoint could be unequivocally assigned to band q15. Similarly, band 12q15 was also rearranged in two other lipomas with translocations (12;14)(q15;q32) and (12;20)(q15;q13.1). Our results support the hypothesis that the chromosome 12 breakpoint in lipomas is located more distally than the breakpoint in myxoid liposarcomas and some other soft-tissue malignant neoplasms and that it is cytogenetically identical with breakpoints detected in such benign tumors as uterine leiomyoma and pleomorphic adenoma of the salivary gland.
Assuntos
Bandeamento Cromossômico , Cromossomos Humanos Par 12 , Lipoma/genética , Lipossarcoma/genética , Neoplasias Cutâneas/genética , Translocação Genética , Adulto , Idoso , Cromossomos Humanos Par 1 , Cromossomos Humanos Par 14 , Cromossomos Humanos Par 16 , Cromossomos Humanos Par 20 , Cromossomos Humanos Par 3 , Cromossomos Humanos Par 7 , Feminino , Humanos , Cariotipagem , Masculino , Pessoa de Meia-IdadeRESUMO
Glucocorticoid receptors were studied in leukemic cells from 14 newly diagnosed adults with acute lymphoblastic leukemia. Receptors were found in all cases; total receptor levels ranged from 1,348 to 18,697 sites per cel (median, 7,553). To determine the utility of glucocorticoid receptor levels for predicting response to glucocorticoid therapy, lymphoblasts were studied for receptors on nine occasions, and the patients were then treated with dexamethasone as a single agent. Response to glucocorticoid therapy appeared to correlate with lymphoblast receptor level. Five of six patients with greater than 4,500 total receptor sites per cell responded; all three patients with less than 4,500 receptors failed to respond. Glucocorticoid receptor level did not correlate with response to combination chemotherapy, duration of remission, or survival. Glucocorticoid receptors were studied at the initial presentation and following relapse in five patients. In two patients, the receptor level did not change following therapy; both patients at diagnosis had leukemias resistant to glucocorticoids. Three patients at relapse developed leukemias with lower receptor levels; these patients had leukemias sensitive to glucocorticoids at diagnosis. Our data suggest that the glucocorticoid receptor levels of lymphoblasts may be useful in predicting response to glucocorticoid therapy in adult acute lymphoblastic leukemia. They also suggest that, following relapse, cases initially sensitive to glucocorticoid may develop resistance by selection of cells with fewer receptors.
Assuntos
Leucemia Linfoide/análise , Receptores de Glucocorticoides/análise , Receptores de Esteroides/análise , Doença Aguda , Adolescente , Adulto , Idoso , Antineoplásicos/uso terapêutico , Dexametasona/uso terapêutico , Quimioterapia Combinada , Feminino , Humanos , Leucemia Linfoide/tratamento farmacológico , Masculino , Pessoa de Meia-Idade , Prednisona/uso terapêutico , PrognósticoRESUMO
The partial tandem duplication (PTD) of ALL1 (MLL) is one of the more common molecular abnormalities in adult de novo acute myeloid leukemia (AML) and carries a poor prognosis. The PTD of ALL1 is identified in leukemic blasts at the genomic level by ALL1 rearrangement upon Southern analysis and by genomic fusion following DNA PCR. The genomic defect encodes for a unique fusion transcript that is readily detected by reverse transcription-PCR (RT-PCR). To determine if the ALL1 fusion transcript is specific for leukemic blasts or instead can be found with any frequency in normal cells, we analyzed 52 bone marrow and 8 peripheral blood samples from 60 normal donors by nested RT-PCR. Ten of 60 samples (16%; 7 bone marrow and 3 peripheral blood) contained a unique transcript showing a fusion of two ALL1 exons that was consistent with the PTD of ALL1. However, a corresponding genomic rearrangement or a unique genomic fusion of ALL1 could not be demonstrated by Southern analysis or DNA PCR, respectively. Marked differences were observed in the size and sequence of the ALL1 fusion transcripts detected in normal donors, compared to those detected in leukemic patients with a PTD of ALL1. Moreover, although the ALL1 fusion transcripts seen in AML always maintain the open reading frame, the open reading frame was preserved in only 5 of 10 fusion transcripts from normal donors. Finally, in contrast to leukemic blasts with the PTD of ALL1, the fusion transcripts in normal cells could not be detected in the poly(A)+ RNA fraction by RT-PCR. In summary, the origin and the composition of the ALL1 fusion transcripts found in normal cells appear to be distinct from those found in the leukemic cells. The data accumulated thus far suggest that the ALL1 fusion product detected in normal tissue results from the process of differential mRNA splicing rather than true ALL1 gene rearrangement. These findings also suggest caution in the use of RT-PCR for detection of minimal residual disease in AML patients with the PTD of ALL1.
Assuntos
Medula Óssea/química , Proteínas de Ligação a DNA/sangue , Leucemia Mieloide/metabolismo , Família Multigênica , Proto-Oncogenes , Fatores de Transcrição , Doença Aguda , Antígenos CD34 , Sequência de Bases , Proteínas de Ligação a DNA/análise , Células-Tronco Hematopoéticas/química , Histona-Lisina N-Metiltransferase , Humanos , Leucemia Mieloide/genética , Proteína de Leucina Linfoide-Mieloide , RNA/análise , Mapeamento por RestriçãoRESUMO
Leukemic cells from 32 cases of acute leukemia were cultured in vitro with the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) to study their differentiative potential. Three cases of acute undifferentiated leukemia (AUL) were studied intensively. We found that culturing of leukemic cells with TPA can induce changes in cell surface antigens. In particular, MCS-2, a "pan" granulocyte/monocyte marker, was inducible in vitro in AUL and in acute myelogenous leukemia, while it was not inducible in acute lymphoblastic leukemia. BA-2 (recognizing the Mr 24,000 protein) and TA-1 (recognizing the Mr 170,000 and Mr 95,000 proteins) were also inducible in cases of AUL, acute myelocytic leukemia, and acute monoblastic leukemia, although these antigens are not limited only to leukemias of the myelomonocytic lineage. Our studies also indicate that undifferentiated cells could be induced to nonspecific esterase and sometimes to chloroacetate esterase reactivity while losing terminal deoxynucleotidyl transferase. Morphological studies in these cases revealed cytological maturation following TPA treatment. In most cases, these changes were also partially inducible by culturing cells in medium alone or with the addition of dimethyl sulfoxide but not to the extent that was demonstrated by TPA. Our studies showed that MCS-2 is a very good, specific marker of acute nonlymphocytic leukemia. A potential use for TPA to aid in the subclassification of patients with AUL is also suggested.
Assuntos
Anticorpos Monoclonais , Antígenos de Superfície/análise , Leucemia/fisiopatologia , Leucócitos/imunologia , Forbóis/toxicidade , Acetato de Tetradecanoilforbol/toxicidade , Doença Aguda , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Imunofluorescência , Humanos , Leucemia/imunologia , Leucócitos/efeitos dos fármacosRESUMO
We have correlated immunological characteristics and karyotypic abnormalities from lymphomas in 118 patients. T-lymphomas differed significantly from B- and non-B-, non-T-lymphomas in having more normal metaphases, trisomy 19, and breaks at 1q21, 2q21, 3q27, 4q21, and 17q21 (P less than or equal to 0.03). Non-T-lymphomas had breaks in 18q in one-half the cases, but only one of 11 T-lymphomas had such breaks (P = 0.02). Among B-lymphomas, specific chromosome abnormalities were associated with the type of immunoglobulin heavy but not light chain expressed. A break at 14q22 or q24 was associated with surface delta mu-immunoglobulin (P = 0.02); trisomy 22 or a break in 22q and a break at 2q32 was associated with surface gamma-immunoglobulin (P less than 0.001); and trisomy 12 and a break at 2p13 was associated with cytoplasmic gamma-immunoglobulin (P less than 0.01). Among B-lymphomas, several cytogenetic abnormalities were associated (P less than or equal to 0.02) with expression of CD24 or CD9 surface antigens. Lack of CD24 was associated with breaks in 2p25, 5q, and 6q21; CD9 was associated with a break at 6q15. Associations with a specific immunological phenotype were not identified for cytogenetic abnormalities involving a band to which genes encoding immunoglobulin or the T-cell receptor have been localized. Breaks were common at 14q32, the genomic site of the immunoglobulin heavy chain loci, in B-, non-B-, non-T-, and T-lymphomas. In T-lymphomas this may be because this is the site of the AKT1 oncogene. Breaks were uncommonly found at the light chain loci or the genomic sites encoding the T-cell receptor. However, the recurring breakpoints associated with T-lymphomas were commonly found on chromosomes to which genes coding for various T-cell antigens have recently been provisionally assigned.
Assuntos
Aberrações Cromossômicas , Linfoma/imunologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígenos de Diferenciação de Linfócitos T , Antígenos de Superfície/análise , Criança , Cromossomos Humanos Par 14 , Cromossomos Humanos Par 19 , Cromossomos Humanos Par 2 , Feminino , Humanos , Imunoglobulina G/análise , Imunoglobulina M/análise , Imunoglobulinas/genética , Cariotipagem , Linfoma/genética , Masculino , Pessoa de Meia-Idade , Fenótipo , Receptores de Antígenos de Linfócitos B/análise , Receptores de Antígenos de Linfócitos T/genética , TrissomiaRESUMO
Rearrangements of the ALL-1 gene by reciprocal translocations involving chromosome band 11q23 are frequently associated with human acute leukemia. We have previously reported the detection of ALL-1 gene rearrangements in adult patients with acute myeloid leukemia lacking cytogenetic evidence of 11q23 translocations. These included 2 of 19 patients with normal karyotypes as well as 3 of 4 patients with trisomy 11 as a sole cytogenetic abnormality. Rearrangement of the ALL-1 genes in two of the patients with trisomy 11 was shown to result from a direct tandem duplication of a portion of the gene spanning exons 2-6. Here we report the characterization of the ALL-1 gene rearrangement in one of the previously reported acute myeloid leukemia patients with a normal karyotype. ALL-1 rearrangement in this patient results from a direct tandem duplication of a portion of the gene spanning exons 2-8. RNA polymerase chain reaction and DNA sequence analysis show that the partially duplicated ALL-1 gene is transcribed into mRNA capable of encoding a partially duplicated protein. Sequence analysis of the genomic fusion region provides evidence for Alu-mediated homologous recombination as a mechanism for partial duplication of the ALL-1 gene.
Assuntos
Cromossomos Humanos Par 11 , Éxons/genética , Rearranjo Gênico/genética , Leucemia Mieloide/genética , Proto-Oncogenes/genética , Sequências Repetitivas de Ácido Nucleico/genética , Doença Aguda , Sequência de Bases , Southern Blotting , Humanos , Dados de Sequência Molecular , Reação em Cadeia da PolimeraseRESUMO
The 11q23 chromosome band is frequently associated with chromosomal aberrations in human leukemias. We have previously cloned a DNA fragment derived from chromosome 11 which could be used as a probe to detect rearrangements in DNAs from the leukemic cells of patients with the t(4;11), t(9;11), and t(11;19) translocations. In this study we now show that the same probe detects DNA rearrangements in malignant cells from patients with the t(1;11), t(6;11), t(10;11), and del (11q23) chromosomal abnormalities. A second probe obtained from a region located centrometric to the breakpoint cluster detects major and minor transcripts of 12.5 and 11.5 kilobases, respectively, in all cell lines examined. The same probe identifies an altered 11-kilobase RNA in all three independent cell lines with the t(4;11)(q21;q23) chromosome translocation.
Assuntos
Cromossomos Humanos Par 11 , Cromossomos Humanos Par 4 , Leucemia/genética , Transcrição Gênica , Translocação Genética , Humanos , Células Tumorais CultivadasRESUMO
Expression of P-glycoprotein has been linked to multidrug resistance in cancer cell lines and human tumors. We investigated the frequency and clinical significance of P-glycoprotein immunoreactivity in 57 previously untreated diffuse large cell and immunoblastic lymphomas. Banked frozen tissue, which had been obtained prior to chemotherapy, was tested for reactivity with 2 monoclonal antibodies (MRK16 and C219) that recognize different domains of P-glycoprotein, using an immunoperoxidase technique. Thirteen of 57 lymphomas (23%) showed strong staining of greater than 50% of neoplastic cells; 15 of 57 (26%) showed labeling of a minority (11-50%) of neoplastic lymphocytes; 14 of 57 (25%) yielded equivocal results (reactivity in less than 10% of cells); and 15 of 57 (26%) were negative for P-glycoprotein. The 2 monoclonal antibodies were comparable in reactivity. Expression of MDR-1 mRNA was determined in 6 cases with sufficient available tissue, and did not correlate well with the percentages of cells reactive for P-glycoprotein by immunohistochemistry. Thirty-nine of our 57 patients completed multiagent chemotherapy. Contrary to our expectations, we found that P-glycoprotein immunoreactivity did not decrease the likelihood of response to induction chemotherapy. Median survival also was not adversely affected.
Assuntos
Resistência a Medicamentos , Linfoma Difuso de Grandes Células B/metabolismo , Linfoma Imunoblástico de Células Grandes/metabolismo , Glicoproteínas de Membrana/análise , Sistema ABO de Grupos Sanguíneos , Membro 1 da Subfamília B de Cassetes de Ligação de ATP , Resistência a Medicamentos/genética , Humanos , Imuno-Histoquímica , Linfoma Difuso de Grandes Células B/tratamento farmacológico , Linfoma Imunoblástico de Células Grandes/tratamento farmacológico , Glicoproteínas de Membrana/imunologia , RNA Mensageiro/análiseRESUMO
G-banded chromosomes were studied from involved lymph nodes or other tumor masses in 94 patients with malignant lymphoma. Clonal chromosome abnormalities were identified in 91 patients including all 81 B-lymphomas but only 6 of 9 T-lymphomas. Many recurring chromosome abnormalities were found. Most common numerical alterations involved gains of chromosomes 12 (19% of patients), 18 (13%), 7 (12%), and 21 (10%). Structural abnormalities, which were more frequent than numerical alterations, most commonly involved chromosome regions 14q (71% of patients), 18q (36%), 6q (31%), 1p (24%), and 8q (19%). Seven recurring translocations were identified, and all except one involved 14q32. The most frequent were t(14;18)(q32;q21) in 22 patients, t(8;14)(q24;q32) in 9 patients, and t(1;14)(q42;q32) in 3 patients. Deletions most frequently involved the long arm of chromosome 6 at band q21 (11 patients) or q23 (7 patients). The common recurring chromosome abnormalities were correlated with histology (International Working Formulation for Clinical Usage) and immunological phenotype. Four abnormalities were significantly associated with specific histologies. Eighteen (82%) patients with t(14;18)(q32;q21) were follicular. Similarly, 82% of patients with del(6)(q21) were large cell lymphomas. Lymphomas with trisomy 7 were either diffuse large cell or follicular, while patients with t(8;14)(q24;q32) were either diffuse large cell or small noncleaved cell. A significant association with immunological phenotype was seen for t(14;18) only. All patients were either B- or complement lymphomas, and the heavy chain(s) was more commonly gamma and less frequently delta mu than among the total B-lymphoma population. We conclude that essentially all lymphomas have cytogenetic abnormalities; further study is required to determine their significance. Particularly, it will be of interest to see if oncogenes are found in the regions of these chromosome abnormalities.