RESUMO
The diagnosis of malignant lymphoma is a recognized difficult area in histopathology. Therefore, detection of clonality in a suspected lymphoproliferation is a valuable diagnostic criterion. We have developed primer sets for the detection of rearrangements in the B- and T-cell receptor genes as reliable tools for clonality assessment in lymphoproliferations suspected for lymphoma. In this issue of Leukemia, the participants of the BIOMED-2 Concerted Action CT98-3936 report on the validation of the newly developed clonality assays in various disease entities. Clonality was detected in 99% of all B-cell malignancies and in 94% of all T-cell malignancies, whereas the great majority of reactive lesions showed polyclonality. The combined BIOMED-2 results are summarized in a guideline, which can now be implemented in routine lymphoma diagnostics. The use of this standardized approach in patients with a suspect lymphoproliferation will result in improved diagnosis of malignant lymphoma.
Assuntos
Linfoma/genética , Linfoma/patologia , Reação em Cadeia da Polimerase/métodos , Reações Falso-Negativas , Rearranjo Gênico , Humanos , Linfoma de Células B/genética , Linfoma de Células B/patologia , Linfoma de Células T/genética , Linfoma de Células T/patologia , Receptores de Antígenos de Linfócitos T/genética , Reprodutibilidade dos TestesRESUMO
The flow cytometric detection of minimal residual disease (MRD) in precursor-B-acute lymphoblastic leukemias (precursor-B-ALL) mainly relies on the identification of minor leukemic cell populations that can be discriminated from their normal counterparts on the basis of phenotypic aberrancies observed at diagnosis. This technique is not very complex, but discordancies are frequently observed between laboratories, due to the lack of standardized methodological procedures and technical conditions. To develop standardized flow cytometric techniques for MRD detection, a European BIOMED-1 Concerted Action was initiated with the participation of laboratories from six different countries. The goal of this concerted action was to define aberrant phenotypic profiles in a series of 264 consecutive de novo precursor-B-ALL cases, systematically studied with one to five triple-labelings (TdT/CD10/CD19, CD10/CD20/CD19, CD34/CD38/CD19, CD34/CD22/CD19 and CD19/CD34/CD45) using common flow cytometric protocols in all participating laboratories. The use of four or five triple-stainings allowed the identification of aberrant phenotypes in virtually all cases tested (127 out of 130, 98%). These phenotypic aberrancies could be identified in at least two and often three triple-labelings per case. When the analysis was based on two or three triple-stainings, lower incidences of aberrancies were identified (75% and 81% of cases, respectively) that could be detected in one and sometimes two triple-stainings per case. The most informative triple staining was the TdT/CD10/CD19 combination, which enabled the identification of aberrancies in 78% of cases. The frequencies of phenotypic aberrations detected with the other four triple-stainings were 64% for CD10/CD20/CD19, 56% for CD34/CD38/CD19, 46% for CD34/CD22/CD19, and 22% for CD19/CD34/CD45. In addition, cross-lineage antigen expression was detected in 45% of cases, mainly coexpression of the myeloid antigens CD13 and/or CD33 (40%). Parallel flow cytometric studies in different laboratories finally resulted in highly concordant results (>90%) for all five antibody combinations, indicating the high reproducibility of our approach. In conclusion, the technique presented here with triple-labelings forms an excellent basis for standardized flow cytometric MRD studies in multicenter international treatment protocols for precursor-B-ALL patients.
Assuntos
Linfócitos B/imunologia , Linfoma de Burkitt/imunologia , Antígenos CD/imunologia , Linfócitos B/patologia , Linfoma de Burkitt/patologia , Citometria de Fluxo/normas , Humanos , Imunofenotipagem , Padrões de Referência , Valores de ReferênciaRESUMO
The European BIOMED-1 Concerted Action was initiated in 1994 to improve and standardize the flow cytometric detection of minimal residual disease (MRD) in acute leukemia (AL). Three different protocols were defined to identify the normal subsets of B, T and myeloid cells in bone marrow (BM), and were applied to the different types of AL in order to study aberrant immunophenotypes. Using sensitive acquisition methods ('live gate') T cell subsets in normal BM could be identified with five triple-stains: CD7/CD5/CD3, CD7/CD4/CD8, CD7/CD2/CD3, CD7/CD38/CD34 and TdT/CD7/surface or cytoplasmic (cy)CD3 (antibodies conjugated with FITC/PE/PECy5 or PerCP, respectively). The identification of T cell subsets in BM allowed definition of 'empty spaces' (ie areas of flow cytometric plots where normally no cells are found). All studied T-ALL cases (n = 65) were located in 'empty spaces' and could be discriminated from normal T cells. The most informative triple staining was TdT/CD7/cyCD3, which was aberrant in 91% of T-ALL cases. In most cases, two or more aberrant patterns were found. Apparently the immunophenotypes of T-ALL differ significantly from normal BM T cells. This is mostly caused by their thymocytic origin, but also the neoplastic transformation might have affected antigen expression patterns. Application of the five proposed marker combinations in T-ALL contributes to standardized detection of MRD, since cells persistent or reappearing in the 'empty spaces' can be easily identified in follow-up BM samples during and after treatment.
Assuntos
Antígenos CD/análise , Citometria de Fluxo/normas , Leucemia-Linfoma de Células T do Adulto/diagnóstico , Neoplasia Residual , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/patologia , Biomarcadores/análise , Células da Medula Óssea/imunologia , Células da Medula Óssea/patologia , Citometria de Fluxo/métodos , Humanos , Leucemia-Linfoma de Células T do Adulto/imunologia , Leucemia-Linfoma de Células T do Adulto/patologia , Controle de QualidadeRESUMO
Prospective studies on the detection of minimal residual disease (MRD) in acute leukemia patients have shown that large-scale MRD studies are feasible and that clinically relevant MRD-based risk group classification can be achieved and can now be used for designing new treatment protocols. However, multicenter international treatment protocols with MRD-based stratification of treatment need careful standardization and quality control of the MRD techniques. This was the aim of the European BIOMED-1 Concerted Action 'Investigation of minimal residual disease in acute leukemia: international standardization and clinical evaluation' with participants of 14 laboratories in eight European countries (ES, NL, PT, IT, DE, FR, SE and AT). Standardization and quality control was performed for the three main types of MRD techniques, ie flow cytometric immunophenotyping, PCR analysis of antigen receptor genes, and RT-PCR analysis of well-defined chromosomal aberrations. This study focussed on the latter MRD technique. A total of nine well-defined chromosome aberrations with fusion gene transcripts were selected: t(1;19) with E2A-PBX1, t(4;11) with MLL-AF4, t(8;21) with AML1-ETO, t(9;22) with BCR-ABL p190 and BCR-ABL p210, t(12;21) with TEL-AML1, t(15;17) with PML-RARA, inv (16) with CBFB-MYH11, and microdeletion 1p32 with SIL-TAL1. PCR primers were designed according to predefined criteria for single PCR (external primers A <--> B) and nested PCR (internal primers C <--> D) as well as for 'shifted' PCR with a primer upstream (E5' primer) or downstream (E3' primer) of the external A <--> B primers. The 'shifted' E primers were designed for performing an independent PCR together with one of the internal primers for confirmation (or exclusion) of positive results. Various local RT and PCR protocols were compared and subsequently a common protocol was designed, tested and adapted, resulting in a standardized RT-PCR protocol. After initial testing (with adaptations whenever necessary) and approval by two or three laboratories, the primers were tested by all participating laboratories, using 17 cell lines and patient samples as positive controls. This testing included comparison with local protocols and primers as well as sensitivity testing via dilution experiments. The collaborative efforts resulted in standardized primer sets with a minimal target sensitivity of 10-2 for virtually all single PCR analyses, whereas the nested PCR analyses generally reached the minimal target sensitivity of 10-4. The standardized RT-PCR protocol and primer sets can now be used for molecular classification of acute leukemia at diagnosis and for MRD detection during follow-up to evaluate treatment effectiveness.
Assuntos
Aberrações Cromossômicas , Leucemia/genética , Proteínas de Fusão Oncogênica/genética , RNA Mensageiro/análise , Reação em Cadeia da Polimerase Via Transcriptase Reversa/normas , Doença Aguda , Proteínas de Fusão bcr-abl/genética , Humanos , Leucemia/diagnóstico , Neoplasia ResidualRESUMO
During the last two decades, major progress has been made in the technology of flow cytometry and in the availability of a large series of monoclonal antibodies against surface membrane and intracellular antigens. Flow cytometric immunophenotyping has become a diagnostic tool for the analysis of normal and malignant leukocytes and it has proven to be a reliable approach for the investigation of minimal residual disease (MRD) in leukemia patients during and after treatment. In order to standardize the flow cytometric detection of MRD in acute leukemia, a BIOMED-1 Concerted Action was initiated with the participation of six laboratories in five different European countries. This European co-operative study included the immunophenotypic characterization and enumeration of different precursor and mature B cell subpopulations in normal bone marrow (BM). The phenotypic profiles in normal B cell differentiation may form a frame of reference for the identification of aberrant phenotypes of precursor-B cell acute lymphoblastic leukemias (precursor-B-ALL) and may therefore be helpful in MRD detection. Thirty-eight normal BM samples were analyzed with five different pre-selected monoclonal antibody combinations: CD10/CD20/CD19, CD34/CD38/CD19, CD34/CD22/CD19, CD19/CD34/CD45 and TdT/CD10/CD19. Two CD19- immature subpopulations which coexpressed B cell-associated antigens were identified: CD34+/CD22+/CD19- and TdT+/CD10+/CD19-, which represented 0.11 +/- 0.09% and 0.04 +/- 0.05% of the total BM nucleated cells, respectively. These immunophenotypes may correspond to the earliest stages of B cell differentiation. In addition to these minor subpopulations, three major CD19+ B cell subpopulations were identified, representing three consecutive maturation stages; CD19dim/CD34+/TdT+/CD10bright/CD22dim/CD45dim /CD38bright/CD20- (subpopulation 1), CD19+/CD34-/TdT-/CD10+/CD22dim/CD45+/CD38bright/ CD20dim (subpopulation 2) and CD19+/CD34-/TdT-/CD10-/CD22bright/CD45bright/ CD38dim/CD20bright (subpopulation 3). The relative sizes of subpopulations 1 and 2 were found to be age related: at the age of 15 years, the phenotypic precursor-B cell profile in BM changed from the childhood 'immature' profile (large subpopulations 1 and 2/small subpopulation 3) to the adult 'mature' profile (small subpopulation 1 and 2/large subpopulation 3). When the immunophenotypically defined precursor-B cell subpopulations from normal BM samples are projected in fluorescence dot-plots, templates for the normal B cell differentiation pathways can be defined and so-called 'empty spaces' where no cell populations are located become evident. This allows discrimination between normal and malignant precursor-B cells and can therefore be used for MRD detection.
Assuntos
Linfócitos B/patologia , Diferenciação Celular , Neoplasia Residual/diagnóstico , Leucemia-Linfoma Linfoblástico de Células Precursoras B/patologia , Adolescente , Adulto , Linfócitos B/imunologia , Criança , Pré-Escolar , Citometria de Fluxo , Células-Tronco Hematopoéticas/imunologia , Humanos , Imunofenotipagem , Pessoa de Meia-Idade , Leucemia-Linfoma Linfoblástico de Células Precursoras B/imunologia , Reprodutibilidade dos TestesRESUMO
In a European BIOMED-2 collaborative study, multiplex PCR assays have successfully been developed and standardized for the detection of clonally rearranged immunoglobulin (Ig) and T-cell receptor (TCR) genes and the chromosome aberrations t(11;14) and t(14;18). This has resulted in 107 different primers in only 18 multiplex PCR tubes: three VH-JH, two DH-JH, two Ig kappa (IGK), one Ig lambda (IGL), three TCR beta (TCRB), two TCR gamma (TCRG), one TCR delta (TCRD), three BCL1-Ig heavy chain (IGH), and one BCL2-IGH. The PCR products of Ig/TCR genes can be analyzed for clonality assessment by heteroduplex analysis or GeneScanning. The detection rate of clonal rearrangements using the BIOMED-2 primer sets is unprecedentedly high. This is mainly based on the complementarity of the various BIOMED-2 tubes. In particular, combined application of IGH (VH-JH and DH-JH) and IGK tubes can detect virtually all clonal B-cell proliferations, even in B-cell malignancies with high levels of somatic mutations. The contribution of IGL gene rearrangements seems limited. Combined usage of the TCRB and TCRG tubes detects virtually all clonal T-cell populations, whereas the TCRD tube has added value in case of TCRgammadelta(+) T-cell proliferations. The BIOMED-2 multiplex tubes can now be used for diagnostic clonality studies as well as for the identification of PCR targets suitable for the detection of minimal residual disease.
Assuntos
Imunoglobulinas/genética , Transtornos Linfoproliferativos/diagnóstico , Transtornos Linfoproliferativos/genética , Reação em Cadeia da Polimerase/métodos , Reação em Cadeia da Polimerase/normas , Receptores de Antígenos de Linfócitos T/genética , Aberrações Cromossômicas , Células Clonais , Primers do DNA , União Europeia , Rearranjo Gênico do Linfócito T , Humanos , Neoplasia Residual/diagnóstico , Neoplasia Residual/genética , Padrões de Referência , Reprodutibilidade dos TestesRESUMO
Peripheral blood stem cells (PBSC) are increasingly being used as an alternative to autologous bone marrow (BM) for hematologic rescue after high-dose chemoradiotherapy in the treatment of hematologic and nonhematologic malignancies. Mobilization procedures such as chemotherapy and/or hematopoietic growth factor administration are employed to allow for the graft enrichment in hematopoietic stem cells and progenitors and to accelerate trilineage recovery after transplant. The influence of these mobilization procedures on the lymphoid populations in the graft and on immunologic recovery after transplant remains to be determined. We studied six consecutive patients undergoing PBSC high-volume collections after cyclophosphamide (Cyc) and granulocyte colony-stimulating factor (G-CSF) administration and observed that NK cell numbers (phenotypically defined as CD3-CD56+ by flow cytometry) and activity (evaluated by a 51Cr release assay) fully recovered after 4-5 weeks; high numbers of functionally active NK cells (42.1-212.1 x 10(6)/kg b.w.) were present in the grafts, and their percentage and cytotoxic activity rose from the beginning to the end of the harvesting procedure in most cases. CD3-CD56+ and CD34+ cell numbers peaked at the same time point during harvesting, which differed from one patient to another. T (CD3+) cells were always present during harvest, and CD4 and CD8 numbers showed interdonor variability. When we cultured leukapheresed PBSC in the presence of interleukin-2 (IL-2) (10-1000 U/mL) for 6-8 days, we were able to expand the NK population three- to 5.4-fold; 100 U/mL appears to be the best concentration to generate high numbers of cytotoxic NK cells. Pilot studies also suggest that this short exposure to IL-2 does not affect the CD34+ cells. We conclude that PBSC grafts mobilized by combined Cyc and G-CSF and harvested through high-volume leukapheresis contain high numbers of cytotoxic NK cells that can be expanded in vitro by exposure to IL-2. In the setting of PBSC transplant, ex vivo immunomodulation aimed at increasing the NK cell numbers and activity is feasible and may prove to be useful in inducing a graft-vs.-tumor effect, thereby decreasing the relapse rate after transplant.
Assuntos
Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/patologia , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/patologia , Contagem de Linfócitos , Adulto , Neoplasias da Mama/terapia , Células Cultivadas , Ciclofosfamida/uso terapêutico , Feminino , Fator Estimulador de Colônias de Granulócitos/uso terapêutico , Humanos , Interleucina-2/farmacologia , Leucaférese , Linfoma não Hodgkin/terapia , Masculino , Pessoa de Meia-Idade , Mieloma Múltiplo/terapia , Transplante AutólogoRESUMO
Cells of the human promyelocytic leukaemia line HL-60 may be induced to differentiate along the monocytic lineage by the seco-steroid hormone 1,25-dihydroxyvitamin D3. We used fluorescence-activated cell sorting to show that, in addition to inducing differentiation, the hormone caused HL-60 cells to accumulate in G1/G0 phases of the cell cycle. We also labelled differentiating cells with both a monocyte-specific antibody and a DNA stain simultaneously. Analysis of these cell populations showed that, although cells acquired the differentiated phenotype irrespective of their position in the cell cycle, they eventually became arrested in G1/G0 as a consequence of differentiation. The normal relationship between differentiation and growth which is lost on malignant transformation is therefore restored by treatment of HL-60 cells with 1,25-dihydroxyvitamin D3.
Assuntos
Calcitriol/farmacologia , Leucemia Mieloide Aguda/patologia , Diferenciação Celular/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Linhagem Celular , Citometria de Fluxo , Humanos , InterfaseRESUMO
A cytochemical study using: Sudan black B; alpha-naphthyl acetate (ANAE) staining; estimation of alpha-naphthyl butyrate (ANBE) esterase activity; acid phosphatase activity; and 5' nucleotidase activity was carried out in 15 cases of megakaryoblastic leukaemia. These included cases of M7 acute myeloid leukaemia and blast crises of chronic granulocytic leukaemia. The megakaryoblastic nature of the blasts was first established using two monoclonal antibodies against platelet glycoproteins, and by estimating the platelet/peroxidase reaction at ultrastructural level. Our findings suggest that megakaryoblasts have a typical cytochemical profile comprising positive ANAE staining and acid phosphatase activity with a predominant localisation in the Golgi zone and negative or weak ANBE activity. A similar positive cytochemical pattern was also found in five cases of erythroleukaemia (M6). The specificity of the 5'nucleotidase activity for megakaryoblasts was not confirmed. In most cases of megakaryoblastic leukaemia there was no 5'nucleotidase activity only two cases showed positive reactions--reactions were positive in several cases of myeloblastic and lymphoblastic leukaemia. We suggest that cytochemical methods may be useful in diagnosing M6 and M7 acute leukaemia because less than 40% of leukaemic cells react with specific monoclonal antibodies.
Assuntos
Fosfatase Ácida/análise , Hidrolases de Éster Carboxílico/análise , Leucemia Megacarioblástica Aguda/metabolismo , Nucleotidases/análise , 5'-Nucleotidase , Adulto , Idoso , Anticorpos Monoclonais , Antígenos de Superfície/análise , Humanos , Leucemia Megacarioblástica Aguda/imunologia , Masculino , Megacariócitos/análise , Pessoa de Meia-IdadeRESUMO
AIMS: To study the expression of the human T lymphotropic virus (HTLV) Tax gene in peripheral blood mononuclear cells. METHODS: Blood was collected from 72 patients with lymphoproliferative disorders. Serum from all patients was assayed for antibodies directed against HTLV-I structural proteins by ELISA and western blotting. RNA was purified from fresh blood cells and amplified by reverse transcription polymerase chain reaction (RT-PCR). After Southern blotting, the PCR products were hybridised with a 32P end-labelled probe specific for the Tax gene. RESULTS: All samples were seronegative. A specific band for the Tax gene was found in five samples. Each of the patients positive for Tax gene expression had a different type of lymphoproliferative disorder. CONCLUSIONS: Infection by HTLV-I cannot be assessed solely by immunological assays, particularly when only disrupted virions are used. Sensitive molecular biology assays are essential for detecting viral gene expression in fresh blood cells.
Assuntos
Genes pX/fisiologia , Genes pol/fisiologia , Infecções por HTLV-I/virologia , Doença de Hodgkin/virologia , Vírus Linfotrópico T Tipo 1 Humano/isolamento & purificação , Linfoma não Hodgkin/virologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Anticorpos Antivirais/sangue , Vírus Linfotrópico T Tipo 1 Humano/genética , Vírus Linfotrópico T Tipo 1 Humano/imunologia , Humanos , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Transcrição GênicaRESUMO
AIMS: To evaluate the overall incidence of immunoglobulin (Ig) and T cell receptor (TCR) beta and gamma gene rearrangements in a series of 40 cases of acute myeloid leukaemia (AML) and to determine whether structural modifications of these genes could be correlated with the abnormal expression of lymphoid markers in malignant cells. METHODS: All cases were classified according to the criteria of the FAB group and immunophenotyped with a panel of monoclonal antibodies reactive with myeloid and lymphoid differentiation antigens. DNA analysis was performed by the method of Southern using probes for the Ig JH, TCR-C beta 1, and TCR-J tau 1 regions. RESULTS: Phenotypic analysis showed that in addition to myeloid markers, 10 cases expressed lymphoid antigens: CD7 in seven (of which three were TdT positive, one CD2 positive, and one CD19 positive) and CD19 in three. Southern blot analysis showed that bands with sizes different from the germ line control were present in the TCR beta genes in 11 cases: in six of 30 with pure myeloid phenotype and in five of 10 of those expressing lymphoid markers. A close observation of the size and patterns of those bands, however, showed that they could be artefactual. Indeed, further analysis showed that they were either due to resistant Eco RI/Hind III sites at the beta locus or to plasmid contamination. Rearranged genes were eventually found in only two of the 40 cases: at the Ig JH region in one of the 30 with pure myeloid phenotype (3.3%) and at the TCR gamma genes in one of 10 with lymphoid markers (10%). CONCLUSIONS: These observations showed that Ig/TCR gene rearrangements were rare in this AML series (overall incidence of 5%) and that they were not significantly more common in cases with aberrant expression of lymphoid markers. The size and pattern of the potential non-germline bands that can be found in these loci must be carefully evaluated.
Assuntos
Rearranjo Gênico de Cadeia Pesada de Linfócito B/genética , Rearranjo Gênico da Cadeia beta dos Receptores de Antígenos dos Linfócitos T/genética , Rearranjo Gênico da Cadeia gama dos Receptores de Antígenos dos Linfócitos T/genética , Leucemia Mieloide/genética , Doença Aguda , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígenos CD/análise , Antígenos CD7 , Antígenos de Diferenciação de Linfócitos T/análise , Antígenos de Neoplasias/análise , Southern Blotting , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Leucemia Mieloide/imunologia , Masculino , Pessoa de Meia-IdadeRESUMO
The clinical, haematological, morphological and histological features of a series of 22 patients presenting with splenic lymphoma with circulating villous lymphocytes were assessed and compared with those of patients with other forms of chronic B cell leukaemia in an attempt to differentiate this condition from hairy cell leukaemia, prolymphocytic leukaemia, and chronic lymphocytic leukaemia, with which this condition has many features in common. The disease was twice as common in men than in women, with a mean (SD) age at diagnosis of 72 (9) years, and the most consistent presenting feature was massive enlargement of the spleen, which showed white and red pulp disease with a plasmacytic component. Small monoclonal bands were found in 60% of cases.
Assuntos
Leucemia/diagnóstico , Linfoma/diagnóstico , Neoplasias Esplênicas/diagnóstico , Linfócitos B , Medula Óssea/patologia , Diagnóstico Diferencial , Humanos , Leucemia de Células Pilosas/diagnóstico , Leucemia de Células Pilosas/patologia , Leucemia Linfoide/diagnóstico , Leucemia Linfoide/patologia , Linfoma/patologia , Baço/patologia , Neoplasias Esplênicas/patologiaRESUMO
Splenic lymphoma with villous lymphocytes is a new entity characterised by the presence of atypical lymphocytes in the peripheral blood and bone marrow, and splenic infiltration in the white and red pulp. Cell membrane markers are those of a B mature cell, and no particular chromosomal abnormalities have been associated with this disease. A case of this rare lymphoma occurred in two sisters. Histological examination of splenic tissue was identical in both cases, with the same immunological surface markers, although the clinical and laboratory features were different. Karyotype analysis showed an abnormal pattern in one case; no environmental causative factor could be detected. Familial cases of other lymphoproliferative disorders have been reported, but no consistent common link has been found. It is suggested that further reports of this lymphoma, including cytogenetic and molecular studies, may provide a better understanding of the aetiology.
Assuntos
Linfócitos/patologia , Linfoma/patologia , Neoplasias Esplênicas/patologia , Idoso , Saúde da Família , Feminino , Humanos , Cariotipagem , Linfoma/genética , Neoplasias Esplênicas/genéticaRESUMO
An unusual case of acute myeloid leukemia with a standard t(15;17) is described. While light microscopy morphology was suggestive of acute myeloid leukemia M5a and light microscopy cytochemistry showed 80% of blasts to be strongly positive with Sudan Black B--more consistent with a diagnosis of M4--ultrastructural analysis demonstrated that the predominant cells were promyelocytes with immature primary granules hardly visible with the Romanovsky stains by light microscopy. Because typical cytologic and clinical features of M3 or M3 variant were lacking this atypical case would not have been recognized but for the presence of t(15;17) and the demonstration of promyelocytic features by electron microscopy.
Assuntos
Cromossomos Humanos 13-15 , Cromossomos Humanos 16-18 , Granulócitos/ultraestrutura , Leucemia Mieloide Aguda/genética , Translocação Genética , Bandeamento Cromossômico , Grânulos Citoplasmáticos/ultraestrutura , Humanos , Cariotipagem , Leucemia Mieloide Aguda/patologia , Masculino , Microscopia Eletrônica , Pessoa de Meia-IdadeRESUMO
We carried out cytogenetic analysis in 23 patients with Ph-positive chronic granulocytic leukemia in blast crisis. In all cases the type of blast cell was characterized by cytochemistry, immunologic markers, and ultrastructural studies. Twelve cases were classified as myeloid transformation, six as lymphoid, two as mixed (lymphoid and myeloid), and two were unclassifiable. Duplication of Ph was the most frequent abnormality in the whole series. Trisomy 8, i(17q) and trisomy 19 were seen only in patients with myeloid blast crisis (53%, 30%, and 23%, respectively). Our findings suggest that the nature of additional chromosome abnormalities arising in blasts with features of myeloid differentiation are different from those in blasts showing lymphoid differentiation.
Assuntos
Aberrações Cromossômicas , Leucemia Mieloide Aguda/genética , Leucemia Mieloide/genética , Cromossomo Filadélfia , Adulto , Idoso , Feminino , Humanos , Cariotipagem , Leucemia Mieloide/patologia , Leucemia Mieloide Aguda/patologia , Masculino , Pessoa de Meia-Idade , FenótipoRESUMO
Ceftazidime was used in twenty-eight febrile episodes recorded in twenty-two neutropenic patients (eleven of each sex, aged between 15 and 73 years) who were immunologically compromised (nineteen acute leukaemias and three malignant lymphomas treated with cytostatics). Seven pathogens were isolated in six patients (four klebsiella, one proteus, one Escherichia coli and one enterobacter). The clinical response was cure in ten cases, improvement in six, failure in four and unassessable in eight. In this study ceftazidime was as effective as the combinations of antibiotics generally used in these cases and free of adverse events.
Assuntos
Infecções Bacterianas/tratamento farmacológico , Ceftazidima/uso terapêutico , Tolerância Imunológica , Leucemia/complicações , Linfoma/complicações , Doença Aguda , Adolescente , Adulto , Idoso , Agranulocitose/complicações , Infecções Bacterianas/complicações , Infecções Bacterianas/fisiopatologia , Feminino , Febre/etiologia , Febre de Causa Desconhecida/tratamento farmacológico , Febre de Causa Desconhecida/etiologia , Doença de Hodgkin/complicações , Humanos , Terapia de Imunossupressão , Masculino , Pessoa de Meia-Idade , Neutropenia/complicaçõesRESUMO
We report the results of PBPC collection by large-volume leukaphereses and the hematologic recovery after high-dose chemotherapy supported by autologous PBPC reinfusion in a series of cancer patients treated at the Hematological Intensive Care Unit (UCHI) (Portuguese Institute of Oncology, Lisbon). Large volume leukaphereses were used to increase the efficacy of the PBPC collection. This modification of the standard apheresis technique allowed the harvesting, in only one session, of enough progenitors to proceed to transplantation in nearly 2/3 of patients and without significant toxicity. From December 1993 until September 1997, 95 autologous PBSC transplants were performed at the UCHI; 45% were performed in solid tumor patients and 55% in patients with hematologic malignancies. Hematologic recovery was similar to that published in the literature and related to the number of CD34+ cells infused. Patients supported with bone marrow in addition to PBPC showed delayed hematopoietic recovery, probably because the bone marrow harvest was only performed when an insufficient number of PBPC had been collected (2 x 10(6) CD34+ cells/Kg). The speed of hematological recovery differed per diagnosis, being higher in multiple myeloma and solid tumor patients and lower in Hodgkin's disease patients.
Assuntos
Antineoplásicos/administração & dosagem , Transplante de Células-Tronco Hematopoéticas/métodos , Adolescente , Adulto , Transplante de Medula Óssea , Institutos de Câncer , Terapia Combinada , Feminino , Transplante de Células-Tronco Hematopoéticas/estatística & dados numéricos , Humanos , Unidades de Terapia Intensiva , Masculino , Pessoa de Meia-Idade , Neoplasias/sangue , Neoplasias/terapia , Portugal , Transplante AutólogoRESUMO
We review the rationale for PBPC transplantation and the results reported in the literature. In order to prolong complete remissions and increase cure rates, high-dose chemotherapy is frequently used in the treatment of selected neoplasias. Hematological toxicity can be overcome by the infusion of autologous hemopoietic progenitors. Recently, peripheral blood is being used as the preferred source for hemopoietic progenitors, since it allows faster hematopoietic recoveries when compared to progenitors harvested from bone marrow. An adequate graft is defined by its content in clonogenic progenitors (mainly CFU-GM) and CD34 positive cells; these two parameters need to be accurately determined by specific laboratory methods. PBPC grafts are harvested using cell separators during leukaphereses; to increase efficiency, hemopoietic progenitors are first mobilized into the circulation with growth factors and or chemotherapy. PBSC transplantation may have procedure-associated toxicity related to the mobilization, harvest or reinfusion of the graft.
Assuntos
Antineoplásicos/administração & dosagem , Transplante de Células-Tronco Hematopoéticas , Antineoplásicos/efeitos adversos , Terapia Combinada , Mobilização de Células-Tronco Hematopoéticas , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Humanos , Neoplasias/sangue , Neoplasias/terapia , Transplante AutólogoRESUMO
Several sesquiterpene lactone were synthesized and their inhibitive activities on phospholipase A(2) (PLA(2)) from Bothrops jararacussu venom were evaluated. Compounds Lac01 and Lac02 were efficient against PLA(2) edema-inducing, enzymatic and myotoxic activities and it reduces around 85% of myotoxicity and around 70% of edema-inducing activity. Lac05-Lac08 presented lower efficiency in inhibiting the biological activities studied and reduce the myotoxic and edema-inducing activities around only 15%. The enzymatic activity was significantly reduced. The values of inhibition constants (K(I)) for Lac01 and Lac02 were approximately 740 µM, and for compounds Lac05-Lac08 the inhibition constants were approximately 7.622-9.240 µM. The enzymatic kinetic studies show that the sesquiterpene lactones inhibit PLA(2) in a non-competitive manner. Some aspects of the structure-activity relationships (topologic, molecular and electronic parameters) were obtained using ab initio quantum calculations and analyzed by chemometric methods (HCA and PCA). The quantum chemistry calculations show that compounds with a higher capacity of inhibiting PLA(2) (Lac01-Lac04) present lower values of highest occupied molecular orbital (HOMO) energy and molecular volume (VOL) and bigger values of hydrophobicity (LogP). These results indicate some topologic aspects of the binding site of sesquiterpene lactone derivatives and PLA(2).