RESUMO
MOTIVATION: Selecting a small number of signature genes for accurate classification of samples is essential for the development of diagnostic tests. However, many genes are highly correlated in gene expression data, and hence, many possible sets of genes are potential classifiers. Because treatment outcomes are poor in advanced chronic myeloid leukemia (CML), we hypothesized that expression of classifiers of advanced phase CML when detected in early CML [chronic phase (CP) CML], correlates with subsequent poorer therapeutic outcome. RESULTS: We developed a method that integrates gene expression data with expert knowledge and predicted functional relationships using iterative Bayesian model averaging. Applying our integrated method to CML, we identified small sets of signature genes that are highly predictive of disease phases and that are more robust and stable than using expression data alone. The accuracy of our algorithm was evaluated using cross-validation on the gene expression data. We then tested the hypothesis that gene sets associated with advanced phase CML would predict relapse after allogeneic transplantation in 176 independent CP CML cases. Our gene signatures of advanced phase CML are predictive of relapse even after adjustment for known risk factors associated with transplant outcomes.
Assuntos
Algoritmos , Transplante de Células-Tronco Hematopoéticas , Leucemia Mielogênica Crônica BCR-ABL Positiva/diagnóstico , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Teorema de Bayes , Progressão da Doença , Humanos , Leucemia Mielogênica Crônica BCR-ABL Positiva/terapia , Leucemia Mieloide de Fase Crônica/genética , Análise de Sequência com Séries de Oligonucleotídeos , Reação em Cadeia da Polimerase , RecidivaRESUMO
The therapeutic landscape of chronic myeloid leukemia (CML) has profoundly changed over the past 7 years. Most patients with chronic phase (CP) now have a normal life expectancy. Another goal is achieving a stable deep molecular response (DMR) and discontinuing medication for treatment-free remission (TFR). The European LeukemiaNet convened an expert panel to critically evaluate and update the evidence to achieve these goals since its previous recommendations. First-line treatment is a tyrosine kinase inhibitor (TKI; imatinib brand or generic, dasatinib, nilotinib, and bosutinib are available first-line). Generic imatinib is the cost-effective initial treatment in CP. Various contraindications and side-effects of all TKIs should be considered. Patient risk status at diagnosis should be assessed with the new EUTOS long-term survival (ELTS)-score. Monitoring of response should be done by quantitative polymerase chain reaction whenever possible. A change of treatment is recommended when intolerance cannot be ameliorated or when molecular milestones are not reached. Greater than 10% BCR-ABL1 at 3 months indicates treatment failure when confirmed. Allogeneic transplantation continues to be a therapeutic option particularly for advanced phase CML. TKI treatment should be withheld during pregnancy. Treatment discontinuation may be considered in patients with durable DMR with the goal of achieving TFR.
Assuntos
Antineoplásicos/uso terapêutico , Proteínas de Fusão bcr-abl/antagonistas & inibidores , Mesilato de Imatinib/uso terapêutico , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Inibidores de Proteínas Quinases/uso terapêutico , Compostos de Anilina/uso terapêutico , Tomada de Decisão Clínica , Conferências de Consenso como Assunto , Dasatinibe/uso terapêutico , Gerenciamento Clínico , Proteínas de Fusão bcr-abl/genética , Proteínas de Fusão bcr-abl/metabolismo , Expressão Gênica , Humanos , Leucemia Mielogênica Crônica BCR-ABL Positiva/diagnóstico , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/mortalidade , Expectativa de Vida/tendências , Monitorização Fisiológica , Nitrilas/uso terapêutico , Pirimidinas/uso terapêutico , Qualidade de Vida , Quinolinas/uso terapêutico , Análise de SobrevidaRESUMO
Molecular monitoring for patients with chronic myeloid leukaemia (CML) has become an important practice in the era of imatinib therapy. For successful widespread introduction into the mainstream patient monitoring schedule, many procedural aspects of the complex real-time quantitative polymerase chain reaction (RQ-PCR) technique for measuring BCR-ABL transcripts require optimization. Recommendations for harmonizing the differing methodologies have recently been proposed. These recommendations were designed to maximize reliability of analysis for clinical decision making and proposed the adoption of an International Scale of measurement. The purpose of this review is to present the evidence and supporting data for specific recommendations. These recommendations include use of the same source of cells, either blood or marrow, for analysis; for validation of equal PCR amplification efficiencies of cDNA and standards when using a plasmid to construct standard curves and for ensuring ongoing high-level performance by undertaking a quality assurance programme. Clinicians must know the measurement reliability of an RQ-PCR assay to be able to determine the significance of a change in BCR-ABL level. An assay with poor precision limits the clinical usefulness of results. International harmonization should establish RQ-PCR measurement of BCR-ABL as the best method for monitoring treatment response for patients with CML.
Assuntos
Monitoramento de Medicamentos/métodos , Proteínas de Fusão bcr-abl/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/terapia , Reação em Cadeia da Polimerase/métodos , Humanos , Reação em Cadeia da Polimerase/normas , Controle de Qualidade , RNA Mensageiro/análise , Reprodutibilidade dos TestesRESUMO
The single-arm, phase 2 ENESTfreedom trial assessed the potential for treatment-free remission (TFR; i.e., the ability to maintain a molecular response after stopping therapy) following frontline nilotinib treatment. Patients with Philadelphia chromosome-positive chronic myeloid leukemia in chronic phase with MR4.5 (BCR-ABL1⩽0.0032% on the International Scale (BCR-ABL1IS)) and ⩾2 years of frontline nilotinib therapy were enrolled. Patients with sustained deep molecular response during the 1-year nilotinib consolidation phase were eligible to stop treatment and enter the TFR phase. Patients with loss of major molecular response (MMR; BCR-ABL1IS⩽0.1%) during the TFR phase reinitiated nilotinib. In total, 215 patients entered the consolidation phase, of whom 190 entered the TFR phase. The median duration of nilotinib before stopping treatment was 43.5 months. At 48 weeks after stopping nilotinib, 98 patients (51.6%; 95% confidence interval, 44.2-58.9%) remained in MMR or better (primary end point). Of the 86 patients who restarted nilotinib in the treatment reinitiation phase after loss of MMR, 98.8% and 88.4%, respectively, regained MMR and MR4.5 by the data cutoff date. Consistent with prior reports of imatinib-treated patients, musculoskeletal pain-related events were reported in 24.7% of patients in the TFR phase (consolidation phase, 16.3%).
Assuntos
Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Inibidores de Proteínas Quinases/uso terapêutico , Pirimidinas/uso terapêutico , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Leucemia Mielogênica Crônica BCR-ABL Positiva/psicologia , Masculino , Pessoa de Meia-Idade , Pirimidinas/efeitos adversos , Qualidade de VidaRESUMO
A total of 24 patients (median age 58; range, 27-71 years) with chronic myeloid leukemia (CML) in first chronic (CP1) (n=14), second chronic (n=4), or accelerated phase (n=6) who were not candidates for conventional hematopoietic cell transplantation (HCT), received nonmyeloablative HCT from HLA-matched siblings a median of 28.5 (range, 11-271) months after diagnosis. They were conditioned with 2 Gy total body irradiation (TBI) alone (n=8) or combined with fludarabine, 90 mg/m(2) (n=16). Postgrafting immunosuppression included cyclosporine and mycophenolate mofetil. All patients initially engrafted. However, 4 of 8 patients not given fludarabine experienced nonfatal rejection while all others had sustained engraftment. With a median follow-up of 36 (range, 4-49) months, 13 of 24 patients (54%) were alive and in complete remission. There were five (21%) deaths from nonrelapse mortality, one (4%) during the first 100 days after transplant. The proportions of grade II, III, and IV acute GVHD were 38, 4, and 8%, respectively. The 2-year estimate of chronic GVHD was 32%. The 2-year survival estimates for patients in CP1 (n=14) and beyond CP1 (n=10) were 70 and 56%, respectively. This study shows encouraging remission rates for patients with CML not eligible for conventional allografting.
Assuntos
Transplante de Células-Tronco Hematopoéticas , Teste de Histocompatibilidade , Leucemia Mielogênica Crônica BCR-ABL Positiva/terapia , Doadores de Tecidos , Condicionamento Pré-Transplante/métodos , Vidarabina/análogos & derivados , Adulto , Idoso , Antineoplásicos/administração & dosagem , Antineoplásicos/efeitos adversos , Feminino , Doença Enxerto-Hospedeiro/etiologia , Doença Enxerto-Hospedeiro/mortalidade , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Transplante de Células-Tronco Hematopoéticas/mortalidade , Humanos , Leucemia Mielogênica Crônica BCR-ABL Positiva/mortalidade , Masculino , Pessoa de Meia-Idade , Doses de Radiação , Irmãos , Quimeras de Transplante , Condicionamento Pré-Transplante/mortalidade , Transplante Homólogo , Resultado do Tratamento , Vidarabina/administração & dosagem , Vidarabina/efeitos adversos , Irradiação Corporal Total/métodosRESUMO
Measurable ('minimal') residual disease (MRD) before or after hematopoietic cell transplantation (HCT) identifies adults with AML at risk of poor outcomes. Here, we studied whether peri-transplant MRD dynamics can refine risk assessment. We analyzed 279 adults receiving myeloablative allogeneic HCT in first or second remission who survived at least 35 days and underwent 10-color multiparametric flow cytometry (MFC) analyses of marrow aspirates before and 28±7 days after transplantation. MFC-detectable MRD before (n=63) or after (n=16) transplantation identified patients with high relapse risk and poor survival. Forty-nine patients cleared MRD with HCT conditioning, whereas two patients developed new evidence of disease. The 214 MRD(neg)/MRD(neg) patients had excellent outcomes, whereas both MRD(neg)/MRD(pos) patients died within 100 days following transplantation. For patients with pre-HCT MRD, outcomes were poor regardless of post-HCT MRD status, although survival beyond 3 years was only observed among the 58 patients with decreasing but not the seven patients with increasing peri-HCT MRD levels. In multivariable models, pre-HCT but not post-HCT MRD was independently associated with overall survival and risk of relapse. These data indicate that MRD(pos) patients before transplantation have a high relapse risk regardless of whether or not they clear MFC-detectable disease with conditioning and should be considered for pre-emptive therapeutic strategies.
Assuntos
Citometria de Fluxo/métodos , Transplante de Células-Tronco Hematopoéticas/métodos , Leucemia Mieloide Aguda/patologia , Neoplasia Residual/diagnóstico , Adolescente , Adulto , Idoso , Exame de Medula Óssea , Feminino , Transplante de Células-Tronco Hematopoéticas/mortalidade , Humanos , Leucemia Mieloide Aguda/mortalidade , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Neoplasia Residual/mortalidade , Período Pós-Operatório , Período Pré-Operatório , Recidiva , Estudos Retrospectivos , Taxa de Sobrevida , Condicionamento Pré-Transplante , Resultado do Tratamento , Adulto JovemRESUMO
Molecular monitoring of chronic myeloid leukemia patients using robust BCR-ABL1 tests standardized to the International Scale (IS) is key to proper disease management, especially when treatment cessation is considered. Most laboratories currently use a time-consuming sample exchange process with reference laboratories for IS calibration. A World Health Organization (WHO) BCR-ABL1 reference panel was developed (MR(1)-MR(4)), but access to the material is limited. In this study, we describe the development of the first cell-based secondary reference panel that is traceable to and faithfully replicates the WHO panel, with an additional MR(4.5) level. The secondary panel was calibrated to IS using digital PCR with ABL1, BCR and GUSB as reference genes and evaluated by 44 laboratories worldwide. Interestingly, we found that >40% of BCR-ABL1 assays showed signs of inadequate optimization such as poor linearity and suboptimal PCR efficiency. Nonetheless, when optimized sample inputs were used, >60% demonstrated satisfactory IS accuracy, precision and/or MR(4.5) sensitivity, and 58% obtained IS conversion factors from the secondary reference concordant with their current values. Correlation analysis indicated no significant alterations in %BCR-ABL1 results caused by different assay configurations. More assays achieved good precision and/or sensitivity than IS accuracy, indicating the need for better IS calibration mechanisms.
Assuntos
Proteínas de Fusão bcr-abl/análise , Calibragem , Proteínas de Fusão bcr-abl/normas , Genes abl , Humanos , Reação em Cadeia da Polimerase , Proteínas Proto-Oncogênicas c-bcr/genética , Padrões de Referência , Organização Mundial da SaúdeRESUMO
PURPOSE: The impact of a second marrow transplant on long-term disease-free survival (DFS) was evaluated for 77 consecutive patients aged 2 to 51 years who relapsed subsequent to allogeneic marrow transplantation after high-dose chemotherapy and total-body irradiation (TBI). PATIENTS AND METHODS: Patients received a second transplant for recurrent chronic myelogenous leukemia (CML) (n = 28), acute myelogenous leukemia (AML) (n = 32), and acute lymphoblastic leukemia (ALL) (n = 15) or lymphoma (n = 2) that used the same marrow donor as the initial transplant. High-dose chemotherapy of busulfan (BU) and cyclophosphamide (CY), or CY, carmustine (BCNU), and etoposide (VP-16), was used as a preparative regimen for the second transplant. Graft-versus-host disease (GVHD) prophylaxis consisted of the following: no prophylaxis (n = 8), T-cell depletion (n = 36), methotrexate (MTX) only (n = 21), cyclosporine (CSP) only (n = 1), MTX and CSP (n = 9), or anti-thymocyte globulin (ATG) and prednisone (n = 2). RESULTS: Engraftment occurred in the 74 assessable patients. Severe veno-occlusive disease (VOD) was the most frequent cause of grades 3 and 4 regimen-related toxicity (RRT); it occurred in 20 patients. The probability of death before day 100 from nonleukemic causes was 36%. The probability of relapse after second transplant was 70%, and the DFS rate was 14% (median DFS, 36 months; range, 22 to 87). The DFS rates for ALL, AML, and CML were 8%, 10%, and 25%, respectively. Multivariate analysis showed that the risk of relapse was inversely associated with acute GVHD (relative risk [RR] of relapse = 0.2; P = .0009). No other factor was associated with relapse. DFS was associated with the presence of acute GVHD (RR of treatment failure = 0.5; P = .0085), and a reduction of DFS was associated with severe VOD (RR = 10.6; P = .0001) and those patients older than 10 years (RR = 2.5; P = .0337). CONCLUSION: These data show that some patients may benefit from a second marrow transplant for recurrent leukemia after an initial marrow transplant. Younger patients and patients with CML especially should be considered as potential candidates for a second transplant.
Assuntos
Transplante de Medula Óssea , Leucemia/radioterapia , Leucemia/cirurgia , Irradiação Corporal Total , Adolescente , Adulto , Transplante de Medula Óssea/efeitos adversos , Criança , Pré-Escolar , Terapia Combinada , Feminino , Doença Enxerto-Hospedeiro/epidemiologia , Doença Enxerto-Hospedeiro/prevenção & controle , Humanos , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Recidiva , Reoperação , Análise de Sobrevida , Transplante Homólogo , Resultado do TratamentoRESUMO
The p73 gene is a candidate tumor suppressor gene that has significant homology to p53. Thus far, p73 has not been investigated in hematopoietic malignancies. We used single-strand conformation polymorphism analysis to examine 60 de novo acute myelogenous leukemia (AML) patients for p73 mutations in exons 4, 6 and 7, which are homologous to the most frequently mutated exons in p53. Mutations were not found, but we did identify polymorphisms in exons 4 and 7. We also examined p73 RNA expression in 15 AML samples, eight cell lines, and eight normal bone marrows using the reverse transcriptase/polymerase chain reaction assay. All 31 RNA samples had p73 expression. Fourteen RNA samples were informative for allelic expression, being heterozygous for a polymorphism in codon 173 of exon 4. The two normal bone marrows and the K562 cell line had evidence of biallelic expression while six of 10 AML patients and the Kasumi (AML) cell line had monoallelic expression. These data suggest that functional p73 mutations in exons 4, 6 and 7 do not occur in most de novo AML patients. In addition, biallelic expression of p73 occurs in normal bone marrows, some AML samples, and specific cell lines. Lastly, monoallelic p73 expression appears to be common in de novo AML.
Assuntos
Proteínas de Ligação a DNA/genética , Genes Supressores de Tumor , Leucemia Mieloide/genética , Proteínas Nucleares/genética , Doença Aguda , Adolescente , Adulto , Alelos , Humanos , Pessoa de Meia-Idade , Mutação , Polimorfismo Conformacional de Fita Simples , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Homologia de Sequência do Ácido Nucleico , Proteína Tumoral p73 , Proteínas Supressoras de TumorRESUMO
A subset of adult acute lymphoblastic leukemia (ALL) patients have blast cells which co-express myeloid-associated antigens (MY+ ALL). We have analyzed 113 adult ALL cases for expression of MY-associated antigens (MAA). ALL was diagnosed by standard morphology, cytochemistry, and immunophenotype in central review. MY+ ALL was diagnosed when > or = 20% of lymphoblasts co-expressed CD13 and/or CD33. Overall incidence of MY+ was 31/113 (27%). MAA expression was not significantly correlated with WBC, blast count, hemoglobin, or hematocrit. MY+ cases were more likely to express B-associated antigens, especially CALLA, and to be FAB L2, Ph+, or to have the BCR-ABL translocation by PCR, but these differences were not statistically significant. All patients were induced with a L10M regimen, and 67 (59%) achieved CR: 43/66 (65%) of B MY neg; 14/29 (48%) of B MY+; 10/16 (63%) T MY neg; and 0/2 T MY+. In age-adjusted analyses CR rate did not differ significantly between MY+ and MY neg patients or between B- and T-cell patients. Of the 113 patients, 84 have died and the remaining 29 patients have been followed for a median of 49 months. In proportional hazards regression analyses adjusting for age and WBC, heterogeneity of survival among the four groups was statistically significant (p = 0.021), largely due to MY status. The mortality rate was 85% greater for MY+ patients compared to MY neg patients (two-tailed p = 0.013). By contrast, survival did not vary significantly between B- and T-cell patients. The data indicate that MAA expression is useful for predicting overall survival of adult patients with ALL treated in a L10M protocol. As a predictive factor MAA expression is comparable to the WBC and superior to the more standard stratification by B- or T-cell markers for this group of patients.
Assuntos
Antígenos de Diferenciação Mielomonocítica/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras/imunologia , Adolescente , Adulto , Idoso , Antígenos CD/análise , Antígenos de Diferenciação Mielomonocítica/análise , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Antígenos CD13/análise , Feminino , Proteínas de Fusão bcr-abl/genética , Humanos , Masculino , Pessoa de Meia-Idade , Neprilisina/análise , Cromossomo Filadélfia , Reação em Cadeia da Polimerase , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/mortalidade , Prognóstico , Modelos de Riscos Proporcionais , Estudos Prospectivos , Indução de Remissão , Lectina 3 Semelhante a Ig de Ligação ao Ácido Siálico , Taxa de SobrevidaRESUMO
In a 5-year survey of nonpromyelocytic/nonmonocytic acute myeloid leukemias (AMLs) diagnosed in the University of Washington Hematopathology Laboratory, we identified 19 cases containing distinctive, cup-like nuclear indentation in 10% or more of the blasts ('AML-cuplike'). Fourteen of these cases (74%) demonstrated near-complete loss of HLA-DR expression, while the other five cases showed partial loss of HLA-DR. A total of 16 of the cases (84%) demonstrated internal tandem duplication (ITD) of the Flt3 gene. When compared to a selected set of AMLs lacking this nuclear morphology, AML-cuplike was significantly more likely to lack HLA-DR and CD34 expression, to express CD123 without CD133, to have a normal karyotype, and to harbor the Flt3 ITD. To characterize AML-cuplike in an unselected series of AMLs, we analyzed 42 consecutive nonpromyelocytic/nonmonocytic AMLs diagnosed in our laboratory during a 6-month period in 2002. Strikingly, in this unselected series, there was a statistically significant coincidence of invaginated nuclear morphology, loss of HLA-DR, and presence of the Flt3 ITD beyond that expected if these three features were unrelated, suggesting that AMLs with these three features may represent a distinct AML subset.
Assuntos
Núcleo Celular/patologia , Antígenos HLA-DR/metabolismo , Leucemia Mieloide/metabolismo , Leucemia Mieloide/patologia , Proteínas Proto-Oncogênicas/metabolismo , Receptores Proteína Tirosina Quinases/metabolismo , Doença Aguda , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Duplicação Gênica , Humanos , Cariotipagem , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Fator de Células-Tronco , Sequências de Repetição em Tandem , Tirosina Quinase 3 Semelhante a fmsRESUMO
The sensitivity and clinical utility of the polymerase chain reaction (PCR) assay for the detection of BCR-ABL gene rearrangement was compared to conventional cytogenetics for the Philadelphia chromosome (Ph1) in adult acute lymphoblastic leukemia (ALL) patients entered onto a single clinical trial. Ninety-three patients had evaluable PCR assays for both the p190bcr-abl and p210bcr-abl type of BCR-ABL gene rearrangements. Twenty-one of 93 patients (23%) were positive for the BCR-ABL rearrangement by the PCR assay. Fourteen of these patients had the p210brc-abl BCR-ABL rearrangement characteristically seen in CML patients, while seven had the p190bcr-abl rearrangement seen in ALL alone. Of 61 patients analyzed, both with conventional cytogenetics and PCR, eight (13%) were positive for the Ph1, while 14 (23%) were positive for the BCR-ABL rearrangement by the PCR assay. Discordance between the PCR assay and cytogenetics occurred in eight cases where the PCR assay was positive and the cytogenetics negative, and two cases where the PCR assay was negative and cytogenetics positive. PCR positivity did not correlate with treatment response, survival, or relapse-free survival, but there was a higher percentage of L2 FAB morphology in the PCR+ cases compared to the PCR-cases (67 vs. 28%, p = 0.003). In addition, the data suggested that patients with a p190bcr-abl rearrangement have a better response to induction therapy, but a worse relapse-free survival compared to patients with a p210bcr-abl breakpoint, but these differences were not statistically significant. These data suggest that PCR and conventional cytogenetics may provide complementary information, since there appear to be a subset of patients who are Ph1-negative yet BCR-ABL positive by PCR. Further studies will be required to determine the prognostic significance of the detailed information about BCR-ABL breakpoints that is available from the PCR assay.
Assuntos
Proteínas de Fusão bcr-abl/genética , Genes abl , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Adolescente , Adulto , Idoso , Fragilidade Cromossômica , Intervalo Livre de Doença , Feminino , Rearranjo Gênico , Humanos , Cariotipagem , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Cromossomo Filadélfia , Reação em Cadeia da Polimerase , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Prognóstico , Modelos de Riscos Proporcionais , Indução de Remissão , Sensibilidade e EspecificidadeRESUMO
To examine the impact of inactivation of tumor suppressor genes on outcome in adult ALL, we compared two groups of patients registered to SWOG treatment protocols for loss of the Rb gene product and p53 overexpression: (1) 89 patients with de novo ALL, and (2) 26 patients with relapsed/refractory ALL. The groups were comparable with respect to age, sex, and race. Cell lysates (> or = 80% blasts) were analyzed by immunoblotting which enabled detection of Rb or p53 proteins in as little as 1 microg of lysate. Loss of Rb expression (pRbneg) was found in 54/85 (64%) de novo and 11/19 (58%) relapsed patients (P = 0.79). Overexpression of p53 (p53abn), indicative of p53 point mutations, was found in 16/75 (21%) de novo and 8/19 (42%) relapsed patients (P = 0.08). Using a nonisotopic RNase cleavage assay, p53 point mutations in exons 5-9 were confirmed in 14/23 (61%) p53abn specimens. For the de novo ALL group, patients with normal Rb protein had higher WBC and higher peripheral blast and lymphocyte counts. Otherwise neither abnormal Rb or p53 expression correlated with any of a large panel of clinical and laboratory variables including FAB class, blast lineage, expression of myeloid antigens or CD34, and presence of the Ph1 chromosome or BCR-ABL. Analyses of treatment outcomes demonstrated no significant impact of Rb or p53 status alone on CR rates, relapse-free or overall survival. An identical percentage (11%) of both de novo and relapsed/refractory patients had concurrent abnormalities of both Rb and p53 expression (pRbneg/p53abn). The survival curve of these patients suggests an increased rate of early death, but the number of patients in this group was small. Summarizing, (1) loss of Rb expression is common in adult ALL; (2) overexpression of p53 may be more frequent in relapsed/refractory than de novo adult ALL; and (3) although Rb or p53 alterations alone are not strong independent predictors of outcome, their concurrent expression may predict a poor response to therapy.
Assuntos
Genes do Retinoblastoma , Genes p53 , Linfócitos/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/metabolismo , Adolescente , Adulto , Idoso , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Intervalo Livre de Doença , Feminino , Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Mutação Puntual , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Prognóstico , Proteína do Retinoblastoma/biossíntese , Resultado do Tratamento , Proteína Supressora de Tumor p53/biossínteseRESUMO
Chromosomal translocations are commonly found in de novo acute myeloid leukemia (AML) cells, and the fusion proteins produced from these genetic abnormalities are assumed to contribute directly to leukemogenesis and/or progression. The AML1/ETO fusion protein, created by translocations between chromosomes 8 and 21 [t(8;21); G. Nucifora and J. D. Rowley, Leuk. Lymphoma, 14: 353-362, 1994; K. L. Rhoades et al., Proc. Natl. Acad. Sci. USA, 93: 11895-11900, 1996] can induce anti-apoptotic Bcl-2 expression in vitro and was proposed to thereby promote the survival of t(8;21)-bearing AML cells (L. Klampfer et al., Proc. Natl. Acad. Sci. USA, 93: 14059-14064, 1996). We confirm that cells of the t(8;21)-bearing Kasumi cell line do express high levels of Bcl-2 protein, as reported previously. However, we show that primary AML cells with (8;21) chromosomal translocations generally express low levels of Bcl-2 protein relative to normal bone marrow-derived myeloid cells and to AML samples with other simple karyotypic abnormalities. We note that p53 mutations are present in the myeloid cell lines expressing AML-ETO protein from chromosomal translocations (Kasumi and SKNO) or from transfected fusion genes (U937) but were undetected in our analyses of 28 primary t(8;21)-bearing AML cell samples from de novo AMLs. Because wild-type p53 can transcriptionally down-regulate bcl-2, we speculate that p53 mutations may contribute to the association of t(8;21) chromosomal abnormalities with higher Bcl-2 expression levels in leukemia cell lines. We also note that some t(8;21)-bearing samples from pediatric and older adult patients do express somewhat higher levels of Bcl-2 than t(8;21)-bearing samples from young adult patients. This suggests that Bcl-2 overexpression could occur in these AML cells by an as yet undefined, p53-independent mechanism and could contribute to the reported association of t(8;21) karyotypes with poor clinical outcomes in childhood AML patients and/or to typically poor clinical outcomes in elderly AML patients.
Assuntos
Cromossomos Humanos Par 21 , Cromossomos Humanos Par 8 , Leucemia Mieloide/genética , Proteínas Proto-Oncogênicas c-bcl-2/biossíntese , Translocação Genética , Doença Aguda , Adolescente , Adulto , Idoso , Western Blotting , Medula Óssea/metabolismo , Criança , Pré-Escolar , Citometria de Fluxo , Expressão Gênica , Humanos , Imunoensaio , Leucócitos Mononucleares/metabolismo , Pessoa de Meia-Idade , Mutação , Antígeno Nuclear de Célula em Proliferação/análise , Proteínas Proto-Oncogênicas c-bcl-2/genética , Reprodutibilidade dos Testes , Células Tumorais Cultivadas , Proteína Supressora de Tumor p53/genéticaRESUMO
Based on our understanding of CML, we recommend that patients age 65 or less with newly diagnosed CML should be evaluated to determine the stage of their disease and should be human lymphocyte antigen-typed along with their siblings. Patients age 55 years and younger who have matched or single antigen mismatched siblings should be advised to undergo BMT as soon as possible and patients up to age 65 should be considered for BMT early in their disease course if in good health. Patients age 55 years or younger without appropriately matched family member donors should have a search for an unrelated donor initiated and should be offered BMT if an appropriate donor is found. Patients for whom no donor is found and older patients without family matches can be considered for entry onto clinical trials evaluating autologous BMT or other experimental approaches.
Assuntos
Transplante de Medula Óssea , Leucemia Mielogênica Crônica BCR-ABL Positiva/terapia , Idoso , Transplante de Medula Óssea/imunologia , Antígenos HLA/análise , Teste de Histocompatibilidade , Humanos , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Núcleo Familiar , Doadores de Tecidos , Transplante HomólogoRESUMO
Internal tandem duplications (ITDs) of the FLT3 gene occur in approximately 20-30% of acute myeloid leukemia (AML) patients. We investigated if FLT3 ITDs could be used as minimal residual disease (MRD) markers for AML patients. Patient-specific polymerase chain reaction (PCR) assays for FLT3 ITDs were developed for four AML samples that contained FLT3 ITDs of varying size and location. The real-time, quantitative PCR assays for FLT3 ITDs were highly sensitive and specific, detecting between 0.01 and 0.001% of FLT3 ITD positive DNA in a background of 1 microg of normal bone marrow DNA. Our findings suggest that FLT3 ITDs can be used as molecular markers for MRD in patients with AML.
Assuntos
Leucemia Mieloide Aguda/diagnóstico , Reação em Cadeia da Polimerase , Proteínas Proto-Oncogênicas/genética , Receptores Proteína Tirosina Quinases/genética , Sequências de Repetição em Tandem , Humanos , Neoplasia Residual , Sensibilidade e Especificidade , Tirosina Quinase 3 Semelhante a fmsRESUMO
Peripheral blood stem cell support allows dose intensification of multiple cycle chemotherapy for metastatic tumors, including pediatric sarcomas. The VACIME protocol (vincristine, adriamycin, cyclophosphamide, ifosfamide, mesna and etoposide) utilizes peripheral blood stem cells (PBSC) collected following the treatment cycle as support for subsequent dose- and time-intensive chemotherapy. A critical assumption is that PBSC collected in this manner will be purged of residual tumor cells in vivo. We tested this assumption using sensitive reverse-transcriptase polymerase chain reaction (RT-PCR) to assess the presence of the characteristic translocations of the Ewing's sarcoma family of tumors (ESFT) and alveolar rhabdomyosarcoma (ARMS), t(11;22), and t(2;13), respectively. We used RT-PCR to evaluate 122 samples of peripheral blood (PB), bone marrow (BM) and PBSC collected from 12 pediatric patients with metastatic ESFT and ARMS. The samples included pre-therapy BM and PB, as well as BM, PB, and PBSC collections at various times in the VACIME treatment course. Molecular evidence of tumor contamination was detected in 1/40 PBSC collections from 12 patients. In all patients, we documented clearance of disease by RT-PCR in peripheral blood and bone marrow by week 9 of the VACIME protocol. In vivo purging in combination with the intensive VACIME regime appears to be effective in removing tumor cells from PBSC, bone marrow, and peripheral blood as detected by RT-PCR.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Medula Óssea/patologia , Neoplasias Ósseas/tratamento farmacológico , Cromossomos Humanos Par 11/ultraestrutura , Cromossomos Humanos Par 13/ultraestrutura , Cromossomos Humanos Par 22/ultraestrutura , Cromossomos Humanos Par 2/ultraestrutura , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/efeitos dos fármacos , Células Neoplásicas Circulantes , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Rabdomiossarcoma Alveolar/tratamento farmacológico , Sarcoma de Ewing/tratamento farmacológico , Neoplasias de Tecidos Moles/tratamento farmacológico , Translocação Genética , Adolescente , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Purging da Medula Óssea , Neoplasias Ósseas/sangue , Neoplasias Ósseas/genética , Criança , Cromossomos Humanos Par 11/genética , Cromossomos Humanos Par 13/genética , Cromossomos Humanos Par 2/genética , Cromossomos Humanos Par 22/genética , Terapia Combinada , Ciclofosfamida/administração & dosagem , Ciclofosfamida/efeitos adversos , Proteínas de Ligação a DNA/genética , Doxorrubicina/administração & dosagem , Doxorrubicina/efeitos adversos , Etoposídeo/administração & dosagem , Etoposídeo/efeitos adversos , Feminino , Proteína Forkhead Box O1 , Fatores de Transcrição Forkhead , Humanos , Ifosfamida/administração & dosagem , Ifosfamida/efeitos adversos , Masculino , Mesna/administração & dosagem , Mesna/efeitos adversos , Proteínas de Neoplasias/genética , Proteínas de Fusão Oncogênica/genética , Fator de Transcrição PAX3 , Fatores de Transcrição Box Pareados , Proteína Proto-Oncogênica c-fli-1 , Proteína EWS de Ligação a RNA , Rabdomiossarcoma Alveolar/sangue , Rabdomiossarcoma Alveolar/genética , Sarcoma de Ewing/sangue , Sarcoma de Ewing/genética , Sensibilidade e Especificidade , Neoplasias de Tecidos Moles/sangue , Neoplasias de Tecidos Moles/genética , Fatores de Transcrição/genética , Vincristina/administração & dosagem , Vincristina/efeitos adversosRESUMO
The CD34 antigen is expressed by human hematopoietic progenitor and stem cells. These cells are capable of reconstituting marrow function after marrow-ablative chemo-radiotherapy. Several different technologies have been developed for the separation of CD34+ cells from bone marrow or peripheral blood stem cell (PBSC) components. We used an immunomagnetic separation technique to enrich CD34+ cells from PBSC components in anticipation of autologous transplantation for patients with B lymphoid malignancies. Twenty-nine patients enrolled on this study and received mobilization chemotherapy followed by G-CSF. Of these, 21 achieved a peripheral blood CD34+ cell level of at least 2.0 x 10(4)/l required by protocol for separation of the stem cell components. A median of three components per patient was collected for processing. The average CD34+ cell concentration in the components after apheresis was 1.0 +/- 1.2%. After the CD34+ cell selection, the enriched components contained 0.6 +/- 0.6% of the starting nucleated cells. The recovery of CD34+ cells, however, averaged 58.4 +/- 19.2% of the starting cell number, with a purity of 90.8 +/- 6.5%. Overall depletion of CD34- cells was 99.96 +/- 0.06%. Nineteen patients were treated with marrow-ablative conditioning regimens and received an average of 6.2 +/- 2.0 x 10(6) CD34+ cells/kg body weight. These patients recovered to an ANC >0.5 x 10(9)/l at a median of 11 days (range 8-14), and platelet transfusion independence at a median of 9 days (range 5-13). Four patients died of transplant-related complications or relapse before 100 days after transplantation. No patient required infusion of unseparated cells because of failure of sustained bone marrow function. These data demonstrate that peripheral blood-derived CD34+ cells enriched by use of an immunomagnetic separation technique are capable of rapid engraftment after autologous transplantation.
Assuntos
Antígenos CD34/análise , Transplante de Células-Tronco Hematopoéticas , Separação Imunomagnética/métodos , Adulto , Antígenos CD19/análise , Feminino , Humanos , Linfoma não Hodgkin/terapia , Masculino , Pessoa de Meia-Idade , Transplante AutólogoRESUMO
Conventional allografting produces considerable regimen-related toxicities that generally limit this treatment to patients younger than 55 years and in otherwise good medical condition. T cell-mediated graft-versus-tumor (GVT) effects are known to play an important role in the elimination of malignant disease after allotransplants. A minimally myelosuppressive regimen that relies on immunosuppression for allogeneic engraftment was developed to reduce toxicities while optimizing GVT effects. Pre-transplant total-body irradiation (200 cGy) followed by post-transplant immunosuppression with cyclosporine (CSP) and mycophenolate mofetil (MMF) permitted human leukocyte antigen (HLA)-matched sibling donor hematopoietic cell engraftment in 82% of patients (n = 55) without prior high-dose therapy. The addition of fludarabine (90 mg/m2) facilitated engraftment in all 28 subsequent patients. Overall, fatal progression of underlying disease occurred in 20% of patients after transplant. Non-relapse mortality occurred in 11% of patients. Toxicities were low. Grade 2-4 acute graft-versus-host disease (GVHD) associated with primary engraftment developed in 47% of patients, and was readily controlled in all but two patients. Donor lymphocyte infusions (DLI) were not very effective at converting a low degree of mixed donor/host chimerism to full donor chimerism; however, the addition of fludarabine reduced the need for DLI. With a median follow-up of 244 days, 68% of patients were alive, with 42% of patients in complete remission, including molecular remissions. Remissions occurred gradually over periods of weeks to a year. If long-term efficacy is demonstrated, such a strategy would expand treatment options for patients who would otherwise be excluded from conventional allografting.
Assuntos
Efeito Enxerto vs Tumor/imunologia , Transplante de Células-Tronco Hematopoéticas/métodos , Condicionamento Pré-Transplante/métodos , Adulto , Animais , Ensaios Clínicos Fase I como Assunto , Ensaios Clínicos Fase II como Assunto , Ciclosporina/uso terapêutico , Progressão da Doença , Cães , Previsões , Facilitação Imunológica de Enxerto/efeitos adversos , Facilitação Imunológica de Enxerto/métodos , Sobrevivência de Enxerto , Doença Enxerto-Hospedeiro/tratamento farmacológico , Doença Enxerto-Hospedeiro/etiologia , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Histocompatibilidade , Humanos , Imunossupressores/uso terapêutico , Transfusão de Linfócitos , Metotrexato/uso terapêutico , Pessoa de Meia-Idade , Ácido Micofenólico/análogos & derivados , Ácido Micofenólico/uso terapêutico , Neoplasias/terapia , Prednisona/uso terapêutico , Quimera por Radiação , Linfócitos T Citotóxicos/imunologia , Doadores de Tecidos , Condicionamento Pré-Transplante/efeitos adversos , Transplante Homólogo , Resultado do Tratamento , Vidarabina/análogos & derivados , Vidarabina/uso terapêutico , Irradiação Corporal TotalRESUMO
On a molecular and cellular level, Ph+ ALL seems to be a heterogeneous disease. Unfortunately, the unifying theme of Ph positivity is the poor outcome associated with its presence. Further characterization of molecular subtypes of Ph+ ALL may in the future distinguish those few patients with a potentially good outcome from the majority who face inevitable relapse. Also, novel targeted biologic therapy especially in combination with aggressive, early chemotherapy, may soon be able to temper the disease. Most patients who obtain a remission would be best served by transplantation during remission. For those without a donor, following the disease by PCR-based techniques may detect early relapse. For relapsed patients without the option of transplantation, investigative studies are appropriate.