RESUMEN
Minimal residual disease (MRD) detection using quantification of clone-specific Ig or TCR rearrangements before and after transplantation in children with high-risk ALL is an important predictor of outcome. The method and guidelines for its interpretation are very precise to avoid both false-negative and -positive results. In a group of 21 patients following transplantation, we observed detectable MRD positivities in Ig/TCR-based real-time quantitative PCR (RQ-PCR) leading to no further progression of the disease (11 of 100 (11%) total samples). We hypothesized that these positivities were mostly the result of nonspecific amplification despite the application of strict internationally agreed-upon measures. We applied two non-self-specific Ig heavy chain assays and received a similar number of positivities (20 and 15%). Nonspecific products amplified in these RQ-PCR systems differed from specific products in length and sequence. Statistical analysis proved that there was an excellent correlation of this phenomenon with B-cell regeneration in BM as measured by flow cytometry and Ig light chain-kappa excision circle quantification. We conclude that although Ig/TCR quantification is a reliable method for post transplant MRD detection, isolated positivities in Ig-based RQ-PCR systems at the time of intense B-cell regeneration must be viewed with caution to avoid the wrong indication of treatment.
Asunto(s)
Linfocitos B/inmunología , Linfoma de Burkitt/cirugía , Trasplante Homólogo/inmunología , Adolescente , Preescolar , ADN de Neoplasias/genética , Reordenamiento Génico , Humanos , Lactante , Transfusión de Linfocitos , Masculino , Neoplasia Residual/diagnóstico , Neoplasia Residual/genética , Análisis de Secuencia por Matrices de Oligonucleótidos , Reacción en Cadena de la Polimerasa , Trasplante de Células MadreRESUMEN
Data on secondary acute lymphoblastic leukaemia (sALL) following ALL treatment are very rare. However, the incidence might be underestimated as sALLs without a significant lineage shift might automatically be diagnosed as relapses. Examination of immunoglobulin and T-cell receptor gene rearrangements brought a new tool that can help in discrimination between relapse and sALL. We focused on the recurrences of childhood ALL to discover the real frequency of the sALL after ALL treatment. We compared clonal markers in matched presentation and recurrence samples of 366 patients treated according to the Berlin-Frankfurt-Munster (BFM)-based protocols. We found two cases of sALL and another three, where the recurrence is suspicious of being sALL rather than relapse. Our proposal for the 'secondary ALL after ALL' diagnostic criteria is as follows: (A) No clonal relationship between diagnosis and recurrence; (B) significant immunophenotypic shift--significant cytogenetic shift--gain/loss of a fusion gene. For the sALL (A) plus at least one (B) criterion should be fulfilled. With these criteria, the estimated frequency of the sALL after ALL is according to our data 0.5-1.5% of ALL recurrences on BFM-based protocols. Finally, we propose a treatment strategy for the patients with secondary disease.
Asunto(s)
Técnicas de Diagnóstico Molecular/métodos , Neoplasias Primarias Secundarias/diagnóstico , Leucemia-Linfoma Linfoblástico de Células Precursoras/diagnóstico , Antineoplásicos/efectos adversos , Preescolar , Diagnóstico Diferencial , Femenino , Reordenamiento Génico de Linfocito T , Genes de Inmunoglobulinas , Humanos , Inmunofenotipificación , Incidencia , Masculino , Neoplasias Primarias Secundarias/inducido químicamente , Leucemia-Linfoma Linfoblástico de Células Precursoras/inducido químicamente , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamiento farmacológico , RecurrenciaRESUMEN
The presence of TEL/AML1 fusion gene in childhood acute lymphoblastic leukaemia (ALL) defines a subgroup of patients with better than average outcome. However, the prognostic significance of this aberration has recently been disputed by the Berlin-Frankfurt-Münster (BFM) study group due to its relatively high incidence found in relapsed patients (19.6% and 21.9%, in two cohorts). In contrast, only four out of 45 (8.9%) unselected relapsed patients (all of whom had been treated according to BFM protocols) in the Czech Republic carry this fusion. From March 1995 to June 1998, 41 out of 190 (21.6%) newly diagnosed children with ALL were TEL/AML1-positive. There is a statistically significant difference between the incidence of TEL/AML1 fusion at diagnosis and at relapse within our group (P = 0.035). Interim analysis of the minimal residual disease (MRD) detection shows heterogeneity within the group of newly diagnosed TEL/AML1-positive leukaemias--10 out of 24 patients tested at the end of induction therapy had detectable levels of MRD. However, only one of these patients reached relapse-predictive level (10(-3)) of MRD. In conclusion, we corroborate low frequency of TEL/AML1 positivity among relapsed patients with ALL among Czech children who are treated by the BFM protocols. Moreover, we demonstrate different patterns of bone marrow clean-up in TEL/AML1-positive patients.
Asunto(s)
Proteínas de Neoplasias/genética , Proteínas de Fusión Oncogénica , Leucemia-Linfoma Linfoblástico de Células Precursoras/diagnóstico , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Adolescente , Médula Ósea/patología , Niño , Subunidad alfa 2 del Factor de Unión al Sitio Principal , República Checa , Exones , Reordenamiento Génico , Humanos , Recuento de Leucocitos , Proteínas de Neoplasias/análisis , Leucemia-Linfoma Linfoblástico de Células Precursoras/sangre , Leucemia-Linfoma Linfoblástico de Células Precursoras/patología , Pronóstico , Recurrencia , Translocación GenéticaRESUMEN
BACKGROUND: The BCR/ABL and MLL/AF4 fusion genes--resulting from t(9;22)(q34;q11) and t(4;11)(q21;q23) translocations, respectively--are considered as a high risk prognostic factors in children with acute lymphoblastic leukaemia (ALL). Their presence in malignant cells indicates patient for the most intensive antileukaemic therapy regardless of the other criteria. In contrast, the most common non-random chromosomal aberration in paediatric ALL--translocation t(12;21)(q12;q22)--is associated with a favourable prognosis. The examination of these rearrangements is important for the stratification of patients to the risk groups and also provides the most sensitive and specific tool for minimal residual disease (MRD) follow-up. METHODS AND RESULTS: This study comprises 241 patients with ALL from Czech and Slovak Republics younger than 18 years at diagnosis. They were examined for presence of m-RNA of fusion genes BCR/ABL, MLL/AF4 and TEL/AML1 by reverse transcriptase-polymerase chain reaction (RT-PCR) method. Seven out of 197 (3.6%) carried MLL/AF4 fusion gene, but among infants it was 56% (5 out of 9). BCR/ABL positivity was found in 2.5% (7 out of 240) and TEL/AML1 in 21.7% (41 out of 189) cases. Event free survival (EFS) curves demonstrate the clinical impact of these hybrid genes on patients' prognosis. Moreover, we present the possibility of the monitoring of MRD levels in follow-up samples of these patients. CONCLUSIONS: All particular rearrangements were found only in a cohort of patients with B-precursor ALL (or hybrid leukaemia), which constitutes 85% of our group. Presence of BCR/ABL or MLL/AF4 fusion gene is associated with poor prognosis and is indispensable condition for correct stratification of patients to the risk groups according to treatment protocols. Hybrid gene TEL/AML1 defines subgroup of children with better prognosis and due to its high frequency provides us with a very useful tool for MRD detection.
Asunto(s)
Biomarcadores de Tumor/análisis , Proteínas de Fusión bcr-abl/genética , Proteínas de Neoplasias/genética , Proteínas de Fusión Oncogénica/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/diagnóstico , Translocación Genética , Niño , Preescolar , Subunidad alfa 2 del Factor de Unión al Sitio Principal , Femenino , Proteínas de Fusión bcr-abl/análisis , Humanos , Hibridación Fluorescente in Situ , Lactante , Masculino , Proteína de la Leucemia Mieloide-Linfoide , Proteínas de Neoplasias/análisis , Proteínas de Fusión Oncogénica/análisis , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
The level of minimal residual disease is an important prognostic factor in childhood acute lymphoblastic leukaemia. The end of induction therapy is the most significant time-point for prediction of treatment outcome. Within a pilot study covered by the Paediatric Haematology Working Group in the Czech Republic 51 childhood patients were analysed at diagnosis of acute lymphoblastic leukaemia and at the end of induction using method based on detection of clonal rearrangements of immuno-receptor genes. The majority of tested patients (32/51, 63%) had a low or non-detectable levels of residual disease, a group of patients with the highest levels and thus the highest risk of relapse included 10% of patients (5/51). Within each of three risk groups one patient has relapsed so far. Therefore, the relapse rate in particular subgroups is 3% (1/32), 7% (1/14) and 20% (1/5) to date, respectively. The results are compared with these published by the BFM group (van Dongen et al., Lancet 1998). The pilot phase of a new BFM treatment protocols includes examination of residual disease for stratification of patients into the different risk groups.
Asunto(s)
Leucemia-Linfoma Linfoblástico de Células Precursoras/diagnóstico , Niño , Regiones Determinantes de Complementariedad , Reordenamiento Génico de la Cadena gamma de los Receptores de Antígenos de los Linfocitos T , Humanos , Cadenas Pesadas de Inmunoglobulina/genética , Neoplasia Residual , Reacción en Cadena de la Polimerasa , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamiento farmacológico , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Pronóstico , RecurrenciaAsunto(s)
Neoplasia Residual/diagnóstico , Proteínas de Fusión Oncogénica/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/diagnóstico , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Niño , Subunidad alfa 2 del Factor de Unión al Sitio Principal , Humanos , Inmunoglobulinas/genética , Estudios Prospectivos , Receptores de Antígenos de Linfocitos T/genéticaAsunto(s)
Linfoma de Burkitt/genética , Proteínas de Neoplasias/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Fusión Artificial Génica , Linfoma de Burkitt/epidemiología , Checoslovaquia/epidemiología , Humanos , Incidencia , Leucemia-Linfoma Linfoblástico de Células Precursoras/epidemiología , Recurrencia , Translocación GenéticaAsunto(s)
Proteínas de Unión al ADN/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Proteínas Proto-Oncogénicas , Proteínas Represoras , Factores de Transcripción/genética , Adolescente , Distribución por Edad , Fusión Artificial Génica , Niño , Preescolar , Subunidad alfa 2 del Factor de Unión al Sitio Principal , Humanos , Lactante , Recién Nacido , Proteínas Proto-Oncogénicas c-ets , Proteína ETS de Variante de Translocación 6Asunto(s)
Reacción en Cadena de la Polimerasa/instrumentación , Reacción en Cadena de la Polimerasa/métodos , Movimientos del Aire , ADN/análisis , Análisis de Falla de Equipo , Control de Calidad , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Temperatura , Proteína p53 Supresora de Tumor/genética , Microglobulina beta-2/genéticaRESUMEN
Minimal residual disease (MRD) monitoring is an essential tool for risk group stratification in current treatment protocols for childhood acute lymphoblastic leukaemia (ALL). Although quantitative detection of clonal immunoglobulin (Ig) and T-cell receptor (TCR) gene rearrangements is currently considered to be the standard method, leukaemia fusion genes provide other possible targets for MRD follow-up, as already demonstrated in TEL/AML1-positive ALLs. We analysed and compared MRD levels quantified by BCR/ABL transcript detection and by the standard Ig/TCR-based method in 218 bone marrow specimens from 17 children with BCR/ABL-positive ALL. We found only a limited overall correlation of MRD levels as assessed by the two methods (correlation coefficient R(2)=0.64). The correlation varied among patients from excellent (R(2)=0.99) to very poor (R(2)=0.17). Despite identical sensitivity of the approaches, 20% of the samples were negative by the Ig/TCR approach whereas positive by the BCR/ABL method. We show that multilineage involvement is at least partly responsible for the discrepancy. Moreover, our data demonstrate that BCR/ABL monitoring enables better and earlier prediction of relapse compared to the standard Ig/TCR methodology. We conclude that BCR/ABL-based MRD monitoring of childhood ALL is a clinically relevant tool and should be performed in parallel with the standard Ig/TCR follow-up.