RESUMEN
BACKGROUND: The study of lineage markers by real-time quantitative polymerase chain reaction (RT-qPCR) at diagnosis enables differentiation between acute myeloblastic leukemia, B- or T-lineage acute lymphoblastic leukemia, without cell sorting. Our objective was to assess the relationship between protein expression and the amount of lineage marker mRNA in acute leukemia samples and to determine whether four lineage markers could be used to differentiate between normal and acute leukemia bone marrow (BM) without cell sorting. METHODS: Quantification of the mRNA of CD19, CD79a, CD3e, and myeloperoxidase was performed by RT-qPCR on 130 acute leukemia BM samples at diagnosis and on 20 BM samples from healthy donors, without cell sorting. Immunophenotyping of leukemia samples was performed after manual gating around the blastic population. RESULTS: Reference values for the four lineage markers were established by RT-qPCR for normal BM. The mRNA expression levels of these four lineage markers allowed the distinction between normal samples and 100% of acute leukemia samples. CONCLUSIONS: With 92% congruence for protein expression and amount of mRNA in acute leukemias, these four lineage markers, essential for diagnosis and subclassification of acute leukemias by flow cytometry, also represent excellent candidate genes when using RT-qPCR technology as a diagnostic tool for molecular cancer class prediction.
Asunto(s)
Biomarcadores de Tumor/genética , Células de la Médula Ósea/citología , Células de la Médula Ósea/metabolismo , Linaje de la Célula/genética , Regulación Neoplásica de la Expresión Génica/genética , Leucemia/genética , Leucemia/patología , Adolescente , Adulto , Niño , Preescolar , Diagnóstico Diferencial , Salud , Humanos , Leucemia/metabolismo , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
BACKGROUND: Chronic myelogenous leukemia (CML) is a rare disease in children and only few data are available concerning the results of interferon based therapy in this age group. Before the imatinib mesylate era, a prospective phase II trial was conducted to assess the efficacy and tolerance of a combination of interferon-alpha 2b (IFN) and cytarabine in children with CML in first chronic phase without a suitable HLA-identical donor. PROCEDURE: Fourteen consecutive children were recruited from 12 pediatric centers. Children received daily IFN (5 million U/m2) and subcutaneous cytarabine (20 mg/m2) for 10 days every month. RESULTS: The median duration of follow-up is 13 months (range 2-32 months). Seven children achieved a complete hematologic response after a median time of treatment of 3 months (range 1 week-4 months). Three children were not evaluable for the cytogenetic response. A major cytogenetic response was achieved in seven patients (including complete cytogenetic response in two patient) within 12 months. The median time to major cytogenetic response was 7 months (range 3-12 months). Thirteen patients discontinued the treatment protocol after a median time of 11 months. Probability of progression free survival at 11 months was 83% (95% CI, 61%-100%). Grade 3 and 4 toxicity was observed in eight patients. The most frequently reported drug-related events were fever, mucositis, neutropenia, and thrombocytopenia. CONCLUSIONS: The combination of IFN and cytarabine provides hematologic and cytogenetic responses in children and adolescents with CML. In the imatinib mesylate era, the role of this combination as second line therapy in children with CML remains to be determined.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Citarabina/administración & dosificación , Interferón-alfa/administración & dosificación , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Adolescente , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Niño , Preescolar , Citarabina/efectos adversos , Análisis Citogenético , Relación Dosis-Respuesta a Droga , Femenino , Estudios de Seguimiento , Humanos , Interferón alfa-2 , Interferón-alfa/efectos adversos , Masculino , Estudios Prospectivos , Proteínas Recombinantes , Tasa de Supervivencia , Resultado del TratamientoRESUMEN
The molecular basis of susceptibility to childhood malignant hemopathy remains largely unknown. An excess of skeletal congenital anomalies has been reported among children with hematological malignancy and points towards involvement of developmental genes, like those belonging to the HOX gene family. In addition to their role in embryogenesis, HOX transcription factors are known to be regulators of proliferation and differentiation of hematopoietic cells. We aimed to explore the possibility that germline alterations of HOX genes might be involved in childhood acute lymphoid malignancies. A cohort of 86 children diagnosed with acute lymphoid malignancy was studied, 20 of them concurrently presenting a congenital anomaly of the skeleton. First, we screened for nucleotide changes throughout the HOX genes of paralogous groups 4 to 13 in the 20 patients with skeletal defects, following a skeletal phenotype-based strategy. Subsequently, we extended the HOX mutation screening to the other 66 children having a malignant lymphoproliferative disorder, but without skeletal defects. In total, 16 germline mutations were identified. While 13 changes were also observed in healthy controls, three variants were exclusively found in acute lymphoid malignancy cases. These comprised the germline c.242A>T (p.Glu81Val) missense mutation of HOXD4, detected in two children diagnosed with acute lymphoblastic leukemia (ALL). Furthermore, this mutation was found in association with other specific HOX variants of cluster D (2q31-q37), defining a unique haplotype. Functional analysis of the murine Hoxd4 homolog revealed that mutant Hoxd4 protein had lower transcriptional activity than wild-type protein in vitro. The p.Glu81Val mutation of HOXD4 thus results in a partial loss-of-function, which might be involved in childhood ALL.
Asunto(s)
Mutación de Línea Germinal , Proteínas de Homeodominio/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Factores de Transcripción/genética , Adolescente , Animales , Niño , Preescolar , Estudios de Cohortes , Análisis Mutacional de ADN , Femenino , Haplotipos , Humanos , Lactante , Masculino , Ratones , Datos de Secuencia MolecularRESUMEN
BACKGROUND: Flow cytometry of lineage markers is useful in the classification of leukemias. Our aim was to assess whether the study of lineage genes at the RNA level would enable differentiation of acute myeloid leukemias (AMLs) from B-and T-lineage acute lymphoid leukemias (ALLs). METHODS: We measured mRNA of four lineage markers [CD19, CD79a, CD3e, and myeloperoxidase (MPO)] by reverse transcription followed by real-time quantitative (RTQ)-PCR. We investigated 72 acute leukemias (40 AMLs with 23-93% blast cells plus 27 B-lineage ALLs and 5 T-lineage ALLs) defined by morphologic criteria at diagnosis. RTQ-PCR analysis was performed on bone marrow without cell sorting. The expression of each gene was calculated as the difference in the threshold cycle [DeltaCT; CT value of target gene minus CT value of housekeeping gene (Abelson)]. RESULTS: Three patterns of expression were detected. In the first, CD19, CD79a, and MPO mRNAs were less abundant than CD3e. In the second pattern, MPO mRNA was more abundant than the other three mRNAs. In the third, CD19 or CD79a was more highly expressed than CD3e and MPO. The three patterns corresponded to T-ALL, AML, and B-ALL, respectively. The use of cutoffs to establish qualitatively the pattern of coexpression of the four lineage markers provided the same information as the comparison among the four DeltaCT values. Prospective use of the scoring system correctly classified each of 13 additional cases (8 AML, 4 B-lineage ALL, and 1 T-lineage ALL). CONCLUSION: Study of lineage markers at diagnosis by RTQ-PCR allows differentiation of AML from B-ALL or T-ALL without cell sorting, even when the bone marrow contains few blast cells.
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Linfoma de Burkitt/diagnóstico , Leucemia Mieloide Aguda/diagnóstico , Leucemia-Linfoma de Células T del Adulto/diagnóstico , ARN Mensajero/análisis , Adulto , Antígenos CD/análisis , Antígenos CD/genética , Antígenos CD19/análisis , Antígenos CD19/genética , Biomarcadores/análisis , Médula Ósea/química , Linfoma de Burkitt/clasificación , Complejo CD3/análisis , Complejo CD3/genética , Antígenos CD79 , Linaje de la Célula , Niño , Diagnóstico Diferencial , Humanos , Inmunofenotipificación , Leucemia Mieloide Aguda/clasificación , Leucemia-Linfoma de Células T del Adulto/clasificación , Peroxidasa/análisis , Peroxidasa/genética , Receptores de Antígenos de Linfocitos B/análisis , Receptores de Antígenos de Linfocitos B/genética , Reproducibilidad de los Resultados , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
A yellow-pigmented coryneform rod was isolated from the blood of a child with acute lymphoblastic leukemia who was perfused with a central venous catheter. The culture bottles were positive twice, at a 2-month interval. The isolate was identified as a Microbacterium sp. and studied along with five other similar strains. Phenotypic, chemotaxonomic, and genetic characteristics indicated that they are closely related to Microbacterium oxydans but that they belong to a distinct species, for which the name Microbacterium paraoxydans sp. nov. is proposed. The type strain of M. paraoxydans is CF36(T) = DSM 15019(T). The G+C content of its DNA is 69.9 mol%.
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Infecciones por Actinomycetales/complicaciones , Infecciones por Actinomycetales/microbiología , Actinomycetales/patogenicidad , Bacteriemia/complicaciones , Bacteriemia/microbiología , Leucemia-Linfoma Linfoblástico de Células Precursoras/complicaciones , Actinomycetales/clasificación , Actinomycetales/genética , Actinomycetales/aislamiento & purificación , Composición de Base , Preescolar , ADN Bacteriano/química , ADN Bacteriano/genética , ADN Ribosómico/genética , Humanos , Masculino , Datos de Secuencia Molecular , Fenotipo , Filogenia , ARN Bacteriano/genética , ARN Ribosómico 16S/genética , Especificidad de la EspecieRESUMEN
BACKGROUND: To assess the efficacy of chemotherapy (chemoreduction) plus local treatments as an alternative to external beam and enucleation for intraocular retinoblastoma. MATERIALS AND METHODS: A prospective study was performed on 21 patients with retinoblastoma treated in our institution from September 1997 to December 2000 to study the ocular outcome of those 33 eyes. RESULTS: There were 9 unilateral and 12 bilateral retinoblastoma cases. There were 12 eyes with Reese-Ellsworth group I-IV and 21 eyes with group V. Among 33 eyes, nine eyes (27%) were initially managed by enucleation. The remaining 24 eyes (73%) were initially treated with chemoreduction (maximum of six cycles of carboplatin, vincristine, etoposide) or chemothermotherapy. Among those 24 eyes, 20 were successfully treated with local treatments (thermotherapy plus cryotherapy in 16 eyes and thermotherapy plus cryotherapy plus (125)I plaque radiotherapy in 4 eyes), enucleation eventually underwent in two eyes and was proposed but refused in one child with bilateral group V retinoblastoma. With a median follow-up of 21 months, conservative management without external beam radiation was successful in all 12 eyes with group I-IV and in a total of 20/33 eyes (60%). Among the nine cases of unilateral retinoblastoma, eight were enucleated but among the 24 eyes with bilateral retinoblastoma, 19 (79%) were successfully treated with conservative therapy. CONCLUSIONS: It may be possible to eradicate viable tumor in all eyes with Reese-Ellsworth group I-IV retinoblastoma by chemoreduction followed by local treatments. Although 8 out of 21 eyes (38%) with group V retinoblastoma may be salvaged after chemoreduction and local therapies, enucleation remained the treatment of choice in those eyes with total retinal detachment and diffuse vitreous seeding.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Neoplasias de la Retina/terapia , Retinoblastoma/terapia , Carboplatino/administración & dosificación , Preescolar , Terapia Combinada , Crioterapia , Etopósido/administración & dosificación , Enucleación del Ojo , Femenino , Humanos , Hipertermia Inducida , Lactante , Masculino , Estudios Prospectivos , Neoplasias de la Retina/tratamiento farmacológico , Retinoblastoma/tratamiento farmacológico , Vincristina/administración & dosificaciónRESUMEN
A 10-month-old girl with a history of recurrent candidiasis, developmental delay, and a fulminant varicella infection is described. The diagnosis of purine nucleoside phosphorylase (PNP) deficiency was suggested by a reduced level of serum uric acid and confirmed by measurement of PNP activity. A human leukocyte antigen-matched bone marrow transplantation resulted in immune reconstitution, but poor neurodevelopmental progression.
Asunto(s)
Trasplante de Médula Ósea , Discapacidades del Desarrollo/fisiopatología , Purina-Nucleósido Fosforilasa/deficiencia , Errores Innatos del Metabolismo de la Purina-Pirimidina/terapia , Empalme Alternativo , Sustitución de Aminoácidos , Aberraciones Cromosómicas , Discapacidades del Desarrollo/genética , Discapacidades del Desarrollo/inmunología , Progresión de la Enfermedad , Exones , Femenino , Humanos , Lactante , Activación de Linfocitos , Recuento de Linfocitos , Mutación Missense , Errores Innatos del Metabolismo de la Purina-Pirimidina/genética , Errores Innatos del Metabolismo de la Purina-Pirimidina/inmunología , Eliminación de Secuencia , Factores de Tiempo , Resultado del Tratamiento , Ácido Úrico/sangreRESUMEN
OBJECTIVE: To evaluate the incidence of hemolytic uremic syndrome (HUS) in Belgium and to determine the role of verocytotoxin-producing Escherichia coli O157:H7 and other serotypes (non-O157 VTEC). METHODS: Twenty-two centers, including the seven university hospitals, registered prospectively all cases of HUS; they collected clinical samples for isolation of VTEC strains and serum for detection of specific O-lipopolysaccharide antibodies. RESULTS: Forty-seven cases of HUS (including five incomplete cases) were recorded. Three cases were seen in non-residents. The incidence of complete HUS in Belgian residents was 4.3 cases/100 000 in children <5 years old, 1.8 cases/100 000 when all children <15 years were considered, and 0.42/100 000 when patients of all ages were taken into account. By combining bacteriologic and serologic results, evidence of VTEC infection was obtained in 64% of the patients, mainly but not exclusively in children with prodromal diarrhea. The 13 VTEC isolates belonged to serotypes O157:H7 (nine isolates), O26:H11, O121:H---, O145:H--- and O172:H--- (one each) and all produced VT2 (+VT2vh-a in three O157 strains) and were positive for the eaeA gene. CONCLUSIONS: The incidence rate found in this study and the high mortality and morbidity linked with this syndrome warrant further registration of pediatric and post-diarrheic adult HUS cases and also examination of stools for both O157 and non-O157 VTEC strains. For effective prevention of this disease, further study of the serotypes and accessory virulence factors associated with HUS is needed.