RESUMEN
Around 40-50% of patients with chronic myeloid leukemia (CML) who achieve a stable complete molecular response (CMR; undetectable breakpoint cluster region-Abelson leukemia gene human homolog 1 (BCR-ABL1) mRNA) on imatinib can stop therapy and remain in CMR, at least for several years. This raises the possibility that imatinib therapy may not need to be continued indefinitely in some CML patients. Two possible explanations for this observation are (1) CML has been eradicated or (2) residual leukemic cells fail to proliferate despite the absence of ongoing kinase inhibition. We used a highly sensitive patient-specific nested quantitative PCR to look for evidence of genomic BCR-ABL1 DNA in patients who sustained CMR after stopping imatinib therapy. Seven of eight patients who sustained CMR off therapy had BCR-ABL1 DNA detected at least once after stopping imatinib, but none has relapsed (follow-up 12-41 months). BCR-ABL1 DNA levels increased in all of the 10 patients who lost CMR soon after imatinib cessation, whereas serial testing of patients in sustained CMR showed a stable level of BCR-ABL1 DNA. This more sensitive assay for BCR-ABL1 provides evidence that even patients who maintain a CMR after stopping imatinib may harbor residual leukemia. A search for intrinsic or extrinsic (for example, immunological) causes for this drug-free leukemic suppression is now indicated.
Asunto(s)
Antineoplásicos/uso terapéutico , ADN de Neoplasias/genética , Proteínas de Fusión bcr-abl/genética , Enfermedad Injerto contra Huésped/genética , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Piperazinas/uso terapéutico , Reacción en Cadena de la Polimerasa/métodos , Pirimidinas/uso terapéutico , Adulto , Anciano , Benzamidas , Femenino , Estudios de Seguimiento , Enfermedad Injerto contra Huésped/prevención & control , Humanos , Mesilato de Imatinib , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , Leucemia Mielógena Crónica BCR-ABL Positiva/patología , Masculino , Persona de Mediana Edad , Inducción de Remisión , Sensibilidad y Especificidad , Tasa de Supervivencia , Resultado del TratamientoRESUMEN
Increasing numbers of patients with chronic myeloid leukaemia (CML) treated with tyrosine kinase inhibitors achieve undetectable levels of BCR-ABL mRNA using sensitive quantitative real-time reverse transcriptase PCR (RT-qPCR) methods and a method to measure minimal residual disease (MRD) in patients with low levels could be of value. Following isolation and sequencing of the patient-specific BCR-ABL breakpoint, a DNA-based nested qPCR assay was established, and MRD was measured by this method and one-round RT-qPCR in 38 samples from 24 patients with CML. Mixing experiments using patient DNA in normal DNA indicated that DNA qPCR could detect BCR-ABL sequences at a limit of approximately 10â»6. In 22 samples in which MRD was detectable by both methods, comparison of the results of DNA qPCR with the results obtained on the same sample by RT-qPCR showed good correlation. In another 16 samples, BCR-ABL mRNA was not detectable by RT-qPCR. In 8 of the 16 samples, BCR-ABL DNA was detected at levels ranging from 1.1 × 10â»5 up to 2.8 × 10â»4 and in the remaining eight samples BCR-ABL was not detected by either method. In one patient, who had stopped imatinib, an almost 1000-fold rise in MRD, to 5.2 × 10â»4 was observed in sequential samples. Nested DNA qPCR was more sensitive than one-round RT-qPCR and could be used for the monitoring of patients with CML with very low levels of MRD.
Asunto(s)
Proteínas de Fusión bcr-abl/análisis , Leucemia Mielógena Crónica BCR-ABL Positiva/diagnóstico , Neoplasia Residual/diagnóstico , Antineoplásicos/uso terapéutico , Benzamidas , ADN de Neoplasias/análisis , Proteínas de Fusión bcr-abl/genética , Humanos , Mesilato de Imatinib , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Piperazinas/uso terapéutico , Reacción en Cadena de la Polimerasa/métodos , Inhibidores de Proteínas Quinasas/uso terapéutico , Pirimidinas/uso terapéutico , ARN Mensajero/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Sensibilidad y EspecificidadRESUMEN
While a considerable number of candidate Myb target genes have been reported to date, most of these are likely to play little or no role in transformation by myb oncogenes. Here we have used a conditionally myb-transformed myeloid cell line (ERMYB) to further examine Myb regulation of one candidate target gene--c-myc--that has the potential to affect cell proliferation. It was found that the major influence on c-myc expression was the presence of cytokine (GM-CSF) rather than Myb activity. We also describe the application of PCR-based subtractive hybridization and low-density cDNA array screening, in conjunction with the ERMYB line, to the identification of additional Myb target genes. Preliminary identification of a number of candidates is reported; these include myeloperoxidase, which is known to have essential Myb-binding sites in its regulatory region.