It is not clear if absence of BCR-ABLtranscripts-complete molecular response (CMR)-is synonymous with, or required for, cure of chronic myeloid leukemia (CML). Some patients achieve CMR with imatinib (IM), but most relapse shortly after treatment discontinuation. Furthermore, most patients in long-term remission (LTR) post-stem cell transplantation (SCT) are considered functionally cured, although some remain occasionally positive for low-level BCR-ABL mRNA. Interpretation of the latter is complicated because it has been observed in healthy subjects. We designed a patientspecific, highly sensitive, DNA quantitative polymerase chain reaction to test follow-up samples for the original leuke-
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 10⁻⁶. 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 × 10⁻⁵ up to 2.8 × 10⁻⁴ 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 × 10⁻⁴ 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.
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