Molecular measurement of BCR-ABL transcript variations in chronic myeloid leukemia patients in cytogenetic remission

Abstract

BackgroundThe monitoring of BCR-ABL transcript levels by real-time quantitative polymerase chain reaction (RT-qPCR) has become important to assess minimal residual disease (MRD) and standard of care in the treatment of chronic myeloid leukemia (CML). In this study, we performed a prospective, sequential analysis using RT-qPCR monitoring of BCR-ABL gene rearrangements in blood samples from 91 CML patients in chronic phase (CP) who achieved complete cytogenetic remission (CCyR) and major molecular remission (MMR) throughout imatinib treatment.MethodsThe absolute level of BCR-ABL transcript from peripheral blood was serially measured every 4 to 12 weeks by RT-qPCR. Only level variations > 0.5%, according to the international scale, was considered positive. Sequential cytogenetic analysis was also performed in bone marrow samples from all patients using standard protocols.ResultsBased on sequential analysis of BCR-ABL transcripts, the 91 patients were divided into three categories: (A) 57 (62.6%) had no variation on sequential analysis; (B) 30 (32.9%) had a single positive variation result obtained in a single sample; and (C) 4 (4.39%) had variations of BCR-ABL transcripts in at least two consecutive samples. Of the 34 patients who had elevated levels of transcripts (group B and C), 19 (55.8%) had a < 1% of BCR-ABL/BCR ratio, 13 (38.2%) patients had a 1% to 10% increase and 2 patients had a >10% increase of RT-qPCR. The last two patients had lost a CCyR, and none of them showed mutations in the ABL gene. Transient cytogenetic alterations in Ph-negative cells were observed in five (5.5%) patients, and none of whom lost CCyR.ConclusionsDespite an increase levels of BCR-ABL/BCR ratio variations by RT-qPCR, the majority of CML patients with MMR remained in CCyR. Thus, such single variations should neither be considered predictive of subsequent failure and nor an indication for altering imatinib dose or switching to second generation therapy. Changing of imatinib on the basis of BCR-ABL/BCR% sustained increase and mutational studies is a prudent approach for preserving other therapeutic options in imatinib-resistant patients.

Pretherapeutic Expression of the <i>hOCT1</i> Gene Predicts a Complete Molecular Response to Imatinib Mesylate in Chronic-Phase Chronic Myeloid Leukemia

“…To detect the expression of the BCR , the primer set and probe from Branford et al [23] were used. Amplification reactions were carried out with a Rotor-Gene RG-3000 (Corbett Biosciences, San Francisco, Calif., USA) in a total volume of 15 µl using a TaqMan Fast Universal Master Mix (Applied Biosystems, Foster City, Calif., USA) with 5 µl of the cDNA synthesis reaction following the protocol and conditions previously described [25]. A negative PCR control and an RT control, both containing nuclease-free water instead of cDNA, were included.…”

Investigation of human parvovirus B19 occurrence and genetic variability in different leukaemia entities

“…The diagnosis of ALL and AML was made through a morphological evaluation of BM biopsies for which immunophenotypic analysis for surface B-lymphocytic and T-lymphocytic markers was also performed. The CML diagnosis was confirmed by the presence of the BCR-ABL translocation by cytogenetic analysis, fluorescence in situ hybridization analysis, or molecular analysis as previously reported [39].…”

Mutations Associated with Imatinib Mesylate Resistance - Review

“…3 Despite this progress, up to 27% of patients who achieve complete cytogenic response (CCyR) and a few patients in the advanced phases of the disease show relapse or resistance while undergoing imatinib therapy. 4 Intrinsic resistance is also known as primary resistance and is caused in the absence of initial effi cacy of drug. Acquired loss of response or relapse is also known as secondary resistance and is caused when the drug loses its initial effi cacy on the patient.…”