Assessment of minimal residual disease (MRD) in CBFbeta/MYH11-positive acute myeloid leukemias by qualitative and quantitative RT-PCR amplification of fusion transcripts

Abstract

The inv(16)(p13q22) chromosomal rearrangement associated with FAB M4Eo acute myeloid leukemia (AML) subtype is characterized by the presence of the CBFbeta/MYH11 fusion transcript that can be used to detect minimal residual disease (MRD). However, qualitative RT-PCR studies of MRD have so far produced conflicting results and seem of limited prognostic value. We have evaluated retrospectively MRD in a large series of CBFbeta/MYH11-positive patients employing both qualitative and quantitative (real-time PCR) approaches. 186 bone marrow samples from 36 patients were examined with a median followup of 27.5 months; 15 patients relapsed during follow-up. In qualitative studies, carried out by 'nested' RT-PCR assay, all patients in complete remission (CR) immediately after induction/consolidation therapy were found to be PCR positive. However, follow-up samples at later time points were persistently negative (except one case) in patients remaining in continuous CR (CCR) for more than 12 months. 16 patients were evaluated by quantitative real-time PCR assay: CBFbeta/MYH11 transcript copy number was normalized for expression of the housekeeping gene ABL, expressed as fusion gene copy number per 10 4 copies of ABL. A 2-3 log decline in leukemic transcript copy number was observed after induction/consolidation therapy. After achieving CR, the mean copy number was significantly higher in patients destined to relapse compared to patients remaining in CCR (151 vs 9, P Ͻ 0.0001 by Mann-Whitney test). Moreover, in CCR patients, the copy number dropped below the detection threshold after the treatment protocol was completed and remained undetectable in subsequent MRD analysis in accordance with results obtained by qualitative RT-PCR. On the contrary, in the seven patients who relapsed, the copy number in CR never declined below the detection threshold; thus a cut-off value discriminating these two groups of patients could be established. The findings of our study, if confirmed, might confer an important predictive value to quantitative real-time PCR determinations of MRD in patients with inv(16) leukemia.

Standardization and quality control studies of ‘real-time’ quantitative reverse transcriptase polymerase chain reaction of fusion gene transcripts for residual disease detection in leukemia – A Europe Against Cancer Program

“…33,45,46,176,177 However, due to the low number of patients so far examined, it was not possible to define a kinetic or a cutoff level for predicting relapse. 178 …”

Prognostic impact of RT-PCR-based quantification of WT1 gene expression during MRD monitoring of acute myeloid leukemia

“…Recently, molecular monitoring of minimal residual disease was shown to early identify patient groups at high risk of relapse and therefore provide a window for therapeutic intervention. [1][2][3] Monitoring of MRD is mostly based on quantitative PCR. Common targets include specific fusion transcripts, for example, AML1-ETO, CBFB-MYH11, PML-RARA, and MLL gene fusions.…”

Prognostic value of minimal residual disease by real-time quantitative PCR in acute myeloid leukemia with CBFB-MYH11 rearrangement: the French experience

“…Low MRD level after induction or following completion of the entire chemotherapy program was associated with a lower risk of relapse. [2][3][4] Schnittger et al 5 previously described a strong prognostic impact of the initial fusion transcript level by creating a score based on the median expression ratio after consolidation therapy and the 75th percentile of the expression ratio at diagnosis. However, no study has defined an absolute threshold for the early molecular response, which could be used for outcome prediction.…”

“…Furthermore, in previous series, MRD was assessed at different follow-up time points, most frequently after the end of treatment. [2][3][4] An early prediction of prognosis is needed to quickly propose alternative strategies of consolidation. A significant association between relapse risk and CBFB-MYH11 transcript levels after induction or after the first course of consolidation therapy (threshold at 0.1%) has been reported, 3 but no survival analysis was performed.…”

“…[2][3][4] An early prediction of prognosis is needed to quickly propose alternative strategies of consolidation. A significant association between relapse risk and CBFB-MYH11 transcript levels after induction or after the first course of consolidation therapy (threshold at 0.1%) has been reported, 3 but no survival analysis was performed. In contrast, Schnittger et al 5 reported that the kinetics of reduction of the transcript level after the first and second induction cycle and after consolidation treatment had no impact on outcome.…”