Detection of bcr/abl transcripts by RT-PCR and their colorimetric evaluation in chronic myeloid leukemia patients receiving allogeneic bone marrow transplantation

Abstract

BACKGROUND. Chronic myeloid leukemia (CML) is a disease characterized by the presence of a unique molecular marker, i.e. the fusion gene bcr-abl and its mRNA and protein products. This marker permits minimal residual disease follow-up after bone marrow transplantation (BMT) through cytogenetic or molecular analysis. Although the reverse transcriptase polymerase chain reaction (RT-PCR) method is largely employed, the clinical value and impact of a positive RT-PCR as a herald of hematological relapse has not yet been definitively ascertained. METHODS. In order to verify the frequency of bcr-abl positivity in CML patients who underwent alloBMT, we performed serial two-step RT-PCR on 63 peripheral blood and bone marrow specimens obtained at different times after non T-cell-depleted BMT from 16 CML patients treated in our Institution. After amplification, RT-PCR products were always checked by liquid hybridization with a specific probe. Median molecular follow-up after BMT was 38 months (range 2-144 months). RESULTS. None of the patients studied presented clinical hematological relapse after BMT. Six out of sixteen patients were found to be positive for bcr-abl. PCR positivity appeared in 4/6 patients more than one year post-BMT and in 2/6 patients within one year post BMT. In both instances PCR was an isolated finding in 5/6 patients and reverted to negativity in subsequent analysis; only one case was PCR-positive twice. It is noteworthy that RT-PCR positivity appeared in five patients presenting acute or chronic graft versus host disease (GVHD) and in one patient who had received MUD-BMT. CONCLUSIONS. In our cohort of patients, transient bcr-abl positivity had no clinical relevance and was also found in MUD-BMT without heralding hematological relapse. Our observations further stress the importance of applying only quantitative PCR methods during the post BMT follow-up of CML patients.