Rapid screening of leukemia fusion transcripts in acute leukemia by real-time PCR

Abstract

We established a real-time PCR method that can simultaneously detect 10 different fusion transcripts (major, minor and micro BCR/ABL, AML1/MTG8, PML/RARalpha, CBFbeta/MYH11, TEL/AML1, E2A/PBX1, MLL/AF4, and MLL/AF9) together with Wilms' tumor gene (WT1) transcripts. This screening method allowed the processing of six specimens concomitantly and required only one working day from RNA extraction to final results. Fifty-seven bone marrow (BM) samples from patients with acute leukemia were retrospectively screened for the presence of fusion and WT1 transcripts without knowledge of the cytogenetic data, and the fusion transcripts were detected in 20 of 57 samples (35.1%). The concordance between the present method and cytogenetic analysis was examined in 38 samples in which the cytogenetic data were available. In 12 of 38 samples, the PCR results agreed with the cytogenetic data, whereas in 4 of the remaining 26 samples, the translocations were detected by real-time PCR alone because of the insufficient number of metaphases obtained and presumably the submicroscopic or masked translocations. The WT1 levels ranged from 400 to 690,000 copies/microg RNA in BM from leukemia patients, whereas 0-470 copies/microg RNA were found in BM cells from BMT donors. This real-time PCR method enables rapid and efficient characterization of acute leukemia in addition to subsequent evaluation of minimal residual diseases.