The amount of BCR-ABL fusion transcripts detected by the real-time quantitative polymerase chain reaction method in patients with Philadelphia chromosome positive chronic myeloid leukemia correlates with the disease stage

The use of the real-time reverse-transcription polymerase-chain reaction (R T-PCR) method to quantify BCR-ABL transcripts before and after allogeneic t ransplant was prospectively studied in 65 patients with chronic myeloid leu kemia (CML). The expression of the BCR-ABL transcript was determined and no rmalized using the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) houseke eping gene product as an endogenous reference. In the single step real-time PCR assay, tenfold serial dilutions of cDNA of the K5652 cell line remaine d positive down to 100 pg cDNA only. However, molecular relapses of CML aft er transplant were only safely detectable when a nested real-time PCR assay was performed, which was able to detect 1-10 pg cDNA from a tenfold serial dilution. The median normalized BCR-ABL transcript level was measured as 0 .004% in 17 patients with a molecular relapse, 0.4% in 7 patients with a cy togenetic relapse, 2.6% in 36 patients with a stable phase of CML, and 36% in 5 patients with a relapse in a blast crisis. The analyzed median normali zed amount of BCR-ABL transcript differed significantly (P<0.001) between t he various disease stages. In ten CML patients with relapse, the real-time PCR method was used to monitor the response of various immunotherapies as d onor leukocyte infusions, withdrawal of immunosuppression, or interferon-a application. The results of the quantitative evaluation of BCR-ABL transcri pts reflected very well the clinical effect of the different applied immuno therapies. The new real-time PCR method seems to be a suitable technique fo r the early detection of relapse after allogeneic transplant in patients wi th the BCR-ABL transcript. Its ability to distinguish between molecular and cytogenetic relapse (P<0.001) allows early therapeutic decisions.