Quantitative polymerase chain reaction (PCR)
• Quantitative real-time PCR is used to monitor residual disease and results are reported as
the BCR-ABL to a reference gene (recommended genes include ABL, BCR and GUSB).
Several technical aspects lack standardization like sample source (blood or marrow) and the
effects of shipping and storage, the amount of sample required, the techniques of RNA extraction
and the preferred reference gene. Variations in all of these will give rise to variation in
comparative results between labs.
• In the IRIS trials, the standardized baseline was 36% and a PCR 3-log reduction was
therefore 0.036%.
• Standardized baseline values have to be determined by individual laboratories. The
baseline value can vary from laboratory to laboratory and is not always included in the report.
• When negative results for PCR are reported, the sensitivities of the assay are not always
reported.
Advantages:
• Good correlation between blood and marrow samples
• Detection of very low levels of residual disease
Disadvantages:
• A substantial incidence of false negative results due to RNA degradation or low sensitivity
of the assay.
• Variability that is up to 0.5log.
• Poor results reproducibility
• Wide variation in transcript levels may be due to technical reasons rather than CML
changes.
• Due to the variation in PCR results, the PCR should be monitored from same source
(blood or marrow), by the same reliable high quality laboratory and using high-quality shipping
and collection procedures.
Recommendations: From the IRIS trials, there is 100% progression-free for patients reaching
MMR at 12 months of Gleevec in the 5-year follow-up and 95% progression-free survival for
those in CCR but not in MMR. Since the difference is low, a major change of therapy due to
absence of achievement of MMR at 12 months is not recommended until more data is there for
long-term outcome. PCR is monitored to detect relapses and MDACC has found incidences of
cytogenetic relapse among patients who never achieved a MMR and who had a 10-fold increase
in PCR levels. The second major reason to monitor PCR levels is to have serial monitoring to
measure Gleevec response. While a mutations analysis can be done when the PCR rises 2-5
fold, the yield of detection will be low in many labs compared to searching for mutations when
there is clinical and cytogenetic evidence of relapse.