Detecting PML-RARα transcript in acute promyelocytic leukemia using real-time quantitative RT-PCR

摘要

＜正＞ Background Real-time quantitative RT-PCR（RQ-PCR）assay has become a vital tool to monitor residual disease ofleukemia.However,the complexity and standardization of RQ-PCR should never be overlooked and the results shouldbe interpreted cautiously in clinical conditions.We aimed to assess the methodology of RQ-PCR and its clinicalapplications in monitoring molecular kinetics of 36 newly diagnosed cases of acute promyelocytic leukemia patients with t（15;17）from October 2004 to December 2005.Methods All the TaqMan probe-based RQ-PCR reactions and analysis were performed on an ABI-PRISM 7500platform.The quantitation of PML-RARα transcripts was represented by the normalized quotient,that is,PML-RARαtranscript copies divided by ABL transcript copies.According to induction therapy,the patients were classed into twogroups:group 1（n=23）,three-drug combination including arsenics,all-trans retinoic acid and mitoxantrone;and group 2（n=13）,two-drug combination from all-trans retinoic acid,arsenics and mitoxantrone.Results The sensitivity of RQ-PCR was 1 per 10~5 cells and 5 copies of the PML-RARα transcript could be reproduciblydetected.No false positive results occurred in 40 non-acute promyelocytic leukemia samples.Optimal amplificationefficiency could be attained,which was determined by the slope of the standard curves（slope:-3.2- -3.7）.Theinter-assay and intra-assay variation coefficients of the method were 1.01% and 0.56% respectively.Although the time toattain hematological complete remission was similar in both groups,the time to achieve molecular remission of group 1was significantly shorter than that of group 2（61 days vs 75 days,P=0.034）.The rate of molecular remission within 70days was higher in group 1 than in group 2（75.00% vs 38.46%,P=0.036）.Compared with pretreatment,medianreduction of the PML-RARα transcript before first consolidation therapy differed significantly between group 1 and group2（log scale,3.15 vs 2.31,P=0.024）.Interestingly,we found that PML-RARα transcript levels temporarily increased inbone marrow（7 patients）and peripheral blood（22 patients）samples of patients during induction therapy in both groups.Conclusions The RQ-PCR assay is reliable for the detection of PML-RARα transcripts.Arsenics,all-trans retinoic acidand mitoxantrone triad induction treatment of acute promyelocytic leukemia is superior to two-drug combination inductiontherapy in terms of the molecular response.