Monitoring minimal residual disease and predicting relapse in APL by quantitating PML-RAR alpha transcripts with a sensitive competitive RT-PCR method

Qualitative RT-PCR methods used for monitoring minimal residual disease (NI RD) in APL patients fail to predict relapse in up to 25% of patients in rem ission. We report here the development and evaluation of a highly sensitive (10(-5) and 10(-6) with one round and two rounds of PCR, respectively) com petitive RT-PCR method to quantitate the PML-RAR alpha: fusion transcripts. PML-RAR alpha transcript's levels were normalised to 10(5) copies of ABL t ranscript. Serial BM and PB samples from 16 patients with APL and t(15;17) were examined. Presentation samples from three patients (three BM, one PB) showed levels in the range of 0.7 x 10(6)-3.5 x 10(6) and 1.2 x 105 molecul es in BM and PB samples respectively. Serial quantitation of MRD in both BM and PB samples showed significantly lower levels of PML-RAR alpha transcri pts in remission, although the majority of samples remain positive for the PML-RAR alpha transcripts even those in long-term remission (up to 94 month s). Levels of PML-RAR alpha in remission samples were up to 2 x 10(2) and u p to 5.2 x 10(1) molecules in BM and PB respectively. BM and PB samples tak en from two patients 2-4 months before relapse showed significantly higher levels of PML-RAR alpha transcripts (1.2 x 10(4) molecules in BM; 3.5 x 10( 2), 1.2 x 10(2) and 1.2 x 10(3) in PB). The same samples, when tested with a standard qualitative PT-POP for the amplification of PML-RAR alpha (with a sensitivity of 10(-4)) produced negative results. This indicates that the qualitative methods would not have predicted relapse in these patients. Ou r data show that quantitating PML-RAR alpha transcripts with a sensitive me thod may provide a superior approach for monitoring MRD in APL and identify ing patients at high risk of relapse.