Abstract
Assessment of MRD in APL by Q-PCR analysis for the presence or absence of PML-RARα mRNA after the completion of consolidation therapy in treatment regimens employing all-trans retinoic acid (ATRA) and chemotherapy (CT) has provided a reasonably accurate but not infallible prognostic test for disease recurrence (DR). This assessment has been made at detection sensitivity levels up to 1 in 104, which is considered to have optimal predictive value, since non-quantified lower levels of PML-RARα mRNA may not be relevant to clinical outcome. In the current study, we compared the results of Q-PCR (touchdown PCR for 40 cycles followed by post-PCR blot hybridization with a RARα chemiluminescent probe) to results from RQ-PCR in 15 patients on a Phase II clinical trial designed to eliminate CT (low-risk/LR patients, WBC <10K) or minimize CT (high-risk/HR patients, WBC >10K), using combined ATRA and arsenic trioxide therapy. All patients achieved complete remission (CR) and have been followed for a median of 1.2 years. DR occurred in 0/8 LR, 0/1 unclassified and 2/6 HR patients. At CR, all Q-PCR and RQ-PCR assays were positive. For RQ-PCR, the normalized quotient (NQ) value of PML-RARα, i.e., the PML-RARα copy number divided by the housekeeping gene glyceraldehyde-3′-phosphate dehydrogenase (GAPDH) copy number, ranged widely from 1.5 x 10−6 to 7.0 x 10−3. Post-CR follow-up data were:Patient GroupNumberQ-PCR+RQ-PCR+NQ RangeLR/Unclassified90/2416/249 x 10−9 – 1 x 10−7HR/no DR40/238/232 x 10−9 – 3 x 10−7HR/DR21/75/71 x 10−8 – 5 x 10−5For the 2 HR/DR patients, the last pre-DR results were, respectively: Q-PCR, positive and negative; RQ-PCR, NQ = 5 x 10−5 and NQ = 2.3 x 10−7. In overall comparisons to RQ-PCR results, all Q-PCR assays were positive for NQ >10−5, 3/4 were positive in the NQ 10−6 to 10−5 range, and all were negative for NQ <10−6. Additionally, dilution-reconstruction experiments confirmed the stochastic nature of PCR assays near the detection limit and indicated that RQ-PCR was about 10-fold more sensitive than Q-PCR. Although larger studies are needed, our results suggest that HR patients who are Q-PCR-negative but RQ-PCR-positive in the >10−7 – <10−5 range at a critical post-CR checkpoint may be at increased risk of DR and, more generally, they suggest that the results of RQ-PCR testing at a critical checkpoint can be used in combination with clinical risk assessment to stratify the intensity of subsequent MRD monitoring in individual patients.
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have