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N.J. Short and F. Ravandi et al.
tectable in long-term survivors (with rare exceptions60,61) and persist or re-emerge in the setting of relapsed dis- ease.19 In acute promyelocytic leukemia, the persistence of PML-RARA transcripts strongly predicts for relapse, and patients with persistent transcripts invariably relapse without intervention.30,31,62 Among patients with acute promyelocytic leukemia treated with chemotherapy plus all-trans retinoic acid, initiation of arsenic trioxide for those with positive MRD prevents morphological relapse in the majority of cases.31 However, in this context, it is important to confirm a positive MRD assessment with repeat testing, given the potential for false positives. Consensus guidelines have therefore defined molecular remission as an important therapeutic milestone and rec- ommend molecular assessment in routine clinical practice.63 The link between MRD status and prognosis, as well as an effective salvage therapy for MRD-positive dis- ease, makes acute promyelocytic leukemia a model for MRD-guided therapies, although it is unclear whether routine, universal monitoring is still required in the era of regimens capable of curing >95% of non-high-risk patients.64,65
Core-binding factor AML accounts for 20-25% of cases of AML in younger patients, but <10% in older adults.2,66 PCR-based MRD assessment of CBFB-MYH11 and RUNX1-RUNX1T1 transcripts for patients with inv(16) and t(8;21), respectively, has been consistently associated with risk of relapse across several studies.32,33,67-70 In the largest study of core-binding factor AML and MRD, 278 patients [163 with t(8;21) and 115 with inv(16)] were treat- ed.33 A >3 log reduction in RUNX1-RUNX1T1 in the bone marrow and CBFB-MYH11 copy number >10 in the peripheral blood after induction were the most prognostic measures of MRD for risk of relapse. Levels of MRD above these thresholds were associated with overall sur- vival in the t(8;21) group but not in the inv(16) group, pos- sibly due to better salvage options fpr patients with the latter subtype. Thresholds of MRD were identified that could predict relapse in 100% of patients in both groups and, notably, rising MRD levels on sequential samples pre- dicted morphological relapse, suggesting a potential role for pre-emptive therapy in this population.
Monitoring of mutant NPM1 throughout treatment may also provide important prognostic information in AML.15,34-36 Mutations in NPM1 are present in approxi- mately 30% of younger patients with AML and in up to 60% of cases of AML with normal karyotype.71 Like PML- RARA, CBFB-MYH11 and RUNX1-RUNX1T1, these muta- tions are stable at diagnosis and relapse, making them ideal targets for molecular MRD assessment.72 In the largest study of NPM1-based MRD (the National Cancer Research Institute AML17 trial), 346 patients with NPM1- mutated AML received intensive chemotherapy and were monitored for mutant NPM1 levels by PCR in the bone marrow and peripheral blood after each cycle of chemotherapy.15 With a median sensitivity of 10-5, MRD negativity in the peripheral blood after two cycles of chemotherapy was achieved by approximately 85% of patients and was the most prognostic MRD measure. Lack of achievement of this MRD milestone was the only inde- pendent prognostic factor for death in multivariate analy- sis [hazard ratio (HR) 4.84; 95% confidence interval (95% CI): 2.57-9.15; P<0.001). When MRD was assessed in the peripheral blood at this time point, the 3-year overall sur- vival rate was 24% for those who were MRD-positive and
75% for those who were MRD-negative (P<0.001). Similar to the experience with acute promyelocytic leukemia and core-binding factor AML, rising levels of mutant NPM1 transcripts on sequential testing reliably predicted relapse. In a separate study, patients with NPM1 transcripts detectable by PCR immediately prior to HSCT had significantly worse 5-year overall survival rates than those who were MRD-negative (40% vs. 89%: P=0.007); furthermore, the overall survival of those in complete remission prior to HSCT but with detectable NPM1 MRD was similar to that of patients transplanted with active disease.73 Together, these studies suggest that PCR-based MRD for mutant NPM1 is prognostic across clinical con- texts, both after induction/early consolidation and before HSCT.
Next-generation sequencing-based measurable residual disease studies
Compared to both MFC and PCR, the use of NGS as an instrument for detecting MRD is relatively new and con- sequently there are fewer studies to guide its use in clinical practice.17,45,46 In a study of 50 patients who underwent paired whole genome or exon sequencing at diagnosis and remission, 48% had persistent leukemia-associated muta- tions in at least 5% of bone marrow cells (i.e. variant allel- ic frequency ≥2.5%).45 These patients with persistent mutations had shorter event-free survival (median: 6.0 vs. 17.9 months, P<0.001) and overall survival (median: 10.5 vs. 42.2 months, P=0.004), compared to those of patients with lower post-remission mutation burden. In a study from the MD Anderson Cancer Center of 131 patients undergoing induction for newly diagnosed AML, com- plete molecular clearance (i.e. absence of detectable muta- tions by NGS) was independently associated with decreased risk of relapse and better event-free survival even when MFC-based MRD status was also available.46 These results were strengthened by removing mutations associated with CHIP (i.e. DNMT3A, TET2, and ASXL1, or “DTA” mutations) from the analysis, suggesting that persistence of these preleukemic mutations may not be associated with worse outcomes. In the largest study of NGS MRD in AML, 482 patients were evaluated with NGS at diagnosis and remission, 430 (89.2%) of whom had a leukemia-associated mutation detected at baseline.17 Mutations persisted in 51% of patients at variant allelic frequencies ranging from 0.02-47%. CHIP-associated DTA mutations were not associated with relapse. When DTA mutations were excluded from analysis, residual dis- ease, as detected by NGS, was independently associated with risk of relapse, relapse-free survival and overall sur- vival. Importantly, NGS MRD added prognostic informa- tion to MFC MRD status. Four-year relapse rates were similar among patients who were MRD-positive by NGS/MRD-negative by MFC and those who were MRD- negative by NGS/MRD-positive by MFC (52.3% vs. 49.8%, respectively), whereas patients who were MRD- positive or MRD-negative by both assays had highly divergent risks of relapse (73.3% vs. 26.7%, respectively). Together, these studies suggest that NGS-based MRD may complement MFC MRD assays, as these two approaches evaluate different targets (i.e. leukemia-associ- ated mutations and aberrant surface protein expression), and also that reduction and/or clearance of mutations associated with CHIP may not be necessary for cure.17,46
Due to the background error rate of commonly used
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haematologica | 2019; 104(8)