A compelling case for central pathology review of multiparameter flow cytometry measurable residual disease results in Acute Myeloid Leukemia (AML) clinical trials: Data from the PALG-AML1/2016 multicenter randomized study

Abstract BACKGROUND: Measurable residual disease (MRD) is increasingly recognized as a critical prognostic factor in AML for guiding therapeutic decisions and predicting patient outcomes. Multiparametric flow cytometry (MFC) is the most widely used MRD assessment method, with applicability in about 90% of AML patients (pts). However, obtaining accurate and reproducible MFC-MRD results requires substantial experience and expertise. Local MRD testing is commonly used in clinical practice and investigational trials but the correlation between MRD and outcomes may be confounded by interlaboratory variability. AIM: In this analysis, we evaluated the concordance between local and central MFC-MRD assessments and its impact on relapse-free survival (RFS) and overall survival (OS) in AML patients in first complete remission (CR1) using data from a prospective, multicenter, randomized phase 3 trial comparing two intensive chemotherapy induction regimens in AML (NCT03257241). PATIENTS AND METHODS: Pts with newly-diagnosed, untreated AML, ECOG performance status 0–2 and HCT-CI≤ 3 were randomized to Daunorubicin+Ara-C (DA-90) (n=220) or Daunorubicin+Ara-C+Cladribine (DAC) (n=219) induction chemotherapy (IC). Pts with >10% blasts in non-aplastic bone marrow at day 14 received early second IC with D-45 and DAC, respectively. Pts who achieved a CR/CRi/CRp (cCR) were offered IDAC consolidation with or without alloSCT according to predefined risk groups. Serial samples for multi-modality MRD assessment were collected at cCR after 1 or 2 inductions (MRD1), and after each consolidation cycle (MRD2-4). MRD evaluation using 6-8-colour MFC with LAIP-based analysis was performed at 16 local MFC labs and MRD-1 and MRD-2 FCS files were subsequently independently evaluated by a central reviewer. The ELN threshold of <0.1% for MRD negativity (MRD-) was used by the local laboratories and central reviewer. RESULTS: MRD results were available from local laboratories for 279 (87.7%) pts in cCR at MRD1 and for 267 (84%) pts at MRD2). Centrally reviewed MRD1 data were available for 173 (54.9%) pts in cCR and for 139 (43.7%) pts in MRD2. Reasons for missing MRD results from central review were: missing or inaccessible FCS files (n=69); insufficient cell acquisition or inadequate antibody panel selection identified by central assessment (n=54); lack of LAIP target by central review (n=22). Of the MRD1 results classified locally as MRD negative (MRD1-), 97.6% (81/83 pts) were concordant with central assessment. However, 74/90 pts (82.2%) considered MRD1+ by local evaluation were reclassified as MRD1- after central review, leading to an overall MRD1 concordance rate of 56.1%. The overall concordance between local and central evaluation of MRD2 was 70.7%, (96.7% and 22.4% concordance with central assessment for MRD2- and MRD2+ results, respectively). Cohen's kappa coefficient was 0.15 for MRD1 and 0.23 for MRD2, indicating poor agreement. Centrally reviewed MRD2 demonstrated strong prognostic value for overall survival, with a hazard ratio of 2.32 (95% CI: 1.1–5.1; p=0.034) in multivariable analysis adjusted for clinical factors. In contrast, MRD1 did not reach statistical significance (p=0.250). Locally assessed MRD1 and MRD2 results were not predictive of overall survival (p=0.829 and p=0.523, respectively) in the cohort with centrally reviewed data. This was also the case when multivariable analysis included all MRD results submitted by local laboratories. CONCLUSIONS: These findings underscore the critical need for expert central verification of MFC-MRD results in multicenter AML trials, as local MRD analysis showed limited prognostic value and substantial discordance with centralized evaluation.