Next Generation Flow and Next Generation Sequencing for Measurable Residual Disease Assessment in Multiple Myeloma...

The level of measurable residual disease (MRD) is one of the most important features correlating depths of response and long-term outcomes in multiple myeloma (MM) and MRD evaluation is currently the gold standard tool for assessing treatment response. Nevertheless, reproducibility across laboratories is a major concern, as discrepancies among results make comparability impractical.

herein, we report preliminary results from the "Italian MM-MRD network" project.

MRD in bone marrow (BM) samples have been measured from newly diagnosed MM patients using next-generation flow-cytometry (NGF) or next-generation sequencing (NGS) approaches in different laboratories.

The NGF workgroup (7 laboratories) implemented the Euro-Flow Standard-Operating-Protocol to reach minimum 1 × 10^-5 sensitivity. The inter-operator retrospective study (Stage 1) showed high inter-center concordance in monoclonal plasma cells detection (ICC = 0.90, p < 0.001), whereas moderate concordance was observed in the inter-laboratory correlation (Stage 2) in in-vivo samples (ICC = 0.63, p < 0.001), reaching a median limit-of-detection (LOD) and limit-of-quantification (LOQ) of 8 × 10-6 and 2 × 10^-5, respectively. Greater variability was also observed in the analysis of other BM cell populations. The NGS workgroup (4 laboratories) employed a targeted amplicon-based approach to detect clonotypic IGH/IGK gene rearrangements in diagnostic samples, subsequently used to track MRD in mock samples. The experimental design was divided into three quality-control (QC) rounds, focused on finding a shared strategy for clonotype identification (QC1: 100% concordance among centers), or quantifying MRD in mock samples (concordance: 81% [QC2]; 91% [QC3]). The 10^-5-sensitivity level was successfully reached in most of tested dilutions (QC2: 19/20 = 95%; QC3: 19/23 = 83%).

Overall, this pilot study provided preliminary data for MRD harmonization across Italian centers, paving the way for an expanded network, aiming at reducing variability, improving comparability, and enabling broader use of MRD-monitoring in clinical practice.