Minimally invasive characterization of peripheral blood measurable residual disease in multiple myeloma using high‐sensitivity detection of ctDNA by next‐generation sequencing

Measurable residual disease assessment in bone marrow (BM-MRD) is a crucial prognostic factor in multiple myeloma (MM), but its clinical use has some caveats, such as invasiveness, poor spatial representativity, and serial sampling. Novel minimally invasive approaches for monitoring peripheral residual disease (PRD) could facilitate its clinical use. We aimed to investigate a sensitive method for detecting PRD in patients with MM. This study included 84 patients enrolled in PETHEMA/GEM clinical trials, and a total of 292 longitudinal samples were analyzed. BM-MRD in these patients was assessed using EuroFlow-based next-generation flow cytometry (NGF) and PRD was simultaneously examined in cell-free DNA (cfDNA) using AltumTrackSeq®, a highly sensitive next-generation sequencing (NGS) method based on patient-specific multiplexed amplicon mini-panels that target somatic mutations identified at diagnosis. Our findings revealed that automated isolation methods enabled standardization and yielded cfDNA levels comparable to manual protocols. A high median cfDNA concentration of 393.2 ng was obtained, with a range from 17.3 to 6300 ng. Furthermore, we demonstrated that compared with traditional BM-MRD by NGF, our ctDNA-NGS approach showed a higher positive predictive value (PPV, 88.9% vs. 34.5%) and specificity (Sp, 98.4% vs. 70.3%) for predicting relapse during the follow-up period. Detectable ctDNA was associated with a high risk of progression and/or death (HR, 11.5; 95% CI, 2.66-49.4), and this was confirmed in the multivariate analysis. In conclusion, this assay offers a method for detecting imminent relapse risk in the peripheral blood of MM patients.