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    MRD and Biomarkers

    (OA-22) Comparative Analysis of Four NGS Protocols for Measurable Residual Disease Assessment Testing ctDNA and Bone Marrow Samples in Multiple Myeloma

    Saturday, September 20, 2025
    08:30 - 08:40 East Coast USA Time
    Location: Room 718

      Abstract Presenter(s)

    • Silvia Armuzzi, MS (she/her/hers)

      PhD Student
      Department of Medical and Surgical Sciences - University of Bologna; IRCCS Azienda Ospedaliero-Universitaria di Bologna, Seràgnoli Institute of Hematology, Bologna
      Bologna, Emilia-Romagna, Italy

    Introduction: MRD is a strong prognostic marker in Multiple Myeloma (MM) and MRD-negativity is recognized as an early endpoint for drug approval. NGS is a sensitive and robust method for MRD detection, but its use is limited by assay variability and reliance on non-universal proprietary platforms. Additionally, bone marrow (BM)–based MRD testing is invasive and not suitable for frequent monitoring.

    Here we report our experience on the application of different assays for patient (pts)-specific clonotype (ID) identification and MRD measurement by NGS, including a liquid biopsy approach, aiming at comparing the concordance between the different approaches and MRD results, in terms of specificity and sensitivity.


    Methods:

    The study included 50 NDMM pts. ID identification was performed by NGS with 4 different assays (clonoSEQ, EuroClonality (EC)-NGS, EC-NDC and LymphoTrack-Dx) on BM enriched PCs and on ctDNA (only EC-NDC). In 11 pts who achieved complete response, MRD was measured at the end of first line therapy both in nucleate cells (NCs) and in ctDNA.

    Results: At baseline, all samples were successfully analysed using the 5 different methods. A total of 929 rearrangements were identified, of which 605 (65%) were considered valid IDs, according to the assay-specific analytical algorithm. The specificity and originality of each sequence was assessed by an originality score, defined considering V and J regions specificity, CDR3 insertions/deletions and the clonality frequencies, leading to the recognition of 364/605 (60%) MRD-trackable IDs. Trackable ID sequences were compared using IMGT/V-QUEST and BLAST. Concordance was considered acceptable when a full or a partial match (median 55 bp) was identified by at least 3 methods for IgH and Igk, or at least 2 methods for IgH-D and IgL. Overall, concordance rates were 90% for IgH, 80% for Igk, 57.1% for Igl and 74% for IgH-D. The genomic profile was highly comparable between PCs and ctDNA.

    MRD was measured by Lymphotrack, EC-NGS and ClonoSEQ assays in NCs and by EC-NDC in ctDNA. Overall, 10/11 (91%) measurements were concordant in NCs. Overall, clonoSEQ and EC-NGS proved higher sensitivity and provided more precise quantification. Genomic aberrations were detected in 7/11 ctDNA samples, with 63% concordance with BM-MRD assessments. Moreover, ctDNA was informative in most cases. The results appear broadly consistent and overlapping across methods, supporting the applicability of these assays for reliable MRD monitoring.


    Conclusions:

    In conclusion, all assays showed high concordance, confirming their validity for ID evaluation with no discrepancies. Different approaches can yield consistent ID results. However, MRD assessment showed variability, with differences in sensitivity and quantification that may affect clinical decisions. Low-invasive methods proved feasible. These findings support the need for harmonized MRD guidelines with standardized criteria for ID identification and analysis.
    Acknowledgments: BoAIL, IMS-Career Award to S.A., AIRC22059