Tracking peripheral residual disease in myeloma: Clonotypic sequencing and mass spectrometry in the teclistamab era

Abstract Introduction Multiple myeloma (MM) is an incurable hematological cancer and is characterized by the proliferation of a plasma cell clone secreting a monoclonal protein. Teclistamab, a BCMA-directed bispecific antibody, has demonstrated improved outcomes in relapsed and refractory MM in the MajesTEC-1 trial. Standard serum electrophoretic methods, such as serum protein electrophoresis (SPEP) and immunofixation (IFE), lack the analytical sensitivity to detect low level of residual monoclonal immunoglobulins, limiting their utility for measurable residual disease (MRD) assessment. Two emerging mass spectrometry-based techniques are revolutionizing the detection and quantification of monoclonal immunoglobulins : the intact immunoglobulin mass spectrometry (MS) and the clonotypic assay (EssayM). This study aimed to assess the performance of MS and EasyM for disease monitoring and early relapse detection in patients treated with teclistamab. Methods In this retrospective study conducted between December 2024 and July 2025, we assessed all consecutive RRMM patients with relapsed MM receiving teclistamab. Inclusion was limited to patients with measurable disease as determined by mass spectrometry-based assays. Serial monitoring with MS was performed monthly, while a clonotypic assessment using EasyM was conducted at baseline and every 2 months thereafter. In parallel, MRD was evaluated by next-generation flow cytometry (NGF) performed at the time MS negativity was documented, according to the Euroflow protocol (sensitivity 10-5). Monoclonal protein detection and quantification via MS was carried out using MALDI-TOF, with analytical validation based on the reference assay established by Mayo Clinic (Mass-Fix). EasyM, a peripheral clonotypic assay (Rapid Novor, Canada), involves de novo amino acid sequencing of the full-length M-protein and quantification of unique peptides with parallel reaction monitoring. Results We retrospectively evaluated 23 RRMM patients, with a median age of 72 years (range 31-91). High-risk cytogenetics were identified in 33%, and 37% presented with extramedullary disease at the initiation of teclistamab therapy. Patients received a median of 3 prior lines of treatment (range 1-8) and all patients were triple-class refractory. Sequencing success rate for EasyM was 92% (n=23/25). The two patients that were not sequenced had no detectable monoclonal protein and were excluded from this study. Best responses based on IMWG criteria were complete response (CR) in 8 patients, very-good partial response (VGPR) in 7 patients, and partial response (PR) in 5 patients. With a median follow-up of 11 months and the median PFS was 172 days. Notably, patients achieving sustained MS negativity demonstrated a 1-year PFS of 100% compared to 58% among those who remained MS positive. Eight patients achieved MS negativity. Among them, 3 also became negative by EasyM, whereas the remaining 5 retained detectable residual clones, with a median of 2.64% residual clone (range 1.17 -13.42%). Similarly, patients achieving CR showed a median residual clonal population of 1.93% (range 0.0-13.42%), despite being uniformly negative by MS. In patients who experienced progressive disease (n=3), a sharp increase in clonal burden was detected by EasyM within the first month of relapse, with a median residual clone level of 138% (range 105-632%). Notably, both SPEP and MS remained below the conventional 25% threshold for biochemical progression during this early relapse period. Regarding bone marrow MRD evaluation, 3 patients achieved concordant negativity by both NGF and EasyM. However, in 5 patients, MRD negativity was discordant with persistent clonal detection by EasyM, indicating residual disease at the molecular level despite immunophenotypic bone marrow clearance. Conclusion The clonotypic mass spectrometry assay (EasyM) demonstrated its ability to detect residual clonal disease undetected by conventional methods such as SPEP, MS and even bone marrow NGF MRD assessment. To our knowledge, this is the first study integrating bispecific antibody therapy with high resolution mass spectrometry profiling for response monitoring in MM. These findings suggest that EasyM is highly sensitive to detect residual monoclonal protein, highlighting its potential to improve monitoring and response assessment in relapsed and refractory multiple myeloma receiving with teclistamab.