Clonotypic Mass Spectrometry with Easym Assay for Detection of Measurable Residual Disease in Multiple Myeloma

The presence of measurable residual disease (MRD) is an important prognostic factor for multiple myeloma. However, MRD tests in current clinical use require bone marrow (BM) samples that are uncomfortable, expensive, and time-consuming for patients. BM-based assays are also limited by sampling of a single location within the marrow, which may not accurately represent overall disease burden. Consequently, there is an urgent unmet need for peripheral blood-based MRD testing that would allow for convenient, repeated assessment of MRD.

M-protein quantification using mass spectrometry (MS) is promising candidate methodology for peripheral blood MRD monitoring due to its exceptional analytical sensitivity. Specifically, "clonotypic" MS techniques can identify and track a specific M-protein "peptide fingerprint", allowing sensitive and specific measurement of MRD that may exceed BM-based techniques. However, clinical data regarding this approach is limited and no clonotypic MS assays are currently in clinical practice.

Consequently, we performed a pilot study to assess the use of clonotypic MS-based MRD testing using the EasyM system (Rapid Novor, Kitchener, ON). We retrospectively identified a subset of patients in our institutional tissue bank who met the following criteria: 1) IgG multiple myeloma, 2) M-protein ≥0.5 g/dL by serum protein electrophoresis (SPEP) at diagnosis, and 3) adequate samples for analysis. Baseline and follow up serum samples were analyzed by the study sponsor in a blinded fashion. EasyM results were subsequently compared to SPEP and, when available, next generation sequencing MRD testing with the clonoSeq assay (Adaptive, Seattle, WA). We used receiver-operator curve analysis to identify cutoffs for percent reduction in EasyM to predict BM-based MRD using the R package "cutpointr".

19 patients were identified for analysis. Median age was 62 and 52% were male. All patients received proteasome inhibitor-based induction and 17 patients received lenalidomide. All underwent autologous stem cell transplant (ASCT) in first remission. A clonotypic signature was identified in all 19 patients from the baseline sample. 54 follow up samples were analyzed. Percent change in EasyM and SPEP were well correlated (Pearson R = 0.808; p < 0.01). 37 timepoints had MRD testing available. Of these, 14 and 9 timepoints were negative at 10^-5 and 10^-6, respectively; only 3 (8%) had undetectable M-protein by EasyM, all MRD negative at 10^-6.

Using receiver-operator curve analysis, we identified optimal cutoffs for percent reduction in EasyM by area under the curve for BM-based MRD at 10^-5 (99.0%) and 10^-6 (99.3%) thresholds. The PPV and NPV for EasyM at 10^-5 were 91% and 80%. The PPV and NPV for EasyM at 10^-6 were 92% and 58%. Given the half-life of IgG (2 - 3 weeks), we also performed an exploratory analysis for MRD at 10^-5 separating day 100 samples (n = 18) from later samples (n = 18). This analysis yielded slightly different optimal cutoffs for percent reduction by EasyM for day 100 (98.2%) versus later timepoints (99.4%), with similar PPV (91% and 92%) and improved NPV (100% for both), suggesting time of sample collection may impact the predictive value of EasyM.

At day 100 after ASCT, 7 patients were in CR, 9 in VGPR and 3 in PR. 7 patients progressed during the follow up period. One patient had undetectable EasyM at day 100 after ASCT and received only 4 months of post-ASCT lenalidomide, but remains in CR over 3 years after transplant. Two patients achieved undetectable EasyM at 9 and 55 months, respectively, post-ASCT and remain in CR at 52 and 64 months.

Our study provides initial data regarding clonotypic MS with the EasyM assay for peripheral blood MRD testing. We demonstrated excellent correlation between percent reduction in M-protein by SPEP and EasyM. Interestingly, 79% of samples that were MRD negative at 10^-5 by clonoSeq had detectable M-protein by clonotypic MS, suggesting EasyM may be a more sensitive test for residual disease. After optimization, EasyM showed excellent PPV and NPV for BM-based MRD negativity, suggesting clonotypic MS be able to identify patients who can forego a bone marrow biopsy for MRD monitoring. We are currently validating this assay in non-IgG isotypes and light chain disease. Overall, clonotypic MS represents a promising new tool for myeloma monitoring and should be performed in larger retrospective cohorts and integrated into prospective trials for further validation.

Khalid:Rapid Novor: Current Employment. Liyasova:Rapid Novor: Current Employment. Zaydman:Siemens Healthineers: Honoraria. McDonald:Rapid Novor: Current Employment. Crees:BioLineRx Ltd.: Research Funding. Schroeder:Cellect Inc: Research Funding; Fortis: Research Funding; Genentech Inc: Research Funding; Incyte: Research Funding; Seagen Inc.: Research Funding; Amgen: Research Funding; Celgene: Research Funding. Yang:Rapid Novor: Current Employment. Vij:Takeda: Honoraria, Research Funding; Oncopeptides: Honoraria; Harpoon: Consultancy; Adaptive: Honoraria; Sanofi: Honoraria, Research Funding; GSK: Honoraria; Pfizer: Honoraria; BMS: Honoraria, Research Funding; Legend: Honoraria; Biegene: Honoraria; Janssen: Honoraria; CareDx: Honoraria.