Detection of M-Protein in Transplant-Ineligible Multiple Myeloma Patients on Daratumumab Using Liquid Chromatography Quadruple Time of Flight Mass Spectrometry

Background

Given the recent transition to novel front-line combinations capable of achieving a deeper response, the incorporation of measurable residual disease (MRD) into the response criteria for multiple myeloma (MM) could allow for better prognostication and potentially introduces MRD-adaptive treatment approaches. Liquid chromatography high-resolution mass spectrometry (LC-HRMS) is a powerful analytical tool that combines physical separation with high-resolution mass spectrometric detection. Mass spectrometry (MS) has emerged as a new diagnostic platform for monitoring M-proteins in MM, offering enhanced sensitivity for detecting M-proteins from peripheral blood compared to conventional serum protein electrophoresis/immunofixation analysis. Its utility has been endorsed in the 2021 IMWG guidelines.

Methods

Peripheral blood samples from MM patients were collected and used for validation of the MS. Samples were prepared in a multistep analytical process involving sample preparation by affinity purification targeting the conserved regions of immunoglobulins G, A, M, kappa, and lambda to enrich immunoglobulins and wash away serum proteins. Samples were analyzed on an Agilent 6545XT AdvanceBio LC/Q-TOF. M-proteins were identified by intact light chain masses above the polyclonal background. Samples were analyzed and compared to immunofixation electrophoresis (IFE) and flow cytometry (FC) (preliminary 10^-4, forthcoming 10^-5). Results were grouped into 4 categories: IFE+/QTOF+, IFE+/QTOF-, IFE-/QTOF+, IFE-/QTOF-. Limit of detection (LOD) studies were conducted by spiking IgG-kappa biologics into serum for both LC/Q-TOF and IFE.

Active and ongoing results

The LC/Q-TOF was able to isotype M-proteins similarly to IFE but with a mass accuracy approximately 1 Da. The LC/Q-TOF LOD was ~35 mg/L. 183 patient serum samples were analyzed. 134 were QTOF+/IFE+, 23 were QTOF-/IFE-, 23 were QTOF+/IFE-, and 3 were QTOF-/IFE+. When QTOF+/IFE- results were excluded due to the improved sensitivity of the LC/Q-TOF method, the concordance of LC/Q-TOF with IFE was ~98%.

Conclusions

Accurate mass determination of the M-protein intact light chain can readily differentiate daratumumab from IgG-kappa M-proteins. Excluding QTOF+/IFE-, the concordance rate was 98% between IFE and QTOF. QTOF demonstrated a LOD for IgG-kappa ~ 35 mg/L which is ~ 5-10x lower than most IFE estimated LODs. Future samples will compare QTOF to MRD assessments using next generation flow cytometry at a detection level of 10^-5.

Phua:Sanofi: Honoraria, Other: Travel support; EusaPharma: Honoraria; CSL: Honoraria, Other: Travel support; Pfizer: Honoraria; Beigene: Honoraria, Other: Travel support; Recordati: Honoraria, Other: Travel support; Bayer: Honoraria; AstraZeneca: Honoraria, Other: Travel support; Johnson & Johnson: Honoraria, Other: Travel support; FORUS Therapeutics: Honoraria, Other: Travel support; Octapharma: Honoraria, Other: Travel support; Roche: Research Funding; Abbvie: Honoraria, Other: Travel support.