Targeted mass spectrometric detection of serum paraproteins with unique V-Region peptide surrogates: a paradigm shift in assessing minimal residual disease in multiple myeloma

Abstract

Aims Monitoring of minimal residual disease (MRD) in multiple myeloma (MM) by conventional serum protein electrophoresis (SPEP) and immunofixation (IFX) is hampered by poor sensitivity, and bone marrow-based techniques have significant drawbacks. We used high resolution Orbitrap mass spectrometry (MS) to derive variable (V)-region molecular signatures of serum para-proteins, and compared a targeted MS approach for paraprotein detection with SPEP/IFX. Methods De novo sequencing of paraprotein V-regions was performed from solution digests of SDS-PAGE gel plugs and whole serum by liquid chromatography tandem MS. Mutated V-region peptides were used to interrogate serum for paraprotein by targeted MS, and sensitivity compared with SPEP/IFX. Results Remarkably, there was extensive somatic mutation throughout the heavy and light chain constant regions, in addition to the expected V-region mutations. Targeted MS detected pico-grams of paraprotein per millilitre of serum, being at least 100 000fold more sensitive than IFX. Discussion This study shows for the first time that the secreted immunoglobulin proteome in MM is significantly more mutated than predicted from genomic studies, potentially bridging the gap between genotype and clinical disease in MM. Furthermore, targeted MS is a novel, individualised, proteomic approach to MRD monitoring, offering vastly increased sensitivity over conventional techniques, without compromising specificity.