Plasmonic exosomes for enhanced monitoring of monoclonal gammopathies

Introduction Exosomes are vesicles with a size ranging from 30 to 150 nm secreted by most cells into the extracellular environment. These soft nano-objects carry rich molecular information of the parental cell and constitute an important route of intercellular communication. Exosomes participate in regulation of physiological as well as pathological processes, including the progression of different types of tumors. Recent studies suggest that exosomes carry determinant information about monoclonal gammopathies, like Monoclonal Gammopathy of Unpredicted Significance (MGUS) and Multiple Myeloma (MM). MGUS is a plasma cell disorder present in more than 3 % of the population aged over 50 years, and often asymptomatic MGUS stages precede MM. New biomarkers are needed to better predict MGUS' progression to MM. MM exosomes' colloidal properties have not yet been investigated to this purpose. Results from this project will contribute to address this problem, strengthening the toolbox to elucidate MGUS-MM switching. Materials and Methods We purified exosomes from serum of healthy donors, MGUS and MM patients. We then evaluated the purity and molar concentration of the exosome solutions by using a colorimetric nanoplasmonic assay we recently introduced. Relying on the discovery that cell surface's heparan sulfate proteoglycans (HSPGs) mediate exosome docking and processing by cells and that MM-derived exosomes experience a higher internalization rate compared to exosomes from MGUS and healthy individuals, we tested the ability of a Surface Plasmon Resonance (SPR) biosensor chip functionalized with heparin (a structural analogous of heparan sulfates), which constitute the side polysaccharide chains of HSPGs - to sort exosomes derived from healthy donors, MGUS and MM patients. Results The presence of typical exosomal markers in our sample indicates that we are actually dealing with exosomes. Over all, our analysis showed that MM patients produce more exosomes than MGUS and healthy individuals. In addition, we discovered that among the analyzed exosomes, only the MM-derived ones strongly bind heparin with an apparent dissociation constant equal to about 1 nM, indicating a high affinity binding. Discussion The analytical efforts and related biosensing strategies to study exosomes have been focused on their molecular profile, while colloidal properties, including size, molar concentration and cell membrane interactions, have been far less investigated.This work contributes to fill the gap combining colloidal gold nanoplasmonics and SPR biosensing. Our nanoplasmonic assay allowed to discover that serum from MM patients contain about four folds more exosomes than serum from MGUS and healthy individuals; SPR spectroscopy showed that among the analyzed exosomes, only the MM-derived ones have a significant binding affinity for heparin. Heparin, a structural analog of HSPGs, influences endocytosis through the interaction with several proteins. Findings lead us to think that antibody Free Light Chains (FLCs, produced and secreted via exosomes in both MM and MGUS) could play a major role in exosomes' internalization. SPR spectroscopy predicted that different exosomes, possibly with different FLC decorations, have different binding affinities for HSPGs. Since contradictory results are reported on the ability of FLCs to bind to HSPGs, further investigations are needed.