Exosomen, die von Myelomzellen und Knochenmarkstromazellen sezerniert werden, können die Expression von Adhäsionsmolekülen gegenseitig modulieren und MM-Chemotaxis und Bortezomib-induzierte Apoptose beeinflussen

UDI J., VOLL R., URETA D., LÓPEZ M., DUYSTER J., ENGELHARDT M., ÁLVAREZ É.

Hamburg

Congreso; Jahrestagung der Deutschen, Österreichischen und Schweizerischen Gesellschaften für Hämatologie und Medizinische Onkologie; 2014

Introduction: In MM pathogenesis, the interaction of plasma cells and the BM microenvironment plays a crucial role. Exosomes are secreted by nearly all cell types,including cancer cells, and have been implicated in intercellular communication, drug resistance and tumor progression. We here purified, characterized and studied theinfluence of exosomes secreted by various MM cell lines (MMCLs) and BMSCs on cell adhesion, migration and bortezomib-induced apoptosis.Methods: Exosomes were purified from the supernatants of MMCLs (IM-9, MM.1S, U266, RPMI8226) and M2-10B4 BMSCs after 72h of culture with RPMI1640 + 10%exosome-depleted FBS by differential centrifugation. Purity and quality of exosomes was confirmed by electron microscopy and western blot for CD63. Exosomal proteinconcentration was quantified by Bradford. CD44 and ICAM1 were evaluated by flow cytometry. Apoptosis was assessed via annexinV/7-AAD-staining. Chemotaxis to M2-10B4 exosomes (100-500μg/ml) was studied using chemotaxis chambers. Migrated cells were counted by flow cytometry.Results: Exosomes from IM-9 cells were identified by electron microscopy, showing the characteristic saucer-like morphology and ranging in diameter between 30-120nm.Both MM cell- and M2-10B4-exosomes expressed CD63. Mean exosomal protein concentration from MMCLs varied between 1.2-1.7μg/μl and was 1.4μg/μl for M2-10B4. M2-10B4 cells when incubated with exosomes from MMCLs vs. without expressed higher levels of both adhesion molecules CD44 and ICAM-1, however failing to reach statisticalsignificance. Conversely, M2-10B4 exosomes induced a significant increase in CD44 expression on IM-9 cells (p=0.03), reflecting an increased adhesion to components ofthe extracellular matrix that might contribute to cell-adhesion-mediated drug resistance. Indeed, IM-9 apoptotic cells after 24h with 10nM bortezomib decreased from 72%to 61% (-11%) in the presence of M2-10B4 exosomes and similarly from 74% to 62% (-12%) with IM-9 exosomes, suggesting an exosome-induced bortezomib resistance.Of note, migration of IM-9 cells increased with 200μg/ml M2-10B4 exosomes.Conclusions: Our findings underline the role of M2-10B4 exosomes on adhesion and migration of MM cells, e.g. IM-9, and the protection of M2-10B4 and IM-9 exosomes onbortezomib-induced cytotoxicity, suggesting their involvement in a paracrine/autocrine loop modulating treatment response and MM cell survival.