Multiple Myeloma-Derived Extracellular Vesicles Impair Normal Hematopoiesis by Acting on Hematopoietic Stem and Progenitor Cells

Abstract

Multiple myeloma (MM) is characterized by the abnormal proliferation of clonal plasma cells (PCs) in bone marrow (BM). MM-PCs progressively occupy and likely alter BM niches where reside hematopoietic stem and progenitor cells (HSPCs) whose viability, self-renewal, proliferation, commitment, and differentiation are essential for normal hematopoiesis. Extracellular vesicles (EVs) are particles released by normal and neoplastic cells, such as MM cells. They are important cell-to-cell communicators able to modify the phenotype, genotype, and the fate of the recipient cells. Investigation of mechanisms and mediators underlying HSPC-MM-PC crosstalk is warranted to better understand the MM hematopoietic impairment and for the identification of novel therapeutic strategies against this incurable malignancy. This study is aimed to evaluate whether EVs released by MM-PCs interact with HSPCs, what effects they exert, and the underlying mechanisms involved. Therefore, we investigated the viability, cell cycle, phenotype, clonogenicity, and microRNA profile of HSPCs exposed to MM cell line-released EVs (MM-EVs). Our data showed that: (i) MM cells released a heterogeneous population of EVs; (ii) MM-EVs caused a dose-dependent reduction of HSPCs viability; (iii) MM-EVs caused a redistribution of the HSPC pool characterized by a significant increase in the frequency of stem and early precursors accompanied by a reduction of late precursor cells, such as common myeloid progenitors (CMPs), megakaryocyte erythroid progenitors (MEPs), B and NK progenitors, and a slight increase of granulocyte macrophage progenitors (GMPs); (iv) MM-EVs caused an increase of stem and early precursors in S phase with a decreased number of cells in G0/G1 phase in a dose-dependent manner; (v) MM-EVs reduced the HSPC colony formation; and (vi) MM-EVs caused an increased expression level of C-X-C motif chemokine receptor type 4 (CXCR4) and activation of miRNAs. In conclusion, MM cells through the release of EVs, by acting directly on normal HSPCs, negatively dysregulate normal hematopoiesis, and this could have important therapeutic implications.

Acute Myeloid Leukemia Cells Functionally Compromise Hematopoietic Stem/Progenitor Cells Inhibiting Normal Hematopoiesis Through the Release of Extracellular Vesicles

“…Of note, we think that AML-EV treatment induced specific effects on HSPCs. In fact, performing similar experiments in another neoplastic context, such as MM, we obtained different results in HSPC differentiation and related functions ( 19 ).…”

“…Our hypothesis supported by literature data reporting that AML-EVs contain RNA, DNA and proteins involved in the pathobiology of AML and that their signaling induces change in the proliferative, angiogenic and migratory characteristics of stromal cells and HSPCs ( 13 ). Recently, we demonstrated that the transfer of specific miRNAs by multiple myeloma (MM)-EVs in HSPCs also occur in MM setting reporting their increased levels in HSPCs after 20 h of MM-EV treatment ( 19 ).…”

“…Concentration of EV proteins was obtained by Bicinchoninic Acid (BCA) Protein Assay Kit (Thermo Scientific, Rockford, IL USA) as previously reported ( 19 ). The absorbance at 560 nm was measured using the VICTOR Nivo (Perkin Elmer, Waltham, MA, USA) and protein concentration was determined.…”

“…Then, droplets were generated as previously described ( 19 ) and PCR amplification was carried out on a thermal cycler at 95°C for 10 min, then 40 cycles of 95°C for 15 seconds and 58°C (for miRNAs) or 60°C (for mRNAs) for 1 min, and finally 98°C for 10 min and 4°C infinite hold. A ramping rate of 2°C/sec was used in every step.…”

Multiple Myeloma-Derived Extracellular Vesicles Modulate the Bone Marrow Immune Microenvironment

“…showed for the first time that MM-derived EV can inhibit normal hematopoietic stem and progenitor cells and dysregulate hematopoiesis. These results suggest that MM-derived EV might have an important impact on stem cells ( 15 ). Furthermore, we and others have demonstrated that EV content in MM patients is different from healthy subjects, revealing its significance in the MM setting ( 16 – 19 ).…”