Chronic Lymphocytic Leukemia-Exosomes Switch Endothelial and Mesenchymal Stromal Cells into Cancer-Associated Fibroblasts to Sustain Leukemic Cell Survival
Authors: Paggetti Jerome , Haderk Franziska , Seiffert Martina , Janji Bassam , Kim Yeoun Jin , Diestler Ute , Ammerlaan Wim , Lichter Peter , Solary Eric , Berchem Guy , Moussay Etienne 
Published: November 2019

Abstract
Chronic lymphocytic leukemia (CLL), the most common hematologic malignancy in Western countries, is mostly affecting the elderly over 65 year-old. CLL is characterized by the accumulation of mature but non-functional B lymphocytes of clonal origin in the blood and the primary lymphoid organs. CLL was previously considered as a relatively static disease resulting from the accumulation of apoptosis-resistant but quiescent B lymphocytes. However, recent studies using heavy water labeling indicated that CLL is in fact a very dynamic disease with alternation of proliferation phases and peripheral circulation. A focus on the trafficking of CLL cells in vivo has shown that leukemic cells circulate between the blood and the lymphoid organs but have a preference for the bone marrow.
Recent next-generation sequencing of CLL cells indicated the presence of different genetic subclones. This intraclonal heterogeneity observed in CLL subpopulations may be in part determined by the interactions that leukemic cells entertain with their microenvironment when B cells migrate into the lymph nodes and the bone marrow. Indeed, tumor-stroma interactions are not only providing signals necessary for leukemic cells survival but may also influence the clonal architecture and evolution. One of these interactions involves CLL-derived exosomes.
Here, we show that CLL-exosomes efficiently transfer nucleic acids, including functional microRNAs, and proteins, including MHC-Class II molecules and B-cell specific proteins, to bone marrow mesenchymal stem cells and endothelial cells. CLL-exosomes also activate signaling pathways, including PI3K and NF-κB pathways, in these stromal cells. As a consequence, gene expression is strongly modified indicating a switch towards a cancer-associated fibroblast phenotype. Functionally, exosome-stimulated stromal cells show a striking actin cytoskeleton remodeling characterized by the formation of stress fibers, and enhanced proliferation, motility and angiogenic properties. We also identified several proteins synthesized and secreted by stromal cells that promote leukemic cell adhesion and survival ex vivo.
To confirm the involvement of CLL-exosomes in CLL pathology in vivo, MEC-1-eGFP cells were subcutaneously injected into immunocompromised NSG mice together with CLL-exosomes. We observed a significant increase in tumor size and a reduction in survival of exosome-treated animals. Flow cytometry analysis of selected organs indicated an enrichment in leukemic cells in the kidney, providing a potential explanation to the renal failures observed in CLL patients.
In conclusion, the communication between CLL cells and stromal cells may be a critical factor influencing CLL progression by promoting leukemic cell survival. This study demonstrates the crucial role of exosomes as mediators of the communication between leukemic cells and their microenvironment. Exosomes could thus represent a suitable target for therapeutic intervention in CLL.

Disclosures
No relevant conflicts of interest to declare.