Impact of Chronic Lymphocytic Leukemia-derived extracellular vesicles onthe microenvironment immune landscape

The bidirectional cross-talk between chronic lymphocytic leukemia (CLL) B-cells and the immune cells of the microenvironment (ME) is leading toimmunosuppression, CLL cell survival and resistance to treatment.Recently, extracellular vesicles (EVs) have been shown to participate to asubstantial part of the intercellular communication by their ability to transfergenetic material. We hypothesize that CLL-EVs could have an impact onME immune cells. In the present project, we will characterize EVs producedby CLL cells obtained from CLL patients in (1) unstimulated, (2) BCR-stimulated, and (3) under inflammation conditions. First, we will determineCLL-EV content in these 3 conditions by next-generation sequencing(NGS), with a particular focus on small RNA (microRNA and Y RNA). Wewill then investigate the impact of these 3 EV types on gene expressionprofiles of healthy immune cells (CD4/CD8 T cells, B cells and monocytes)in order to evaluate the pathways deregulated by these CLL-EVs. Thedifferentially expressed genes after EV treatment will be correlated with thecontent of each EV type. We will investigate the functional impact of the EVtypes on healthy immune cells in vitro, with a particular focus on immunesynapse formation, migration, proliferation, apoptosis and chemoresistanceto new small molecules, in addition to the relevant pathways identifiedabove. In parallel, we will study EVs in an in vivo context using a newmethod of EV isolation, directly from the spleen ME of the Eμ-TCL1transgenic CLL mouse model. We will perform transcriptomic and proteomicanalysis of these EVs as well as track the immune target cells of these EVsin vivo. Effect of (1), (2) and (3) EVs will be studied in vivo by monitoringmouse survival, cytokine production in the plasma, leukocytosis andresistance to new treatment. Finally, pertinent RNA or protein discoveredwill be evaluated as prognostic biomarkers using serums samples of a well-characterized CLL patient cohort.