role of CML exosomes in the crosstalk between chronic myelogenous leukaemia and bone marrow-derived cells

Abstract

Exosomes are small membrane vesicles (30–100 nm) derived from the luminal membranes of multivesicular bodies and constitutively released by fusion with the cell membrane (1). Exosomes mediate local and systemic cell communication through the presence of cytokines, growth factors and others molecules. It is well recognized that bone marrow–derived cells (BMDCs) are crucial for the generation of a suitable microenvironment for the primary tumor and the development of metastasis through a process called pre-metastatic niche formation. Secreted factors are known contributors to BMDC recruitment to both the primary tumor and to pre-metastatic niches (2) and in particular exosomes may have a role in the crosstalk between the primary tumor and BMDCs, leading to the homing of both cell types to sites of metastasis (3). Chronic myelogenous leukemia (CML) is a myeloproliferative disorder characterized by the (9;22) reciprocal chromosomal translocation that produces the so-called Philadelphia (Ph) chromosome. This rearrangement results in the production of the chimeric bcr-abl oncoprotein, p210, with a constitutive tyrosine kinase activity. Taking these data into account, the aims of our study were (i) to clarify the role of CML-derived exosomes as new factors that promote metastatic niche formation by educating BMDCs toward a new phenotype. (ii) to better understand the molecular composition of CML-derived exosomes and the possible role of molecules, such as microRNAs (miRNAs), transferred by exosomes and eventually involved in this crosstalk. Here we investigated the role of LAMA84R-derived exosomes in the crosstalk between CML cell line, LAMA84R, resistant to imatinib, and the bone marrow stromal cell line, HS-5. Our preliminary results, obtained by real time PCR, show that pretreatment of bone marrow stromal cell line HS-5 with LAMA84R-derived exosomes induce, in a time dependent fashion, the expression of cytokines involved in angiogenesis such as IL-6 and IL-8. Moreover treatment of LAMA84R with recombinant IL-8 induce the activation of Akt and Erk ½, principal mediators of cell proliferation, survival, and chemotaxis. It is conceivable to hypothesize that IL-8 released by HS-5 cells after stimulation with CML exosomes, may modulate both myeloid malignant cells and bone marrow microenvironment. In this paracrine pathway CML exosomes could modulate the angiogenic process through a reciprocal stimulatory loop. Our previous paper showed that CML exosomes promote angiogenesis (4, 5) and other works indicate that hypoxia triggers a pro-angiogenic pathway involving cancer cell exosomes (6). Because of exosomes are involved in the horizontal transfer of information, through export of mRNAs and miRNAs (7), we focus on their miRNAs composition. miRNAs are short non- coding RNAs that regulate preferentially gene expression by inhibiting translation of specific target mRNAs. Our preliminary data indicate that LAMA84R exosomes transport miRNAs involved in angiogenesis and hypoxia. Further functional assays need to clarify the role of exosomes in the crosstalk between leukaemia and bone marrow cells and if miRNAs transferred by exosomes have a role in this process.