Proteomic characterization of CLL exosomes during disease evolution

SOTELO NATALIA; PRIETO DANIEL; MORANDE PABLO; ABREU CECILIA; ORTEGA CLAUDIA; CORREA AGUSTÍN; DURAN ROSARIO; IRIGOIN VICTORIA; LANDONI ANA INÉS; GABÚS RAÚL; DIGHIERO GUILLERMO; OPPEZZO PABLO

Sydney

Workshop; XVI International Workshop on Chronic Lymphocytic Leukaemia; 2015

Chronic Lymphocytic Leukemia (CLL) is an incurable disease characterized by theaccumulation of clonal B lymphocytes with a mature phenotype, resulting from a complexbalance between cell proliferation and apoptotic death. It is widely accepted that a continuouscrosstalk between cancer cells and local/distant host environment is required for effectivetumor growth. The interaction with accessory stromal cells within a specialized tissuemicroenvironment favors disease progression, by promoting malignant B-cell growth anddrug resistance. Among of the main actors of this dynamic and reciprocal interplay betweentumoral cells and their microenvironment are the nano-sized vesicles called exosomes.Emerging evidence indicates that exosomes play a key role in tumor-host crosstalk and theirsecretion, composition, and functional capacity are altered as tumors progress to anaggressive phenotype. The exosomes released by cancer cells can also signal to stromal cellswithin the cancer microenvironment, thus impacting tumor cell growth, metastasis, andangiogenesis. In CLL, it has been shown that plasma derived MVs mediate AKT activation inbone marrow stromal cells. Moreover, a recent work of Yeh et al. identified a specificexosomal microRNA signature and demonstrated that the high amount of exosomes found inCLL could be related to BCR signaling. Since several works state that exosomes promotetumor progression and that these small vesicles are increased in CLL plasma samples, wewondered: a) Is there a common proteomic profile on exosomes from CLL? b) Whichproteins are differentially expressed in exosomes during CLL evolution? and c) how couldthese proteins affect CLL progression? To this aim we studied 12 CLLs which weresegregated into indolent UM and Mut CLLs and progresssive UM CLLs (n=4 for eachgroup). Indolent and progressive patients were characterized by IgVH gene profile, AID andlipoprotein lipase expression. Samples from progressive patients were further segregatedfrom indolent CLLs by clinical parameters, such as a highly progressive disease, rapidlymphocyte doubling time (1 year), lymphocytosis >50.000/mm3 and FCR treatmentindicated before 3 years after debut. PB from progressive CLLs was obtained at debut andbefore treatment, whereas for indolent patients, PB was taken at debut and after 3-4 years offollow-up. Exosomes were purified from plasma on an iodixanol gradient under a differentialcentrifugation protocol, and their protein expression profile analyzed by nano liquidchromatography/Mass Spectrometry (nano LC-MS). Our results show for the first time acommon proteomic signature on exosomes from CLL and describe a differential expressionpattern between indolent and progressive CLLs. Taking into account that exosomes have arole in oncogenic transformation and cancer progression, we focused on the proteins thatwere overexpressed in plasma-derived exosomes from those patients whose CLL hadundergone clinical progression. Our data underline three proteins, i.e. gamma-catenin,thioredoxin-1, and S100-A9 that appear to be only in the plasma of patients with active CLLprogression. Interestingly, these proteins are associated with key signaling pathways ascancer progression, cellular proliferation and tissue inflammation which lead us to speculateabout the involvement of these proteins in CLL progression.