Metastatic breast cancer and circulating exosomes. Hints from an exploratory analysis

Abstract

Background: Exosomes are nanovesicles secreted by living cells. Depending on their origin, exosomes could carry different biological molecules capable of influencing tumor microenvironment. Aim of the study is to characterize circulating exosomes and to explore their possible association with clinicopathological features of metastatic breast cancer (MBC). Patients and methods: The study enrolled 56 MBC patients ( pts) treated at the University Hospital of Udine between 2013 and 2015, regardless line of treatment. Exosomes were isolated from plasma by ExoQuick solution. After this enrichment step, exosomes were conjugated with anti-CD63 coated beads to assess their protein expression profiles by flow cytometry. The fluorescence intensities of different antibodies were tested on single bead-exosomes decorated with CD63 and CD9 antibodies. The differences in exosomal subpopulation distribution between controls and MBC patients and their association with clinicopathological features were analyzed through Wilcoxon-Mann-Whitney test. Results: MBC pts differed significantly in specific exosomes subpopulations. In particular, compared to controls, a higher expression of CD44 (P = 0.0372), HGFR (P = 0.0311), CXCR4 (P = 0.0094), CD49d (P = 0.0441), E-cadherin (P = 0.0003) and HER2 (P < 0.0001) was observed. Exosomes positive for CXCR4 characterized pts affected by luminal or HER2 positive disease (P = 0.0199 and P = 0.0227, respectively), and among the latter subgroup a lower expression of E-cadherin (P = 0.0137) was also observed. A disease with KI67 >14% was associated with higher expression of CD49D (P = 0.0487). Pts with multiple metastatic sites had a higher fraction of HER2 (P = 0.0376) and marginally of KDR-positive exosomes. Visceral localizations were associated with higher expression of KDR (P = 0.041) and, in particular, lung involvement was associated with a lower expression of CD49d (P = 0.0438). Of note, liver localizations were marginally associated with higher expression of CXCR4 and KDR. Pts beyond 1st line of treatment showed a higher proportion of CD44 positive exosomes (P = 0.0379), whereas a marginally lower expression of EGFR was observed in pts beyond the 3rd line. Conclusions: In pts with MBC, distinct subpopulations of exosomes were observed according to tumor biology, disease burden and therapeutic history. The capability of exosomes to be a proxy of disease characteristics and a potential predictor of metastatization spread warrants further investigation through larger trials.