Session Item

The induction of wound healing response is believed to be one of the reasons for local relapse in breast cancer (BC) patients. In normal tissue, it is necessary for tissue repair. However, induction of adaptive and innate immune response in tumor tissue stimulates cell survival, induction of angiogenesis, and extravasation of circulating tumor cells. Previous studies have shown that surgical wound fluids (SWF) obtained from BC patients after breast-conserving surgery (BCS) stimulated the motility and invasiveness of tumor cells in vitro. Moreover, we have demonstrated that intraoperative radiation therapy (IORT) significantly inhibits the stimulatory effect of SWF on tumor cells in vitro. This effect may be due to both direct cell killing by ionizing radiation and by modulating the tumor microenvironment. Nowadays exosomes are believed to be the most important molecules responsible for cell-to-cell contact and mediate the changes in tumor microenvironment connected to further tumor progression and therapy resistance. We speculate that exosomes are one of the key components of SWF that distinguish BC cells response to BCS and IORT group of SWF. Thus the main aim of this study was to isolate and analyze the exosomes from BCS and IORT patients.

SWF were isolated from BC patients after BCS and BCS followed by IORT. At least 20 SWF were pooled and exosomes were isolated using ultracentrifugation. The electron microscopy, the vesicle range, and the western blot for exosome-specific markers were determined for their validation. In order to determine the miRNA expression profile in exosomes, SWF from 3 BCS and 3 IORT patients were subjected to high-throughput next-generation sequencing.

We confirmed the presence of exosomes in both SWF from BCS and IORT group. Principal component analysis (PCA) of all differentially expressed miRNAs, revealed that samples from IORT clearly clustered separately from BCS samples. We found 74 miRNA which significantly differentiates both groups of exosomes. Using miRNA databases, we matched miRNA with genes and then with processes and found, that processes which differentiate BCS and IORT exosomes are: Wnt signaling pathway, cell migration, TGFβ receptor signaling pathway, cell-cell adhesion, and cell cycle arrest.

This is the first report documenting the miRNA cargo of exosomes in surgical wound fluids in BCS and IORT patients. This comprehensive approach which allows for a deeper and broader analysis of miRNA cargo of EXO-SWF enables a better understanding of their molecular and functional significance. Moreover, the presented results may be a starting point for modifying the treatment scheme by targeting specific molecules and signaling pathways.