microRNA-mediated intercellular communication in the primary breast tumour microenvironment

Abstract

microRNAs (miRNA) have been highlighted as potential circulating biomarkers and as therapeutic targets for breast cancer. Recent evidence suggests that miRNAs are transported in protective microvesicles called exosomes, which play an important role in intercellular communication. This study aimed to validate the expression of miRNAs in patient breast tumours, and to investigate their secretion by breast cancer cells, identifying a specific profile of miRNAs packaged into exosomes. Two miRNAs, miR-106a and miR-191, were shown to be significantly elevated in patient breast tumours and may represent oncomiRs, while one miRNA, miR-504, was found to be significantly decreased and may represent a tumour suppressor. The relationship between circulating miRNA expression levels and menstrual cycle hormones was investigated, where miRNA expression was found to remain stable, with the exception of one miRNA. miRNAs were found to be secreted by breast cancer cells into conditioned media, where a specific miRNA profile was actively and selectively packaged into exosomes. Approximately 390 miRNAs were detectable in exosomes collected from a range of breast cancer cell lines, with four miRNAs, miR-10b/miR-145/miR-492/miR-498, further validated by RQ-PCR. The ability to enrich exosomes with a particular miRNA, miR-504 (previously identified as a potential tumour suppressor in breast cancer), was achieved by transducing a breast cancer cell line to overexpress the miRNA. The impact of overexpressing miR-504 was investigated in vitro, with no significant alterations observed in cell proliferation or migration. Furthermore, the impact was also investigated in vivo, where miR-504 overexpression was not found to suppress tumour formation. Exosome transfer between cell populations was visualised using confocal microscopy and the impact of transfer on the recipient cells was investigated, where a trend towards increased miR-504 expression was observed. In addition, a significant increase in cell proliferation as a result of the whole exosome was observed, with no significant impact on cell migration identified. The data presented here is a contribution to the ongoing research of exosome-encapsulated miRNAs in the cancer research field. Identification and functional characterisation of exosome-encapsulated miRNAs in cancer may be central to the generation of novel biomarkers and therapeutic strategies for this disease.