Exosomes are extracellular vesicles that are present in body fluids and known to play key roles in intercellular signaling communication. Exosomes carry several types of molecules of their tumor of origin, including miRNAs, and therefore present potential as biomarkers for cancer prognosis and treatment. Triple-negative breast cancer (TNBC) is a clinically aggressive subtype of breast cancer, associated with treatment resistance, recurrence and high mortality rates. These tumors are molecularly heterogeneous, which impairs the identification of effective prognostic molecular markers and target therapies. The main objective of this study was to characterize the miRNA expression profile of exosomes derived from TNBC cells in comparison to normal controls and to the luminal A (LA) breast cancer subtype. The exosomal miRNAs were isolated from 19 serum samples of breast cancer patients (10 with the LA and 9 with the TNBC subtype) and 10 serum samples of healthy women (control group) using the exosome precipitation method. Exosome characterization and quantification was assessed by Western blot using the exosomal surface markers, CD6 and CD9, and Nanotracking analysis. The tumor and normal exosomal samples were combined into 6 different pools (of at least 4 samples in each group) and sequenced in the Ion Torrent Proton platform (Applied Biosystems). RNA-seq analysis revealed differentially expressed (DE) levels of exosomal miRNAs, with log2FC ranging from 1.2 to 3.6, among the four groups studied (control, cancer (TNBC+LA), TNBC and LA groups): 22 DE miRNAs were observed in the cancer vs. control, 19 in the LA vs control, 30 in the TNBC vs. control and 7 in the LA vs. TNBC groups. Eleven miRNAs were commonly DE in the cancer, LA and TNBC groups when compared to the control group, which included miRNAs, such as miR-103a-3p, miR-107 and miR-423-5p, involved in the biosynthesis of unsaturated fatty acids and neutrophin signaling pathways. Nine miRNAs were only observed DE in the TNBC group (vs. control), with the majority of them being downregulated. The top 10 significant functional enriched pathways affected by these miRNAs included proteoglycans, pathways in cancer and TGF-beta signaling pathways. Only three miRNAs were observed DE in the TNBC vs. LA groups, all with downregulated expression in the TNBC group. These miRNAs were observed mostly involved in fatty acid biosynthesis, proteoglycans in cancer and TGF-beta signaling pathways. In conclusion, our results indicate that expression alterations of exosomal miRNAs affect critical cancer-associated signaling pathways, suggesting their potential use as noninvasive cancer biomarkers. Additional studies should be performed to determine whether the expression patterns of these exosomal miRNAs reflect the ones observed in the corresponding patients' TNBC tissue.

Funding: CAPES, PPSUS-Fundação Araucária, Brazil.

Citation Format: Patricia M. Ozawa, Débora S. Lemos, Evelyn Vieira, Ingrid L. Souza, Silvio M. Zanata, Vânia C. Pankievicz, Thalita R. Tuleski, Emanuel M. Souza, Rosiane V. Silva, Pryscilla F. Wowk, Rodrigo C. Almeida, Iglenir J. Cavalli, Danielle M. Ferreira, Luciane R. Cavalli, Enilze M. Ribeiro. Exosomal miRNA expression profiling in triple-negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5392.