Abstract PS16-14: Eribulin and paclitaxel differentially affect exosome formation and release from triple negative breast cancer cells

• Published in: Cancer research (Chicago, Ill.) Ill.), 2021-02, Vol.81 (4_Supplement), p.PS16-14-PS16-14
• Main Authors: Pederson, Petra EJ, Liang, Huiyun, Mooberry, Susan L
• Format: Article
• Language: English
• ISSN: 0008-5472; 1538-7445
• DOI: 10.1158/1538-7445.SABCS20-PS16-14

Extracellular vesicles (EVs) containing proteins, DNA, and RNA facilitate cellular crosstalk within and among tissues and are involved in a variety of physiologic and disease processes. Cancer cells secrete EVs that promote multiple aspects of cancer progression, including epithelial-mesenchymal transition (EMT), angiogenesis, metastasis, and immune suppression. EVs are formed by two different mechanisms, budding from the plasma membrane and through the multivesicular endosome pathway, and these subpopulations of EVs are commonly referred to as microvesicles and exosomes respectively. Notably, exosomes range in size from 50-150 nm, while microvesicles can range from 100-1000 nm. Microtubules are important for the vesicular trafficking involved in exosome formation, cargo loading, and release. Therefore, we hypothesized that microtubule targeting drugs would affect the formation and release of exosomes from triple negative breast cancer (TNBC) cells and alter their cargo. We investigated whether eribulin and paclitaxel differently affect exosome formation and release because they have highly divergent effects on microtubules, in that eribulin causes microtubule loss and paclitaxel increases microtubule density. Exosome formation was evaluated by immunofluorescence of the multivesicular endosome/exosome marker, CD63, in multiple TNBC cell lines. Eribulin and paclitaxel significantly altered the cellular localization of CD63 after 4 h. High-content imaging analysis revealed a significant intracellular accumulation of CD63, suggesting that eribulin and paclitaxel could impair exosome release. Small EVs/exosomes were then isolated from TNBC cells treated for 8 h with concentrations of eribulin and paclitaxel that maximally disrupted CD63 without causing significant mitotic accumulation or cell death. Small EVs/exosomes isolated from cell-conditioned media were characterized by electron microscopy, immunoblotting and nanoparticle tracking analysis (NTA). The results of NTA show that while paclitaxel did not change the number of EVs released after an 8 h treatment, eribulin caused a modest, but consistent decrease in the number of EVs released by MDA-MB-231 cells. Immunoblot analysis further revealed a significant reduction in CD63 content of EVs from eribulin, but not paclitaxel-treated MDA-MB-231 cells. Additional fluorescent NTA using CD63 antibody labeling showed that CD63+ EVs were substantially reduced by eribulin but unchanged by paclitaxel. Interestingly,