Abstract P6-07-02: Evaluating the effects of eribulin and paclitaxel on exosome formation and release from triple negative breast cancer cells
Authors: Petra Elizabeth Jans Pederson, Susan L Mooberry
Journal: Cancer Research (2020)

Abstract Extracellular vesicles (EVs) are small membrane-bound vesicles released from cells that constitute an important mode of intercellular communication. These vesicles contain bioactive molecules, including proteins, DNA, microRNAs, and lipids that facilitate cellular cross-talk within and among tissues, and they 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 multiple mechanisms, including budding from the plasma membrane and through the multivesicular endosome pathway, and the latter are commonly referred to as exosomes. 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. We investigated whether the microtubule targeting drugs, eribulin and paclitaxel, differently affect exosome formation and release because they have highly divergent effects in cells, in that eribulin causes loss of microtubules and paclitaxel increases microtubule density. Furthermore, eribulin, but not paclitaxel, has been shown to reverse EMT1 and decrease TGF-β in breast cancer patients’ serum.2 TGF-β is a known exosome-associated protein capable of promoting EMT, and therefore, we hypothesize that altered exosome production could contribute to these clinical observations. The effects of eribulin and paclitaxel on the localization of multivesicular endosomes in HCC1937 triple negative breast cancer (TNBC) cells was evaluated by immunofluorescence and cellular CD63 quantified using high content imaging. The results show that a 4-hour treatment of TNBC cells with eribulin caused a rapid and a statistically significant accumulation of the exosome and multivesicular endosome associated protein CD63. Interestingly, paclitaxel had a different effect on CD63 localization in that in some cells there was an accumulation of CD63 in the cell periphery, but in the population, there was no significant difference in CD63 intensity. These results show that eribulin rapidly alters the localization of multivesicular endosomes, which could indicate impaired exosome release. Exosome-enriched extracellular vesicles were isolated from TNBC cells treated with eribulin and paclitaxel for 8 hours at concentrations which maximally disrupt CD63 without significant mitotic accumulation or cell death. Electron microscopy and immunoblot analyses confirmed the isolation of exosome-enriched extracellular vesicles. Nanoparticle tracking analysis is being used to evaluate the number and size of the exosomes to test whether eribulin and paclitaxel affect their release. Future studies will involve proteomic analysis of exosomes from cells treated with eribulin and paclitaxel to determine whether the drugs differentially affect exosome cargo, such as TGF-β. Understanding the effects of diverse MTAs on exosomes and their cargo has the potential to inform a more targeted use of these drugs in the clinic. These studies were funded by Eisai Inc. 1. Yoshida, T. et al. Eribulin mesilate suppresses experimental metastasis of breast cancer cells by reversing phenotype from epithelial–mesenchymal transition (EMT) to mesenchymal–epithelial transition (MET) states. Br. J. Cancer 110, (2014). 2. Ueda, S. et al. In vivo imaging of eribulin-induced reoxygenation in advanced breast cancer patients: a comparison to bevacizumab. Br. J. Cancer 114, 1212–8 (2016). Citation Format: Petra Elizabeth Jans Pederson, Susan L Mooberry. Evaluating the effects of eribulin and paclitaxel on exosome formation and release from triple negative breast cancer cells [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P6-07-02.