Homosalate boosts the release of tumor-derived Extracellular Vesicles with anti-anoikis properties

Abstract

Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular vesicles (EVs). EVs could have different subcellular origin, composition and functional properties, but tools to distinguish between EV subtypes are scarce. Here, we tagged CD63- or CD9-positive EVs secreted by triple negative breast cancer cells with Nanoluciferase enzyme, to set-up a miniaturized method to quantify secretion of these two EV subtypes directly in the supernatant of cells. We performed a cell-based high-content screening to identify clinically-approved drugs able to affect EV secretion. One of the identified hits is Homosalate, an anti-inflammatory drug found in sunscreens which robustly increased EVs release. Comparing EVs induced by Homosalate with those induced by Bafilomycin A1, we discovered that: 1) the two drugs act on EVs generated in distinct subcellular compartments and 2) EVs released upon treatment with Homosalate, but not with Bafilomycin A1, conferred anti-anoikis properties to another recipient tumor cell line. In conclusion, we identified a new drug modifying EV release and demonstrated that under influence of different drugs, triple negative breast cancer cells release EV subpopulations from different subcellular origins harboring distinct functional properties.

Detection of tumor-derived extracellular vesicles interactions with immune cells is dependent on EV-labelling methods

“…Pooled EVs (fractions 7-11), intermediate (fractions [12][13][14][15][16] and soluble (fractions 17-21) SEC fractions were compared by Western Blot following MISEV2018 guidelines (19). EV fractions from both cell lines contained CD63, CD9, Alix EV markers and were devoid of calreticulin (which was not detected in any fraction), and of 14-3-3, which was present in soluble fractions according to previous publications (8,20). To label EVs with a fluorescent reporter protein (sfGFP-EVs), sfGFP tagged with myristoylation and palmitoylation signals (MyrPalm-sfGFP) was stably expressed in MDA-MB-231 by lentiviral transduction.…”