<p>Mast cells contribute to the pathology of various diseases, in particular allergic conditions. Therefore, it is essential to develop strategies that efficiently prevent their harmful effects under such circumstances. Here, we sought to evaluate the possibility of cell death induction as a potential means of selectively depleting mast cells. Previous work has suggested that mast cells are sensitive to regimes that target their acidic secretory granules, and the aim of this study was therefore to identify novel anti-mast cell compounds that act via a granule-mediated pathway. To this end, we evaluated trifluoperazine, an antipsychotic drug known to present lysosomotropic properties. We demonstrate that trifluoperazine is cytotoxic for mast cells, whereas multiple other cell types were resistant. Trifluoperazine induced mainly apoptotic cell death in mast cells. Further, our data indicate that trifluoperazine acts on mast cells by inducing secretory granule permeabilization. In support of this, trifluoperazine caused granule deacidification, accompanied by cytosolic acidification as well as translocation of tryptase from the secretory granules into the cytosol. Trifluoperazine-induced cell death and subsequent DNA degradation were profoundly abrogated when granule acidification was inhibited by the V-ATPase inhibitor bafilomycin A1, suggesting that the granule acidity has a key role in the cell death mechanism. Moreover, mast cell death in response to trifluoperazine was largely caspase-independent, whereas serine protease activity was shown to promote apoptosis-like vs. necrosis-like cell death. Overall, these findings introduce trifluoperazine as a novel anti-mast cell agent that induces cell death through granule permeabilization. Trifluoperazine may thus be evaluated for therapeutic intervention to ameliorate mast cell-mediated detrimental effects.</p>