Herein we demonstrate that exposure of human breast tumor cells to several structurally unrelated selective antagonists of 5-HT5A reduced BTIC frequency and that this effect was phenocopied by a CRISPR-Cas9-mediated knockout of HTR5A. We used a phosphoproteomic approach to establish that exposure of human breast tumor cells to SB-699551 disrupts signaling via the Gαi/o-coupled pathway and the PI3K/AKT/mTOR axis, consistent with antagonism of 5-HT5A. Here, AKT1 is linked to breast neoplasm.